Role of NO in Modulating Neuronal Activity in Superficial Dorsal Horn of Spinal Cord During Exercise Pressor Reflex

American Journal of Physiology. Heart and Circulatory Physiology. Sep, 2002  |  Pubmed ID: 12181131

Static contraction of hindlimb skeletal muscle in cats induces a reflex pressor response. The superficial dorsal horn of the spinal cord is the major site of the first synapse of this reflex. In this study, static contraction of the triceps surae muscle was evoked by electrical stimulation of the tibial nerve for 2 min in anesthetized cats (stimulus parameters: two times motor threshold at 30 Hz, 0.025-ms duration). Ten stimulations were performed and 1-min rest was allowed between stimulations. Muscle contraction caused a maximal increase of 32 +/- 5 mmHg in mean arterial pressure (MAP), which was obtained from the first three contractions. Activated neurons in the superficial dorsal horn were identified by c-Fos protein. Distinct c-Fos expression was present in the L6-S1 level of the superficial dorsal horn ipsilateral to the contracting leg (88 +/- 14 labeled cells per section at L7), whereas only scattered c-Fos expression was observed in the contralateral superficial dorsal horn (9 +/- 2 labeled cells per section, P < 0.05 compared with ipsilateral section). A few c-Fos-labeled cells were found in control animals (12 +/- 5 labeled cells per section, P < 0.05 compared with stimulated cats). Furthermore, double-labeling methods demonstrated that c-Fos protein coexisted with nitric oxide (NO) synthase (NOS) positive staining in the superficial dorsal horn. Finally, an intrathecal injection of an inhibitor of NOS, N-nitro-L-arginine methyl ester (5 mM), resulted in fewer c-Fos-labeled cells (58 +/- 12 labeled cells per section) and a reduced maximal MAP response (20 +/- 3 mmHg, P < 0.05). These results suggest that the exercise pressor reflex induced by static contraction is mediated by activation of neurons in the superficial dorsal horn and that formation of NO in this region is involved in modulating the activated neurons and the pressor response to contraction.

Effect of Nitric Oxide on Release of Glutamate in the Subretrofacial Nucleus (SRF) During the Exercise Pressor Reflex in Cats

Brain Research. Sep, 2002  |  Pubmed ID: 12231244

The subretrofacial nucleus (SRF) has been known to play a crucial role in the expression of the exercise pressor reflex. Previously, we have reported that the release of glutamate (Glu) in the SRF was increased during muscle contraction in anesthetized cats. In this study, static muscle contraction of the triceps surae for 4 min was induced by electrical stimulation of L7 and S1 ventral roots. Endogenous release of Glu and citrulline (Cit) from the SRF was recovered by microdialysis and measured by HPLC. The microdialysis probes were also used to deliver L-arginine and L-NAME to test the effect of nitric oxide (NO) on release of Glu in the SRF and on the cardiovascular responses during muscle contraction. During control, muscle contraction significantly increased mean arterial pressure (MAP) from 98+/-8 to 151+/-9 mmHg, and the extracellular concentration of Glu from 610+/-120 to 1280+/-290 nM. Dialyzing 2 mM L-arginine into the SRF increased basal Cit concentration from 260+/-50 to 760+/-210 nM (P<0.05). During contraction after L-arginine, the increases in MAP and Glu concentration were significantly attenuated (86+/-3-124+/-6 mmHg and 300+/-60-460+/-100 nM, respectively). Dialysis of 0.5 mM L-NAME into the SRF decreased Cit concentration from 340+/-40 to 180+/-20 nM (P<0.05). During contraction after dialyzing L-NAME, the increases in MAP and Glu concentration were significantly potentiated (93+/-6-154+/-9 mmHg and 520+/-80-1290+/-380 nM, respectively). These results suggest that endogenous NO modulates the cardiovascular responses to static muscle contraction by affecting the release of Glu in the SRF.

Nitric Oxide Synthase (NOS) Coexists with Activated Neurons by Skeletal Muscle Contraction in the Brainstem of Cats

Life Sciences. Nov, 2002  |  Pubmed ID: 12377266

Contraction of skeletal muscle evokes increases in arterial blood pressure and heart rate. Some regions of the brainstem have been implicated for expression of the cardiovascular responses to muscle contraction. Previous studies have reported that static muscle contraction induced c-Fos protein in the nucleus of tractus solitarii (NTS), lateral reticular nucleus (LRN), lateral tegmental field (FTL), subretrofacial nucleus (SRF), A1 region and periaqueductal gray (PAG) of the brainstem. Furthermore, neuronal NADPH-diaphorase (NADPH-d), which is considered as a marker of neuronal nitric oxide synthase (nNOS), has been localized in those same regions. In this study, static muscle contraction was induced by electrical stimulation of the L7 and S1 ventral roots in anaesthetized cats. Distribution of c-Fos protein within neurons containing nNOS was evaluated by double labeling methods in order to determine if nNOS containing neurons in the brainstem were activated during muscle contraction. The results indicate that c-Fos protein colocalized with NADPH-d positive staining within the neurons of the SRF and PAG, but not within the NTS neurons. Distinct number of neurons with c-Fos protein was in close proximity to NADPH-d positive staining in the NTS, SRF, and PAG. Coexisting of c-Fos protein and NADPH-d positive staining was not observed in the LRN, FTL and A1 region. These findings demonstrate that nNOS containing neurons were activated by muscle contraction in the selective regions of the brainstem, and nNOS positive staining had close anatomic contacts with the neurons activated by contraction. This result provides neuroanatomic evidence suggesting that nitric oxide modulates the cardiovascular responses to muscle contraction within the NTS, SRF and PAG of the brainstem.

Phylogenetic Relationships of Empetraceae Inferred from Sequences of Chloroplast Gene MatK and Nuclear Ribosomal DNA ITS Region

Molecular Phylogenetics and Evolution. Nov, 2002  |  Pubmed ID: 12414312

We used sequences of nrDNA ITS and chloroplast gene matK to evaluate the monophyly of Empetrum and Corema and to examine phylogenetic relationships of the Empetraceae. Sequences of these two DNA markers were obtained for 11 plant samples, representing species of Empetrum from both the Southern and Northern Hemispheres, species and subspecies of Corema, and the monotypic Ceratiola. Sequences of four species of Rhododendron were used for rooting purposes. Our results show that species of Empetrum form a clade sister to the clade containing both Corema and Ceratiola. These two clades are strongly supported in both the matK and ITS trees, suggesting that Ceratiola is more closely related to Corema than to Empetrum, and is not of a hybrid origin between the ancestors of the latter two genera. In the matK tree, Corema conradii is more closely related to Ceratiola than to Corema album and C. album subsp. azoricum, whereas in the ITS tree, Ceratiola is allied with Corema album and C. album subsp. azoricum. This suggests that C. conradii might be a hybrid between ancestral populations of Ceratiola and C. album. The monophyly of Empetrum rejects the hypothesis of its independent origin in the two Hemispheres. Our trees also suggest the fact that the modern amphitropical distribution of Empetrum is the result of long distance dispersal, not of the vicarious events.

ATP Stimulates Chemically Sensitive and Sensitizes Mechanically Sensitive Afferents

American Journal of Physiology. Heart and Circulatory Physiology. Dec, 2002  |  Pubmed ID: 12427602

We examined whether ATP stimulation of P2X purinoceptors would raise blood pressure in decerebrate cats. Femoral arterial injection of the P2X receptor agonist alpha,beta-methylene ATP into the blood supply of the triceps surae muscle induced a dose-dependent increase in arterial blood pressure. The maximal increase in mean arterial pressure (MAP) evoked by 0.1, 0.2, and 0.5 mM alpha,beta-methylene ATP (0.5 ml/min injection rate) was 6.2 +/- 2.5, 22.5 +/- 4.4, and 35.2 +/- 3.9 mmHg, respectively. The P2X receptor antagonist pyridoxal phosphate-6-azophenyl-2',4'-disulfonic acid (2 mM ia) attenuated the increase in MAP elicited by intra-arterial alpha,beta-methylene ATP (0.5 mM), whereas the P2Y receptor antagonist reactive blue 2 (2 mM ia) did not affect the MAP response to alpha,beta-methylene ATP. In a second group of experiments, we tested the hypothesis that ATP acting through P2X receptors would sensitize muscle afferents and, thereby, augment the blood pressure response to muscle stretch. Two kilograms of muscle stretch evoked a 26.5 +/- 4.3 mmHg increase in MAP. This MAP response was enhanced when 2 mM ATP or 0.1 mM alpha,beta-methylene ATP (0.5 ml/min) was arterially infused 10 min before muscle stretch. Furthermore, this effect of ATP on the pressor response to stretch was attenuated by 2 mM pyridoxal phosphate-6-azophenyl-2',4'-disulfonic acid (P < 0.05) but not by the P1 purinoceptor antagonist 8-(p-sulfophenyl)-theophylline (2 mM). These data indicate that activation of ATP-sensitive P2X receptors evokes a skeletal muscle afferent-mediated pressor response and that ATP at relatively low doses enhances the muscle pressor response to stretch via engagement of P2X receptors.

Community-based Trial to Prevent Drug Use Among Youths in Yunnan, China

American Journal of Public Health. Dec, 2002  |  Pubmed ID: 12453815

This study evaluated a community-based program in China to prevent initiation of drug use in young men.

Theoretical, Empirical and Practical Approaches to Resolving the Unmet Information Needs of Clinical Information System Users

Proceedings / AMIA ... Annual Symposium. AMIA Symposium. 2002  |  Pubmed ID: 12463809

We hypothesize that when clinicians review clinical data in an electronic medical record, the information needs that arise are predictable, based on a number of situational factors. Because our theory does not say, exactly, what those needs are, we are using an empirical approach (observation) to detecting and categorizing them. For each need, we can construct an "infobutton" that links the clinical data to an on-line information resource. We have constructed an Infobutton Manager to match the data being reviewed by clinicians with context-appropriate infobuttons. This paper describes how the theory, observations, and practical solutions can come together to improve clinician decision making by resolving information needs.

Interventional and Synthetic Therapy of Advanced Hepatocellular Carcinoma

Chinese Medical Journal. Dec, 2002  |  Pubmed ID: 12622945

To evaluate the clinical efficacy of interventional therapy in the treatment of hepatocellular carcinoma (HCC).

IFN-gamma Induces High Mobility Group Box 1 Protein Release Partly Through a TNF-dependent Mechanism

Journal of Immunology (Baltimore, Md. : 1950). Apr, 2003  |  Pubmed ID: 12646658

We recently discovered that a ubiquitous protein, high mobility group box 1 protein (HMGB1), is released by activated macrophages, and functions as a late mediator of lethal systemic inflammation. To elucidate mechanisms underlying the regulation of HMGB1 release, we examined the roles of other cytokines in induction of HMGB1 release in macrophage cell cultures. Macrophage migration inhibitory factor, macrophage-inflammatory protein 1beta, and IL-6 each failed to significantly induce the release of HMGB1 even at supraphysiological levels (up to 200 ng/ml). IFN-gamma, an immunoregulatory cytokine known to mediate the innate immune response, dose-dependently induced the release of HMGB1, TNF, and NO, but not other cytokines such as IL-1alpha, IL-1beta, or IL-6. Pharmacological suppression of TNF activity with neutralizing Abs, or genetic disruption of TNF expression (TNF knockout) partially (50-60%) inhibited IFN-gamma-mediated HMGB1 release. AG490, a specific inhibitor for Janus kinase 2 of the IFN-gamma signaling pathway, dose-dependently attenuated IFN-gamma-induced HMGB1 release. These data suggest that IFN-gamma plays an important role in the regulation of HMGB1 release through a TNF- and Janus kinase 2-dependent mechanism.

Glutamate Release in Midbrain Periaqueductal Gray by Activation of Skeletal Muscle Receptors and Arterial Baroreceptors

American Journal of Physiology. Heart and Circulatory Physiology. Jul, 2003  |  Pubmed ID: 12649075

We have previously reported that both skeletal muscle receptor and arterial baroreceptor afferent inputs activate neurons in the dorsolateral (DL) and lateral regions of the midbrain periaqueductal gray (PAG). In this study, we determined whether the excitatory amino acid glutamate (Glu) is released to mediate the increased activity in these regions. Static contraction of the triceps surae muscle for 4 min was evoked by electrical stimulation of the L7 and S1 ventral roots in cats. Activation of arterial baroreceptor was induced by intravenous injection of phenylephrine. The endogenous release of Glu from the PAG was recovered with the use of a microdialysis probe. Glu concentration was measured by the HPLC method. Muscle contraction increased mean arterial pressure (MAP) from 98 +/- 10 to 149 +/- 12 mmHg (P < 0.05) and increased Glu release in the DL and lateral regions of the middle PAG from 0.39 +/- 0.10 to 0.73 +/- 0.12 microM (87%, P < 0.05) in intact cats. After sinoaortic denervation and vagotomy were performed, contraction increased MAP from 95 +/- 12 to 158 +/- 15 mmHg, and Glu from 0.34 +/- 0.08 to 0.54 +/- 0.10 microM (59%, P < 0.05). The increases in arterial pressure and Glu were abolished by muscle paralysis. Phenylephrine increased MAP from 100 +/- 13 to 162 +/- 22 mmHg and increased Glu from 0.36 +/- 0.10 to 0.59 +/- 0.18 microM (64%, P < 0.05) in intact animals. Denervation abolished this Glu increase. Summation of the changes in Glu evoked by muscle receptor and arterial baroreceptor afferent inputs was greater than the increase in Glu produced when both reflexes were activated simultaneously in intact state (123% vs. 87%). These data demonstrate that activation of skeletal muscle receptors evokes release of Glu in the DL and lateral regions of the middle PAG, and convergence of afferent inputs from muscle receptors and arterial baroreceptors in these regions inhibits the release of Glu. These results suggest that the PAG is a neural integrating site for the interaction between the exercise pressor reflex and the arterial baroreceptor reflex.

ATP Concentrations and Muscle Tension Increase Linearly with Muscle Contraction

Journal of Applied Physiology (Bethesda, Md. : 1985). Aug, 2003  |  Pubmed ID: 12716867

Previous studies have suggested that activation of ATP-sensitive P2X receptors in skeletal muscle play a role in mediating the exercise pressor reflex (Li J and Sinoway LI. Am J Physiol Heart Circ Physiol 283: H2636-H2643, 2002). To determine the role ATP plays in this reflex, it is necessary to examine whether muscle interstitial ATP (ATPi) concentrations rise with muscle contraction. Accordingly, in this study, muscle contraction was evoked by electrical stimulation of the L7 and S1 ventral roots of the spinal cord in 12 decerebrate cats. Muscle ATPi was collected from microdialysis probes inserted in the muscle. ATP concentrations were determined by the HPLC method. Electrical stimulation of the ventral roots at 3 and 5 Hz increased mean arterial pressure by 13 +/- 2 and 16 +/- 3 mmHg (P < 0.05), respectively, and it increased ATP concentration in contracting muscle by 150% (P < 0.05) and 200% (P < 0.05), respectively. ATP measured in the opposite control limb did not rise with ventral root stimulation. Section of the L7 and S1 dorsal roots did not affect the ATPi seen with 5-Hz ventral root stimulation. Finally, ventral roots stimulation sufficient to drive motor nerve fibers did not increase ATP in previously paralyzed cats. Thus ATPi is not largely released from sympathetic or motor nerves and does not require an intact afferent reflex pathway. We conclude that ATPi is due to the release of ATP from contracting skeletal muscle cells.

Structural Basis for the Proinflammatory Cytokine Activity of High Mobility Group Box 1

Molecular Medicine (Cambridge, Mass.). Jan-Feb, 2003  |  Pubmed ID: 12765338

High mobility group box 1 (HMGB), a ubiquitous DNA-binding protein, has been implicated as a proinflammatory cytokine and late mediator of lethal endotoxemia. HMGB1 is released by activated macrophages. It amplifies and extends the inflammatory response by inducing cytokine release and mediating acute lung injury, anorexia, and the inflammatory response to tissue necrosis. The kinetics of HMGB1 release provide a wide therapeutic window for endotoxemia because extracellular levels of HMGB1 begin to increase 12 to 24 h after exposure to inflammatory stimuli. Here, we demonstrate that a DNA-binding domain of HMGB1, the B box, recapitulates the cytokine activity of full length HMGB1 and efficiently activates macrophages to release tumor necrosis factor (TNF) and other proinflammatory cytokines. Truncation of the B box revealed that the TNF-stimulating activity localizes to 20 amino acids (HMGB1 amino acids 89 to 108). Passive immunization of mice with antibodies raised against B box conferred significant protection against lethal endotoxemia or sepsis, induced by cecal perforation. These results indicate that a proinflammatory domain of HMGB1 maps to the highly conserved DNA-binding B box, making this primary sequence a suitable target in the design of therapeutics.

Transaxillary Minithoracotomy in Intrathoracic Surgery for 316 Infants and Children

Chinese Medical Journal. Jul, 2003  |  Pubmed ID: 12890373

To introduce the technique of intrathoracic surgery performed through vertical transaxillary minithoracotomy.

Use of Online Resources While Using a Clinical Information System

AMIA ... Annual Symposium Proceedings / AMIA Symposium. AMIA Symposium. 2003  |  Pubmed ID: 14728157

Clinical information system (CIS) use is likely to evoke information needs, yet information resources use during CIS use has not been studied.

Sharing Infobuttons to Resolve Clinicians' Information Needs

AMIA ... Annual Symposium Proceedings / AMIA Symposium. AMIA Symposium. 2003  |  Pubmed ID: 14728320

Attempts to link clinical information systems to on-line information resources date back over a decade. The World Wide Web presents new opportunities to create such links, which we refer to as "infobuttons". This capability is partly due to the ease with which a link (called a Uniform Resource Locator, or URL) in one Web-based system can take users to another Web-based system, and partly due to the proliferation of high-quality Web-based resources. Typically, contextual information from the clinical system (such as patient data) is passed to a search engine that is evoked and, in turn, presents search results to the user. Impediments to infobutton development include customized programming to pass context information to the information resource, lack of standards for types and values of context information, variability of search engine interfaces, and a lack of understanding of clinicians' information needs. We are addressing these limitations with an Infobutton Manager (IM) that provides a standardized interface for matching user contexts to information resources.

Epidemiology of Nosocomial Pneumonia in Infants After Cardiac Surgery

Chest. Feb, 2004  |  Pubmed ID: 14769717

The pattern of nosocomial pneumonia (NP) in infants in a pediatric surgical ICU after cardiac surgery may differ from that seen in adult ICUs.

Central Integration of Muscle Reflex and Arterial Baroreflex in Midbrain Periaqueductal Gray: Roles of GABA and NO

American Journal of Physiology. Heart and Circulatory Physiology. Sep, 2004  |  Pubmed ID: 15087292

It has been suggested that the midbrain periaqueductal gray (PAG) is a neural integrating site for the interaction between the muscle pressor reflex and the arterial baroreceptor reflex. The underlying mechanisms are poorly understood. The purpose of this study was to examine the roles of GABA and nitric oxide (NO) in modulating the PAG integration of both reflexes. To activate muscle afferents, static contraction of the triceps surae muscle was evoked by electrical stimulation of the L7 and S1 ventral roots of 18 anesthetized cats. In the first group of experiments (n = 6), the pressor response to muscle contraction was attenuated by bilateral microinjection of muscimol (a GABA receptor agonist) into the lateral PAG [change in mean arterial pressure (DeltaMAP) = 24 +/- 5 vs. 46 +/- 8 mmHg in control]. Conversely, the pressor response was significantly augmented by 0.1 mM bicuculline, a GABAA receptor antagonist (DeltaMAP = 65 +/- 10 mmHg). In addition, the effect of GABAA receptor blockade on the reflex response was significantly blunted after sinoaortic denervation and vagotomy (n = 4). In the second group of experiments (n = 8), the pressor response to contraction was significantly attenuated by microinjection of L-arginine into the lateral PAG (DeltaMAP = 26 +/- 4 mmHg after L-arginine injection vs. 45 +/- 7 mmHg in control). The effect of NO attenuation was antagonized by bicuculline and was reduced after denervation. These data demonstrate that GABA and NO within the PAG modulate the pressor response to muscle contraction and that NO attenuation of the muscle pressor reflex is mediated via arterial baroreflex-engaged GABA increase. The results suggest that the PAG plays an important role in modulating cardiovascular responses when muscle afferents are activated.

High Mobility Group Box Protein 1: an Endogenous Signal for Dendritic Cell Maturation and Th1 Polarization

Journal of Immunology (Baltimore, Md. : 1950). Jul, 2004  |  Pubmed ID: 15210788

High mobility group box protein 1 (HMGB1), a DNA binding nuclear and cytosolic protein, is a proinflammatory cytokine released by monocytes and macrophages. This study addressed the hypothesis that HMGB1 is an immunostimulatory signal that induces dendritic cell (DC) maturation. We show that HMGB1, via its B box domain, induced phenotypic maturation of DCs, as evidenced by increased CD83, CD54, CD80, CD40, CD58, and MHC class II expression and decreased CD206 expression. The B box caused increased secretion of the proinflammatory cytokines IL-12, IL-6, IL-1alpha, IL-8, TNF-alpha, and RANTES. B box up-regulated CD83 expression as well as IL-6 secretion via a p38 MAPK-dependent pathway. In the MLR, B box-activated DCs acted as potent stimulators of allogeneic T cells, and the magnitude of the response was equivalent to DCs activated by exposure to LPS, nonmethylated CpG oligonucleotides, or CD40L. Furthermore, B box induced secretion of IL-12 from DCs as well as IL-2 and IFN-gamma secretion from allogeneic T cells, suggesting a Th1 bias. HMGB1 released by necrotic cells may be a signal of tissue or cellular injury that, when sensed by DCs, induces and/or enhances an immune reaction.

Muscle Pressor Reflex: Potential Role of Vanilloid Type 1 Receptor and Acid-sensing Ion Channel

Journal of Applied Physiology (Bethesda, Md. : 1985). Nov, 2004  |  Pubmed ID: 15220301

Reflex cardiovascular responses to muscle contraction are mediated by mechanical and metabolic stimulation of thin muscle afferent fibers. Metabolic stimulants and receptors involved in responses are uncertain. Capsaicin depolarizes thin sensory afferent nerves that have vanilloid type 1 receptors (VR1). Among potential endogenous ligands of thin fibers, H+ has been suggested as a metabolite mediating the reflex muscle response as well as a potential stimulant of VR1. It has also been suggested that acid-sensing ion channels (ASIC) mediate H+, evoking afferent nerve excitation. We have examined the roles of VR1 and ASIC in mediating cardiovascular reflex responses to acid stimulation of muscle afferents in a rat model. In anesthetized rats, injections of capsaicin into the arterial blood supply of triceps surae muscles evoked a biphasic response (n = 6). An initial fall in mean arterial pressure (from baseline of 95.8 +/- 9.5 to 70.4 +/- 4.5 mmHg, P < 0.05 vs. baseline) was followed by an increase (to 131.6 +/- 11.3 mmHg, P < 0.05 vs. baseline). Anandamide (an endogenous substance that activates VR1) induced the same change in blood pressure as did capsaicin. The pressor (but not depressor) component of the response was blocked by capsazepine (a VR1 antagonist) and section of afferent nerves. In decerebrate rats (n = 8), H+ evoked a pressor response that was not blocked by capsazepine but was attenuated by amiloride (an ASIC blocker). In rats (n = 12) pretreated with resiniferatoxin to destroy muscle afferents containing VR1, capsaicin and H+ responses were blunted. We conclude that H+ stimulates ASIC, evoking the reflex response, and that ASIC are likely to be frequently found on afferents containing VR1. The data also suggest that VR1 and ASIC may play a role in processing of muscle afferent signals, evoking the muscle pressor reflex.

Recombinant HMGB1 with Cytokine-stimulating Activity

Journal of Immunological Methods. Jun, 2004  |  Pubmed ID: 15251426

We describe methods for the isolation, purification, and characterization of full-length high-mobility group box 1 (HMGB1) and truncated mutants expressed in bacteria and in mammalian Chinese Hamster Ovary (CHO) cells. HMGB1 is an abundant nuclear and cytoplasmic protein, highly conserved across species and widely distributed in eukaryotic cells from yeast to man. As a ubiquitous nuclear DNA binding protein, HMGB1 binds DNA, facilitates gene transcription, and stabilizes nucleosome structure. In addition to these intracellular roles, HMGB1 can be released into the extracellular milieu by activated innate immune cells (i.e., macrophages, monocytes) and functions as a mediator of lethal endotoxemia and sepsis. The proinflammatory cytokine activity of HMGB1 has become an intense area of research and recombinant protein can be a useful tool to probe HMGB1 functions. Due to its dipolar charged properties, HMGB1 isolated by some methods can be contaminated with bacterial products (such as CpG DNA or lipopolysaccharide [LPS]) that may interfere with immunological analyses. Here we report our newly developed methods for the isolation and purification of biologically active HMGB1 from bacteria or mammalian CHO cells that is essentially free of contaminants. This strategy provides an important advance in methodology to facilitate future HMGB1 studies.

Pathogenic Role of HMGB1 in SARS?

Medical Hypotheses. 2004  |  Pubmed ID: 15325019

High mobility group box 1 protein (HMGB1) is released by necrotic cells or activated macrophages/monocytes, and functions as a late mediator of lethal systemic and local pulmonary inflammation. Passive immunization with anti-HMGB1 antibodies confers significant protection against lethal endotoxemia, sepsis, and acute lung injury, even when antibodies are administered after the onset of these diseases. In light of observations that three Chinese herbal formulations recommended for treatment of severe acute respiratory syndrome (SARS) specifically inhibited the release of HMGB1 from innate immune cells, we hypothesize that HMGB1 might occupy a pathogenic role in SARS by mediating an injurious pulmonary inflammatory response.

Bacterial Endotoxin Stimulates Macrophages to Release HMGB1 Partly Through CD14- and TNF-dependent Mechanisms

Journal of Leukocyte Biology. Nov, 2004  |  Pubmed ID: 15331624

Bacterial endotoxin [lipopolysaccharide (LPS)] stimulates macrophages to sequentially release early [tumor necrosis factor (TNF)] and late [high mobility group box 1 (HMGB1)] proinflammatory cytokines. The requirement of CD14 and mitogen-activated protein kinases [MAPK; e.g., p38 and extracellular signal-regulated kinase (ERK)1/2] for endotoxin-induced TNF production has been demonstrated previously, but little is known about their involvement in endotoxin-mediated HMGB1 release. Here, we demonstrated that genetic disruption of CD14 expression abrogated LPS-induced TNF production but only partially attenuated LPS-induced HMGB1 release in cultures of primary murine peritoneal macrophages. Pharmacological suppression of p38 or ERK1/2 MAPK with specific inhibitors (SB203580, SB202190, U0126, or PD98059) significantly attenuated LPS-induced TNF production but failed to inhibit LPS-induced HMGB1 release. Consistently, an endogenous, immunosuppressive molecule, spermine, failed to inhibit LPS-induced activation of p38 MAPK and yet, still significantly attenuated LPS-mediated HMGB1 release. Direct suppression of TNF activity with neutralizing antibodies or genetic disruption of TNF expression partially attenuated HMGB1 release from macrophages induced by LPS at lower concentrations (e.g., 10 ng/ml). Taken together, these data suggest that LPS stimulates macrophages to release HMGB1 partly through CD14- and TNF-dependent mechanisms.

