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Find video protocols related to scientific articles indexed in Pubmed.
Evaluation of memory endophenotypes for association with CLU, CR1, and PICALM variants in black and white subjects.
Alzheimers Dement
PUBLISHED: 11-18-2014
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Genetic variants at the CLU, CR1, and PICALM loci associate with risk for late-onset Alzheimer's disease (LOAD) in genomewide association studies. In this study, our aim was to determine whether the LOAD risk variants at these three loci influence memory endophenotypes in black and white subjects.
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Akt1 and Akt3 exert opposing roles in the regulation of vascular tumor growth.
Cancer Res.
PUBLISHED: 11-13-2014
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Vascular tumors are endothelial cell neoplasms whose mechanisms of tumorigenesis are poorly understood. Moreover, current therapies, particularly those for malignant lesions, have little beneficial effect on clinical outcomes. In this study, we show that show that endothelial activation of the Akt1 kinase is sufficient to drive de novo tumor formation. Mechanistic investigations uncovered opposing functions for different Akt isoforms in this regulation, where Akt1 promotes and Akt3 inhibits vascular tumor growth. Akt3 exerted negative effects on tumor endothelial cell growth and migration by inhibiting activation of the translation regulatory kinase S6K through modulation of Rictor expression. S6K in turn acted through a negative feedback loop to restrain Akt3 expression. Conversely, S6K signaling was increased in vascular tumor cells where Akt3 was silenced, and the growth of these tumor cells was inhibied by a novel S6K inhibitor. Overall, our findings offer a preclinical proof of concept for the therapeutic utility of treating vascular tumors such as angiosarcomas with S6K inhibitors.
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Pilot Study of a Novel Population of Head and Neck Cancer Patients in the CRN.
Otolaryngol Head Neck Surg
PUBLISHED: 11-12-2014
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Head and neck cancers are challenging to study because of their relatively low incidence. A large, novel population of patients with head and neck cancers that has not been previously studied and distinct from the referral populations has been identified. The National Cancer Institute-funded Health Maintenance Organization Cancer Research Network is a consortium of 15 nonprofit research centers based in large, vertically integrated health care delivery organizations across the United States. They represent a geographically, racially, and socioeconomically diverse population. These community-based organizations provide care to approximately 10 million individuals and 57,692 patients with head and neck cancer. This pilot study and preliminary analysis seeks to demonstrate the potential this network holds as a resource for clinical cancer research and to identify it as a unique resource that allows for more detailed queries than are currently available to researchers.
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Genomicus update 2015: KaryoView and MatrixView provide a genome-wide perspective to multispecies comparative genomics.
Nucleic Acids Res.
PUBLISHED: 11-08-2014
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The Genomicus web server (http://www.genomicus.biologie.ens.fr/genomicus) is a visualization tool allowing comparative genomics in four different phyla (Vertebrate, Fungi, Metazoan and Plants). It provides access to genomic information from extant species, as well as ancestral gene content and gene order for vertebrates and flowering plants. Here we present the new features available for vertebrate genome with a focus on new graphical tools. The interface to enter the database has been improved, two pairwise genome comparison tools are now available (KaryoView and MatrixView) and the multiple genome comparison tools (PhyloView and AlignView) propose three new kinds of representation and a more intuitive menu. These new developments have been implemented for Genomicus portal dedicated to vertebrates. This allows the analysis of 68 extant animal genomes, as well as 58 ancestral reconstructed genomes. The Genomicus server also provides access to ancestral gene orders, to facilitate evolutionary and comparative genomics studies, as well as computationally predicted regulatory interactions, thanks to the representation of conserved non-coding elements with their putative gene targets.
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F3 -Isoprostanes as a Measure of in vivo Oxidative Damage in Caenorhabditis elegans.
Curr Protoc Toxicol
PUBLISHED: 11-08-2014
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Oxidative stress has been implicated in the development of a wide variety of disease processes, including cardiovascular disease, cancer, and neurodegenerative diseases, as well as progressive and normal aging processes. Isoprostanes (IsoPs) are prostaglandin-like compounds that are generated in vivo from lipid peroxidation of arachidonic acid (AA, C20:4, ?-6) and other polyunsaturated fatty acids (PUFA). Since the discovery of IsoPs by Morrow and Roberts in 1990, quantification of IsoPs has been shown to be an excellent source of biomarkers of in vivo oxidative damage. Eicosapentaenoic acid (EPA, C20:5, ?-3) is the most abundant PUFA in Caenorhabditis elegans and gives rise to F3 -IsoPs upon nonenzymatic free-radical-catalyzed lipid peroxidation. The protocol presented is the current methodology that our laboratory uses to quantify F3 -IsoPs in C. elegans using gas chromatography/mass spectrometry (GC/MS). The methods described herein have been optimized and validated to provide the best sensitivity and selectivity for quantification of F3 -IsoPs from C. elegans lysates. © 2014 by John Wiley & Sons, Inc.
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Objective Measures of Activity Level and Mortality in Older Men.
J Am Geriatr Soc
PUBLISHED: 11-03-2014
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To examine associations between objective measures of activity level and mortality risk in older men.
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Natural History and Malignant Transformation in Recurrent Respiratory Papillomatosis: Human Papillomavirus (HPV), Dysplasia and an Autopsy Review.
Fetal Pediatr Pathol
PUBLISHED: 10-30-2014
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Introduction: Recurrent respiratory papillomatosis (RRP) is a human papillomavirus (HPV) related disease in both children and adults, characterized by recurrent benign squamous papillomas of the respiratory mucosa. Malignant transformation is rare. The present report concerns the natural history of RRP in two children. Materials and Methods: Clinical records, autopsy material and tissue from previous surgical excisions were reviewed in both cases. Select surgical and autopsy specimens were examined using p16 immunohistochemistry and in-situ hybridization for low and high risk HPV. Results: Both children had pulmonary involvement with incidental invasive keratinizing squamous carcinoma of the lung at autopsy. Low-risk HPV was present in the papillomas and carcinoma at autopsy in both cases. Conclusions: The autopsy examinations in these two cases emphasize the serious, if uncommon, pulmonary complications of this disease. In conjunction with previously reported autopsies, destructive lung disease may be as frequent a cause of death as disseminated malignancy.
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Application of crude and recombinant ELISAs and immunochromatographic test for serodiagnosis of animal trypanosomosis in the Umkhanyakude district of KwaZulu-Natal province, South Africa.
J. Vet. Med. Sci.
PUBLISHED: 10-25-2014
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A total of 231 serum samples were collected from sheep (n=9), goats (n=99) and cattle (n=123) in northeastern KwaZulu-Natal, South Africa. Trypanosome infection was detected using Trypanosoma brucei brucei crude antigen (TbbCA) and T. congolense crude antigen (TcoCA) ELISA assays. Recombinant antigen (T. evansi GM6 which consisted of 4 repeat domains, TeGM6-4r) ELISA and immunochromatographic test (ICT) were also used. Crude antigen ELISA, TeGM6-4r-ELISA and ICT detected 27.3%, 29% and 19.9% of trypanosome seropositive samples, respectively. Trypanosome infection prevalence in cattle and goats was 35.8-46.3% and 0-9.1%, respectively. Out of 9 sheep serum samples, 2-4 sera (22.2-44.4%) were positive. The detection performance of crude and recombinant antigen ELISAs was relatively similar (K=0.6-0.7); both are recommended for reference diagnosis and large scale epidemiological surveys. There is potential application for ICT in on-site diagnosis, but its sensitivity should be improved.
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A Nationally-Representative Epidemiological and Risk Factor Assessment of Child Mental Health in Vietnam.
Int Perspect Psychol
PUBLISHED: 10-21-2014
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As part of the global mental health movement's focus on identifying and reducing international disparities, this study conducted the first nationally representative child mental health epidemiological survey in Vietnam. We assessed as risk/protective factors several family social structure characteristics (e.g., presence of grandparents, number of siblings in the home) of particular relevance to non-Western countries. Epidemiological data using the Child Behavior Checklist and the Strengths and Difficulties Questionnaire were collected at 60 sites in 10 of Vietnam's 63 provinces selected to provide a nationally representative sample, which included 1,314 adult informants of children 6-16 years of age, and 591 children aged 12-16. Vietnamese children's mental health functioning was reported overall to be better by approximately a third standard deviation than the international average; this international difference was particularly large for externalizing (behavior) problems as compared to internalizing (emotional) problems, suggesting that a cultural problem suppression model may be operating in Vietnam. Significant variability in mental health problems was found across provinces, emphasizing the need for nationally representative samples when conducting child mental health epidemiological surveys. Contrary to many other studies, in Vietnam higher SES was found to be a risk factor for attention/hyperactivity problems.
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Declining Prevalence of Tobacco Smoking in Vietnam.
Nicotine Tob. Res.
PUBLISHED: 10-19-2014
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To supplement limited information on tobacco use in Vietnam, data from a nationally-representative population-based survey was used to estimate the prevalence of smoking among 25-64 year-olds.
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Deletion of ADORA2B from myeloid cells dampens lung fibrosis and pulmonary hypertension.
FASEB J.