Practical Considerations for Exploiting the World Wide Web to Create Infobuttons

Studies in Health Technology and Informatics. 2004  |  Pubmed ID: 15360818

We are studying ways to provide automated, context-specific links (called "infobuttons") between clinical information systems (CIS) and other information resources available on the World Wide Web. As part of this work, we observed the information needs that arose when clinicians used a CIS and we classified those needs into generic questions. We then sought general methods for accessing information resources to answer the questions.

Muscle Mechanoreflex and Metaboreflex Responses After Myocardial Infarction in Rats

Circulation. Nov, 2004  |  Pubmed ID: 15520319

During exercise, the sympathetic nervous system is activated and blood pressure and heart rate increase. In heart failure (HF), the muscle metaboreceptor contribution to sympathetic outflow is attenuated and the mechanoreceptor contribution is accentuated. Previous studies suggest that (1) capsaicin stimulates muscle metabosensitive vanilloid receptor subtype 1 (VR1), inducing a neurally mediated pressor response, and (2) activation of ATP-sensitive P2X receptors enhances the pressor response seen when muscle mechanoreceptors are engaged by muscle stretch. Thus, we hypothesized that the pressor response to VR1 stimulation would be smaller and the sensitizing effects of P2X stimulation greater in rats with HF due to chronic myocardial infarction (MI) than in controls.

Reversing Established Sepsis with Antagonists of Endogenous High-mobility Group Box 1

Proceedings of the National Academy of Sciences of the United States of America. Jan, 2004  |  Pubmed ID: 14695889

Despite significant advances in intensive care therapy and antibiotics, severe sepsis accounts for 9% of all deaths in the United States annually. The pathological sequelae of sepsis are characterized by a systemic inflammatory response, but experimental therapeutics that target specific early inflammatory mediators [tumor necrosis factor (TNF) and IL-1beta] have not proven efficacious in the clinic. We recently identified high mobility group box 1 (HMGB1) as a late mediator of endotoxin-induced lethality that exhibits significantly delayed kinetics relative to TNF and IL-1beta. Here, we report that serum HMGB1 levels are increased significantly in a standardized model of murine sepsis, beginning 18 h after surgical induction of peritonitis. Specific inhibition of HMGB1 activity [with either anti-HMGB1 antibody (600 microg per mouse) or the DNA-binding A box (600 microg per mouse)] beginning as late as 24 h after surgical induction of peritonitis significantly increased survival (nonimmune IgG-treated controls = 28% vs. anti-HMGB1 antibody group = 72%, P < 0.03; GST control protein = 28% vs. A box = 68%, P < 0.03). Animals treated with either HMGB1 antagonist were protected against the development of organ injury, as evidenced by improved levels of serum creatinine and blood urea nitrogen. These observations demonstrate that specific inhibition of endogenous HMGB1 therapeutically reverses lethality of established sepsis indicating that HMGB1 inhibitors can be administered in a clinically relevant time frame.

Low Temperature Bonding of Poly(methylmethacrylate) Electrophoresis Microchips by in Situ Polymerisation

Journal of Chromatography. A. Nov, 2005  |  Pubmed ID: 16257300

A novel method for bonding poly(methyl methacrylate) (PMMA) electrophoresis microchips at the temperature below the glass transition temperature of PMMA based on in situ polymerization has been demonstrated. Methyl methacrylate (MMA) containing initiators was allowed to prepolymerize in an 85 degrees C water bath for 8 min and 15 min to produce a bonding solution and a dense molding solution, respectively. The channel plate of the PMMA microchip was fabricated by the UV-initiated polymerization of the molding solution between a nickel template and a PMMA plate at room temperature. Prior to bonding, the blank cover was coated with a thin layer of the bonding solution and was bonded to the channel plate at 95 degrees C for 20 min under the pressure of binder clips. The attractive performance of the PMMA chips bonded by the new approach has been demonstrated by separating and detecting dopamine, catechol, three cations, and three organic acids in connection with end-column amperometric detection and contactless conductivity detection.

Independent Modification of Baroreceptor and Exercise Pressor Reflex Function by Nitric Oxide in Nucleus Tractus Solitarius

American Journal of Physiology. Heart and Circulatory Physiology. May, 2005  |  Pubmed ID: 15604127

It has been suggested that nitric oxide (NO) is a key modulator of both baroreceptor and exercise pressor reflex afferent signals processed within the nucleus tractus solitarius (NTS). However, studies investigating the independent effects of NO within the NTS on the function of each reflex have produced inconsistent results. To address these concerns, the effects of microdialyzing 10 mM L-arginine, an NO precursor, and 20 mM N(G)-nitro-L-arginine methyl ester (L-NAME), an NO synthase inhibitor, into the NTS on baroreceptor and exercise pressor reflex function were examined in 17 anesthetized cats. Arterial baroreflex regulation of heart rate was quantified using vasoactive drugs to induce acute changes in mean arterial pressure (MAP). To activate the exercise pressor reflex, static hindlimb contractions were induced by electrical stimulation of spinal ventral roots. To isolate the exercise pressor reflex, contractions were repeated after barodenervation. The gain coefficient of the arterial cardiac baroreflex was significantly different from control (-0.24 +/- 0.04 beats.min(-1).mmHg(-1)) after the dialysis of L-arginine (-0.18 +/- 0.02 beats.min(-1).mmHg(-1)) and L-NAME (-0.29 +/- 0.02 beats.min(-1).mmHg(-1)). In barodenervated animals, the peak MAP response to activation of the exercise pressor reflex (change in MAP from baseline, 39 +/- 7 mmHg) was significantly attenuated by the dialysis of L-arginine (change in MAP from baseline, 29 +/- 6 mmHg). The results demonstrate that NO within the NTS can independently modulate both the arterial cardiac baroreflex and the exercise pressor reflex. Collectively, these findings provide a neuroanatomical and chemical basis for the regulation of baroreflex and exercise pressor reflex function within the central nervous system.

Spinal P2X Receptor Modulates Reflex Pressor Response to Activation of Muscle Afferents

American Journal of Physiology. Heart and Circulatory Physiology. May, 2005  |  Pubmed ID: 15615840

Static contraction of skeletal muscle evokes increases in blood pressure and heart rate. Previous studies suggested that the dorsal horn of the spinal cord is the first synaptic site responsible for those cardiovascular responses. In this study, we examined the role of ATP-sensitive P2X receptors in the cardiovascular responses to contraction by microdialyzing the P2X receptor antagonist pyridoxal phosphate-6-azophenyl-2',4'-disulfonic acid (PPADS) into the L7 level of the dorsal horn of nine anesthetized cats. Contraction was elicited by electrical stimulation of the L7 and S1 ventral roots. Blockade of P2X receptor attenuated the contraction induced-pressor response [change in mean arterial pressure (delta MAP): 16 +/- 4 mmHg after 10 mM PPADS vs. 42 +/- 8 mmHg in control; P < 0.05]. In addition, the pressor response to muscle stretch was also blunted by PPADS (delta MAP: 27 +/- 5 mmHg after PPADS vs. 49 +/- 8 mmHg in control; P < 0.05). Finally, activation of P2X receptor by microdialyzing 0.5 mM alpha,beta-methylene into the dorsal horn significantly augmented the pressor response to contraction. This effect was antagonized by prior PPADS dialysis. These data demonstrate that blockade of P2X receptors in the dorsal horn attenuates the pressor response to activation of muscle afferents and that stimulation of P2X receptors enhances the reflex response, indicating that P2X receptors play a role in mediating the muscle pressor reflex at the first synaptic site of this reflex.

Suppression of HMGB1 Release by Stearoyl Lysophosphatidylcholine:an Additional Mechanism for Its Therapeutic Effects in Experimental Sepsis

Journal of Lipid Research. Apr, 2005  |  Pubmed ID: 15687351

Stearoyl lysophosphatidylcholine (LPC) has recently been proven protective against lethal sepsis by stimulating neutrophils to eliminate invading pathogens through an H2O2-dependent mechanism. Here, we demonstrate that stearoyl LPC, but not caproyl LPC, significantly attenuates circulating high-mobility group box 1 (HMGB1) levels in endotoxemia and sepsis by suppressing endotoxin-induced HMGB1 release from macrophages/monocytes. Neutralizing antibodies against G2A, a potential cell surface receptor for LPC, partially abrogated stearoyl LPC-mediated suppression of HMGB1 release. Thus, stearoyl LPC confers protection against lethal experimental sepsis partly by facilitating the elimination of the invading pathogens and partly by inhibiting endotoxin-induced release of a late proinflammatory cytokine, HMGB1.

Giardia Lamblia: Stable Expression of Green Fluorescent Protein Mediated by Giardiavirus

Experimental Parasitology. Mar, 2005  |  Pubmed ID: 15713450

Giardia lamblia, an early diverging eukaryote that infects several species including humans and a major agent of water-borne diarrhea throughout the world, can be infected with a double-stranded RNA virus, giardiavirus (GLV). A chimeric GLV cDNA and green fluorescent protein (GFP) according to the cis-acting signals of the GLV genome required for expression of foreign gene was constructed and its in vitro transcript was electroporated into GLV-infected G. lamblia trophozoites, GFP was expressed transiently. pGDH5/NEO/GLV was constructed by combining the neomycin resistance cassette in which the neomycin phosphotransferase gene was flanked by Giardia glutamate dehydrogenase (GDH) uncoding regions and the transcription cassette in which the chimera of GLV cDNA and GFP was located downstream from GDH gene promoter on a single plasmid. This plasmid was electroporated into G. lamblia and the transfectants persistently expressed GFP under G418 selection. This stable transfection system should provide a valuable tool for genetic study of G. lamblia.

The Nuclear Factor HMGB1 Mediates Hepatic Injury After Murine Liver Ischemia-reperfusion

The Journal of Experimental Medicine. Apr, 2005  |  Pubmed ID: 15795240

High-mobility group box 1 (HMGB1) is a nuclear factor that is released extracellularly as a late mediator of lethality in sepsis as well as after necrotic, but not apoptotic, death. Here we demonstrate that in contrast to the delayed role of HMGB1 in the systemic inflammation of sepsis, HMGB1 acts as an early mediator of inflammation and organ damage in hepatic ischemia reperfusion (I/R) injury. HMGB1 levels were increased during liver I/R as early as 1 h after reperfusion and then increased in a time-dependent manner up to 24 h. Inhibition of HMGB1 activity with neutralizing antibody significantly decreased liver damage after I/R, whereas administration of recombinant HMGB1 worsened I/R injury. Treatment with neutralizing antibody was associated with less phosphorylation of c-Jun NH(2)-terminal kinase and higher nuclear factor-kappaB DNA binding in the liver after I/R. Toll-like receptor 4 (TLR4)-defective (C3H/Hej) mice exhibited less damage in the hepatic I/R model than did wild-type (C3H/HeOuj) mice. Anti-HMGB1 antibody failed to provide protection in C3H/Hej mice, but successfully reduced damage in C3H/Ouj mice. Together, these results demonstrate that HMGB1 is an early mediator of injury and inflammation in liver I/R and implicates TLR4 as one of the receptors that is involved in the process.

Development of a New Animal Model of Chronic Mitral Regurgitation in Rats Under Transesophageal Echocardiographic Guidance

Journal of the American Society of Echocardiography : Official Publication of the American Society of Echocardiography. May, 2005  |  Pubmed ID: 15891757

Large animal models (dog and sheep) are often used for the investigation of the pathophysiology of chronic mitral regurgitation (MR). A major limitation of large animal models is cost. The aim of this study was to develop a new animal model of chronic MR. Left thoracotomy was performed in 34 rats. Under the guidance of transesophageal echocardiography, a fine needle was inserted into the left ventricle (LV) to damage the mitral leaflets and produce MR. Serial transthoracic echocardiography was performed to assess LV remodeling and function. Left atrial and LV diameters were significantly larger, and LV fractional shortening was lower in the MR group than in the sham group. The 150-day survival was 59% in the MR group and 100% in the sham group (P < .01). This new animal model of chronic MR may be used in the study of the pathophysiology of chronic MR and pharmacologic therapies.

Interstitial ATP and Norepinephrine Concentrations in Active Muscle

Circulation. May, 2005  |  Pubmed ID: 15911708

Sympathetic nervous system activity increases with exercise in normal subjects. Heightened peripheral sympathetic nervous activity and the resultant increased neurovascular levels of norepinephrine (NE) evoke vasoconstriction and serve to maintain blood pressure and perfusion to vital organs. Previous work demonstrated that the interstitial ATP concentrations ([ATP]i) rise in contracting skeletal muscle, and it is known that sympathetic nerves have purinergic P2X receptors. Thus, in this report we tested the hypothesis that elevated ATP would stimulate these receptors and increase interstitial NE concentrations ([NE]i).

An Initial Application of Transesophageal Doppler Echocardiography in Experimental Small Animal Models

Journal of the American Society of Echocardiography : Official Publication of the American Society of Echocardiography. Jun, 2005  |  Pubmed ID: 15947763

This study examined whether an intracardiac echocardiography catheter could be used for transesophageal echocardiography (TEE) examinations in normal rats, and intraoperative TEE in small animal models of disease. The study used 30 Sprague-Dawley normal rats, 10 rats undergoing coronary artery ligation, and 10 rats with experimentally induced mitral regurgitation. The rats were anesthetized with isoflurane and intubated. An intracardiac echocardiographic catheter was inserted into the esophagus. M-mode, 2-dimensional, and Doppler studies were performed in multiple views. TEE probe insertions were successful in all animals. Intraoperative TEE was safely performed in the rat models of myocardial infarction or mitral regurgitation. Mitral regurgitation was well assessed using color Doppler and pulmonary venous flow. This study demonstrates that TEE (including intraoperative TEE) can be safely performed in rats using an intracardiac echocardiographic catheter. It provides a new approach to the assessment of cardiac function and valvular regurgitation in small animals.

A Perspective on the Muscle Reflex: Implications for Congestive Heart Failure

Journal of Applied Physiology (Bethesda, Md. : 1985). Jul, 2005  |  Pubmed ID: 16036901

In this review we examine the exercise pressor reflex in health and disease. The role of metabolic and mechanical stimulation of thin fiber muscle afferents is discussed. The role ATP and lactic acid play in stimulating and sensitizing these afferents is examined. The role played by purinergic receptors subdivision 2, subtype X, vanilloid receptor subtype 1, and acid-sensing ion channels in mediating the effects of ATP and H+ are discussed. Muscle reflex activation in heart failure is then examined. Data supporting the concept that the metaboreflex is attenuated and that the mechanoreflex is accentuated are presented. The role the muscle mechanoreflex plays in evoking renal vasoconstriction is also described.

The Effects of CpG DNA on HMGB1 Release by Murine Macrophage Cell Lines

Journal of Leukocyte Biology. Oct, 2005  |  Pubmed ID: 16081598

DNA containing cytosine-guanine dinucleotide (CpG) motifs (CpG DNA) has potent immunostimulatory activities that resemble those of lipopolysaccharide (LPS) in its effects on the innate immune system. Among its activities, LPS can induce the release of high mobility group protein (HMGB1) by macrophages, a dual function molecule that can mediate the late effects of LPS. To determine whether CpG DNA can also induce HMGB1 release, the effects of a synthetic CpG oligonucleotide (ODN) on HMGB1 release from RAW 264.7 and J774A.1 cells were assessed by Western blotting of culture supernatants. Under conditions in which the CpG ODN activated the cell lines, as assessed by stimulation of tumor necrosis factor alpha and interleukin-12, it failed to cause HMGB1 release into the media. Although unable to induce HMGB1 release by itself, the CpG ODN nevertheless potentiated the action of LPS. With RAW 264.7 cells, lipoteichoic acid and polyinosinic-polycytidylic acid, like LPS, stimulated HMGB1 release as well as cytokine production. These results indicate that the effects of CpG DNA on macrophages differ from other ligands of Toll-like receptors and may lead to a distinct pattern of immune cell activation in the context of infection or its use as an immunomodulatory agent.

[Determination of Thiophanate-methyl Residue in Peppers by Solid Phase Extraction-HPLC]

Se Pu = Chinese Journal of Chromatography / Zhongguo Hua Xue Hui. May, 2005  |  Pubmed ID: 16124590

Composite Fiber of Poly D,L-lactic Acid/hydroxyapatite Produced by Melt Spinning Technology

Bio-medical Materials and Engineering. 2005  |  Pubmed ID: 16179753

In this paper, preparation technologies for the compound fibers of poly D,L-lactic acid (PDLLA)/hydroxyapatite (HA) were investigated. Starting with PDLLA of weight average molecular mass 94,200-1,130,000 and HA powders of diameter 4-10 microm, the compound fibers of PDLLA/HA were obtained through a two stage process: first the adsorption of HA particles on the surface of PDLLA flakes using the liquid-phase adsorption method then melt-extrusion, and second the spinning collection. Experimental result was showed that the high performance composite fibers of PDLLA/HA with diameter of 15-30 micrometer could be produced by the PDLLA of weight average molecular mass 150,000-300,000, the HA powders content 5 wt%, the melt extrusion temperature below 160 degrees C and the screw rotating speed of 10-15 r/min, the spinning collection speed of 2-5 m/min.

Analysis of Polarity Information in Medical Text

AMIA ... Annual Symposium Proceedings / AMIA Symposium. AMIA Symposium. 2005  |  Pubmed ID: 16779104

Knowing the polarity of clinical outcomes is important in answering questions posed by clinicians in patient treatment. We treat analysis of this information as a classification problem. Natural language processing and machine learning techniques are applied to detect four possibilities in medical text: no outcome, positive outcome, negative outcome, and neutral outcome. A supervised learning method is used to perform the classification at the sentence level. Five feature sets are constructed: unigrams, bigrams, change phrases, negations, and categories. The performance of different combinations of feature sets is compared. The results show that generalization using the category information in the domain knowledge base Unified Medical Language System is effective in the task. The effect of context information is significant. Combining linguistic features and domain knowledge leads to the highest accuracy.

Sympathetic Responses to Exercise in Myocardial Infarction Rats: a Role of Central Command

American Journal of Physiology. Heart and Circulatory Physiology. Dec, 2006  |  Pubmed ID: 16844916

In congestive heart failure (CHF), exaggerated sympathetic activation is observed during exercise, which elicits excess peripheral vasoconstriction. The mechanisms causing this abnormality are not fully understood. Central command is a central neural process that induces parallel activation of motor and cardiovascular systems. This study was undertaken to determine whether central command serves as a mechanism that contributes to the exaggerated sympathetic response to exercise in CHF. In decerebrated rats, renal and lumbar sympathetic nerve responses (RSNA and LSNA, respectively) to 30 s of fictive locomotion were examined. The fictive locomotion was induced by electrical stimulation of the mesencephalic locomotor region (MLR). The study was performed in control animals (fractional shortening > 40%) and animals with myocardial infarctions (MI; fractional shortening < 30%). With low stimulation of the MLR (current intensity = 20 microA), the sympathetic responses were not significantly different in the control (RSNA: +18 +/- 4%; LSNA: +3 +/- 2%) and MI rats (RSNA: +16 +/- 5%; LSNA: +8 +/- 3%). With intense stimulation of the MLR (50 microA), the responses were significantly greater in MI rats (RSNA: +127 +/- 15%; LSNA: +57 +/- 10%) than in the control rats (RSNA: +62 +/- 5%; LSNA: +21 +/- 6%). In this study, the data demonstrate that RSNA and LSNA responses to intense stimulation of the MLR are exaggerated in MI rats. We suggest that intense activation of central command may play a role in evoking exaggerated sympathetic activation and inducing excessive peripheral vasoconstriction during exercise in CHF.

Genome-wide Linkage Analysis of Heroin Dependence in Han Chinese: Results from Wave One of a Multi-stage Study

American Journal of Medical Genetics. Part B, Neuropsychiatric Genetics : the Official Publication of the International Society of Psychiatric Genetics. Sep, 2006  |  Pubmed ID: 16856125

The contribution of genes to the etiology of heroin dependence is greater than for any other illicit drug. The specific genes mediating this effect remain unknown, despite several candidate gene association studies of the condition. Here we report the results of a genome-wide search for heroin dependence susceptibility loci using multipoint linkage analysis. In phase I, we ascertained 207 independent affected sibling pairs from 202 Han Chinese families from Yunnan Province, China (near Asia's "Golden Triangle"). After data-cleaning, 194 fully independent sibling pairs (i.e., with no overlapping individuals) from 192 families were genotyped on 404 short tandem-repeat markers spaced at an average inter-marker distance of 9 cM. Although none of our findings achieved genome-wide significance, we found two regions with non-parametric linkage (NPL) Z-scores greater than 2.0. An NPL Z-score of 2.19 (uncorrected P-value = 0.014) was observed at D4S1644, located at 143.3 cM on chromosomal region 4q31.21. The highest NPL Z-score of 2.36 (uncorrected P-value = 0.009) was observed at 53.4 cM on chromosomal region 17q11.2 at marker D17S1880. This is among the first published reports of a genome-wide linkage analysis of heroin dependence. Forthcoming results from other groups and from two additional waves of ascertainment (one planned, one currently ongoing) for our own study should be able to support or refute the putative susceptibility loci we have identified, after which positional candidate genes can be further evaluated as risk factors for the illness.

Dendritic Cell Activating Peptides Induce Distinct Cytokine Profiles

International Immunology. Nov, 2006  |  Pubmed ID: 16966494

High-mobility group box 1 protein (HMGB1), a DNA-binding nuclear and cytosolic protein, is a pro-inflammatory cytokine released by monocytes and macrophages. HMGB1 as well as its B box domain induce maturation of human dendritic cells (DCs). This report demonstrates that the B box domain induces phenotypic maturation of murine bone marrow-derived dendritic cells (BM-DCs) as evidenced by increased CD86, CD40 and MHC-II expression. The B box domain enhanced secretion of pro-inflammatory cytokines and chemokines: IL-1beta, IL-2, IL-5, IL-8, IL-12 and tumor necrosis factor (TNF)-alpha, but not IL-6 and IL-10. Furthermore, four peptides whose sequences correspond to different regions of HMGB1 induced production of IL-1beta, IL-2 and IL-12 (p70), but not IL-10 and IL-6 in mouse BM-DCs. Interestingly, these peptides differed in their capacity to induce TNF-alpha, IL-5, IL-18 and IL-8. B box domain as well as peptide-activated DCs acted as potent stimulators of allogeneic T cells in a mixed leukocyte reaction. DCs exposed to HMGB1 peptides induced proliferation of ovalbumin-specific syngeneic T cells. These DC-activating peptides could serve as an adjuvant in immunotherapeutic or vaccine context and the selective activity of these different peptides suggests a means to customize the functional properties of DCs.

Requirements Specification for Automated Fall and Injury Risk Assessment

Studies in Health Technology and Informatics. 2006  |  Pubmed ID: 17102234

Fall and injury prevention continues to be a challenge in the acute care environment. Identification of patients at risk can guide preventive care for these individuals. The following study employed usability engineering methods via a series of focus groups, to specify functional and design requirements for an automated Fall-Injury Risk Assessment Instrument. Focus groups were held with interdisciplinary decision makers and end-users to identify functional and design specifications for the automated instrument. The results were mapped to usability heuristics, which were used to guide design decisions. The main elements identified were data completeness, workflow processes, resource access, and cognitive burden. The main usability factors identified were efficiency of user, match with real world, error prevention, recognition not recall and minimalist design. Focus groups are a useful methodology to specify requirements for healthcare applications. Outcomes evaluation of the automated instrument is in process.

Chemistry of the Silica Surface: Liquid-solid Reactions of Silica Gel with Trimethylaluminum

Journal of the American Chemical Society. Dec, 2006  |  Pubmed ID: 17177462

The reaction of trimethylaluminum and dry, high-surface-area (500 m2/g) silica gel in a mixed slurry was studied using multinuclear, solid-state NMR spectroscopy. The products of the initial reaction were characterized, and their progress through subsequent washing with diethyl ether and reactions with measured amounts of water was followed. The quantitative distribution of different chemical forms of carbon deposited on the silica surface by the initial reaction was measured. The products of the initial reaction are dominated by methyl species of the types Al(CH3)n (with Si-O-Al linkages), Si-O-CH3, and (Si-O)4-nSi(CH3)n; aluminum is seen to exist predominantly as a five-coordinate species. Subsequent treatment with diethyl ether fails to remove any surface species, but instead the ether becomes strongly associated with the surface and highly resistant to removal. Stepwise additions of water hydrolyze the Al-CH3 and Si-O-CH3 moieties, leading to conversion of five-coordinate aluminum to four- and six-coordinate aluminum, and affect the partial release of the surface-associated diethyl ether; Si-CH3 moieties remain. The effect of aromatic and saturated solvents on the initial reaction was examined and found to cause a small but significant change in the distribution of products. Structures of aluminum-centered species on the silica surface consistent with the spectroscopic data are proposed.

Vanilloid Type 1 Receptor and the Acid-sensing Ion Channel Mediate Acid Phosphate Activation of Muscle Afferent Nerves in Rats

Journal of Applied Physiology (Bethesda, Md. : 1985). Feb, 2006  |  Pubmed ID: 16210435

Reflex cardiovascular responses to contracting skeletal muscle are mediated by mechanical and metabolic stimulation of thin-fiber muscle afferents. Diprotonated phosphate (H2PO4-) excites those thin-fiber nerves and evokes the muscle pressor reflex. The receptors mediating this response are unknown. Thus we examined the role played by purinergic receptors, vanilloid type 1 receptors (VR1), and acid-sensing ion channels (ASIC) in mediating H2PO4- -evoked pressor responses. Phosphate and blocking agents were injected into the arterial blood supply of the hindlimb muscles of 53 decerebrated rats. H2PO4- (86 mM, pH 6.0) increased mean arterial pressure by 25 +/- 2 mmHg, whereas monoprotonated phosphate (HPO4(2-), pH 7.5) had no effect. Pyridoxalphosphate-6-azophenyl-2',4'-disulfonic acid (a purinergic receptor antagonist, 2 mM) did not block the response. However, capsazepine (a VR1 antagonist, 1 mg/kg) attenuated the reflex by 60% and amiloride (an ASIC blocker, 6 microg/kg) by 52%. Of note, the H2PO4- -induced pressor response was attenuated by 87% when both capsazepine and amiloride were injected before the H2PO4-. In conclusion, VR1 and ASIC mediate the pressor response due to H2PO4-. The H2PO4- -evoked response was greater when VR1 and ASIC blockers were given simultaneously than when the respective blockers were given separately. Our laboratory's previous study has shown that H+ stimulates ASIC (but not VR1) on thin-fiber afferent nerves in evoking the reflex response. Thus VR1 and ASIC are likely to play a coordinated and interactive role in processing the muscle afferent response to H2PO4-. Furthermore, the physiological mechanisms mediating the response to H+ and H2PO4- are likely to be different.

Aging Augments Interstitial K+ Concentrations in Active Muscle of Rats

Journal of Applied Physiology (Bethesda, Md. : 1985). Apr, 2006  |  Pubmed ID: 16322369

Skeletal muscle performance declines with advancing age, and the underlying mechanism is not completely understood. A large body of convincing evidence has demonstrated a crucial role for interstitial K+ concentration ([K+]o) in modulating contractile function of skeletal muscle. The present study tested the hypothesis that during muscle contraction there is a greater accumulation of [K+]o in aged compared with adult skeletal muscle. Twitch muscle contraction was induced by electrical stimulation of the sciatic nerves of 8- and 32-mo-old Fischer 344 x Brown Norway rats. Levels of [K+]o were measured continuously by a microdialysis technique with the probes inserted into the gastrocnemius muscle. Stimulation at 1, 3, and 5 Hz elevated muscle [K+]o by 52, 64, and 88% in adult rats, and by 78, 98, and 104% in aged rats, respectively, and the increase was significantly higher in aged than in adult rats. Recovery for [K+]o, as measured by the time for [K+]o to recover by 20 and 50% from peak response after stimulation, was slower in aged rats. Ouabain (5 mM), a specific inhibitor of the Na+-K+ pump, was added in the perfusate to inhibit the reuptake of K+ into the cells to assess the role of the pump in the overall K+ balance. Ouabain elevated muscle [K+]o at rest, and the effect was significantly attenuated in aged animals. The present data demonstrated an augmented [K+]o in aged skeletal muscle compared with adult skeletal muscle, and the data suggested that an alteration in the function of the Na+-K+ pump may contribute, in part, to the deficiency in K+ balance in skeletal muscle of aged rats.

Methylation Analysis of HMLH1 Gene Promoter by a Bisulfite-sensitive Single-strand Conformation Polymorphism-capillary Electrophoresis Method

Biomedical Chromatography : BMC. Aug, 2006  |  Pubmed ID: 16358356

DNA methylation is an important epigenetic modification that alters transcription in those genes containing CpG islands. In this report, a novel DNA methylation analysis method was developed employing bisulfite-single strand conformation polymorphism combined with capillary electrophoresis (bisulfite-SSCP-CE). During the bisulfite treatment of genomic DNA, a high concentration of sodium bisulfite (4.8 mol/L) was preferred in order to shorten reaction time and minimize template degradation. The methylated and unmethylated ssDNA of hMLH1 promoter were simultaneously separated under the optimized CE conditions, including 6% SLPA with 10% glycerol as sieving medium, 25 degrees C as separation temperature and 12 kV as running voltage. The heterogeneous methylation of hMLH1 promoter was identified in 13 of 64 colorectal cancer patients. Moreover, hMLH1 promoter methylation had a significant relationship with protein expression loss and increased with the age of patients. Our results indicated that DNA methylation analysis for a large number of clinical samples would be facilitated by use of the bisulfite-SSCP-CE method.

Electrophoresis Microchips with Sharp Inlet Tips, for Contactless Conductivity Detection, Fabricated by In-situ Surface Polymerization

Analytical and Bioanalytical Chemistry. Feb, 2006  |  Pubmed ID: 16372178

A novel method based on in-situ surface polymerization of methyl methacrylate (MMA) has been developed for rapid fabrication of poly(methyl methacrylate) (PMMA) electrophoresis microchips with sharp inlet tips. Prepolymerized MMA containing an ultraviolet (UV) initiator was directly sandwiched between a nickel template and a PMMA plate. The image of the relief on the nickel template was precisely replicated in the synthesized PMMA layer on the surface of the commercially available PMMA plate during UV-initiated polymerization at room temperature. The chips were subsequently assembled by thermal bonding of channel plates and cover sheets. The sample was directly introduced into the separation channel through a sharp inlet tip, which was placed in the sample vial, without use of an injection cross. The attractive performance of the novel PMMA microchips has been demonstrated by using contactless conductivity detection for determination of several inorganic ions. Such rapid and simple sample introduction leads to highly reproducible signals with relative standard deviations of less than 5% for peak responses. These new approaches significantly simplify the process of fabricating PMMA devices and show great promise for high-speed microchip analysis.

The Aqueous Extract of a Popular Herbal Nutrient Supplement, Angelica Sinensis, Protects Mice Against Lethal Endotoxemia and Sepsis

The Journal of Nutrition. Feb, 2006  |  Pubmed ID: 16424112

Despite recent advances in antibiotic therapy and intensive care, sepsis remains a widespread problem in critically ill patients. The high mortality from sepsis is in part mediated by bacterial endotoxin, which stimulates macrophages/monocytes to sequentially release early (e.g., tumor necrosis factor, interleukin-1, and interferon-gamma) and late [e.g., high mobility group box 1 protein (HMGB1)] proinflammatory cytokines. Our discovery of HMGB1 as a late mediator of lethal systemic inflammation has initiated a new field of investigation for the development of experimental therapeutics. A popular Chinese herb, Angelica sinensis (also known as Dang Gui or Dong Quai) has been used traditionally for treating women with gynecological disorders (such as dysmenorrheal and hot flashes). Here we examined the effect of Angelica sinensis extract on endotoxin-induced HMGB1 release in vitro, and explored its therapeutic potential in animal models of lethal endotoxemia and sepsis [induced by cecal ligation and puncture (CLP)] in vivo. We demonstrated that a low-molecular-weight (<10 kDa) fraction of A. sinensis extract significantly attenuated endotoxin-induced HMGB1 release in part through interfering with its cytoplasmic translocation in macrophage cultures. Prophylactic administration of an aqueous extract of A. sinensis significantly attenuated systemic HMGB1 accumulation in vivo, and conferred a dose-dependent protection against lethal endotoxemia. Furthermore, delayed administration of A. sinensis extract beginning 24 h after CLP attenuated systemic HMGB1 accumulation, and significantly rescued mice from lethal sepsis. Taken together, these data suggest that A. sinensis contains water-soluble components that exert protective effects against lethal endotoxemia and experimental sepsis in part by attenuating systemic accumulation of a late proinflammatory cytokine, HMGB1.

Temperature Modulates P2X Receptor-mediated Cardiovascular Responses to Muscle Afferent Activation

American Journal of Physiology. Heart and Circulatory Physiology. Sep, 2006  |  Pubmed ID: 16501013

Static muscle contraction increases ATP release into the muscle interstitial space. Elevated ATP in muscle stimulates thin fiber muscle afferents and increases blood pressure via engagement of purinergic P2X receptors. In addition, ATP activates P2X receptors and enhances cardiovascular responses induced by stimulation of muscle mechanoreceptors. In this study, we examined whether elevated muscle temperature would attenuate and whether reduced temperature would potentiate P2X effects on reflex muscle responses. alpha,beta-Methylene ATP (alpha,beta-MeATP) was injected into the arterial blood supply of hindlimb muscle to stimulate P2X receptors, and muscle stretch was induced to activate mechanically sensitive muscle afferents as alpha,beta-MeATP was injected in 10 anesthetized cats. Femoral arterial injection of alpha,beta-MeATP (1.0 mM) increased mean arterial pressure (MAP) by 35+/-5 (35 degrees C), 26+/-3 (37 degrees C), and 19+/-3 mmHg (39 degrees C; P<0.05 vs. 35 degrees C), respectively. Muscle stretch (2 kg) elevated MAP. The MAP response was significantly enhanced 34% and 36% when alpha,beta-MeATP (0.2 mM) was arterially infused 5 min before muscle stretch at 35 degrees and 37 degrees C, respectively. However, as muscle temperature reached 39 degrees C, the stretch-evoked response was augmented only 6% by alpha,beta-MeATP injection, and the response was significantly attenuated compared with the response with muscle temperature of 35 degrees and 37 degrees C. In addition, we also examined effects of muscle temperature on alpha,beta-MeATP enhancement of the cardiovascular responses to static muscle contraction while the muscles were freely perfused and the circulation to the muscles was occluded. Because muscle temperature was 37 degrees C, arterial injections of alpha,beta-MeATP significantly augmented contraction-evoked MAP response by 49% (freely perfused) and 53% (ischemic condition), respectively. It is noted that this effect was significantly attenuated at a muscle temperature of 39 degrees C. These data indicate that the effect of P2X receptor on reflex muscle response is sensitive to alternations of muscle temperature and that elevated temperature attenuates the response.

Eimeria Tenella: Cloning of a Novel Eimeria Tenella CDNA Encoding a Protein Related to Rhomboid Family from F2 Hybrid Strain

Experimental Parasitology. Aug, 2006  |  Pubmed ID: 16545808

A novel cDNA sequence with an open reading frame of 774 bp from Eimeria tenella F2 hybrid strain (ETRH01) was isolated from a lambda cDNA library with a monoclonal antibody against sporozoite. Analysis of the genomic sequence suggests that this is an intronless gene. The deduced protein sequence has 257 amino acids with a calculated molecular weight of 28.349 kDa and an isoelectric point of 8.56. Sequence analysis revealed seven transmembrane domains and a rhomboid domain within the protein. RT-PCR result indicates that this gene was expressed in all of the five E. tenella isolates analyzed. To further study the role of this novel gene in the life cycle of E. tenella, ETRH01 was successfully expressed using pET28b(+) expression system.

[A Study on the Mechanical Properties of Poly-D,L-lactic Acid/hydroxyapatite Compound Fibers]

Sheng Wu Yi Xue Gong Cheng Xue Za Zhi = Journal of Biomedical Engineering = Shengwu Yixue Gongchengxue Zazhi. Apr, 2006  |  Pubmed ID: 16706370

The PDLLA/HA compound fibers with excellent properties could be obtained by melt-spinning. This study inquired about the mechanical properties, change rule and influencing factors. The results showed that the composites fibers of mechanical properties were good in the PDLLA(molecular weight 120,000) to which were added the 4-20 microm HA particles. In the matrix of PDLLA with accretion (10%) of HA particles, the tensile strength of the compound fibers was the highest, compared with the others. The tensile strength of the compound fibers of PDLLA (molecular weight 20,000-300,000) was high. The tensile strength of the compound fibers decreased with the increase of the fiber diameter, and the PDLLA/HA fibers 40-60 micrometers in diameter had the highest elongation at break.

The QSPR (quantitative Structure-property Relationship) Study About the Anaerobic Biodegradation of Chlorophenols

Chemosphere. Dec, 2006  |  Pubmed ID: 16750555

In this study, based on quantum chemical and physicochemical descriptors, by the use of partial least squares analysis, a good prediction quantitative structure-property relationship for the disappearance rate constant (logK) of chlorophenols (CPs) in the anaerobic culture was obtained. It was found that the resonance energy of the two-center term (J), which described the character of the weakest carbon-chlorine bond, played an important role in the reductive chlorine processes, the greater the sizes of CPs molecules, the higher the logK values. Increasing energy of the lowest unoccupied molecular orbital (E(lumo)) values of the CPs lead to decreasing logK values, and CPs with large absolute hardness values tended to have big logK values.

Cutting Edge: High-mobility Group Box 1 Preconditioning Protects Against Liver Ischemia-reperfusion Injury

Journal of Immunology (Baltimore, Md. : 1950). Jun, 2006  |  Pubmed ID: 16751357

High mobility group box 1 (HMGB1) is a NF released extracellularly as a late mediator of lethality in sepsis and as an early mediator of inflammation following injury. Here we demonstrate that in contrast to the proinflammatory role of HMGB1, preconditioning with HMGB1 results in protection following hepatic ischemia/reperfusion (I/R). Pretreatment of mice with HMGB1 significantly decreased liver damage after I/R. The protection observed in mice pretreated with HMGB1 was associated with a higher expression of IL-1R-associated kinase-M, a negative regulator of TLR4 signaling, compared with controls. We thus explored the possibility that HMGB1 preconditioning was mediated through TLR4 activation. HMGB1 preconditioning failed to provide protection in TLR4 mutant (C3H/HeJ) mice, but successfully reduced damage in TLR4 wild-type (C3H/HeOuj) mice. Our studies demonstrate that in contrast to the role of HMGB1 as an early mediator of inflammation and organ damage in hepatic I/R, HMGB1 preconditioning can be protective.

[Clinical Value of Liquid-based Cytologic Test in Sputum Examination of Patients with Lung Cancer.]

Zhongguo Fei Ai Za Zhi = Chinese Journal of Lung Cancer. Apr, 2006  |  Pubmed ID: 21144309

BACKGROUND: There are lots of mucus, blood, inflammatory cells and necrotic material in the pick-and-smear slides, resulting in a low detection rate. Liquid-based cytologic test (LCT) has been applied for cervical cytology diagnosis successfully and widely, however it is few reported yet for sputum cytology diagnosis at present. The aim of this study is to evaluate the clinical value of LCT in sputum examination of patients with lung cancer, and to find a novel method of early diagnosis of lung cancer. METHODS: The cytologic findings and the diagnostic rate for lung cancer were compared between LCT and conventional pick-and-smear method. RESULTS: There were smaller area of smear membrane, clearer background, more distinctly cytologic picture and stereoscopic fell by LCT comparing with pick-and-smear method. The diagnostic rate for small cell lung cancer by LCT was significantly higher than that by pick-and-smear method (P < 0.05). After combined detection of the two methods, the diagnostic rate for lung cancer was obviously improved (85.1%), which was remarkably higher than that by pick-and-smear method alone (P < 0.01). CONCLUSIONS: It is operated easily for LCT to be well controlled in making smear and dyeing. LCT may be a novel technique worthy of wide use. Combination of LCT with pick-and-smear method appears to be of great value in clinical application.

TSP50 Encodes a Testis-specific Protease and is Negatively Regulated by P53

Cancer Research. Feb, 2007  |  Pubmed ID: 17283160

Earlier studies suggested that TSP50 is a testis-specific gene that encodes a protein, which is homologous to serine proteases but differs in that threonine replaces serine in its catalytic triad. Most importantly, it was abnormally reactivated in many breast cancer biopsies tested. While further investigating its biochemical and cell biological natures, we found that TSP50 exhibited enzyme activity and was located in the endoplasmic reticulum and cytosol membrane. During our studies to elucidate the regulatory mechanisms related to its differential expression, we discovered a putative p53-binding site and several Sp1-binding sites in the TSP50 promoter, which led us to test if it was regulated by the p53 gene. We found that the p53 transgene negatively regulated the TSP50 promoter in diverse types of cell lines. This result was consistent with other observations: (a) p53 overexpression reduced endogenous TSP50 expression; and (b) breast cancer cell lines containing mutated p53, such as MCF7/Adr, or normal p53, such as MCF7, produced high or low levels of TSP50 transcripts, which was consistent with the fact that TSP50 promoter activity was much higher in MCF7/Adr than that in MCF7 cells. We also found that the quantity of Sp1 transcription factor was lower in MCF7/Adr than in MCF7 cells, which suggested that another mechanism (i.e., transcription factor modulation) was also involved in TSP50 differential expression.

A Cardiovascular Drug Rescues Mice from Lethal Sepsis by Selectively Attenuating a Late-acting Proinflammatory Mediator, High Mobility Group Box 1

Journal of Immunology (Baltimore, Md. : 1950). Mar, 2007  |  Pubmed ID: 17339485

The pathogenesis of sepsis is mediated in part by bacterial endotoxin, which stimulates macrophages/monocytes to sequentially release early (e.g., TNF, IL-1, and IFN-gamma) and late (e.g., high mobility group box 1 (HMGB1) protein) proinflammatory cytokines. The recent discovery of HMGB1 as a late mediator of lethal sepsis has prompted investigation for development of new experimental therapeutics. We found that many steroidal drugs (such as dexamethasone and cortisone) and nonsteroidal anti-inflammatory drugs (such as aspirin, ibuprofen, and indomethacin) failed to influence endotoxin-induced HMGB1 release even at superpharmacological concentrations (up to 10-25 microM). However, several steroid-like pigments (tanshinone I, tanshinone IIA, and cryptotanshinone) of a popular Chinese herb, Danshen (Salvia miltiorrhiza), dose dependently attenuated endotoxin-induced HMGB1 release in macrophage/monocyte cultures. A water-soluble tanshinone IIA sodium sulfonate derivative (TSNIIA-SS), which has been widely used as a Chinese medicine for patients with cardiovascular disorders, selectively abrogated endotoxin-induced HMGB1 cytoplasmic translocation and release in a glucocorticoid receptor-independent manner. Administration of TSNIIA-SS significantly protected mice against lethal endotoxemia and rescued mice from lethal sepsis even when the first dose was given 24 h after the onset of sepsis. The therapeutic effects were partly attributable to attenuation of systemic accumulation of HMGB1 (but not TNF and NO) and improvement of cardiovascular physiologic parameters (e.g., decrease in total peripheral vascular resistance and increase in cardiac stroke volume) in septic animals. Taken together, these data re-enforce the pathogenic role of HMGB1 in lethal sepsis, and support a therapeutic potential for TSNIIA-SS in the treatment of human sepsis.

Effect of Muscle Interstitial PH on P2X and TRPV1 Receptor-mediated Pressor Response

Journal of Applied Physiology (Bethesda, Md. : 1985). Jun, 2007  |  Pubmed ID: 17379752

Activation of purinergic P2X receptors and transient receptor potential vanilloid type 1 (TRPV1) on muscle afferent nerve evokes the pressor response. Because P2X and TRPV1 receptors are sensitive to changes in pH, the aim of this study was to examine the effects of muscle acidification on those receptor-mediated cardiovascular responses. In decerebrate rats, the pH in the hindlimb muscle was adjusted by infusing acidic Ringer solutions into the femoral artery. Dialysate was then collected using microdialysis probes inserted into the muscles, and pH was measured. The interstitial pH was 7.53+/-0.01, 7.22+/-0.02, 6.94+/-0.04, and 6.59+/-0.03 in response to arterial infusion of the Ringer solution at pH 7.4, 6.5, 5.5, and 4.5, respectively. Femoral arterial injection of alpha,beta-methylene-ATP (P2X receptor agonist) in the concentration of 0.25 mM (volume, 0.15-0.25 ml; injection duration, 1 min) at the infused pH of 7.4, 6.5, and 5.5 increased mean arterial pressure (MAP) by 29+/-2, 24+/-3, and 21+/-3 mmHg, respectively (P<0.05, pH 5.5 vs. pH 7.4). When pH levels in the infused solution were 7.4, 6.5, 5.5, and 4.5, capsaicin (1 microg/kg), a TRPV1 agonist, was injected into the artery. This elevated MAP by 29+/-4, 33+/-2, 35+/-3, and 40+/-3 mmHg, respectively (P<0.05, pH 4.5 vs. pH 7.4). Furthermore, blocking acid-sensing ion channel (ASIC) blunted pH effects on TRPV1 response. Our data indicate that 1) muscle acidosis attenuates P2X-mediated pressor response but enhances TRPV1 response; 2) exaggerated TRPV1 response may require lower pH in muscle, and the effect is likely to be mediated via ASIC mechanisms. This study provides evidence that muscle pH may be important in modulating P2X and TRPV1 responsiveness in exercising muscle.

Evaluation of OPRM1 Variants in Heroin Dependence by Family-based Association Testing and Meta-analysis

Drug and Alcohol Dependence. Oct, 2007  |  Pubmed ID: 17416470

OPRM1, which codes for the mu-opioid receptor, is the most frequently studied candidate gene for opioid dependence. Despite numerous allelic association studies, no definitive conclusion has been reached regarding the role of OPRM1 polymorphisms in determining risk for opioid dependence. We attempted to resolve this by conducting a family-based association study and meta-analysis which may be more robust and powerful, respectively, than traditional case-control analyses. First, we genotyped three single nucleotide polymorphisms (SNPs) of OPRM1 in 1208 individuals from 473 Han Chinese families ascertained on the basis of having two or more siblings with DSM-IV-defined opioid dependence. The Val6Ala and Arg111His SNPs were detected, but with low minor allele frequencies (0.002 and 0.001, respectively). The Asn40Asp SNP was more informative (minor allele frequency: 0.419), but no significant evidence was observed for either a dominant (p=0.810) or additive (p=0.406) effect of this polymorphism on risk for opioid dependence. In addition, a meta-analysis of case-control studies of opioid dependence was performed, and found a similar lack of evidence for an association with the Asn40Asp SNP (p=0.859). Although a role of OPRM1 polymorphisms in determining risk for opioid dependence cannot be entirely discounted, a major contribution of the Asn40Asp polymorphism seems unlikely. Further analysis is warranted in samples from specific ancestral groups. In addition, it is critical that other OPRM1 variants, including all haplotype-tagging and amino-acid-coding SNPs, be tested for an influence on risk for opioid dependence, since the Asn40Asp polymorphism is only one of several hundred known mutations in the gene.

Interstitial Norepinephrine Concentrations in Skeletal Muscle of Ischemic Heart Failure

American Journal of Physiology. Heart and Circulatory Physiology. Aug, 2007  |  Pubmed ID: 17449553

During exercise, sympathetic nerve responses are accentuated in heart failure (HF), and this enhances norepinephrine (NE) release and evokes vasoconstriction. Two key pathophysiological responses could contribute to the greater NE release: 1) increased sympathetic nerve discharge and 2) increased NE in the neurovascular junction for a given level of sympathetic discharge. In this report, we focus on the second of these two general issues and test the following hypotheses: 1) in HF for a given level of sympathetic nerve stimulation, NE concentration in the interstitium (an index of neurovascular NE) would be greater, and 2) the greater interstitial NE concentration would be linked to reduced NE uptake. Studies were performed in rats 8-10 wk after induction of myocardial infarction (MI). Interstitial NE samples were collected from microdialysis probes inserted into the hindlimb muscle. Dialysate concentration of NE was determined by the HPLC method. First, interstitial NE concentration increased during electrical stimulation of the lumbar sympathetic nerves in eight control rats. An increase in interstitial NE concentration was significantly greater in 10 rats with severe MI. Additionally, an NE uptake-1 inhibitor (desipramine, 1 microM) was injected into the arterial blood supply of the muscle in six control and eight MI rats. Desipramine increased interstitial NE concentration by 24% in control and by only 3% (P < 0.05 vs. control) in MI rats. In conclusion, given levels of electrical stimulation of the lumbar sympathetic nerve lead to higher interstitial NE concentration in HF. This effect is due, in part, to reduced NE uptake-1 in HF.

Differential Sympathetic Outflow Elicited by Active Muscle in Rats

American Journal of Physiology. Heart and Circulatory Physiology. Oct, 2007  |  Pubmed ID: 17573458

The present study was undertaken to test the hypothesis that activation of the muscle reflex elicits less sympathetic activation in skeletal muscle than in internal organs. In decerebrate rats, we examined renal and lumbar (mainly innervating hindlimb blood vessels) sympathetic nerve activities (RSNA and LSNA, respectively) during 1 min of 1) repetitive (1- to 4-s stimulation-to-relaxation) contraction of the triceps surae muscle, 2) repetitive tendon stretch, and 3) repetitive contraction with hindlimb circulatory occlusion. During these interventions, RSNA and LSNA responded synchronously as tension developed. The increase was greater in RSNA than in LSNA [+51 +/- 14 vs. +24 +/- 5% (P < 0.05) with contraction, +46 +/- 8 vs. +17 +/- 4% (P < 0.05) with stretch, +76 +/- 20 vs. 39 +/- 7% (P < 0.05) with contraction during occlusion] during all three interventions: repetitive contraction (n = 10, +508 +/- 48 g tension from baseline), tendon stretch (n = 12, +454 +/- 34 g), and contraction during occlusion (n = 9, +473 +/- 33 g). Additionally, hindlimb circulatory occlusion significantly enhanced RSNA and LSNA responses to contraction. These data demonstrate that RSNA responses to muscle contraction and stretch are greater than LSNA responses. We suggest that activation of the muscle afferents induces the differential sympathetic outflow that is directed toward the kidney as opposed to the limbs. This differential outflow contributes to the distribution of cardiac output observed during exercise. We further suggest that as exercise proceeds, muscle metabolites produced in contracting muscle sensitize muscle afferents and enhance sympathetic drive to limbs and renal beds.

Prostaglandin E2 (PGE2) Inhibits Glutamatergic Synaptic Transmission in Dorsolateral Periaqueductal Gray (dl-PAG)

Brain Research. Aug, 2007  |  Pubmed ID: 17612511

The purpose of this study was to determine the role of prostaglandin E(2) (PGE(2)) in modulating neuronal activity of the dorsolateral periaqueductal gray (dl-PAG) through excitatory and inhibitory synaptic inputs. First, whole cell voltage-clamp recording was performed to obtain excitatory and inhibitory postsynaptic currents (EPSCs and IPSCs) of the dl-PAG neurons. Our results show that PGE(2) significantly decreased the frequency of miniature EPSCs and amplitude of evoked EPSCs. The effects were mimicked by sulprostone, an agonist to PGE(2) EP(3) receptors. In contrast, PGE(2) had no distinct effect on IPSCs. In addition, spontaneous action potential of the dl-PAG neurons was recorded using whole cell current-clamp methods. PGE(2) significantly attenuated the discharge rate of the dl-PAG neurons. The decreased firing activity was abolished in the presence of glutamate NMDA and non-NMDA receptor antagonists. The results from the current study provide the first evidence indicating that PGE(2) inhibits the neuronal activity of the dl-PAG via selective attenuation of glutamatergic synaptic inputs, likely due to the activation of presynaptic EP(3) receptors.

Medication Reconciliation Using Natural Language Processing and Controlled Terminologies

Studies in Health Technology and Informatics. 2007  |  Pubmed ID: 17911803

Medication reconciliation (MR) is a process that seeks to assure that the medications a patient is supposed to take are the same as what they are actually taking. We have developed a method in which medication information (consisting of both coded data and narrative text) is extracted from twelve sources from two clinical information systems and assembled into a chronological sequence of medication history, plans, and orders that correspond to periods before, during and after a hospital admission. We use natural language processing, a controlled terminology, and a medication classification system to create matrices that can be used to determine the initiation, changes and discontinuation of medications over time. We applied the process to a set of 17 patient records and successfully abstracted and summarized the medication data. This approach has implications for efforts to improve medication history-taking, order entry, and automated auditing of patient records for quality assurance.

Inhibition of Krr1 Gene Expression in Giardia Canis by a Virus-mediated Hammerhead Ribozyme

Veterinary Parasitology. Jan, 2007  |  Pubmed ID: 16982153

Giardia, a most primitive eukaryote, infects several species including human and it is a major agent of waterborne outbreak of diarrhea. It has been difficult to employ standard genetic methods in the study of Giardia, but the RNA virus-based transfection system has been developed and used for the genetic manipulation. KRR1 protein is responsible for ribosome biosynthesis in Giardia. In this study, cDNA encoding hammerhead ribozyme flanked with various lengths of antisense Krr1 RNA were cloned into a viral vector pGCV634/GFP/GCV2174 derived from the genome of Giardia canis virus (GCV). RNA transcripts of the plasmids showed high cleavage activities on Krr1 mRNA in vitro. They were electroporated into GCV-infected G. canis trophozoites and Krr1 mRNA level was decreased by 72% with the ribozyme KRzS and 86% with the ribozyme KRzL, while the control ribozyme TRzS showed no effect on the level of Krr1 mRNA. The two hammerhead ribozyme transfected cells grew slowly, their internal structures got blurred and the cells were deformed. These results indicated that GCV could be useful tool for gene manipulation of G. canis.

P2X Receptor-mediated Muscle Pressor Reflex in Myocardial Infarction

American Journal of Physiology. Heart and Circulatory Physiology. Feb, 2007  |  Pubmed ID: 17012345

A previous report from this laboratory demonstrated that the ATP-sensitive P2X receptor-mediated muscle pressor reflex was augmented in rats with heart failure (HF). The purpose of this study was to better understand the underlying mechanisms for this greater response in HF rats. We examined 1) responsiveness of the P2X receptor to alpha,beta-methylene ATP (alpha,beta-me-ATP), a P2X receptor agonist, in control and HF rats induced by myocardial infarction (MI); 2) the relationship between P2X-induced blood pressure response and left ventricular (LV) function; and 3) the expression of P2X receptors in the dorsal root ganglion (DRG) of control rats and rats with HF. Eight to 14 wk after coronary artery ligation, the severity of the MI was determined by echocardiography. In the first group of the experiment, alpha,beta-me-ATP (0.0625, 0.125, 0.25, and 0.5 mM) was injected into the arterial blood supply of the hindlimb muscles to evoke a pressor response in 17 decerebrated rats (6 controls, 6 small MIs with infarcts of the LV between 10 and 35%, and 5 large MIs with infarcts >35%). The P2X agonist increased blood pressure, and the effect was significantly accentuated in large MI rats compared with small MI rats and control rats. A significant correlation was observed between alpha,beta-me-ATP-evoked pressor response and the LV fractional shortening, an index of LV function. In the second group of the experiment, immunocytochemistry was used to examine the immunoreactivity of P2X receptor in the DRG neurons of small diameter fibers in six healthy control rats, five small MI, and five large MI rats. The percentage of P2X immunostaining-positive neurons in the DRG was markedly greater in large MI rats (52% vs. 29% in controls and 34% in small MIs, P < 0.05). In conclusion, our findings demonstrate that 1) muscle afferent-mediated pressor response of P2X activation was exaggerated in MI animals, and the responsiveness was related to the degree of LV dysfunction; and 2) augmented reflex response was associated with upregulated P2X receptors in the DRG neurons of thin fiber afferent nerves following MI. The data suggest that P2X-mediated responsiveness in the processing of muscle afferent signals may have important implications for understanding cardiovascular responses to exercise in HF.

Interstitial K+ Concentration in Active Muscle After Myocardial Infarction

American Journal of Physiology. Heart and Circulatory Physiology. Feb, 2007  |  Pubmed ID: 17012361

Previous work demonstrated that Na(+)-K(+) pump activity within skeletal muscle is attenuated in myocardial infarction (MI). This may lead to enhanced interstitial K(+) concentration ([K(+)](o)) in the muscle. We tested the hypothesis that [K(+)](o) rises with muscle contraction and that, in rats with MI, the rate of rise in [K(+)](o) is greater than it is in control animals. Microdialysis probes were inserted in the skeletal muscle of six healthy control and six MI rats. The ends of the probes were then attached to the K(+) electrodes, and [K(+)](o) was continuously measured. Muscle contraction was induced by electrical stimulation of the sciatic nerves for 1 min. Stimulation at 1 and 3 Hz increased muscle [K(+)](o) by 14.2% and 44.7% in controls and by 22.9% and 62.8% in MI rats (P < 0.05 vs. controls), respectively. When ouabain, an inhibitor of Na(+)-K(+) pump, was added to the perfusate, muscle [K(+)](o) rose significantly. This effect of ouabain was significantly attenuated in MI animals. In conclusion, when compared with that in control animals, an increase of [K(+)](o) in exercising muscle is augmented in MI rats, likely due to an attenuation of Na(+)-K(+) pump activity.

TRPV1 Receptor Mediates Glutamatergic Synaptic Input to Dorsolateral Periaqueductal Gray (dl-PAG) Neurons

Journal of Neurophysiology. Jan, 2007  |  Pubmed ID: 17065246

The purpose of this study was to determine the role of transient receptor potential vanilloid type 1 (TRPV1) receptor in modulating neuronal activity of the dorsolateral periaqueductal gray (dl-PAG) through excitatory and inhibitory synaptic inputs. First, whole cell voltage-clamp recording was performed to obtain the spontaneous miniature excitatory postsynaptic currents (mEPSCs) and inhibitory postsynaptic currents (mIPSCs) of the dl-PAG neurons. As 1 microM of capsaicin was applied into the perfusion chamber, the frequency of mEPSCs was increased from 3.21 +/- 0.49 to 5.64 +/- 0.64 Hz (P < 0.05, n = 12) without altering the amplitude and the decay time constant of mEPSCs. In contrast, capsaicin had no distinct effect on mIPSCs. A specific TRPV1 receptor antagonist, iodo-resiniferatoxin (i-RTX, 300 nM), decreased the frequency of mEPSCs from 3.51 +/- 0.29 to 2.01 +/- 0.2 Hz (P < 0.05, n = 8) but did not alter the amplitude and decay time. In addition, i-RTX applied into the chamber abolished the effect of capsaicin on mEPSC of the dl-PAG. In another experiment, spontaneous action potential of the dl-PAG neurons was recorded using whole cell current-clamp methods. Capsaicin significantly elevated the discharge rate of the dl-PAG neurons from 3.03 +/- 0.38 to 5.96 +/- 0.87 Hz (n = 8). The increased firing activity was abolished in the presence of glutamate N-methy-D-aspartate (NMDA) and non-NMDA antagonists, 2-amino-5-phosphonopentanoic acid, and 6-cyano-7-nitroquinoxaline-2,3-dione. The results from this study provide the first evidence indicating that activation of TRPV1 receptors increases the neuronal activity of the dl-PAG through selective potentiation of glutamatergic synaptic inputs.

Cryptosporidium Parvum: Identification of a New Surface Adhesion Protein on Sporozoite and Oocyst by Screening of a Phage-display CDNA Library

Experimental Parasitology. Apr, 2007  |  Pubmed ID: 17097085

Cryptosporidium parvum is a significant cause of diarrheal disease worldwide. The specific molecules that mediate C. parvum-host interaction and the molecular mechanisms involved in the pathogenesis are unknown. In this study we described a novel phage display method to identify surface adhesion proteins of C. parvum. A cDNA library of the sporozoite and oocyst stages of C. parvum expressed on the surface of T7 phage was screened with intestinal epithelial cells (IECs) from the newborn Cryptosporidium-free Holstein calves. Proteins that selectively and specifically bound to IECs were then enriched using a multi-step panning procedure. Two proteins of C. parvum were selected, one was previously reported (p23), which was an important surface adhesion protein; the other was a novel surface adherence protein (CP12). Sequence analysis showed that CP12 has a N-terminal signal peptide, a transmembrane region, a N-glycosylation site, a casein kinase II phosphorylation site and two N-myristoylation sites. Immunofluorescence assay (IFA) using antibody specific for rCP12 demonstrated that the antibody can specifically bind the surface of sporozoite and oocyst, especially apical region of sporozoite. The surface localization of CP12 and its involvement in the host-parasite interaction suggest that it may serve as an effective target for specific preventive and therapeutic measures for cryptosporidiosis.

Effectiveness of First Eight Methadone Maintenance Treatment Clinics in China

AIDS (London, England). Dec, 2007  |  Pubmed ID: 18172377

To evaluate the effectiveness of the first phase of eight methadone maintenance treatment (MMT) clinics in China.

Gene Polymorphisms of the Renin-angiotensin-aldosterone System and Angiotensin II Type 1-receptor Activating Antibodies in Renal Rejection

The Tohoku Journal of Experimental Medicine. Nov, 2007  |  Pubmed ID: 17984617

Steroid refractory acute rejection (SRAR) is a major vital factor in renal transplantation recipients. The pathogenesis of SRAR may involve both immune and non-immune mechanisms. A decreased renal allograft function has also been associated with increased activity of renin-angiotensin-aldosterone system (RAS), which may be genetically determined. A total 206 renal transplant recipients, 116 males and 90 females, were included. The effects of gene polymorphisms of the four components of RAS including angiotensinogen (AGT), angiotensin-converting enzyme (ACE), angiotensin type 1 receptor (AT1R), and aldosterone synthase (CYP11B2) were investigated in 19 cases of renal transplant patients with SRAR. The association between SRAR and the activating antibodies targeting the AT1R were also investigated. Genotyping was performed for the M235T-AGT, the I/D-ACE, the A1166C-AT1R, and the -344T/C-CYP11B2 gene polymorphisms using polymerase chain reaction. Our results showed that renal allograft recipients with SRAR had significantly higher occurrences of the DD genotype of ACE and CC genotype of AT1R than recipients without SRAR. The other genetic polymorphisms of the RAS were not associated with SRAR. Activating antibodies targeting the AT1R were detected in the sera from 14 SRAR victims with malignant hypertension and without anti-HLA antibodies. This study provides evidence that determination before transplantation of the polymorphism of the gene encoding components of RAS may help identify patients who are at risk for SRAR. The detection of the antibodies of AT1R may contribute to the prevention of SRAR.

A Major Ingredient of Green Tea Rescues Mice from Lethal Sepsis Partly by Inhibiting HMGB1

PloS One. 2007  |  Pubmed ID: 17987129

The pathogenesis of sepsis is mediated in part by bacterial endotoxin, which stimulates macrophages/monocytes to sequentially release early (e.g., TNF, IL-1, and IFN-gamma) and late (e.g., HMGB1) pro-inflammatory cytokines. Our recent discovery of HMGB1 as a late mediator of lethal sepsis has prompted investigation for development of new experimental therapeutics. We previously reported that green tea brewed from the leaves of the plant Camellia sinensis is effective in inhibiting endotoxin-induced HMGB1 release.

Microbial Transformation of Cephalomannine by Luteibacter Sp

Journal of Natural Products. Dec, 2007  |  Pubmed ID: 18001087

Luteibacter sp., a new bacterium isolated from the soil around a Taxus cuspidata Sieb. et Zucc plant, was studied for its capability to metabolize cephalomannine (1). After preparative fermentation, eight metabolites were obtained and characterized as baccatin III (2), baccatin V (3), 10-deacetylbaccatin III (4), 10-deacetyl-10-oxobaccatin V (5), 7-epicephalomannine (6), 10-deacetylcephalomannine (7), 10-deacetyl-7-epicephalomannine (8), and 3'-N-debenzoyl-3'-N-(2-methylbutyryl)-7-epitaxol (9). Among these metabolites, 9 is a new compound. Epimerization of the 7beta-OH group and hydrolysis of the C-13 side-chain were the two major reactions in this bioprocess. However, the biotransformation of 7beta-D-xylosyl-10-deacetyltaxol (10) with the same strain yielded a C-13 side-chain eliminated product without epimerization at C-7 (11). Metabolites 5-9 and 11, together with 1 and paclitaxel, were evaluated for their inhibitory activities against five human cancer cell lines (HCT-8, Bel-7402, BGC-823, A549, and A2780). All these compounds showed less potent activities than paclitaxel, which is currently used in clinical chemotherapy.

Redesign of the Columbia University Infobutton Manager

AMIA ... Annual Symposium Proceedings / AMIA Symposium. AMIA Symposium. 2007  |  Pubmed ID: 18693813

The Infobutton Manager (IM) is an application that provides clinical users with context-specific links to health information resources. Usage of the first version (IM-1) suggested that the user interface was suboptimal.

Digital Partnerships for Health: Steps to Develop a Community-specific Health Portal Aimed at Promoting Health and Well-being

AMIA ... Annual Symposium Proceedings / AMIA Symposium. AMIA Symposium. 2007  |  Pubmed ID: 18693872

We describe the steps taken by the Harlem Health Promotion Center to develop a community-specific health web portal aimed at promoting health and well-being in Harlem. Methods and results that begin with data collection and move onto elucidating requirements for the web portal are discussed. Sentiments of distrust in medical institutions, and the desire for community specific content and resources were among the needs emanating from our data analysis. These findings guided our decision to customize social software designed to foster connections, collaborations, flexibility, and interactivity; an "architecture of participation". While, we maintain that the leveraging of social software may indeed be the way to build healthy communities and support learning and engagement in underserved communities, our conclusion calls for careful thinking, testing and evaluation research to establish best practice models for leveraging these emerging technologies to support health improvements in the community.

Auditing Dynamic Links to Online Information Resources

AMIA ... Annual Symposium Proceedings / AMIA Symposium. AMIA Symposium. 2007  |  Pubmed ID: 18693876

The Columbia University Infobutton Manager (IM) is a system that provides dynamically generated, context-specific links between clinical information systems and online information resources. The resources range from local documents, to commercially available document sets and search engines. The links provided by the IM can be reliably created, but there is no guarantee that they will function reliably, since the resources to which they point are subject to unannounced changes and failures. We have developed a set of tools to audit the links periodically to determine if the resources are available and if the IM has sufficient information to generate all the links needed by its users. These tools have been in use since February, 2006 and have provided timely warnings on many occasions. These warnings have allowed us to correct problems with resource access before they became apparent to our users and often before the resource maintainers were aware of the problems. The tools have thus helped us provide clinicians with a dependable level of service.

Healthy Harlem: Empowering Health Consumers Through Social Networking, Tailoring and Web 2.0 Technologies

AMIA ... Annual Symposium Proceedings / AMIA Symposium. AMIA Symposium. 2007  |  Pubmed ID: 18694106

Consumer health informatics has emerged as a strategy to inform and empower patients for self management of their health. The emergence of and explosion in use of user-generated online media (e.g.,blogs) has created new opportunities to inform and educate people about healthy living. Under a prevention research project, we are developing a website that utilizes social content collaboration mediums in conjunction with open-source technologies to create a community-driven resource that provides users with tailored health information.

Functional Grouping of Osteoclast Genes Revealed Through Microarray Analysis

Biochemical and Biophysical Research Communications. Feb, 2008  |  Pubmed ID: 18060857

We describe for the first time functional clusters of genes that are modulated during the differentiation of osteoclasts. Pathway analysis was applied to gene array data generated from affymetrix chips hybridized to RNA isolated from RAW264.7 cells exposed to RANK-ligand (RANK-L) for 5 days. This analysis revealed major functional gene clusters that were either up- or down-regulated during osteoclastogenesis. Some of the genes within the clusters have known functions, while others do not. We discuss herein the relevance of these functional gene clusters and their modulation to biological processes underlying the formation, function, and fate of osteoclasts.

Role of GABA Receptors in Nitric Oxide Inhibition of Dorsolateral Periaqueductal Gray Neurons

Neuropharmacology. Mar, 2008  |  Pubmed ID: 18222497

Nitric oxide (NO) affects neuronal activity of the midbrain periaqueductal gray (PAG). The purpose of this report was to investigate the role of GABA receptors in NO modulation of neuronal activity through inhibitory and excitatory synaptic inputs within the dorsolateral PAG (dl-PAG). First, spontaneous miniature inhibitory postsynaptic currents (mIPSCs) and excitatory postsynaptic currents (mEPSCs) were recorded using whole cell voltage-clamp methods. Increased NO by either S-nitroso-N-acetyl-penicillamine (SNAP, 100 microM) or L-arginine (50 microM) significantly augmented the frequency of mIPSCs of the dl-PAG neurons without altering their amplitudes or decay time constants. The effects were eliminated after bath application of carboxy-PTIO (NO scavenger), and 1-(2-trifluorom-ethylphenyl) imidazole (NO synthase inhibitor). In contrast, SNAP and L-arginine did not alter mEPSCs in dl-PAG neurons. However the frequency of mEPSCs was significantly increased with prior application of the GABA(B) receptors antagonist, CGP55845. In addition, NO significantly decreased the discharge rate of spontaneous action potentials in the dl-PAG neurons and the effect was reduced in the presence of the GABA(A) receptor antagonist, bicuculline. Our data show that within the dl-PAG NO potentiates the synaptic release of GABA, while NO-induced GABA presynaptically inhibits glutamate release through GABA(B) receptors. Overall, NO suppresses neuronal activity of the dl-PAG via a potentiation of GABAergic synaptic inputs and via GABA(A) receptors.

Eimeria Tenella: Construction of a Recombinant Fowlpox Virus Expressing Rhomboid Gene and Its Protective Efficacy Against Homologous Infection

Experimental Parasitology. May, 2008  |  Pubmed ID: 18261729

A recombinant fowlpox virus (rFPV) expressing the Eimeria tenella rhomboid gene was constructed and its protective efficacy against homologous infection in chickens determined. Three-day-old-specific pathogen free (SPF) chickens were immunized s.c. with 10(2) plaque forming units (PFU), 10(4) PFU, or 10(6) PFU of rFPV-rhomboid, and challenged with 5x10(4) homologous sporulated oocysts 14 days post-immunization (p.i.). The specific antibody response and lymphocyte proliferation were measured 1, 2, 3 and 4 weeks p.i. Oocyst output, body weight gains and lesion scores were measured to evaluate the protective effects of immunization. rFPV-rhomboid elicited a specific humoral immune response and stimulated proliferation of peripheral blood lymphocytes. The lesion scores in groups vaccinated with rFPV-rhomboid were significantly higher than in other groups. At the same time, rFPV-rhomboid improved body weight significantly compared with other groups. Immunization with rFPV-rhomboid reduced oocyst shedding significantly, resulting in a protection rate of 39.6%, 41.1% or 41.7% given a dose of 10(2) PFU, 10(4) PFU, or 10(6) PFU of rFPV-rhomboid, respectively. These results indicated that rFPV can induce immune responses and offer partial protection of chickens against E. tenella challenge.

20-HETE Increases Renal Sympathetic Nerve Activity Via Activation of Chemically and Mechanically Sensitive Muscle Afferents

The Journal of Physiology. May, 2008  |  Pubmed ID: 18372304

Arachidonic acid and its metabolites produced via cyclooxygenase (COX) and lipoxygenase pathways have been reported to contribute to the cardiovascular reflexes evoked by stimulating thin fibre muscle afferents during muscle contraction. 20-Hydroxyeicosatetraenoic acid (20-HETE), a primarily metabolized product of arachidonic acid by cytochrome P450 enzymes, can be accumulated in contracting muscles. Thus, the purpose of this study was to determine the role of 20-HETE in modulating the reflex sympathetic responses to activation of chemically and mechanically sensitive muscle afferents. The renal sympathetic nerve activity (RSNA) and cardiovascular responses were examined after injections of 20-HETE into the arterial blood supply of the hindlimb muscles of decerebrated rats. This induced a dose-dependent increases in RSNA and mean arterial pressure (MAP). We also tested the hypothesis that 20-HETE would sensitize muscle afferents and, thereby, augment the RSNA and blood pressure response to muscle stretch. The results show that arterial infusion of 20-HETE significantly enhanced the RSNA and MAP responses to muscle stretch. In contrast, N-hydroxy-N'-(4-butyl-2-methylphenyl)formamidine, a potent inhibitor of 20-HETE production, attenuated the reflex muscle responses. Furthermore, the sensitizing effect of 20-HETE on the muscle reflex was significantly attenuated after blocking COX activity with indomethacin. Our data suggest that 20-HETE plays a role in modulating muscle afferent-mediated sympathetic responses, probably through engagement of a COX-dependent mechanism.

Stable Expression of Green Fluorescent Protein Mediated by GCV in Giardia Canis

Parasitology International. Sep, 2008  |  Pubmed ID: 18378489

Giardia canis can be infected with a double-stranded RNA virus, that is giardiavirus (G. canis virus, GCV). In this study, green fluorescent protein (GFP) was stably expressed in G. canis mediated by GCV. The plasmid pNEO/GDH/MCS/GFP, containing the neomycin phosphotransferase (NEO) encoding region flanked by the 636 nt of 5'-terminus and the 2174 nt of 3'-terminus from GCV positive strand RNA, was constructed by inserting GFP gene into downstream from the NEO gene and glutamate dehydrogenase (GDH) 5'-terminus uncoding regions on a single plasmid, and its in vitro transcript was introduced into GCV-infected G. canis by electroporation. The transfectants expressed GFP persistently under G418 selection. This stable transfection system should provide a valuable tool for genetic study of G. canis.

Purinergic P2X Receptors Presynaptically Increase Glutamatergic Synaptic Transmission in Dorsolateral Periaqueductal Gray

Brain Research. May, 2008  |  Pubmed ID: 18395189

Purinergic P2X receptors have been reported to be present in regions of the midbrain periaqueductal gray (PAG). The purpose of this study was to determine the role of presynaptic P2X receptors in modulating excitatory and inhibitory synaptic inputs to the dorsolateral PAG (dl-PAG), which has abundant neuronal connections. First, whole cell voltage-clamp recording was performed to obtain excitatory and inhibitory postsynaptic currents (EPSCs and IPSCs) of the dl-PAG neurons. Our data show that alpha, beta-methylene ATP (a P2X receptor agonist), in the concentration of 50 microM, significantly increased the frequency of miniature EPSCs without altering the amplitude of miniature EPSCs in eight tested neurons. The effects were attenuated by PPADS, an antagonist to P2X receptors. Furthermore, alpha, beta-methylene ATP increased the amplitude of evoked EPSCs, and decreased the paired-pulse ratio of eEPSCs in ten neurons. In contrast, activation of P2X had no distinct effect on IPSCs. In addition, immunofluorescent methods demonstrate that P2X labeling was co-localized with a presynaptic marker, synaptophysin, in the dl-PAG. The results of the current study provide the first evidence indicating that P2X receptors facilitate glutamatergic synaptic transmission in the dl-PAG via presynaptic mechanisms.

Differential Responses of Sensory Neurones Innervating Glycolytic and Oxidative Muscle to Protons and Capsaicin

The Journal of Physiology. Jul, 2008  |  Pubmed ID: 18450773

Activation of thin fibre muscle afferent nerves by metabolic by-products plays a critical role in the initiation and maintenance of the autonomic response to exercise and the metabolic profile of active muscle can influence the response. The purpose of this report was to determine the responsiveness of sensory neurones innervating muscles comprising predominantly glycolytic and oxidative fibres to protons and capsaicin using whole-cell patch clamp methods. Dorsal root ganglion (DRG) neurones from 4- to 6-week-old rats were labelled by injecting the fluorescence tracer DiI into the muscle 3-5 days prior to the recording experiments. The percentage of the DRG neurones innervating glycolytic and oxidative muscle was similar in response to both protons and capsaicin. However, the neurones innervating glycolytic muscle had greater inward current amplitude responses to protons and capsaicin as compared with oxidative muscle. The peak current amplitudes to pH 6.0 were 0.84 +/- 0.06 nA (oxidative muscle) versus 1.36 +/- 0.07 nA (glycolytic muscle, P < 0.05). The capsaicin-induced current amplitudes were 2.3 +/- 0.15 nA (oxidative muscle) versus 3.1 +/- 0.21 nA (glycolytic muscle, P < 0.05). Of neurones that responded to pH 6.0 with a sustained current, 88% also responded to capsaicin. Capsaicin exposure enhanced the proton responsiveness in the neurones innervating the muscle; and protons also increased the capsaicin response. These data suggest that (1) receptors mediating protons and capsaicin responses coexist in the DRG neurones innervating muscle; (2) the responsiveness of acidosis and capsaicin can be sensitized by each other; and (3) DRG neurones with nerve endings in a glycolytic muscle developed greater inward current responses to protons and capsaicin than did those with nerve endings in an oxidative muscle.

A Brief HIV Stigma Reduction Intervention for Service Providers in China

AIDS Patient Care and STDs. Jun, 2008  |  Pubmed ID: 18462076

This study assessed the effect of a brief intervention aimed at reducing HIV-related stigma among service providers in China. From December 2005 to June 2006, 138 service providers from four county hospitals in the Yunnan province of China were randomly assigned into either an intervention or a control condition. HIV stigma reduction concepts were conveyed through participatory small group activities, including role-plays, games, group discussions, and testimony by an HIV advocate. Participants were assessed at baseline before the intervention, and at 3- and 6-month follow-ups. Data were analyzed using a logistic regression mixed-effects model. Service providers in the brief intervention condition were significantly more likely to report better protection of patients' confidentiality and right to HIV testing, lower levels of negative feelings toward people living with HIV/AIDS, and more accurate understanding and practice of universal precautions. This brief intervention pilot showed potential in reducing HIV stigma and discrimination among service providers in China. Further intervention trials are needed to test the efficacy and long-term outcomes of this intervention.

Genome-wide Linkage Analysis of Heroin Dependence in Han Chinese: Results from Wave Two of a Multi-stage Study

Drug and Alcohol Dependence. Nov, 2008  |  Pubmed ID: 18538955

Previously we reported the results of Wave One of a genome-wide search for heroin dependence susceptibility loci in Han Chinese families from Yunnan Province, China, near Asia's "Golden Triangle". Our initial analysis of 194 independent affected sibling-pairs from 192 families identified two regions with nonparametric linkage (NPL) Z-scores greater than 2.0, which were suggestive of linkage. Presently we have supplemented our sample with additional individuals and families, bringing the total number of genotyped individuals to 1513 and the number of independent sibling-pairs to 397. Upon repeating our analyses with this larger sample, we found that the evidence for linkage at our most strongly implicated locus from Wave One (marker D17S1880; 53.4cM on 17q11.2; NPL Z=2.36; uncorrected p=0.009) was completely abolished (Z=-1.13; p=0.900). In contrast, the evidence for linkage at the second-most strongly implicated locus from Wave One (D4S1644; 143.3cM on 4q31.21; NPL Z=2.19; uncorrected p=0.014) increased in its magnitude and significance (Z=2.64; uncorrected p=0.004), becoming the most strongly implicated locus overall in our full sample. Other loci on chromosomes 1, 2, 4, 12, 16, and X also displayed nominally significant evidence for linkage (p< or =0.05). These loci appear to be entirely distinct from opioid-linked loci reported by other groups; however, meta-analyses of all available linkage data may reveal common sites of interest and promising candidate genes that can be further evaluated as risk factors for the illness.

Interstitial Adenosine Triphosphate Modulates Muscle Afferent Nerve-mediated Pressor Reflex

Muscle & Nerve. Aug, 2008  |  Pubmed ID: 18570238

Previous work has shown that muscle contraction elevates interstitial adenosine triphosphate concentration ([ATP]i), which is likely due to the release of ATP from active skeletal muscle. ATP activation of purinergic receptors P2X on thin muscle afferent fibers further enhances cardiovascular responses to contraction. Thus, the purposes of this study were: (1) to examine the mechanisms by which ATP is released from muscle in response to mechanical stimulation; and (2) to study the effects of interstitial ATP concentrations on modulating pressor response to muscle contraction. Static contraction of the triceps surae muscle was evoked by electrical stimulation (at 5 HZ and 2.5 times motor threshold) of the tibial nerve in 9 anesthetized cats. Muscle interstitial ATP samples were collected from microdialysis probes inserted into the muscles. Dialysate ATP concentrations were determined using the luciferin-luciferase assay. In a control experiment, contraction was induced after 0.5 ml of saline was injected into the arterial blood supply of the hindlimb muscles. This increased [ATP]i by 220% (P < 0.05 vs. baseline). After gadolinium (1 mM), a blocker of mechanically sensitive channels, was injected into the muscles, contraction increased [ATP]i by 112% (P < 0.05 vs. control). In contrast, glibenclamide (an inhibitor of the ATP-binding cassette protein), monensin, and brefeldin A, which interfere with vesicular formation (or trafficking) and inhibit exocytosis, did not significantly affect ATP release by muscle contraction. In addition, a regression analysis showed that [ATP]i was linearly related to the pressor response to muscle contraction. The data suggest that ATP release from skeletal muscle is mediated via involvement of mechanosensitive channels. These findings further support a physiological role for release of ATP in modulating cardiovascular responses during static muscle contraction.

A Preliminary Study of Methamphetamine Use in Yunnan, China

AIDS Patient Care and STDs. Jul, 2008  |  Pubmed ID: 18601581

Femoral Artery Occlusion Augments TRPV1-mediated Sympathetic Responsiveness

American Journal of Physiology. Heart and Circulatory Physiology. Sep, 2008  |  Pubmed ID: 18660449

Muscle metabolic by-products stimulate thin fiber muscle afferent nerves and evoke reflex increases in blood pressure and sympathetic nerve activity. Previous studies reported that chemically sensitive transient receptor potential vanilloid type 1 (TRPV1) channels present on sensory muscle afferent neurons have an important impact on sympathetically mediated cardiovascular responses. The reflex-mediated reduction in blood flow to skeletal muscle leads to limited exercise capacity in patients with peripheral arterial occlusive disease. Thus, in this study, we tested the hypothesis that the expression of enhanced TRPV1 receptor and its responsiveness in primary afferent neurons innervating muscles initiate exaggerated reflex sympathetic responses after vascular insufficiency to the muscle. Muscle vascular insufficiency was induced by the femoral artery ligation in rats for 24 h. Our data show that 1) the ligation surgery leads to the upregulation of TRPV1 expression in the dorsal root ganglion; 2) the magnitude of the dorsal root ganglion neuron TRPV1 response induced by capsaicin is greater in vascular insufficiency (4.0 +/- 0.31 nA, P < 0.05 vs. sham-operated control) than that in sham-operated control (2.9 +/- 0.23 nA); and 3) renal sympathetic nerve activity and mean arterial pressure responses to capsaicin (0.5 microg/kg body wt) are also enhanced by vascular insufficiency (54 +/- 11%, 9 +/- 2 mmHg in sham-operated controls vs. 98 +/- 13%, 33 +/- 5 mmHg after vascular insufficiency, P < 0.05). In conclusion, sympathetic nerve responses to the activation of metabolite-sensitive TRPV1 receptors are augmented in rats with the femoral artery occlusion compared with sham-operated control animals, due to alterations in the expression of TRPV1 receptor and its responsiveness in sensory neurons.

Determination of Huperzine A in Human Plasma by Liquid Chromatography-electrospray Tandem Mass Spectrometry: Application to a Bioequivalence Study on Chinese Volunteers

Biomedical Chromatography : BMC. Apr, 2008  |  Pubmed ID: 17939152

A simple, sensitive and selective LC-MS-MS method has been developed for the quantification of huperzine A in human plasma. Huperzine A and pseudoephedrine hydrochloride (internal standard) were isolated from human plasma by extraction with ethyl acetate, chromatographed on a C(18) column with a mobile phase consisting of 0.2% formic acid-methanol (15:85, v/v) and detected using a tandem mass spectrometer with an electrospray ionization interface. The lower limit of quantification was 0.0508 ng/mL, and the assay exhibited a linear range of 0.0508-5.08 ng/mL (r = 0.9998). The method was successfully applied to investigate the bioequivalence between two kinds of tablets (test vs reference product) in 18 healthy male Chinese volunteers. After a single 0.2 mg dose for the test and reference product, the resulting means of major pharmacokinetic parameters such as AUC(0-24), AUC(0-infinity), C(max), T(max) and t(1/2) of huperzine A were 16.35 +/- 3.42 vs 16.38 +/- 3.61 ng h/mL, 17.53 +/- 3.80 vs 17.70 +/- 3.97 ng h/mL, 2.47 +/- 0.49 vs 2.51 +/- 0.51 ng/mL, 1.3 +/- 0.4 vs 1.2 +/- 0.3 h and 5.92 +/- 0.75 vs 6.18 +/- 0.66 h, respectively, indicating that these two kinds of tablets were bioequivalent.

Sympathetic Nerve Responses to Muscle Contraction and Stretch in Ischemic Heart Failure

American Journal of Physiology. Heart and Circulatory Physiology. Jan, 2008  |  Pubmed ID: 17965282

Congestive heart failure (CHF) induces abnormal regulation of peripheral blood flow during exercise. Previous studies have suggested that a reflex from contracting muscle is disordered in this disease. However, there has been very little investigation of the muscle reflex regulating sympathetic outflows in CHF. Myocardial infarction (MI) was induced by the coronary artery ligation in rats. Echocardiography was performed to determine fractional shortening (FS), an index of the left ventricular function. We examined renal and lumbar sympathetic nerve activities (RSNA and LSNA, respectively) during 1-min repetitive (1- to 4-s stimulation to relaxation) contraction or stretch of the triceps surae muscles. During these interventions, the RSNA and LSNA responded synchronously as tension was developed. The RSNA and LSNA responses to contraction were significantly greater in MI rats (n = 13) with FS <30% than in control animals (n = 13) with FS >40% (RSNA: +49 +/- 7 vs. +19 +/- 4 a.u., P < 0.01; LSNA: +28 +/- 7 vs. +8 +/- 2 a.u., P < 0.01) at the same tension development. Stretch also increased the RSNA and LSNA to a larger degree in MI (n = 13) than in control animals (n = 13) (RSNA: +36 +/- 6 vs. +19 +/- 3 a.u., P < 0.05; LSNA: +24 +/- 3 vs. +9 +/- 2 a.u., P < 0.01). The data demonstrate that CHF exaggerates sympathetic nerve responses to muscle contraction as well as stretch. We suggest that muscle afferent-mediated sympathetic outflows contribute to the abnormal regulation of peripheral blood flow seen during exercise in CHF.

[Determination of Benomyl, Carbendazim and Thiabendazole in Apple Juice Concentrate Using Solid-phase Extraction Coupled with Ion Exchange Chromatography]

Se Pu = Chinese Journal of Chromatography / Zhongguo Hua Xue Hui. Sep, 2008  |  Pubmed ID: 19160754

A method was developed for the determination of benomyl, carbendazim and thiabendazole in apple juice concentrate by solid-phase extraction coupled with ion exchange chromatography (IEC). The sample was diluted with water, and then benomyl was degradated completely to carbendazim at 80 degrees C, and purified by an SCX solid-phase extraction column. Liquid chromatographic analysis was performed on the instrument of Agilent 1200 series equipped with a diode-array detector and autosampler. The column was LC-SCX (25 cm x 4.6 mm, 5 microm). The mobile phase was 0.1 mol/L KH2PO4 (pH 2.5)-acetonitrile (70:30, v/v) with a flow rate of 1.0 mL/min. The presented method showed good linear relationship with good precision and accuracy at the range of 0.02 - 2.0 mg/L for carbendazim and thiabendazole. The detection limits were 0. 004 mg/kg for carbendazim and thiabendazole. The method was characterized with acceptable sensitivity to meet the requirements for monitoring these pesticides in apple juice concentrate.

Caging a Beast in the Inflammation Arena: Use of Chinese Medicinal Herbs to Inhibit a Late Mediator of Lethal Sepsis, HMGB1

International Journal of Clinical and Experimental Medicine. 2008  |  Pubmed ID: 19079688

Sepsis refers to a systemic inflammatory response syndrome resulting from a microbial infection, which kills > 225,000 people annually in the U.S. alone. The high mortality of sepsis is partly mediated by bacterial endotoxin, which stimulates macrophages/monocytes to sequentially release early (e.g., TNF) and late (e.g., HMGB1) pro-inflammatory cytokines. Although early proinflammatory cytokines may be protective against infection, excessive accumulation of late-acting proinflammatory mediators (such as HMGB1) may sustain a potentially injurious inflammatory response. Agents capable of inhibiting HMGB1 activities (e.g., neutralizing antibodies) or release [e.g., Chinese herbs, Danggui (Angelica sinensis), Danshen (Salvia miltiorrhiza) and Green tea (Camellia sinensis)] rescue mice from lethal sepsis even when given 24 hours after onset of the disease. Here we review emerging evidence that support a critical role for extracellular HMGB1 as a late mediator of lethal sepsis, and several commonly used Chinese herbs (Danggui, Danshen and Green tea) as potential HMGB1- targeting therapeutic agents in experimental sepsis.

Protective Immunity of Recombinant Mycobacterium Bovis BCG Expressing Rhomboid Gene Against Eimeria Tenella Challenge

Veterinary Parasitology. Mar, 2009  |  Pubmed ID: 19117681

Two recombinant Mycobacterium bovis BCG (rBCG) strains carrying the Eimeria tenella rhomboid gene (Rho) delivered by extrachromosomal vector pMV261 and integrative vector pMV361 were evaluated for their ability to protect chickens against E. tenella challenge. The chickens were immunized intranasal with BCG, rBCG pMV261-Rho, or rBCG pMV361-Rho twice at a 2-week interval. All the recombinant BCG immunized chickens developed specific immune responses, and there was a significant increases of the percentages of CD4(+) and CD8(+) cells compared to the control (P<0.05). Challenge experiments demonstrated that the two rBCG strains could provide significant protection against E. tenella challenge. But vaccination with rBCG pMV261-Rho induced higher specific antibody titers and produced greater protection rate (56.04%) than rBCG pMV361-Rho group (P<0.05). These results indicated that M. bovis BCG is a novel vaccine vector to express and present antigens of E. tenella, and rBCG has a potential as vaccine in chickens.

Spinal P2X Receptor Modulates Muscle Pressor Reflex Via Glutamate

Journal of Applied Physiology (Bethesda, Md. : 1985). Mar, 2009  |  Pubmed ID: 19131479

Static contraction of skeletal muscle evokes reflex increases in blood pressure and heart rate. Previous studies showed that P2X receptors located at the dorsal horn of the spinal cord play a role in modulating the muscle pressor reflex. P2X stimulation can alter release of the excitatory amino acid, glutamate (Glu). In this report, we tested the hypothesis that stimulation of P2X receptors enhances the concentrations of Glu ([Glu]) in the dorsal horn, and that blocking P2X receptors attenuates contraction-induced Glu increases and the resultant reflex pressor response. Contraction was elicited by electrical stimulation of the L(7) and S(1) ventral roots of 14 cats. Glu samples were collected from microdialysis probes inserted in the L(7) level of the dorsal horn of the spinal cord, and dialysate [Glu] was determined using the HPLC method. First, microdialyzing alpha,beta-methylene ATP (0.4 mM) into the dorsal horn significantly increased [Glu]. In addition, contraction elevated [Glu] from baseline of 536 +/- 53 to 1,179 +/- 192 nM (P < 0.05 vs. baseline), and mean arterial pressure by 39 +/- 8 mmHg in the control experiment. Microdialyzing the P2X receptor antagonist pyridoxal phosphate-6-azophenyl-2',4'-disulfonic acid (10 mM) into the dorsal horn attenuated the contraction induced-Glu increase (610 +/- 128 to 759 +/- 147 nM; P > 0.05) and pressor response (16 +/- 3 mmHg, P < 0.05 vs. control). Our findings demonstrate that P2X modulates the cardiovascular responses to static muscle contraction by affecting the release of Glu in the dorsal horn of the spinal cord.

Ultra-thin Microporous-mesoporous Metal Oxide Films Prepared by Molecular Layer Deposition (MLD)

Chemical Communications (Cambridge, England). Dec, 2009  |  Pubmed ID: 19921009

Porous aluminium oxide films with precisely controlled thickness down to several angstroms are deposited on particle surfaces from dense aluminium alkoxide hybrid polymer films by molecular layer deposition. Porous structures are obtained by either mild water etching at room temperature or calcination in air at elevated temperatures.

Web-based Hazard and Near-miss Reporting As Part of a Patient Safety Curriculum

The Journal of Nursing Education. Dec, 2009  |  Pubmed ID: 20000248

As part of a patient safety curriculum, we developed a Web-based hazard and near-miss reporting system for postbaccalaureate nursing students to use during their clinical experiences in the first year of their combined BS-MS advanced practice nurse program. The 25-week clinical rotations included 2 days per week for 5 weeks each in community, medical-surgical, obstetrics, pediatrics, and psychiatric settings. During a 3-year period, 453 students made 21,276 reports. Of the 10,206 positive (yes) responses to a hazard or near miss, 6,005 hazards (59%) and 4,200 near misses (41%) were reported. The most common reports were related to infection, medication, environmental, fall, and equipment issues. Of the near misses, 1,996 (48%) had planned interceptions and 2,240 (52%) had unplanned interceptions. Types of hazards and near misses varied by rotation. Incorporating hazard and near-miss reporting into the patient safety curriculum was an innovative strategy to promote mindfulness among nursing students.

Upper and Lower Tear Menisci in the Diagnosis of Dry Eye

Investigative Ophthalmology & Visual Science. Jun, 2009  |  Pubmed ID: 19218609

To measure the upper and lower tear menisci in patients with aqueous tear deficiency (ATD) dry eye by real-time optical coherence tomography (OCT) and to determine the most effective meniscus variables for the diagnosis of dry eye.

Lower Volumes of Tear Menisci in Contact Lens Wearers with Dry Eye Symptoms

Investigative Ophthalmology & Visual Science. Jul, 2009  |  Pubmed ID: 19279314

To investigate tear meniscus volumes during short-term lens wear by soft contact lens (SCL) wearers with dryness symptoms.

Contribution of Nerve Growth Factor to Augmented TRPV1 Responses of Muscle Sensory Neurons by Femoral Artery Occlusion

American Journal of Physiology. Heart and Circulatory Physiology. May, 2009  |  Pubmed ID: 19286963

In rats, hindlimb muscle ischemia induced by femoral artery occlusion augments the sympathetic nervous response to stimulation of transient receptor potential vanilloid type 1 (TRPV1) by injection of capsaicin into the arterial blood supply of the hindlimb muscles. The enhanced sympathetic response is due to alterations in TRPV1 receptor expression and its responsiveness in sensory neurons. The underlying mechanism by which TRPV1 receptor responses are increased after muscle vascular insufficiency/ischemia is unclear. In this report we tested the hypothesis that muscle ischemia elevates nerve growth factor (NGF) levels in primary afferent neurons, thereby increasing TRPV1 responsiveness. Muscle vascular insufficiency induced by the femoral artery ligation significantly increased NGF in the dorsal root ganglion (DRG) compared with sham controls. Furthermore, when NGF was infused in the hindlimb muscles of healthy rats (72 h using an osmotic minipump), the magnitude of the DRG neuron response to capsaicin was augmented (5.4 +/- 0.54 nA with NGF infusion vs. 3.0 +/- 0.17 nA in control; P < 0.05). With the addition of NGF in the culture dish containing the DRG neurons, the magnitude of the DRG neuron response to capsaicin was greater (6.4 +/- 0.27 nA; P < 0.05 vs. control) than that seen in control (2.9 +/- 0.16 nA). Note that this NGF effect was seen in isolectin B(4)-negative DRG neurons, a group of thin fiber nerves that contain neuropeptides and depend on NGF for survival. These data suggest that NGF affects a selective subpopulation of the afferent neurons in mediating augmented TRPV1 responses after femoral artery occlusion.

Construction of EGFP-tagged RBCG of E.tenella and Distribution in Chickens

Science in China. Series C, Life Sciences / Chinese Academy of Sciences. Mar, 2009  |  Pubmed ID: 19294353

Chicken coccidiosis is a major parasitic disease with substantial economic burden to the poultry industry. Enhanced green fluorescent protein (EGFP) tagged recombinant Bacille Calmette-Guerin (rBCG), as a fusion protein with coccidian rhomboid antigen was constructed to track rBCG in vivo in chickens in this study. Immunization of chickens with one dose of rBCG pMV361-Rho/EGFP induced humoral immune response. The colonization of rBCG in liver, spleen, lung, kidney and caecum was observed by laser confocal microscopy. Real-time quantitative RT-PCR showed a rise expression level of rhomboid protein on the 7th day and a peak on the 14th day and disappearance on the 28th day after immunization. These results have significant implications for the development of rBCG vaccines against avian coccidiosis.

Oxytocin is an Anabolic Bone Hormone

Proceedings of the National Academy of Sciences of the United States of America. Apr, 2009  |  Pubmed ID: 19369205

We report that oxytocin (OT), a primitive neurohypophyseal hormone, hitherto thought solely to modulate lactation and social bonding, is a direct regulator of bone mass. Deletion of OT or the OT receptor (Oxtr) in male or female mice causes osteoporosis resulting from reduced bone formation. Consistent with low bone formation, OT stimulates the differentiation of osteoblasts to a mineralizing phenotype by causing the up-regulation of BMP-2, which in turn controls Schnurri-2 and 3, Osterix, and ATF-4 expression. In contrast, OT has dual effects on the osteoclast. It stimulates osteoclast formation both directly, by activating NF-kappaB and MAP kinase signaling, and indirectly through the up-regulation of RANK-L. On the other hand, OT inhibits bone resorption by mature osteoclasts by triggering cytosolic Ca(2+) release and NO synthesis. Together, the complementary genetic and pharmacologic approaches reveal OT as a novel anabolic regulator of bone mass, with potential implications for osteoporosis therapy.

Atomic Layer Deposition of Quantum-confined ZnO Nanostructures

Nanotechnology. May, 2009  |  Pubmed ID: 19420639

The modulation of optoelectronic properties, such as the bandgap of a pure-component semiconductor material, is a useful ability that can be achieved by few techniques. Atomic layer deposition (ALD) was used here to experimentally demonstrate the ability to deposit films that exhibit quantum confinement on three-dimensional surfaces. Polycrystalline ZnO films ranging from approximately 1.5 to 15 nm in thickness were deposited via ALD using diethylzinc and hydrogen peroxide at 100 degrees C. Conformal, pinhole-free films were deposited on Si wafers and on nanosized spherical SiO(2) particles using an augmented central composite design strategy. Powder x-ray diffraction was used to measure the crystallite size of the films and monitor size evolution on the basis of the number of ALD cycles and thermal annealing post-treatments. The absorbance of the ZnO films on Si wafers and SiO(2) particles was measured using spectroscopic ellipsometry and diffuse transmittance techniques, respectively. Post-deposition annealing steps increased the crystallite size of the films, independently of the coating thickness. The ZnO bandgap was increasingly blue-shifted for films of decreasing crystallite size, approaching +0.3 eV at dimensions of 2-3 nm. The nonlinear bandgap response correlated well with the Brus model. This work represents an experimental demonstration of quantum confinement using ALD on two- and three-dimensional substrates.

Angiotensin II Inhibits GABAergic Synaptic Transmission in Dorsolateral Periaqueductal Gray Neurons

Neuroscience Letters. May, 2009  |  Pubmed ID: 19429096

The purpose of this study was to determine the role of angiotensin II (Ang II) in modulating inhibitory and excitatory synaptic inputs to the dorsolateral periaqueductal gray (dl-PAG). The whole cell voltage-clamp recording was performed to examine inhibitory and excitatory postsynaptic currents (IPSCs and EPSCs) of the dl-PAG neurons. Ang II, at the concentration of 2microM, decreased the frequency of miniature IPSCs from 0.83+/-0.02 to 0.45+/-0.03Hz (P<0.05) in 10 tested neurons. This did not significantly affect the amplitude and decay time constant. The effect of Ang II on miniature IPSCs was blocked by the prior application of Ang II AT1 receptor antagonist losartan, but not by AT2 receptor antagonist PD123319. Additionally, Ang II decreased the amplitude of evoked IPSCs from 148+/-15 to 89+/-7pA (P<0.05), and increased the paired-pulse ratio from 96+/-5% to 125+/-7% (P<0.05) in eight tested neurons. In contrast, Ang II had no distinct effects on the EPSCs. Our data suggest that Ang II inhibits GABAergic synaptic inputs to the dl-PAG through activation of presynaptic AT1 receptors.

Neuroprotection in a 6-hydroxydopamine-lesioned Parkinson Model Using Lactoferrin-modified Nanoparticles

The Journal of Gene Medicine. Sep, 2009  |  Pubmed ID: 19554623

Nonviral gene therapy of chronic degenerative diseases such as Parkinson's disease (PD) is a great challenge as a result of the low tranfection efficiency of nonviral gene vectors. We previously constructed a lactoferrin (Lf)-modified vector, which was demonstrated to be potential for brain gene delivery both in vitro and in vivo. In the present study, this type of vector was applied to load human glial cell line-derived neurotrophic factor gene (hGDNF).

Spermine Protects Mice Against Lethal Sepsis Partly by Attenuating Surrogate Inflammatory Markers

Molecular Medicine (Cambridge, Mass.). Jul-Aug, 2009  |  Pubmed ID: 19593412

The pathogenesis of sepsis is partly attributable to dysregulated inflammatory response mediated by pathogen-associated molecular patterns (PAMPs) (for example, endotoxin) and damage-associated molecular patterns (DAMPs) (for example, high-mobility group box 1 [HMGB1]). An endogenous ubiquitous polyamine, spermine, inhibits endotoxin-induced cytokine release in vitro, but its capacities to attenuate sepsis- and HMGB1-induced inflammatory responses was previously unknown. We thus tested the hypothesis that spermine protects mice against lethal sepsis by attenuating sepsis-induced local and systemic inflammatory responses. Intraperitoneal (i.p.) administration of spermine (10 mg/kg, twice daily, for 3 d) conferred a significant protection against lethal sepsis. The protective effects were associated with a significant reduction in peritoneal and serum levels of several surrogate markers of sepsis (for example, Interleukin-6 [IL-6], keratinocyte-derived chemokine [KC], monocytes chemoattractant protein-1 [MCP-1], macrophage inflammatory protein-2 [MIP-2], tissue inhibitor of metalloproteinase-1 [TIMP-1], soluble tumor necrosis factor-alpha receptor I [sTNFRI], and soluble tumor necrosis factor-alpha receptor II [sTNFRII]) during a late stage of sepsis. In vitro, spermine effectively inhibited HMGB1-induced release of the above surrogate markers in peritoneal macrophages. Thus, spermine confers protection against lethal sepsis partly by attenuating sepsis- and HMGB1-induced inflammatory responses.

Giardia Canis: Ultrastructural Analysis of G. Canis Trophozoites Transfected with Full Length G. Canis Virus CDNA Transcripts

Experimental Parasitology. Nov, 2009  |  Pubmed ID: 19619539

Giardia canis virus (GCV) is a double-stranded RNA (dsRNA) virus of the family Totiviridae. In this study, the full length cDNA of the G. canis virus was constructed in pPoly2/sfinot vector and RNA was transcribed in vitro. Virus-free G. canis trophozoites were transfected with in vitro transcribed GCV RNA by electroporation. Transfected trophozoites were cultured for 12, 24, 36, 48, 60, or 72h post transfection for analysis. The ultrastructures of the transfected trophozoites were determined by transmission electron microscopy. The viral particles were detectable sporadically in the cytoplasm as early as 24h post transfection, but became evident and wide-spread 36h post transfection. The number of viral particles increased dramatically from 48 to 60h. Viral particles were released into the culture medium starting at about 60h and detectable in nuclei 72h post transfection. Severe vacuolization was seen in transfected G. canis trophozoites as early as 36h post transfection and persisted throughout the course of this study. The results of the present study indicate that in vitro transcribed GCV transcripts were capable of infecting Giardia trophozoites, apparently replicated and packaged into mature infectious viral particles which were released from the host.

Transient Transfection of Cryptosporidium Parvum Using Green Fluorescent Protein (GFP) As a Marker

Molecular and Biochemical Parasitology. Dec, 2009  |  Pubmed ID: 19631239

Cryptosporidium parvum is a protozoan parasite that infects a variety of mammals. The parasite has been shown to harbor a dsRNA virus (CPV) and in the present study, we have developed a CPV transient transfection system for this parasite by using green fluorescent protein (GFP) to replace the partial gene encoding region of the larger dsRNA (CPV-L) and the smaller dsRNA (CPV-S) virus. Two viral RNA-mediated transfection vectors: pCPVL-GFP and pCPVS-GFP were successfully constructed and both in vitro transcripts were electroporated into oocysts and sporozoites. Transient expression of GFP was detected in C. parvum oocysts and excysted sporozoites by fluorescence microscopy and by RT-PCR detection of GFP mRNA and antisense RNA in transfected C. parvum oocysts. Our study provides a new approach for studying gene expression and regulation in C. parvum and will hopefully lead to the construction of a stable CPV transfection system in the future.

Oxytocin Deficiency Impairs Maternal Skeletal Remodeling

Biochemical and Biophysical Research Communications. Oct, 2009  |  Pubmed ID: 19653998

We have reported that the posterior pituitary hormone, oxytocin (OT), known for its effects in inducing parturition, lactation and social bonding, is also a skeletal hormone. Here, we demonstrate that OT plays a key role in enabling maternal skeletal mobilization during pregnancy by enhancing the formation of bone resorbing osteoclasts. Osteoclast formation ex vivo is thus diminished in pregnant mothers with genetic OT-deficiency. OT(-/-) pups at day E20 also show a defect in trabecular bone. microCT measurements reveal normal bone volume, but increased trabecular numbers, suggesting that trabeculae in OT(-/-) pups are hypomineralized. We suggest that OT facilitates intergenerational transfer of calcium ions from a pregnant mother to the pups.

GetHealthyHarlem.org: Developing a Web Platform for Health Promotion and Wellness Driven by and for the Harlem Community

AMIA ... Annual Symposium Proceedings / AMIA Symposium. AMIA Symposium. 2009  |  Pubmed ID: 20351872

GetHealthyHarlem.org is a community website developed on an open-source platform to facilitate collaborative development of health content through participatory action research (PAR) principles. The website was developed to enable the Harlem community to create a shared health and wellness knowledgebase, to enable discourse about local and culturally relevant health information, and to foster social connections between community members and health promotion organizations. The site is gaining active use with more than 9,500 unique site visits in the six months since going live in November, 2008. In ongoing research studies, we are using the website to explore how the PAR model can be applied to the development of a community health website.

TLC: an Informatics Approach to Enable Patients to Initiate Tailored Lifestyle Conversations with Providers at the Point of Care

AMIA ... Annual Symposium Proceedings / AMIA Symposium. AMIA Symposium. 2009  |  Pubmed ID: 20351881

Chronic illness including cardiovascular disease (CVD) is a major burden on the healthcare system. Behavioral and lifestyle changes could significantly reduce the burden of CVD, but provider counseling for behavior change is a very challenging, and often ineffective task. We have developed a patient-centric decision support tool to be incorporated into an Electronic Health Record system (EHR). The tool provides tailored feedback on behavioral risk, readiness and confidence in an effort to empower patients to make decisions about improving health behaviors. In turn, the tool will facilitate an informed and balanced discussion between patients and their providers about behavioral changes, incorporating both the clinical view and the individual's preferences for choosing among multiple behavior change goals based on their psychosocial characteristics, and evaluation of benefits and barriers.

Cryopreservation Has No Effect on Meiotic Recombination and Synapsis in Testicular Tissues

Fertility and Sterility. Apr, 2009  |  Pubmed ID: 18710708

The effects of cryopreservation on meiotic progression, synapsis, recombination, and structure of synaptonemal complexes (SCs) in testicular tissues were evaluated by comparing the above-mentioned parameters in frozen and fresh testicular tissues from the same men. No differences in meiotic progression, the mean number of MLH1 foci per cell, the mean number of autosomal SCs with different numbers of MLH1 foci, or the fidelity of the synapsis were observed between fresh and frozen testicular tissues.

Minimally Invasive Perventricular Device Closure of Perimembranous Ventricular Septal Defect Without Cardiopulmonary Bypass: Multicenter Experience and Mid-term Follow-up

The Journal of Thoracic and Cardiovascular Surgery. Jun, 2010  |  Pubmed ID: 20363483

To summarize the clinical experiences and mid-term follow-up results of perventricular closure of perimembranous ventricular septal defect without cardiopulmonary bypass under transesophageal echocardiography guidance.

Inhibition of Hepatitis B Virus Replication by MyD88 Involves Accelerated Degradation of Pregenomic RNA and Nuclear Retention of Pre-S/S RNAs

Journal of Virology. Jul, 2010  |  Pubmed ID: 20410269

Myeloid differentiation primary response protein 88 (MyD88), which can be induced by alpha interferon (IFN-alpha), has an antiviral activity against the hepatitis B virus (HBV). The mechanism of this antiviral activity remains poorly understood. Here, we report that MyD88 inhibited HBV replication in HepG2.2.15 cells and in a mouse model. The knockdown of MyD88 expression weakened the IFN-alpha-induced inhibition of HBV replication. Furthermore, MyD88 posttranscriptionally reduced the levels of viral RNA. Remarkably, MyD88 accelerated the decay of viral pregenomic RNA in the cytoplasm. Mapping analysis showed that the RNA sequence located in the 5'-proximal region of the pregenomic RNA was critical for the decay. In addition, MyD88 inhibited the nuclear export of pre-S/S RNAs via the posttranscriptional regulatory element (PRE). The retained pre-S/S RNAs were shown to degrade in the nucleus. Finally, we found that MyD88 inhibited the expression of polypyrimidine tract-binding protein (PTB), a key nuclear export factor for PRE-containing RNA. Taken together, our results define a novel antiviral mechanism against HBV mediated by MyD88.

Congenital Left Ventricular Diverticulum Manifested As T-wave Inversion in a Child

Pediatric Cardiology. Aug, 2010  |  Pubmed ID: 20411250

Congenital diverticulum of the ventricle is a rare disease, but most cases of congenital left ventricular diverticula are asymptomatic. We present a child with congenital left ventricular diverticulum whose routine electrocardiographic examination showed T-wave inversion in inferior and V4 to V6 leads. He was successfully repaired surgically.

ACTH Protects Against Glucocorticoid-induced Osteonecrosis of Bone

Proceedings of the National Academy of Sciences of the United States of America. May, 2010  |  Pubmed ID: 20421485

We report that adrenocorticotropic hormone (ACTH) protects against osteonecrosis of the femoral head induced by depot methylprednisolone acetate (depomedrol). This therapeutic response likely arises from enhanced osteoblastic support and the stimulation of VEGF by ACTH; the latter is largely responsible for maintaining the fine vascular network that surrounds highly remodeling bone. We suggest examining the efficacy of ACTH in preventing human osteonecrosis, a devastating complication of glucocorticoid therapy.

Hepatitis C Virus NS5A Activates the Mammalian Target of Rapamycin (mTOR) Pathway, Contributing to Cell Survival by Disrupting the Interaction Between FK506-binding Protein 38 (FKBP38) and MTOR

The Journal of Biological Chemistry. Jul, 2010  |  Pubmed ID: 20439463

Hepatitis C virus (HCV) often establishes a persistent infection that most likely involves a complex host-virus interplay. We previously reported that the HCV nonstructural protein 5A (NS5A) bound to cellular protein FKBP38 and resulted in apoptosis suppression in human hepatoma cell line Huh7. In the present research we further found that NS5A increased phosphorylation levels of two mTOR-targeted substrates, S6K1 and 4EBP1, in Huh7 in the absence of serum. mTOR inhibitor rapamycin or NS5A knockdown blocked S6K1 and 4EBP1 phosphorylation increase in NS5A-Huh7 and HCV replicon cells, suggesting that NS5A specifically regulated mTOR activation. Overexpression of NS5A and FKBP38 mutants or FKBP38 knockdown revealed this mTOR activation was dependent on NS5A-FKBP38 interaction. Phosphatidylinositol 3-kinase (PI3K) inhibitor LY294002 treatment in NS5A-Huh7 showed that the mTOR activation was independent of PI3K. Moreover, NS5A suppressed caspase 3 and poly(ADP-ribose) polymerase activation, which was abolished by NS5A knockdown or rapamycin, indicating NS5A inhibited apoptosis specifically through the mTOR pathway. Further analyses suggested that apoptotic inhibition exerted by NS5A via mTOR also required NS5A-FKBP38 interaction. Glutathione S-transferase pulldown and co-immunoprecipitation showed that NS5A disrupted the mTOR-FKBP38 association. Additionally, NS5A or FKBP38 mutants recovered the mTOR-FKBP38 interaction; this indicated that the impairment of mTOR-FKBP38 association was dependent on NS5A-FKBP38 binding. Collectively, our data demonstrate that HCV NS5A activates the mTOR pathway to inhibit apoptosis through impairing the interaction between mTOR and FKBP38, which may represent a pivotal mechanism for HCV persistence and pathogenesis.

Lentivirus-mediated Knockdown of Cyclin Y (CCNY) Inhibits Glioma Cell Proliferation

Oncology Research. 2010  |  Pubmed ID: 20441050

Cyclin Y (CCNY) is a novel cyclin and almost nothing is known about its expression level and role in human cancers. In the present study, we found that lentivirus-mediated RNA interference (RNAi) of CCNY significantly downregulated its expression level. More importantly, knockdown of CCNY inhibited cell proliferation, colony formation, and cell cycle progression in glioma cells. These data suggest that CCNY might play an important role in glioma tumorigenisis.

A Simple and Sensitive HPLC Method for Quantification of the Metabolin of Meclofenoxate in Human Plasma

Journal of Chromatographic Science. May-Jun, 2010  |  Pubmed ID: 20515527

A simple and sensitive high-performance liquid chromatographic method was developed for quantification of the metabolin of meclofenoxate, chlorophenoxyacetic acid, in human plasma. Ibuprofen was used as an internal standard. The present method used protein precipitation for extraction of chlorophenoxyacetic acid from human plasma. Separation was carried out on a reversed-phase C(18) column. The column effluent was monitored by UV detection at 254 nm. The mobile phase was a mixture of methanol and water containing 1.0% glacial acetic acid (70:30 v/v) at a flow rate of 1.0 mL/min. The column temperature was 20 degrees C. This method was linear over the range of 0.047-28.20 microg/mL with a regression coefficient greater than 0.99. The mean recovery of chlorophenoxyacetic acid and IS were (79.54 +/- 6.33)% and (78.48 +/- 2.14)%, respectively, and the method was found to be precise, accurate, and specific during the study. The method was successfully applied for pharmacokinetic study of chlorophenoxyacetic acid in human.

Induction of Immune Responses in Mice by a DNA Vaccine Encoding Cryptosporidium Parvum Cp12 and Cp21 and Its Effect Against Homologous Oocyst Challenge

Veterinary Parasitology. Aug, 2010  |  Pubmed ID: 20541869

Cp12 and Cp21 surface proteins on the sporozoite of Cryptosporidium parvum have been identified as the immunodominant antigens involved in the immune response to C. parvum infection. In the present study, the efficacy of Cp12 and Cp21 antigens as vaccine candidates was investigated in BALB/c mice that were susceptible to C. parvum infection. DNA sequences of Cp12, Cp21, Cp12-Cp21, and C (CpG oligodeoxynucleotide (ODN))-Cp12-Cp21 were amplified and then cloned into pVAX1 vector to form the four recombinant plasmids pVAX1-Cp12, pVAX1-Cp21, pVAX1-Cp12-Cp21, and pVAX1-C-Cp12-Cp21. Recombinant protein expression from these four plasmids in HeLa cells were confirmed by indirect immunofluorescence staining and Western blot analysis. The in vivo efficacies of the four DNA vaccines were tested in BALB/c mice. The results indicated that the four DNA vaccines elicited significant antibody responses and specific cellular responses when compared to control mice that received vector only or PBS. Among those four plasmids, pVAX1-C-Cp12-Cp21 elicited significantly higher levels of IgG. Also, the percentages of CD4(+) and CD8(+) T cells were significantly higher in the group with pVAX1-C-Cp12-Cp21 nasal sprays. Their efficacy in immunoprotection against homologous challenge was also detected after administration of the four DNA vaccines. The results showed that mice in the pVAX1-C-Cp12-Cp21 nasal group had a 77.5% reduction in the level of oocyst shedding and a significant difference was detected when this group was compared with the pVAX1, PBS, pVAX1-Cp12, and pVAX1-Cp21 groups. The reduction in the level of oocysts shedding from the group of pVAX1-C-Cp12-Cp21 nasal spray was also higher than that of pVAX1-Cp12-Cp21 group. These results suggested that C-Cp12-Cp21-DNA may provide an effective means of eliciting humoral and cellular responses and generating protective immunity against C. parvum infections in BALB/c mice.

A Critical Cysteine is Required for HMGB1 Binding to Toll-like Receptor 4 and Activation of Macrophage Cytokine Release

Proceedings of the National Academy of Sciences of the United States of America. Jun, 2010  |  Pubmed ID: 20547845

During infection, vertebrates develop "sickness syndrome," characterized by fever, anorexia, behavioral withdrawal, acute-phase protein responses, and inflammation. These pathophysiological responses are mediated by cytokines, including TNF and IL-1, released during the innate immune response to invasion. Even in the absence of infection, qualitatively similar physiological syndromes occur following sterile injury, ischemia reperfusion, crush injury, and autoimmune-mediated tissue damage. Recent advances implicate high-mobility group box 1 (HMGB1), a nuclear protein with inflammatory cytokine activities, in stimulating cytokine release. HMGB1 is passively released during cell injury and necrosis, or actively secreted during immune cell activation, positioning it at the intersection of sterile and infection-associated inflammation. To date, eight candidate receptors have been implicated in mediating the biological responses to HMGB1, but the mechanism of HMGB1-dependent cytokine release is unknown. Here we show that Toll-like receptor 4 (TLR4), a pivotal receptor for activation of innate immunity and cytokine release, is required for HMGB1-dependent activation of macrophage TNF release. Surface plasmon resonance studies indicate that HMGB1 binds specifically to TLR4, and that this binding requires a cysteine in position 106. A wholly synthetic 20-mer peptide containing cysteine 106 from within the cytokine-stimulating B box mediates TLR4-dependent activation of macrophage TNF release. Inhibition of TLR4 binding with neutralizing anti-HMGB1 mAb or by mutating cysteine 106 prevents HMGB1 activation of cytokine release. These results have implications for rationale, design, and development of experimental therapeutics for use in sterile and infectious inflammation.

Predictors of Early Dropout in Methadone Maintenance Treatment Program in Yunnan Province, China

Drug and Alcohol Review. May, 2010  |  Pubmed ID: 20565518

The aim of this study is to identify the predictors of early dropout of patients in methadone maintenance treatment (MMT) clinics in Yunnan province, China.

Neospora Caninum: in Vitro Culture of Tachyzoites in MCF-7 Human Breast Carcinoma Cells

Experimental Parasitology. Dec, 2010  |  Pubmed ID: 20566363

The development of Neospora caninum tachyzoites, an apicomplexan protozoan parasite, was studied in vitro using the human breast carcinoma cell 7 (MCF-7) as the host cell line. The extracellular NC-1 tachyzoites in MCF-7 cells were observed and counted daily for 6 consecutive days post-infection to establish the growth curve. The intracellular parasites were observed by acridine orange staining using Laser scanning confocal microscope. The results indicated that NC-1 tachyzoites invaded MCF-7 cells and multiplied intracellularly. The number of extracellular NC-1 tachyzoites started to increase rapidly around day 3 and reached the maximum number around day 4. Results from the present study suggested that MCF-7 cells were susceptible to NC-1 tachyzoites and could be used as an alternative cell line for in vitro studies.

Quantitative Metabolic Flux Analysis Revealed Uneconomical Utilization of ATP and NADPH in Acremonium Chrysogenum Fed with Soybean Oil

Bioprocess and Biosystems Engineering. Nov, 2010  |  Pubmed ID: 20571830

A metabolic network was constructed for the Acremonium chrysogenum cultivation fed with soybean oil. Metabolic flux analysis indicated that the shift from exponential growth to rapid cephalosporin C (CPC) formation was accompanied by 1.63- and 5-fold carbon flux enlargement in TCA cycle and glyoxylate by-pass, respectively. The flux via pentose phosphate pathway branch was little affected during the rapid CPC formation period; the contributory explanation was that 35.6% of NADPH was consumed in the dissimilation of fatty acids. Estimation of NADPH, ATP generation, and consumption demonstrated that, with soybean oil as carbon source in rapid CPC formation phase, the NADPH consumed in fatty acid catabolism was fourfold greater than that used in the CPC biosynthesis-relevant part; simultaneously, more than 90% energy spent was not directly related to the CPC formation. Therefore, the improvement of CPC production yield through optimization of the NADPH, ATP generation, and consumption was put forward.

A Novel Multiplex PCR Coupled with Luminex Assay for the Simultaneous Detection of Cryptosporidium Spp., Cryptosporidium Parvum and Giardia Duodenalis

Veterinary Parasitology. Oct, 2010  |  Pubmed ID: 20594647

Cryptosporidium parvum and Giardia duodenalis are the most frequently identified enteric parasites associated with diarrhea-causing disease outbreaks, and many non-parvum species of Cryptosporidium also can replicate and cause illness in mammals including humans. In this study, we describe a novel multiplex PCR coupled with Luminex assay for the identification of Cryptosporidium spp., C. parvum and G. duodenalis in a rapid manner. The multiplex PCR for the simultaneous detection of Cryptosporidium and Giardia was developed using three pairs of biotinylated primers which amplify 424, 223 and 267 bp products from the U1 small nuclear ribonucleoprotein (U1 snr) gene, 18S rRNA gene of Cryptosporidium and the beta-giardin gene of Giardia, respectively. The genus and species-specific capture probes linked to carboxylated Luminex microspheres hybridized to the multiplex PCR amplicons to enhance sensitivity and specificity. The conditions of multiplex PCR and Luminex hybridization reaction were optimized to enable the minimum detection limits of 5x10(-6), 5x10(-6), and 5x10(-6) ng DNAs (corresponding approximately to 0.1 oocyst/cyst). The Luminex approach proved to be 100% specific and accurate by testing a total of 240 fecal samples compared with microscopic examination of fecal smears and further modified acid-fast staining or iodine-staining observation. The established assay offers the potential for rapid detection of Cryptosporidium spp., C. parvum and G. duodenalis in fecal and environmental samples.

Lifetime Multiple Substance Use Pattern Among Heroin Users Before Entering Methadone Maintenance Treatment Clinic in Yunnan, China

Drug and Alcohol Review. Jul, 2010  |  Pubmed ID: 20636659

Multiple substance use leads to greater levels of psycho-behavioural problems, unsafe sex, and therefore a high risk of contracting sexually transmitted diseases, and is also more difficult to treat. This study aims to determine pattern of lifetime multiple substance use among Chinese heroin users before entering methadone maintenance treatment clinic.

Nano-structure of the Laminin γ-1 Short Arm Reveals an Extended and Curved Multidomain Assembly

Matrix Biology : Journal of the International Society for Matrix Biology. Sep, 2010  |  Pubmed ID: 20688161

Laminins are multidomain glycoproteins that play important roles in development and maintenance of the extracellular matrix via their numerous interactions with other proteins. Several receptors for the laminin short arms revealed their importance in network formation and intercellular signaling. However, both the detailed structure of the laminin γ-1 short arm and its organization within the complexes is poorly understood due to the complexity of the molecule and the lack of a high-resolution structure. The presented data provide the first subatomic resolution structure for the laminin γ-1 short arm in solution. This was achieved using an integrated approach that combined a number of complementary biophysical techniques such as small angle X-ray scattering (SAXS), analytical ultracentrifugation, dynamic light scattering and electron microscopy. As a result of this study, we have obtained a significantly improved model for the laminin γ-1 short arm that represents a major step forward in molecular understanding of laminin-mediated complex formations.

Femoral Artery Occlusion Increases Expression of ASIC3 in Dorsal Root Ganglion Neurons

American Journal of Physiology. Heart and Circulatory Physiology. Nov, 2010  |  Pubmed ID: 20852050

Acid-sensing ion channels (ASICs) in sensory nerves are responsive to increases in the levels of protons in the extracellular medium. Prior studies suggest that the muscle metabolite, lactic acid, plays a role in reflex sympathetic and cardiovascular responses via stimulation of thin muscle afferent nerves. Also, femoral artery occlusion augments the reflex sympathetic nerve response in rats. ASIC3 is a main subtype to appear in sensory nerves in mediating the response induced by increases in protons in the interstitial space of contracting muscles. Thus, in this article, we hypothesized that femoral occlusion increases the expression of ASIC3 in primary afferent neurons innervating muscles, and this contributes to the exaggerated reflex sympathetic responses. Femoral occlusion/vascular insufficiency of the hindlimb muscles was induced by the femoral artery ligation in rats. First, Western blot analysis shows that 24-72 h of femoral artery ligation significantly increased the expression of ASIC3 protein in dorsal root ganglion (optical density, 1.0 ± 0.07 in control vs. 1.65 ± 0.1 after 24 h of occlusion, P < 0.05; n = 6 in each group). There were no significant differences for increases in ASIC3 24 and 72 h postocclusion. Second, experiments using fluorescent immunohistochemistry and retrograde-labeling technique show that a greater percentage of ASIC3 staining neurons are localized in muscle-innervating dorsal root ganglion neurons after the arterial occlusion (78 ± 3% in 24 h post occlusion vs. 59 ± 5% in control, P < 0.05; n = 6 in each group). Third, the reflex responses in renal sympathetic nerve and arterial blood pressure induced by the stimulation of ASIC were examined after an injection of lactic acid into the arterial blood supply of hindlimb muscles of control rats and ligated rats. The results demonstrate that the sympathetic and pressor responses to lactic acid were significantly augmented after femoral occlusion compared with those in the control group. The data of this study suggest that enhanced ASIC3 expression in muscle afferent nerves contributes to the exaggerated reflex sympathetic and pressor responses to lactic acid as seen in arterial occlusion.

[Determination of Arbutin in Apple Juice Concentrate by Ultra Performance Liquid Chromatography with Electrospray Ionization Tandem Mass Spectrometry]

Se Pu = Chinese Journal of Chromatography / Zhongguo Hua Xue Hui. Jun, 2010  |  Pubmed ID: 20873587

An ultra performance liquid chromatography-tandem mass spectrometry (UPLC-MS/ MS) method was developed for the determination of arbutin in apple juice concentrate. Samples were diluted with water, then cleaned-up with a PS-DVB column. Quantitation was carried out using an external standard method. UPLC was performed on an Eclipse Plus C, column (100 mm x 2.1 mm, 1.8 microm) using a gradient solvent system (methanol-water). MS/MS was performed with multiple reaction monitoring (MRM) mode. The detection limit of arbutin was 0.02 mg/L. The method showed good linear relationship at the range of 0.04-2.0 mg/L. The recoveries ranged from 75.2% to 102.7% with relative standard deviations (RSDs) less than 8.9%. The method is simple, fast and sensitive. It's suitable for quantitative and qualitative analysis of arbutin in apple juice concentrate.

Clinical Outcomes for Various Causes of Infertility with Natural-cycle in Vitro Fertilization Combined with in Vitro Maturation of Immature Oocytes

Fertility and Sterility. Jul, 2010  |  Pubmed ID: 19909949

The clinical outcomes of different infertility causes (tubal factor, male factor, unexplained infertility, combination of tubal and male factors, and other/mixed factors) with natural-cycle in vitro fertilization/maturation (IVF/M) treatment were evaluated. There were no significant differences in the rates of IVM, IVF, and cleavage as well as the clinical pregnancy (30.4%-46.9%) and live birth rates among the five subgroups, which suggests that natural-cycle IVF/M is a suitable treatment for infertility of various causes with acceptable pregnancy and live birth rates.

Eimeria Tenella: Cloning and Characterization of Telomerase Reverse Transcriptase Gene

Experimental Parasitology. Apr, 2010  |  Pubmed ID: 20034493

Telomerase is essential for maintaining telomere length and chromosome stability in most eukaryotic organisms. The telomerase ribonucleoprotein complex consists of two essential components, the intrinsic telomerase RNA and the catalytic telomerase reverse transcriptase protein. Here we describe the cloning, sequencing and characterization of the telomerase reverse transcriptase catalytic subunit from the apicomplexan protozoon Eimeria tenella. The amino acid sequence predicted from it has all the signature motifs of the TERT family members. The E. tenella TERT cDNA contains an open reading frame encoding a protein with 1497 amino acids predicted size of 172kDa and isoelectric point of 9.344. It contains the conserved reverse transcriptase motifs 1, 2, A, B, C, D and E as well as the TERT-specific T motif and the N-terminal conserved motifs GQ, CP and QFP. RT-PCR analysis indicated that E. tenella TERT mRNA expressed in sporozoites and merozoites phase while not in unsporulated oocysts. At the same time, the result of TRAP assay indicated that marked telomerase activity were detected in sporozoites and merozoites. In contrast, telomerase activity was not detected in unsporulated oocysts.

Use of the BCR Sequential Extraction Procedure for the Study of Metal Availability to Plants

Journal of Environmental Monitoring : JEM. Feb, 2010  |  Pubmed ID: 20145888

To investigate the mobility and availability of metals from soil to plant, concentrations of zinc (Zn), copper (Cu), lead (Pb), cadmium (Cd), mercury (Hg) and arsenic (As) in topsoils and plants (lettuce, scallion, celery, tomato, carambola, wampee and longan) collected from the area around a petrochemical complex in Guangzhou, China, were analyzed. The modified European Community Bureau of Reference (BCR) three-step sequential extraction procedure was applied to determine the concentration of metal fractions in soils. The results showed that the distribution of Zn, Cu, Pb and Cd in four fractions varied greatly among the soil samples, and 18.8% of vegetable and fruit samples for Cd and 5.8% for Pb exceeded the maximum permissible levels in food of China. Soil-to-plant transfer coefficients were in the order of Cd>Zn>Cu>Hg>As>Pb, suggesting Cd being the most mobile and available to plants among the metals studied. Principal component analysis indicated that metal fractions and soil physicochemical properties (pH, organic matter, cation exchange capacity, clay content and electrical conductivity) affected metal uptake by plants. Furthermore, atmospheric deposition may be another important factor for the accumulation of metals in plants.

Further Evidence for Direct Pro-resorptive Actions of FSH

Biochemical and Biophysical Research Communications. Mar, 2010  |  Pubmed ID: 20171951

We confirm that FSH stimulates osteoclast formation, function and survival to enhance bone resorption. It does so via the activation of a pertussis toxin-sensitive G(i)-coupled FSH receptor that we and others have identified on murine and human osteoclast precursors and mature osteoclasts. FSH additionally enhances the production of several osteoclastogenic cytokines, importantly TNFalpha, likely within the bone marrow microenvironment, to augment its pro-resorptive action. FSH levels in humans rise before estrogen falls, and this hormonal change coincides with the most rapid rates of bone loss. On the basis of accumulating evidence, we reaffirm that FSH contributes to the rapid peri-menopausal and early post-menopausal bone loss, which might thus be amenable to FSH blockade.

The Chinese Government's Response to Drug Use and HIV/AIDS: a Review of Policies and Programs

Harm Reduction Journal. 2010  |  Pubmed ID: 20205729

Illicit drug use has become popular in China. Acknowledging the challenge of illicit drug use, China has adopted several new policies on the management of illicit drug use in recent years. This study reviews the current policies on drug use and assesses the harm reduction interventions among drug users in China. The review documents that the new policies on drug use provide a variety of choices of detoxification treatment for drug users. The methadone maintenance treatment and needle exchange programs have been adopted as harm reduction models in China. Most of the reviewed harm reduction programs have been successfully implemented and yielded positive effects in reducing drug related risk behaviors among drug users. Although there remain barriers to the effective implementation of policies on drug use and harm reduction programs, Chinese government has shown their commitment to support the expansion of harm reduction interventions for drug users throughout the country.

Bradykinin Receptor Blockade Reduces Sympathetic Nerve Response to Muscle Contraction in Rats with Ischemic Heart Failure

American Journal of Physiology. Heart and Circulatory Physiology. May, 2010  |  Pubmed ID: 20207818

Previous animal and human studies have suggested that a muscle reflex engaged during contraction leads to heightened levels of sympathetic activity in congestive heart failure (CHF). The present experiment was designed to test the role for bradykinin, which is produced within contracting skeletal muscle and contributes to the muscle reflex through its action on kinin B(2) receptors located on the endings of thin fiber muscle afferents. CHF was induced in rats by myocardial infarction (MI) after coronary artery ligation. Echocardiography was performed to determine fractional shortening (FS), an index of the left ventricular function. In the decerebrate rats, we examined renal sympathetic nerve activity (RSNA) during 1 min intermittent (1 to 4 s stimulation to relaxation) contraction of left triceps surae muscles. RSNA responded synchronously as tension was developed, and the response was significantly (P < 0.05) greater in MI rats [+39 +/- 9% s(-1) (integrated RSNA over time); n = 16] with 20 +/- 2% of FS than that in control healthy rats (+19 +/- 2% s(-1); n = 16) with 49 +/- 2% of FS. Tension development did not differ significantly between the two groups of rats. Thirty minutes after intra-arterial injection into the hindlimb circulation of the kinin B(2) receptor antagonist, HOE-140 (2 microg/kg), the RSNA response to contraction was significantly reduced in the MI rats (+26 +/- 7% s(-1)) but not in the control rats (+17 +/- 2% s(-1)). These data suggest that bradykinin within contracting muscle is part of the exaggerated muscle reflex seen in CHF.

Peripheral Administration of Fetuin-A Attenuates Early Cerebral Ischemic Injury in Rats

Journal of Cerebral Blood Flow and Metabolism : Official Journal of the International Society of Cerebral Blood Flow and Metabolism. Mar, 2010  |  Pubmed ID: 19953099

Cerebral ischemia-elicited inflammatory responses are driven by inflammatory mediators produced both by central (e.g., neurons and microglia) and infiltrating peripheral immune cells (e.g., macrophage/monocyte), and contribute to the evolution of tissue injury. A ubiquitous molecule, spermine, is released from injured cells, and counter-regulates release of various proinflammatory cytokines. However, the spermine-mediated anti-inflammatory activities are dependent on the availability of fetuin-A, a liver-derived negative acute-phase protein. Using an animal model of focal cerebral ischemia (i.e., permanent middle cerebral artery occlusion, MCAo), we found that levels of fetuin-A in the ischemic brain tissue were elevated in a time-dependent manner, starting between 2 and 6 h, peaking around 24 to 48 h, and returning to baseline 72 h after MCAo. When administered peripherally, exogenous fetuin-A gained entry across the BBB into the ischemic brain tissue, and dose dependently reduced brain infarct volume at 24 h after MCAo. Meanwhile, fetuin-A effectively attenuated (i) ischemia-induced HMGB1 depletion from the ischemic core; (ii) activation of centrally (e.g., microglia) and peripherally derived immune cells (e.g., macrophage/monocytes); and (iii) TNF production in ischemic brain tissue. Taken together, these experimental data suggest that fetuin-A protects against early cerebral ischemic injury partly by attenuating the brain inflammatory response.

Integrating Heterogeneous Knowledge Sources to Acquire Executable Drug-related Knowledge

AMIA ... Annual Symposium Proceedings / AMIA Symposium. AMIA Symposium. 2010  |  Pubmed ID: 21347099

Knowledge of medical entities, such as drug-related information is critical for many automated biomedical applications, such as decision support and pharmacovigilance. In this work, heterogeneous information sources were integrated automatically to obtain drug-related knowledge. We focus on one type of knowledge, drug-treats-condition, in the study and propose a framework for integrating disparate knowledge sources. Evaluation based on a random sample of drug-condition pairs indicated an overall coverage of 96%, recall of 98% and a precision of 87%. In conclusion, the preliminary study demonstrated that the knowledge generated from this study was comparable to the manually curated gold standard and that this method of automatically integrating knowledge sources is effective. The automated method should also be applicable to integrate other clinical knowledge, such as drug-related knowledge with omics information.

[Simultaneous Determination of Four Alternaria Toxins in Apple Juice Concentrate by Ultra Performance Liquid Chromatography-electrospray Ionization Tandem Mass Spectrometry]

Se Pu = Chinese Journal of Chromatography / Zhongguo Hua Xue Hui. Dec, 2010  |  Pubmed ID: 21438363

An ultra performance liquid chromatography-tandem mass spectrometry (UPLC-MS/ MS) method was developed for the determination of altenuene (ALT), alternariol (AOH), tentoxin and alternariol monomethyl ether (AME) in apple juice concentrate (AJC). The sample was diluted with water, and then cleaned up with a PS DVB column. The quantification was carried out using an external standard method. The UPLC was performed on a BEH C18 column (50 mm x 2.1 mm, 1.7 microm) using a gradient elution of acetonitrile and water. The MS/MS was performed with multiple reaction monitoring (MRM) mode. The limits of quantification of the four Alternaria toxins were between 1.0 and 5.0 microg/L. The recoveries were 77.8%-117.2% with the relative standard deviations less than 9.7%. The method is sensitive, stable and reliable. It's suitable for the quantitative and qualitative analyses of the four Alternaria toxins in AJC.

Scaling Up the National Methadone Maintenance Treatment Program in China: Achievements and Challenges

International Journal of Epidemiology. Dec, 2010  |  Pubmed ID: 21113034

China's methadone maintenance treatment program was initiated in 2004 as a small pilot project in just eight sites. It has since expanded into a nationwide program encompassing more than 680 clinics covering 27 provinces and serving some 242 000 heroin users by the end of 2009. The agencies that were tasked with the program's expansion have been confronted with many challenges, including high drop-out rates, poor cooperation between local governing authorities and poor service quality at the counter. In spite of these difficulties, ongoing evaluation has suggested reductions in heroin use, risky injection practices and, importantly, criminal behaviours among clients, which has thus provided the impetus for further expansion. Clinic services have been extended to offer clients a range of ancillary services, including HIV, syphilis and hepatitis C testing, information, education and communication, psychosocial support services and referrals for treatment of HIV, tuberculosis and sexually transmitted diseases. Cooperation between health and public security officials has improved through regular meetings and dialogue. However, institutional capacity building is still needed to deliver sustainable and standardized services that will ultimately improve retention rates. This article documents the steps China made in overcoming the many barriers to success of its methadone program. These lessons might be useful for other countries in the region that are scaling-up their methadone programs.

Protective Effect of Abamectin on Acute Lung Injury Induced by Lipopolysaccharide in Mice

Fundamental & Clinical Pharmacology. Dec, 2011  |  Pubmed ID: 21118302

Abamectin, a broad-spectrum antiparasitic agent, has been shown to exert an anti-inflammatory effect, in vitro, by down regulating both the nuclear transcription factor kappa-B and the mitogen-activated protein kinase (MAPK) activation pathway. In this study, we investigated the role of abamectin in acute lung injury (ALI) induced by Lipopolysaccharide (LPS), and the invovment of MAPK and NF-κB. BALB/C mice were administered abamectin (PBS) orally, followed by a dose of 0.5 mg/kg of LPS. After 10 h, tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) in bronchoalveolar lavage fluid (BALF) were measured using enzyme-linked immunosorbent assay. The number of total cells, neutrophils, and macrophages in the BALF were determined. The right lung was then excised for histological examination and analysis of myeloperoxidase (MPO) activity. Phosphorylation of MAPK family and IκB were detected by western blot. We found that 2 mg/kg of abamectin had significant protective effects on ALI. Mice treated with LPS alone showed markedly increased TNF-α and IL-6 levels in the BALF. In addition, not only was the W/D ratio of lung tissue significantly decreased, the number of total cells, neutrophils and macrophages in the BALF was also significantly reduced 11 h after treatment with abamectin. Furthermore, p38MAPK, ERK, and IκB were activated in 10 h after LPS treatment, which could be blunted by Abamectin. These results indicate that abamectin could attenuate inflammatory injury induced by LPS through MAPK and NF-κB passway.

Diagnostic Significance of CK19, TG, Ki67 and Galectin-3 Expression for Papillary Thyroid Carcinoma in the Northeastern Region of China

Diagnostic Pathology. 2011  |  Pubmed ID: 22188859

To evaluate the expression and differential diagnostic significance of CK19, TG, Ki67 and galectin-3 in papillary thyroid carcinoma (PTC) (metastatic and non metastatic), follicular adenoma and nodular goiter in patients from the northeastern part of China.

Nanopaper Based on Ag/TiO2 Nanobelts Heterostructure for Continuous-flow Photocatalytic Treatment of Liquid and Gas Phase Pollutants

Journal of Hazardous Materials. Dec, 2011  |  Pubmed ID: 21978584

The Ag/TiO(2) nanobelt heterostructures were prepared by the acid-assisted hydrothermal method followed by an in situ photo-reduction process. The photocatalytic activity of TiO(2) nanobelts was evidently enhanced by the heterostructures between Ag nanoparticles and TiO(2) nanobelts. The nanopapers based on Ag/TiO(2) nanobelt heterostructures were fabricated via a modified paper-making process. A novel continuous photocatalytic reactor was designed, and MO removal rate of Ag/C-TiO(2) nanopaper was achieved to 100% in 40 min for single layer and only in 6 min for three layers. The self-supported TiO(2) nanopapers with porous structures also showed an excellent continuous photocatalytic performance for toluene gas under UV light irradiation, and the corresponding degradation rate was 69.5% in 184 min. Moreover, the Ag/TiO(2) nanobelts nanopaper showed a good antibacterial effect. The multifunctional TiO(2) nanopapers modified by the heterostuctures could have potential applications in the environmental and biomaterial fields.

Circulating MicroRNAs in Patients with Active Pulmonary Tuberculosis

Journal of Clinical Microbiology. Dec, 2011  |  Pubmed ID: 21998423

Emerging evidence shows that microRNAs (miRNAs) play an important role in pathogen-host interactions. Circulating miRNAs have been repeatedly and stably detected in blood and hold promise to serve as molecular markers for diverse physiological and pathological conditions. To date, the relationship between circulating miRNAs and active pulmonary tuberculosis (TB) has not been reported. Using microarray-based expression profiling followed by real-time quantitative PCR validation, the levels of circulating miRNAs were compared between patients with active pulmonary tuberculosis and matched healthy controls. The receiver operating characteristic curve was used to evaluate the diagnostic effect of selected miRNA. Bioinformatic analysis was used to explore the potential roles of these circulating miRNAs in active pulmonary tuberculosis infection. Among 92 miRNAs significantly detected, 59 miRNAs were downregulated and 33 miRNAs were upregulated in the TB serum compared to their levels in the control serum. Interestingly, only two differentially expressed miRNAs were increased not only in the serum but also in the sputum of patients with active pulmonary tuberculosis compared to the levels for the healthy controls. Upregulated miR-29a could discriminate TB patients from healthy controls with reasonable sensitivity and specificity. A number of significantly enriched pathways regulated by these circulating miRNAs were predicted, and most of them were involved in acute-phase response, inflammatory response, and the regulation of the cytoskeleton. In all, for the first time our results revealed that a number of miRNAs were differentially expressed during active pulmonary tuberculosis infection, and circulating miR-29a has great potential to serve as a marker for the detection of active pulmonary tuberculosis infection.

Role of Sexual Transmission of HIV Among Young Noninjection and Injection Opiate Users: a Respondent-driven Sampling Study

Sexually Transmitted Diseases. Dec, 2011  |  Pubmed ID: 22082729

Sexual transmissibility of HIV among young drug users in China has been investigated in few studies. The objective of this study was to examine the role of sexual transmission on HIV infection among injection drug users (IDUs) and noninjection drug users (NIDUs).

Gamma-irradiation Increased Meiotic Crossovers in Mouse Spermatocytes

Mutagenesis. Nov, 2011  |  Pubmed ID: 21778358

In mice, the occurrence of immunofluorescent foci for mismatch repair protein MLH1 correlates closely with the occurrence of crossovers, as detected genetically, and MLH1 foci represent virtually all prospective crossover positions. To examine the effects of γ-irradiation on meiotic crossovers in mouse spermatocytes, male mice were subjected to whole-body γ-irradiation at different sub-stages of meiotic prophase and crossovers on synaptonemal complexes (SCs) were analysed by visualising and quantifying the immunofluorescent MLH1 foci. At both 24 and 48 h after exposure, significant dose-dependent increases in the number of total MLH1 foci per spermatocyte were observed at late zygotene-early pachytene with the gradient increase of radiation dose from 0, 1.5, 3-6 Gy. Furthermore, irradiation at preleptotene-leptotene still led to significant dose-dependent increased meiotic crossovers in the spermatocytes analysed 120 h after exposure. In further analysis, these dose-dependent increases in the number of total MLH1 foci per cell were attributed to significant dose-dependent decreases in autosomal SCs with 0 MLH1 focus, and the dose-dependent increases in autosomal SCs with 2 MLH1 foci and the percentage of cells with MLH1 focus on XY bivalent. The increased number of cells with an MLH1 focus on the pseudoautosomal regions (PARs) may indicate that there is a delay in meiotic progression in the irradiated cells. Although significant dose-dependent increases in the number of total MLH1 foci per cell were examined 24, 48 or 120 h after exposure with the gradient increase of radiation doses, these increases were mild compared to the control groups. This suggests that there is tight control of crossover formation (at least with respect to MLH1 foci number). The mechanisms underlying irradiation-induced DNA lesion repair, cellular responses independent of DNA damage and meiotic crossover homeostasis in mammals will be the subjects of future study.

Effects of a Randomized Contingency Management Intervention on Opiate Abstinence and Retention in Methadone Maintenance Treatment in China

Addiction (Abingdon, England). Oct, 2011  |  Pubmed ID: 21793958

Methadone maintenance treatment has been made available in China in response to the rapid spread of human immunodeficiency virus (HIV), but high rates of dropout and relapse are problematic. The aim of this study was to apply and test if a contingency management (or motivational incentives) intervention can improve treatment retention and reduce drug use.

Inhibition of Hepatitis B Virus Replication by CIAP2 Involves Accelerating the Ubiquitin-proteasome-mediated Destruction of Polymerase

Journal of Virology. Nov, 2011  |  Pubmed ID: 21865390

Cellular inhibitor of apoptosis protein 2 (cIAP2) is a potent suppressor of apoptotic cell death. We have shown previously that cIAP2 is involved in the tumor necrosis factor alpha (TNF-α)-induced anti-hepatitis B virus (HBV) response; however, the mechanism for this antiviral effect remains unclear. In the present study, we demonstrate that cIAP2 can significantly reduce the levels of HBV DNA replication intermediates but not the total viral RNA or core protein levels. Domain-mapping analysis revealed that the carboxy-terminal domains of cIAP2 were indispensable for this anti-HBV ability and that an E3 ligase-deficient mutant of cIAP2 (termed cIAP2*) completely lost its antiviral activity. We further identified HBV polymerase as the target of cIAP2. Overexpression of cIAP2 but not cIAP2* reduced polymerase protein levels, while cIAP2 knockdown increased polymerase expression. In addition, we observed that cIAP2 promoted the degradation of the viral polymerase through a proteasome-dependent pathway. Further experiments demonstrated that cIAP2 can bind to polymerase and promote its polyubiquitylation. Finally, we found that cIAP2 downregulated the encapsidation of HBV pregenomic RNA. Taken together, these data reveal a novel mechanism for the inhibition of HBV replication by cIAP2 via acceleration of the ubiquitin-proteasome-mediated decay of polymerase and reduction of the encapsidation of HBV pregenomic RNA, making this mechanism a novel strategy for HBV therapy.

Expression and Characterization of Recombinant Human Milk Fat Globule-EGF Factor VIII

International Journal of Molecular Medicine. Dec, 2011  |  Pubmed ID: 21874225

Apoptosis plays an important role in the patho-biology of sepsis. The opsonizing protein milk fat globule-EGF factor VIII (MFG-E8) is involved in apoptotic cell clearance. Our previous studies have shown that administration of rat MFG-E8-containing exosomes or recombinant murine MFG-E8 (rmMFG-E8) is protective in a rat model of sepsis induced by cecal ligation of puncture (CLP). However, one obstacle hampering the development of MFG-E8 as a therapeutic agent for septic patients is the potential immunogenicity of animal proteins in humans. The purpose of this study, therefore, was to express recombinant human MFG-E8 (rhMFG-E8) and characterize its biological activity. Using an E. coli system, we successfully expressed and purified the mature molecule of human MFG-E8 (Leu24-Cys387). The purity of rhMFG-E8 was over 99% and it was immunoreactive for specific anti-human MFG-E8 antibodies. Amino acid sequence analysis by LC-MS/MS identified the purified protein as human MFG-E8. Using primary rat peritoneal macrophages, we showed that rhMFG-E8 markedly increased peritoneal macrophage phagocytosis of apoptotic thymocytes, which was as effective as commercial rmMFG-E8. To determine the biological activity of rhMFG-E8 in vivo, male adult rats were subjected to sepsis by CLP. rhMFG-E8 or rmMFG-E8 were administered intravenously at the time of CLP. Our results demonstrated that both rhMFG-E8 and rmMFG-E8 reduced thymocyte apoptosis and plasma levels of lactate and IL-6 at 20 h after CLP, and improved the 10-day survival rate. Thus, we have successfully expressed and purified biologically active rhMFG-E8. Our newly-expressed rhMFG-E8 is highly effective in the rat model of sepsis.

Prevalence of Dirofilaria Immitis Infection in Dogs from Dandong, China

Veterinary Parasitology. Dec, 2011  |  Pubmed ID: 21889850

The aim of the present study was to estimate the prevalence and risk factors of Dirofilaria immitis infection in dogs from Dandong, China. A total of 886 dogs were examined for D. immitis infection by microscopic examination and PCR, indicating that the prevalence was 16.6% (213/886) and 24.0% (147/886), respectively. The odds of infection were significantly higher in older dogs and dogs sheltered in outdoor, compared to the younger ones and ones sheltered in indoor. No significant difference of infection was observed in different genders, and between pure breed and cross-breed dogs in the same rearing conditions. These results indicated that the risk of exposure to D. immitis in dogs is high in Dandong, China, and prophylaxis against the parasite is advisable to decrease the incidence of canine dirofilariosis.

T-shaped Arrangement of the Recombinant Agrin G3-IgG Fc Protein

Protein Science : a Publication of the Protein Society. Jun, 2011  |  Pubmed ID: 21448912

Agrin is a large heparin sulphate proteoglycan with multiple domains, which is located in the extracellular matrix. The C-terminal G3 domain of agrin is functionally one of the most important domains. It harbors an α-dystroglycan binding site and carries out acetylcholine receptor clustering activities. In the present study, we have fused the G3 domain of agrin to an IgG Fc domain to produce a G3-Fc fusion protein that we intend to use as a tool to investigate new binding partners of agrin. As a first step of the study, we have characterized the recombinant fusion protein using a multidisciplinary approach using dynamic light scattering, analytical ultracentrifugation and small angle X-ray scattering (SAXS). Interestingly, our SAXS analysis using the high-resolution structures of G3 and Fc domain as models indicates that the G3-Fc protein forms a T-shaped molecule with the G3 domains extruding perpendicularly from the Fc scaffold. To validate our models, we have used the program HYDROPRO to calculate the hydrodynamic properties of the solution models. The calculated values are in excellent agreement with those determined experimentally.

A Recombinant DNA Vaccine Encoding C. Andersoni Oocyst Wall Protein Induces Immunity Against Experimental C. Parvum Infection

Veterinary Parasitology. Jun, 2011  |  Pubmed ID: 21450406

Cryptosporidium andersoni parasited in the abomasum has been demonstrated as a cause of reduction of milk production in dairy cow. In this study, a novel chimeric DNA vaccine pVAX1-AB was constructed and the efficacy against Cryptosporidium parvum was determined. BALB/c mice were divided into 3 groups and immunized with DNA vaccine expressing the oocyst wall protein, AB protein of C. andersoni, the recombinant plasmid containing the AB gene, respectively. After inoculation of 1 × 10(6) oocysts of C. parvum, the humoral and cellular immune responses were detected. Experimental results showed that the recombinant plasmid can induce corresponding specific antibody response, simultaneously influenced cellular immune responses, and provided greater protection rate (48.6%) than the other groups. These results indicated that chimeric DNA vaccine has a potential in Cryptosporidium vaccine development.

Sexual Transmissibility of HIV Among Opiate Users with Concurrent Sexual Partnerships: an Egocentric Network Study in Yunnan, China

Addiction (Abingdon, England). Oct, 2011  |  Pubmed ID: 21457169

To investigate the patterns of concurrent sexual partnerships among young opiate users and sexual transmissibility of human immunodeficiency virus (HIV) in concurrent sexual partnerships in drug-use and sexual networks.

Immune Response and Protective Efficacy Against Homologous Challenge in BALB/c Mice Vaccinated with DNA Vaccine Encoding Toxoplasma Gondii Actin Depolymerizing Factor Gene

Veterinary Parasitology. Jun, 2011  |  Pubmed ID: 21489695

A DNA vaccine (pVAX1-TgADF) encoding Toxoplasma gondii actin depolymerizing factor (ADF) gene was constructed and the immune response and protective efficacy of this vaccine against homologous challenge in BALB/c mice were evaluated. High titers of specific antibody and increases in the percentage of CD4(+) and CD8(+) T lymphocyte cells were observed from BALB/c mice vaccinated with pVAX1-TgADF (P<0.05), when PBS group was used as control. The survival time of BALB/c mice in pVAX1-TgADF group was longer than those in control groups. The numbers of brain cysts in the experimental BALB/c mice immunized with pVAX1-TgADF reduced significantly compared with those in PBS group (P<0.05), and the rate of reduction could reach to around 42.8%. These results suggested that the DNA vaccine pVAX1-TgADF could generate specific humoral and cellular immune responses, prolong survival times, and reduce brain cysts load against T. gondii infection in BALB/c mice.

Subversion of Cellular Autophagy Machinery by Hepatitis B Virus for Viral Envelopment

Journal of Virology. Jul, 2011  |  Pubmed ID: 21507968

Autophagy is a conserved eukaryotic mechanism that mediates the removal of long-lived cytoplasmic macromolecules and damaged organelles via a lysosomal degradative pathway. Recently, a multitude of studies have reported that viral infections may have complex interconnections with the autophagic process. The findings reported here demonstrate that hepatitis B virus (HBV) can enhance the autophagic process in hepatoma cells without promoting protein degradation by the lysosome. Mutation analysis showed that HBV small surface protein (SHBs) was required for HBV to induce autophagy. The overexpression of SHBs was sufficient to induce autophagy. Furthermore, SHBs could trigger unfolded protein responses (UPR), and the blockage of UPR signaling pathways abrogated the SHB-induced lipidation of LC3-I. Meanwhile, the role of the autophagosome in HBV replication was examined. The inhibition of autophagosome formation by the autophagy inhibitor 3-methyladenine (3-MA) or small interfering RNA duplexes targeting the genes critical for autophagosome formation (Beclin1 and ATG5 genes) markedly inhibited HBV production, and the induction of autophagy by rapamycin or starvation greatly contributed to HBV production. Furthermore, evidence was provided to suggest that the autophagy machinery was required for HBV envelopment but not for the efficiency of HBV release. Finally, SHBs partially colocalized and interacted with autophagy protein LC3. Taken together, these results suggest that the host's autophagy machinery is activated during HBV infection to enhance HBV replication.

A Fine Balance Between CCNL1 and TIMP1 Contributes to the Development of Breast Cancer Cells

Biochemical and Biophysical Research Communications. Jun, 2011  |  Pubmed ID: 21586274

Cyclin L1 (CCNL1) and tissue inhibitor of matrix metalloproteinase-1 (TIMP1) are candidate genes involved in several types of cancer. However, the expression of CCNL1 and the relationship between CCNL1 and TIMP1 in breast cancer cells is unknown. Using patients' breast cancer tissues, the expression of CCNL1 and TIMP1 was measured by cDNA microarray and further confirmed by real-time RT-PCR and western blotting. Overexpression or repression of CCNL1 and TIMP1, individually or together, was performed in breast cancer MDA-MB-231 cells by transient transformation methods to investigate their role in breast cancer cell growth. Simultaneously, mRNA and protein expression levels of CCNL1 and TIMP1 were also measured. CCNL1 and TIMP1 expression was significantly elevated in breast cancer tissues compared with that in peri-breast cancer tissues of patients by cDNA microarray and these results were further confirmed by real-time RT-PCR and western blotting. Interestingly, in vitro experiments showed a stimulatory effect of TIMP1 and an inhibitory effect of CCNL1 on growth of MDA-MB-231 cells. Co-expression or co-repression of these two genes did not affect cell growth. Overexpression of CCNL1 and TIMP1 individually induced overexpression of each other. These data demonstrate that there is a fine balance between CCNL1 and TIMP1, which may contribute to breast cancer development.

Hepatitis C Virus NS5B Protein Delays S Phase Progression in Human Hepatocyte-derived Cells by Relocalizing Cyclin-dependent Kinase 2-interacting Protein (CINP)

The Journal of Biological Chemistry. Jul, 2011  |  Pubmed ID: 21628470

Cell cycle dysregulation is a critical event in virus infection-associated tumorigenesis. Previous studies have suggested that hepatitis C virus NS5B modulates cell cycle progression in addition to participating in RNA synthesis as an RNA-dependent RNA polymerase. However, the molecular mechanisms have thus far remained unclear. In this study, a HepG2 Tet-On NS5B stable cell line was generated to confirm the effect of NS5B on the cell cycle. To better understand the role of NS5B in cell cycle regulation, yeast two-hybrid assays were performed using a human liver cDNA library. The cyclin-dependent kinase 2-interacting protein (CINP) was identified. The interaction between NS5B and CINP was further demonstrated by in vivo and in vitro assays, and their association was found to be indispensable for S phase delay and cell proliferation suppression. Further experiments indicated that NS5B relocalized CINP from the nucleus to the cytoplasm. Directly knocking down CINP by specific siRNA resulted in a significant alteration in the DNA damage response and expression of cell cycle checkpoint proteins, including an increase in p21 and a decrease in phosphorylated Retinoblastoma and Chk1. Similar results were observed in cells expressing NS5B, and the effects were partially reversed upon ectopic overexpression of CINP. These studies suggest that the DNA damage response might be exploited by NS5B to hinder cell cycle progression. Taken together, our data demonstrate that NS5B delays cells in S phase through interaction with CINP and relocalization of the protein from the nucleus to the cytoplasm. Such effects might contribute to hepatitis C virus persistence and pathogenesis.

Contribution of Nerve Growth Factor to Upregulation of P2X₃ Expression in DRG Neurons of Rats with Femoral Artery Occlusion

American Journal of Physiology. Heart and Circulatory Physiology. Sep, 2011  |  Pubmed ID: 21642505

Femoral artery occlusion augments the sympathetic nerve and pressor responses to muscle contraction and muscle metabolites injected into the arterial blood supply of the hindlimb muscles in rats. The underlying mechanism by which these reflex responses are enhanced after muscle vascular insufficiency is unclear. Purinergic P2X(3) receptor has been reported to contribute to the metabolic component of the exercise pressor reflex. Thus the purpose of this study was to examine if chronic femoral occlusion would alter the expression of P2X(3) in dorsal root ganglion (DRG) neurons of rats. Also, P2X(3)-mediated sympathetic responsiveness was examined after femoral occlusion. In addition, the role played by nerve growth factor (NGF) in regulating the expression and response of P2X(3) was examined. Western blot analysis showed that 24 h of femoral ligation increased the levels of P2X(3) (optical density: 0.93 ± 0.07 in control and 1.37 ± 0.10 after occlusion; P < 0.05 vs. control). The fluorescence immunohistochemistry further demonstrated that the occlusion elevated P2X(3) expression in DRG neurons (percentage of P2X(3)-positive cells: 33 ± 3% in control and 51 ± 3% in occlusion; P < 0.05 vs. control). Furthermore, the results showed that responses of renal sympathetic nerve activity and blood pressure to stimulation of P2X were greater in occluded rats than responses in control rats by injection of α,β-methylene ATP into the arterial blood supply of the hindlimb muscle. Finally, infusion of NGF in the hindlimb muscles of healthy rats increased P2X(3) (optical density: 0.98 ± 0.12 in control and 1.37 ± 0.16 with NGF; P < 0.05 vs. control). The pressor response to injection of α,β-methylene ATP was increased in the rats with NGF infusion. Likewise, blocking NGF attenuated exaggeration of the reflex response induced by α,β-methylene ATP in occluded rats. The findings of this study suggest that the levels of P2X(3) in primary afferent neurons are upregulated as the blood supply to the hindlimb is deficient under ischemic conditions, leading to augmentation of the muscle reflex. NGF is closely related to increases in P2X(3) receptor expression and response.

Natural Cycle IVF/IVM May Be More Desirable for Poor Responder Patients After Failure of Stimulated Cycles

Journal of Assisted Reproduction and Genetics. Sep, 2011  |  Pubmed ID: 21695516

To describe pregnancies and live births resulted from natural cycle IVF combined with in-vitro maturation (natural-cycle IVF/IVM) for three poor responder women after failure of stimulated cycles.

Regulated Production of the Pituitary Hormone Oxytocin from Murine and Human Osteoblasts

Biochemical and Biophysical Research Communications. Aug, 2011  |  Pubmed ID: 21741363

Oxytocin (OT) is a primitive neurohypophyseal hormone that plays a primary and indispensible role in mammalian lactation. We have shown recently that OT also regulates bone remodeling, mainly bone formation, with remarkable sensitivity. We now show that OT, apart from its neurohypophyseal origin, is produced in abundance by both human and murine osteoblasts. Production of osteoblast OT is under the control of estrogen, which acts by activating the MAP kinase Erk. This non-genomic mechanism of estrogen action is in stark contrast to its genomic control of OT receptor (OTR) expression. We surmise that there is a local feed-forward loop in bone marrow through which the OT so produced from osteoblasts in response to estrogen acts upon its receptor to exert a potent anabolic action.

A Hepatic Protein, Fetuin-A, Occupies a Protective Role in Lethal Systemic Inflammation

PloS One. 2011  |  Pubmed ID: 21347455

A liver-derived protein, fetuin-A, was first purified from calf fetal serum in 1944, but its potential role in lethal systemic inflammation was previously unknown. This study aims to delineate the molecular mechanisms underlying the regulation of hepatic fetuin-A expression during lethal systemic inflammation (LSI), and investigated whether alterations of fetuin-A levels affect animal survival, and influence systemic accumulation of a late mediator, HMGB1.

EGCG Stimulates Autophagy and Reduces Cytoplasmic HMGB1 Levels in Endotoxin-stimulated Macrophages

Biochemical Pharmacology. May, 2011  |  Pubmed ID: 21371444

Historically, consumption of Green tea (Camellia sinensis) has been associated with health benefits against multiple diseases including cancer, atherosclerosis and cardiovascular disorders. Emerging evidence has suggested a pathogenic role for HMGB1, a newly identified "late" mediator of lethal systemic inflammation, in the aforementioned diseases. Here we demonstrated that a major ingredient of Green tea, EGCG, was internalized into HMGB1-containing LC3-positive cytoplasmic vesicles (likely autophagosomes) in macrophages, and induced HMGB1 aggregation in a time-dependent manner. Furthermore, EGCG stimulated LC3-II production and autophagosome formation, and inhibited LPS-induced HMGB1 up-regulation and extracellular release. The EGCG-mediated HMGB1 inhibitory effects were diminished by inhibition of class III phosphatidylinositol-3 kinase (with 3-methyladenine) or knockdown of an essential autophagy-regulating protein, beclin-1. Moreover, the EGCG-mediated protection against lethal sepsis was partly impaired by co-administration of an autophagy inhibitor, chloroquine. Taken together, the present study has suggested a possibility that EGCG inhibits HMGB1 release by stimulating its autophagic degradation.

Patterns of Attendance in Methadone Maintenance Treatment Program in Yunnan Province, China

The American Journal of Drug and Alcohol Abuse. May, 2011  |  Pubmed ID: 21413914

To describe the patterns of patients' attendance over the first three quarters of a year under methadone maintenance treatment (MMT) clinics in Yunnan Province, China.

Role of Social Network Dimensions in the Transition to Injection Drug Use: Actions Speak Louder Than Words

AIDS and Behavior. Oct, 2011  |  Pubmed ID: 21431412

The objective of this study was to examine the influences of social network factors, particularly social support and norms, in the transition from non-injection heroin and/or opiate use to heroin-injection, which is one of the leading causes of the spread of HIV/AIDS in China. Respondent-driven sampling was used to recruit young heroin and/or opiate users in an egocentric network study in Yunnan, China. Multivariate logistic regression using hierarchical combinations of candidate variables was used to analyze network factors for the injection transition. A total of 3,121 social network alters were reported by 403 egos with an average network size of eight. Fifty-eight percent of egos transitioned to heroin-injection from non-injection. This transition was associated with having a larger sex network size, a larger number of heroin injectors in one's network, and a higher network density. The findings enhance our understanding of the influence of social network dimensions on the transition to injection drug use. Accordingly, the development of interventions for heroin and/or opiate users in China should consider social network characteristics.

Intravenous Ibandronate Acutely Reduces Bone Hyperresorption in Chronic Critical Illness

Journal of Intensive Care Medicine. Mar, 2011  |  Pubmed ID: 21436164

Objective: Patients who remain critically ill for prolonged periods and require tracheotomy, defined as chronic critical illness (CCI), display elevated levels of bone resorption. The measurement of bone turnover markers reveals that osteoclastic bone resorption is not only enhanced but also uncoupled from osteoblastic bone formation. We examine the effect of ibandronate on bone turnover in patients with CCI. Methods: This study is a prospective, double-blind, placebo-controlled trial, in which 20 postmenopausal female participants with CCI were followed for an 11-day period after the administration of a single intravenous dose of ibandronate (3 mg). All participants were treated with ergocalciferol (2000 IU daily), calcium carbonate (1250 mg daily), and calcitriol (0.25 μg daily). Results: The ibandronate group showed a 34% decrease in serum C-telopeptide (CTX) levels (a marker of osteoclastic activity) on day 6, while the placebo group showed a 13% increase (P = .01). By day 11, CTX levels in ibandronate group were not significantly different than baseline or from the placebo group. Osteocalcin (OCN) levels (a marker of osteoblast activity) increased by 78% compared to baseline in the ibandronate group (P = .01) and by 42% in the placebo group (P = .05). There were no significant differences in OCN between the 2 groups throughout the study. Parathyroid hormone levels remained constant throughout the study. No adverse events were observed. Conclusion: A single dose of intravenous ibandronate causes a significant but transient reduction in osteoclast activity in patients with CCI, which persists over a 6-day period.

Virus-like Particles in Eimeria Tenella Are Associated with Multiple RNA Segments

Experimental Parasitology. Mar, 2011  |  Pubmed ID: 21184756

Total nucleic acids from sporulated oocysts of Eimeria tenella isolated from Changchun in China were found to contain three extrachromosomal double-stranded RNA segments (dsRNAs) of 1.4, 2.4 and 3.6 kb in sizes. These RNAs were resistant to RNase A digestion under high salt concentration (0.3 M NaCl). RNA-dependent RNA polymerase (RDRP) activity was detected in crude extracts of E. tenella sporulated oocysts containing these nucleic acid species. Virus-like particles (VLPs) were shown to have a diameter of approximately 38 nm under Electron Microscopy (EM) after purification by sucrose density gradient centrifugation. In keeping with the nomenclature generally adopted for protozoan viruses, we have named this isolate as E. tenella virus (ETV) which is the first virus isolated from E. tenella.

Identification and Characterization of Myeloma-associated Antigens in Trichinella Spiralis

Experimental Parasitology. Apr, 2011  |  Pubmed ID: 21232537

To investigate the presence of myeloma-associated antigens in Trichinella spiralis and their anti-tumor effect, cross-immune responses between antigens of the myeloma cell SP2/0 versus positive sera to T. spiralis, and antigens of T. spiralis versus positive sera to myeloma cell SP2/0 were determined using T. spiralis and myeloma specific enzyme-linked immunosorbent assays (ELISA). The myeloma-associated antigens in T. spiralis were separated by ultrafiltration and 2-D electrophoresis, and the amino acid sequences and molecular weights were determined by spectrometry. An obvious reaction was found between a 33 kDa antigen and positive sera, and the major component of the antigen was tropomyosin (TM), which is an surface acidic protein with 284 amino acids. Mice were immunized with TM to determine the anti-tumor effect in vivo. The results showed that CD4(+), CD8(+) T lymphocyte, and CD19(+) B lymphocyte were significantly increased (P<0.05). The anti-tumor effects were significantly different between mice immunized with the antigens or adjuvant alone (P<0.05), while the difference between mice immunized with antigens and whole T. spiralis was not significant (P>0.05). The results indicated that TM identified in this study may play a role in eliciting cross-protective immunity.

Anti-inflammatory Effects of Ivermectin in Mouse Model of Allergic Asthma

Inflammation Research : Official Journal of the European Histamine Research Society ... [et Al.]. Jun, 2011  |  Pubmed ID: 21279416

Asthma is an inflammatory disease of the lungs that is characterised by increased inflammatory cell infiltration into the airways and poor respiratory function. Ivermectin is a semi-synthetic derivative of a family of macrocyclic lactones that shows broad-spectrum anti-parasitic activity. This drug has been shown to possess anti-inflammatory activity, but whether it can be used in asthma treatment has not yet been investigated. In this study, we aimed to investigate the inhibitory effects of ivermectin on allergic asthma symptoms in mice.

Multiple Substance Use Among Heroin-dependent Patients Before and During Attendance at Methadone Maintenance Treatment Program, Yunnan, China

Drug and Alcohol Dependence. Jul, 2011  |  Pubmed ID: 21282020

Multiple substance use is a common problem among heroin users. This study aims to describe patterns of multiple substance use one year before and during attendance at methadone maintenance treatment (MMT) programs and associated variables of continued heroin use in MMT clinics in Yunnan, China.

Efflux Pump Overexpression in Conjunction with Alternation of Outer Membrane Protein May Induce Acinetobacter Baumannii Resistant to Imipenem

Chemotherapy. 2011  |  Pubmed ID: 21346352

To investigate the role of outer membrane proteins in Acinetobacter baumannii resistant to imipenem, 2 strains were procured from the same patient.

Sequence Analysis and Verification of Eimeria Tenella Rhomboid Bait Plasmid Suitability for CytoTrap Yeast Two-hybrid System

Parasitology Research. Feb, 2011  |  Pubmed ID: 20941631

Effects of in Situ and Physical Mixing on Mechanical and Bioactive Behaviors of Nano Hydroxyapatite-chitosan Scaffolds

Journal of Biomaterials Science. Polymer Edition. 2011  |  Pubmed ID: 21067654

Nano hydroxyapatite (HAP) was employed to intensify chitosan (CS) scaffolds by two methods. The first one is nano HAP crystallized in situ from the CS matrix by a biomimetic method (in situ scaffold). In the second method the sol-gel nano HAP powder was added directly to the CS solution (physical mixing scaffold). The distribution status of nano HAP was examined by scanning electron microscopy. The compressive performance was measured by a universal material testing machine. The in vitro study in stimulated body fluid was performed to evaluate the biological properties of both scaffolds. MTT testing and alkaline phosphatase activity from human bone mesenchymal stem cell culture showed differences in biocompatibility and bioactivity between the scaffolds. The results indicated that the in situ scaffold possessed more excellent mechanical and bioactive behaviors than that of the physical mixing scaffold.

[Study on Effect of Fushenkeli on Expression of TAK1 in Human Renal Tubular Epithelial Cells and Its Possible Mechanism]

Zhongguo Zhong Yao Za Zhi = Zhongguo Zhongyao Zazhi = China Journal of Chinese Materia Medica. Sep, 2011  |  Pubmed ID: 22256769

To investigate the effects of fushenkeli on the expression of TAK1 in the proliferation of the renal tubular epithelial cells induced by TGF-beta1 and its possible mechanism.

Responses of Butachlor Degradation and Microbial Properties in a Riparian Soil to the Cultivation of Three Different Plants

Journal of Environmental Sciences (China). 2011  |  Pubmed ID: 22432278

A pot experiment was conducted to investigate the biodegradation dynamics and related microbial ecophysiological responses to butachlor addition in a riparian soil planted with different plants such as Phragmites australis, Zizania aquatica, and Acorus calamus. The results showed that there were significant differences in microbial degradation dynamics of butachlor in the rhizosphere soils among the three riparian plants. A. calamus displays a significantly higher degradation efficiency of butachlor in the rhizosphere soils, as compared with Z. aquatica and P. australis. Half-life time of butachlor degradation in the rhizospheric soils of P. australis, Z. aquatica, and A. calamus were 7.5, 9.8 and 5.4 days, respectively. Residual butachlor concentration in A. calamus rhizosphere soil was 35.2% and 21.7% lower than that in Z. aquatica and P. australis rhizosphere soils, respectively, indicating that A. calamus showed a greater improvement effect on biodegradation of butachlor in rhizosphere soils than the other two riparian plant. In general, microbial biomass and biochemical activities in rhizosphere soils were depressed by butachlor addition, despite the riparian plant types. However, rhizospheric soil microbial ecophysiological responses to butachlor addition significantly (P < 0.05) differed between riparian plant species. Compared to Z. aquatica and P. australis, A. calamus showed significantly larger microbial number, higher enzyme activities and soil respiration rates in the rhizosphere soils. The results indicated that A. calamus have a better alleviative effect on inhibition of microbial growth due to butachlor addition and can be used as a suitable riparian plant for detoxifying and remediating butachlor contamination from agricultural nonpoint pollution.

Effectiveness of TAD-anchored Maxillary Protraction in Late Mixed Dentition

The Angle Orthodontist. Mar, 2012  |  Pubmed ID: 22458766

Abstract Objective: To evaluate the effectiveness of temporary anchorage device (TAD)-anchored maxillary protraction (MP) in terms of the skeletal and dentoalveolar changes and to compare it with traditional tooth-anchored MP. Materials and Methods: A computerized literature search for relative randomized controlled trials and prospective controlled trials was performed in PubMed, MEDLINE, Cochrane Central Register of Controlled Trials, Embase, CNKI, and Google Scholar, complemented with manual search. Data extraction and quality assessment were carried out by two reviewers independently. Meta-analysis was followed when possible; otherwise, description was done. Results: Forty articles were found, among which four trials were qualified for meta-analysis. The results showed that there was significant difference between TAD-anchored MP and untreated control in terms of maxillary advancement (weighted mean differences (WMD) 3.08 mm; 95% CI: 1.61 to approximately 4.56; P < .0001), but there were no consistent points in terms of mandibular rotation. Also, there were significant differences between both treatment patterns regarding maxillary advancement (WMD 1.41 mm; 95% CI: 0.47 to approximately 2.35; P  =  .003), mandibular rotation (WMD -1.39°, 95% CI: -2.47 to approximately -0.31; P  =  .01), proclination of maxillary incisors (WMD -2.29°; 95% CI: -4.41 to approximately -0.17; P  =  .03), and extrusion of maxillary molars (WMD -1.68 mm; 95% CI: -2.51 to approximately -0.85; P < .0001). Conclusions: According to the present results, TAD-anchored MP might have a greater maxillary advancement effect and might reduce skeletal and dental side effects, compared with tooth-anchored MP.

Neuroprotective Effects of Valproic Acid Following Transient Global Ischemia in Rats

Life Sciences. Mar, 2012  |  Pubmed ID: 22285595

A growing number of studies demonstrate that valproic acid (VPA), an anti-convulsant and mood-stabilizing drug, is neuroprotective against various insults. This study investigated whether treatment of ischemic stroke with VPA ameliorated hippocampal cell death and cognitive deficits. Possible mechanisms of action were also investigated.

Inhibition of HMGB1 Enhances Bacterial Clearance and Protects Against P. Aeruginosa Pneumonia in Cystic Fibrosis

Molecular Medicine (Cambridge, Mass.). Feb, 2012  |  Pubmed ID: 22314397

Pulmonary infection with Pseudomonas (P.) aeruginosa and neutrophilic lung inflammation significantly contribute to morbidity and mortality in cystic fibrosis (CF). HMGB1, a ubiquitous DNA binding protein that promotes inflammatory tissue injury, is significantly elevated in CF sputum. However, its mechanistic and potential therapeutic implications in CF were previously unknown. We found that HMGB1 levels were significantly elevated in bronchoalveolar lavage fluids (BAL) of CF patients and CFTR(-/-) mice. Neutralizing anti-HMGB1 mAb conferred significant protection against P. aeruginosa-induced neutrophil recruitment, lung injury and bacterial infection in both CFTR(-/-) and wildtype mice. Alveolar macrophages isolated from mice treated with anti-HMGB1 mAb had improved phagocytic activity, which was suppressed by direct exposure to HMGB1. In addition, BAL from CF patients significantly impaired macrophage phagocytotic function and this impairment was attenuated by HMGB1-neutralizing antibodies. The HMGB1-mediated suppression of bacterial phagocytosis was attenuated in macrophages lacking toll-like receptor 4 (TLR4), suggesting a critical role for TLR4 in signaling HMGB1-mediated macrophage dysfunction. These studies demonstrate that the elevated levels of HMGB1 in CF airways are critical for neutrophil recruitment and persistent presence of P. aeruginosa in the lung. Thus, HMGB1 may provide a therapeutic target for reducing bacterial infection and lung inflammation in CF.

Complex Relationship Between Meiotic Recombination Frequency and Autosomal Synaptonemal Complex Length Per Cell in Normal Human Males

American Journal of Medical Genetics. Part A. Mar, 2012  |  Pubmed ID: 22315211

Although the relationship between meiotic recombination frequency and synaptonemal complex (SC) length has been of interest for a long time, how recombination frequency is related to SC length has not been carefully explored. To address this question, we have measured the meiotic recombination frequency as represented by MLH1 foci in 889 pachytene spermatocytes and measured the length of 19,558 autosomal SCs from 10 human males. A complex relationship between the number of MLH1 foci and total autosomal SC length per cell was observed. A positive correlation with significant correlation coefficients between the two variables was found in eight of the ten donors examined, with three donors showing weak correlation, and five showing moderate correlation. Two donors who did not show any correlation between the two variables were identified for the first time. Moreover, most cells with similar total autosomal SC length showed very different numbers of MLH1 foci both between individuals and even within an individual, and vice versa. Our data provide the first evidence for a complex relationship between the recombination frequency and total length of autosomal SCs per cell in human males.

Orthogonal-reference-pattern-modulated Shift Multiplexing for Collinear Holographic Data Storage

Optics Letters. Mar, 2012  |  Pubmed ID: 22378444

A novel hybrid shift multiplexing method for collinear holographic data storage (CHDS) by using orthogonal reference patterns (RPs) is proposed, analyzed, and demonstrated. For this method, holograms are multiplexed by not only shifting the media but also using different RPs. Compared with the traditional method, the shift pitch for the hybrid method is substantially reduced because of the selectivity introduced by different RPs. The interpage cross talk due to Bragg mismatch and degeneracy for multiplexing holograms in the same volume by using orthogonal RPs is also attenuated by utilizing the shift selectivity of the hologram. A 1.5 μm shift pitch is experimentally achieved by using three amplitude RPs in a system that would be 4.5 μm with only one RP. This new method offers an alternative to significantly increase the data density and transfer rate of the CHDS system given that the media has ideal properties.

Cloning and Characterization of Telomerase Reverse Transcriptase Gene in Trichinella Spiralis

Parasitology Research. Jan, 2012  |  Pubmed ID: 21748355

The telomerase reverse transcriptase (TERT) is primarily known for its ability to elongate telomeres for maintaining chromosomal integrity and delaying cellular senescence. It plays an important role in cell proliferation, differentiation, tumorigenesis, and aging. Telomerase includes two core components-an internal RNA moiety acting as a template of DNA extension and a catalytic subunit (TERT) which provides catalytic activity. Here, we described the cloning, sequence, and characterization of the TERT gene from Trichinella spiralis (T. spiralis). The prediction results of amino acid sequence showed that it possessed all the motifs characteristics of the TERT family members. T. spiralis TERT (Ts_TERT) cDNA contains an open reading frame encoding a protein with 1,201 amino acids with moleculer mass of 139 kDa and isoelectric point of 9.673, and the protein contains the conserved reverse transcriptase motifs 1, 2, A, B, C, D, and E, as well as the TERT-specific T motifs and the N-terminal conserved motifs GQ, CP, and QFP. While RT-PCR analysis indicates that TERT mRNA is expressed in T. spiralis adult worm, newborn larvae, and muscle larvae.

Toxoplasma Gondii Rhomboid Protein 1 (TgROM1) is a Potential Vaccine Candidate Against Toxoplasmosis

Veterinary Parasitology. Mar, 2012  |  Pubmed ID: 21906881

Infection with the intracellular protozoan parasite Toxoplasma gondii causes serious public health problems and is of great economic importance worldwide. The rhomboid proteins which are responsible for adhesion and invasion of host cells have been suggested as vaccine candidates against toxoplasmosis. A DNA vaccine (pVAX-ROM1) encoding T. gondii rhomboid protein 1 (TgROM1) gene was constructed and the immune response and protective efficacy of this vaccine against lethal challenge in BALB/c mice were evaluated. The results indicated that specific antibody and lymphocyte proliferative responses were elicited in mice receiving pVAX-ROM1. The production levels of IFN-γ, IL-2, IL-4, and IL-10, as well as the percentage of CD4(+) cells in mice vaccinated with pVAX-ROM1 were significantly increased respectively, compared to controls receiving either pVAX1 alone or PBS. After lethal challenge, the mice immunized with pVAX-ROM1 showed an increased survival time compared with the mice in the controls. Our data suggested that a DNA vaccine pVAX-ROM1 encoding T. gondii rhomboid protein 1 triggered strong humoral and cellular responses, and prolonged survival time against T. gondii infection in BALB/c mice.

Efficacy of Eimeria Tenella Rhomboid-like Protein As a Subunit Vaccine in Protective Immunity Against Homologous Challenge

Parasitology Research. Mar, 2012  |  Pubmed ID: 21845409

The immune responses and protective efficacy against homologous challenge in chickens elicited by recombinant proteins of a rhomboid-like gene (ETRHO1) from Eimeria tenella was investigated in the present study. When chickens were immunized with the recombinant rhomboid antigen, specific antibody was generated by ELISA assay. In comparison with the PBS group, the expression levels of interleukin-2, interferon-γ, as well as the percentages of CD4⁺ and CD8⁺ cells in the group immunized with the recombinant rhomboid proteins were significantly increased (p < 0.01, p < 0.05, and p < 0.05, respectively). These results suggest that rhomboid was capable of eliciting humoral and cell-mediated immunity response in birds. Challenge experiments demonstrated that the recombinant rhomboid protein could provide chickens with a protection rate around 77.3%. Numbers of oocysts and cecal lesion from chickens in the group immunized with recombinant rhomboid proteins decreased significantly, and the body weight increased significantly when compared with chickens in the PBS group (p < 0.05). These results suggested that the recombinant rhomboid antigen was able to impart partial protection against homologous challenge in chicken and could be a potential candidate for an E. tenella vaccine development.

The Viral RNA-based Transfection of Enhanced Green Fluorescent Protein (EGFP) in the Parasitic Protozoan Trichomonas Vaginalis

Parasitology Research. Mar, 2012  |  Pubmed ID: 21861063

Here we have developed methods to transiently and stably transfect the human pathogenic protist Trichomonas vaginalis. The viral RNA-based transfection vector pTVV-EGFP/NEO was constructed by using enhanced green fluorescent protein gene (EGFP) and neomycin resistance gene (NEO) in tandem to replace the whole gene encoding region of T. vaginalis virus (TVV). The in vitro transcripts of linearized pTVV-EGFP/NEO were electroporated into trophozoites and the transfectants transiently expressed EGFP after 16 h postincubation. Stable expression of EGFP was persistently detected by fluorescence microscopy and by RT-PCR in transfected trophozoites under G418 selection. Our study provides a novel and valuable approach for genetic study of T. vaginalis.

Acid-sensing Ion Channel Subtype 3 Function and Immunolabelling Increases in Skeletal Muscle Sensory Neurons Following Femoral Artery Occlusion

The Journal of Physiology. Mar, 2012  |  Pubmed ID: 22183722

Abstract  Sympathetic nerve activity and arterial blood pressure responses to static hindlimb muscle contractions are greater in rats with femoral arteries that were previously ligated (24-72 h earlier) than in control rats. Studies further demonstrate that acid-sensing ion channel subtype 3 (ASIC(3)) in thin-fibre muscle afferents contributes to the amplified reflex muscle responses observed in occluded rats, probably due to enhanced ASIC(3) expression in muscle sensory neurons. The purpose of this study was to characterize acid-induced current with activation of ASIC(3) in dorsal root ganglion (DRG) neurons of control rats and rats with 24 h of femoral occlusion using whole-cell patch clamp methods. Also, immunohistochemistry was employed to examine existence of ASIC(3) expression in DRG neurons of thin-fibre afferents. DRG neurons from 4- to 6-week-old rats were labelled by injecting the fluorescence tracer DiI into the hindlimb muscles 4-5 days prior to the recording experiments. The results of this study show that ∼90% of current responses evoked by pH 6.7 in DRG neurons innervating the hindlimb muscles are ASIC(3)-like. The peak current amplitude to pH 6.7 is significantly attenuated with application of rAPETx2, a specific ASIC(3) antagonist. In addition, ASIC(3)-like current responses to pH 6.7 are observed in small, medium and large DRG neurons, and size distribution of DRG neurons is similar in control and occluded animals. However, the peak current amplitude of DRG neuron response induced by ASIC(3) stimulation is larger in occluded rats than that in control rats. Moreover, the percentage of DRG neurons with ASIC(3)-like currents is greater after arterial occlusion compared with control. Furthermore, results from double immunofluorescence experiments show that femoral artery occlusion mainly augments ASIC(3) expression within DRG neurons projecting C-fibre afferents. Taken together, these data suggest that (1) the majority of current responses to pH 6.7 are ASIC(3)-like in DRG neurons with nerve endings in the hindlimb muscles, (2) a greater acid-induced current responding to pH 6.7 develops when hindlimb arterial blood supply is deficient under ischaemic conditions, and (3) increased ASIC(3) expression is largely observed in thin C-fibres of DRG neurons after hindlimb ischaemia.

Toxoplasma Gondii: Protective Immunity Against Toxoplasmosis with Recombinant Actin Depolymerizing Factor Protein in BALB/c Mice

Experimental Parasitology. Mar, 2012  |  Pubmed ID: 22248986

Toxoplasmosis is one of the most world-wide spread zoonosis representing a very serious clinical and veterinary problem. There is still need for vaccines for toxoplasmosis. In the present study, we evaluated the protective efficacy of a recombinant actin depolymerizing factor (ADF) subunit vaccine against Toxoplasma gondii infection in BALB/c mice. The recombinant T. gondii ADF protein (rADF) was expressed in Escherichia coli and used as antigens for BALB/c mice immunization. The results indicated that specific antibody and the increased percentage of CD4(+) T lymphocyte were found in vaccinated BALB/c mice with rADF, when compared with adjuvant or PBS groups. After challenged with T. gondii (RH strain) tachyzoites, the survival time of the mice in rADF group was longer than those in the control group. The numbers of brain cysts of the mice in rADF group reduced significantly when compared with those in control groups (P<0.05), and the rate of reduction could reach to around 30%. These results suggest that rADF can generate protective immunity against T. gondii infection in BALB/c mice.