PUBLISHED: 10-17-2014
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Idiopathic pulmonary fibrosis (IPF) is a lethal, fibroproliferative disease. Pulmonary hypertension (PH) can develop secondary to IPF and increase mortality. Alternatively, activated macrophages (AAMs) contribute to the pathogenesis of both IPF and PH. Here we hypothesized that adenosine signaling through the ADORA2B on AAMs impacts the progression of these disorders and that conditional deletion of ADORA2B on myeloid cells would have a beneficial effect in a model of these diseases. Conditional knockout mice lacking ADORA2B on myeloid cells (Adora2B(f/f)-LysM(Cre)) were exposed to the fibrotic agent bleomycin (BLM; 0.035 U/g body weight, i.p.). At 14, 17, 21, 25, or 33 d after exposure, SpO2, bronchoalveolar lavage fluid (BALF), and histologic analyses were performed. On day 33, lung function and cardiovascular analyses were determined. Markers for AAM and mediators of fibrosis and PH were assessed. Adora2B(f/f)-LysM(Cre) mice presented with attenuated fibrosis, improved lung function, and no evidence of PH compared with control mice exposed to BLM. These findings were accompanied by reduced expression of CD206 and arginase-1, markers for AAMs. A 10-fold reduction in IL-6 and a 5-fold decrease in hyaluronan, both linked to lung fibrosis and PH, were also observed. These data suggest that activation of the ADORA2B on macrophages plays an active role in the pathogenesis of lung fibrosis and PH.-Karmouty-Quintana, H., Philip, K., Acero, L. F., Chen, N.-Y., Weng, T., Molina, J. G., Luo, F., Davies, J., Le, N.-B., Bunge, I., Volcik, K. A., Le, T.-T. T., Johnston, R. A., Xia, Y., Eltzschig, H. K., Blackburn, M. R. Deletion of ADORA2B from myeloid cells dampens lung fibrosis and pulmonary hypertension.
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Oxidation of Alcohols and Activated Alkanes with Lewis Acid-Activated TEMPO.
Inorg Chem
PUBLISHED: 10-16-2014
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The reactivity of MCl3(?(1)-TEMPO) (M = Fe, 1; Al, 2; TEMPO = 2,2,6,6-tetramethylpiperidine-N-oxyl) with a variety of alcohols, including 3,4-dimethoxybenzyl alcohol, 1-phenyl-2-phenoxyethanol, and 1,2-diphenyl-2-methoxyethanol, was investigated using NMR spectroscopy and mass spectrometry. Complex 1 was effective in cleanly converting these substrates to the corresponding aldehyde or ketone. Complex 2 was also able to oxidize these substrates; however, in a few instances the products of overoxidation were also observed. Oxidation of activated alkanes, such as xanthene, by 1 or 2 suggests that the reactions proceed via an initial 1-electron concerted proton-electron transfer (CPET) event. Finally, reaction of TEMPO with FeBr3 in Et2O results in the formation of a mixture of FeBr3(?(1)-TEMPOH) (23) and [FeBr2(?(1)-TEMPOH)]2(?-O) (24), via oxidation of the solvent, Et2O.
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Programmable protein-DNA hybrid hydrogels for the immobilization and release of functional proteins.
Chem. Commun. (Camb.)
PUBLISHED: 10-15-2014
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A modular approach for the precise assembly of multi-component hydrogels consisting of protein and DNA building blocks is described for the first time. Multi-arm DNA is designed for crosslinking and stepwise, non-covalent assembly of active proteins inside the hydrogel.
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Promotion and provision of colorectal cancer screening: a comparison of colorectal cancer control program grantees and nongrantees, 2011-2012.
Prev Chronic Dis
PUBLISHED: 10-03-2014
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Since 2009, the Centers for Disease Control and Prevention (CDC) has awarded nearly $95 million to 29 states and tribes through the Colorectal Cancer Control Program (CRCCP) to fund 2 program components: 1) providing colorectal cancer (CRC) screening to uninsured and underinsured low-income adults and 2) promoting population-wide CRC screening through evidence-based interventions identified in the Guide to Community Preventive Services (Community Guide). CRCCP is a new model for disseminating and promoting use of evidence-based interventions. If the program proves successful, CDC may adopt the model for future cancer control programs. The objective of our study was to compare the colorectal cancer screening practices of recipients of CRCCP funding (grantees) with those of nonrecipients (nongrantees).
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Association between chlorinated pesticides in the serum of prepubertal Russian boys and longitudinal biomarkers of metabolic function.
Am. J. Epidemiol.
PUBLISHED: 09-25-2014
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Organochlorine pesticides (OCPs) have been linked to adult metabolic disorders; however, few studies have examined these associations in childhood. We prospectively evaluated the associations of baseline serum OCPs (hexachlorobenzene, ?-hexachlorocyclohexane, and p,p'-dichlorodiphenyldichloroethylene) in Russian boys with subsequent repeated measurements of serum glucose, insulin, lipids, leptin, and calculated homeostatic model assessment of insulin resistance (IR). During 2003-2005, we enrolled 499 boys aged 8-9 years in a prospective cohort; 318 had baseline serum OCPs and serum biomarkers measured at ages 10-13 years. Multivariable generalized estimating equation and mediation regression models were used to examine associations and direct and indirect (via body mass index (BMI) (weight (kg)/height (m)(2))) effects of prepubertal OCP tertiles and quintiles with biomarkers. In multivariable models, higher p,p'-dichlorodiphenyldichloroethylene (quintile 5 vs. quintile 1) was associated with lower leptin, with relative mean decreases of 61.8% (95% confidence interval: 48.4%, 71.7%) in models unadjusted for BMI and 22.2% (95% confidence interval: 7.1%, 34.9%) in models adjusted for BMI; the direct effect of p,p'-dichlorodiphenyldichloroethylene on leptin accounted for 27% of the total effect. IR prevalence was 6.6% at ages 12-13 years. Higher hexachlorobenzene (tertile 3 vs. tertile 1) was associated with higher odds of IR in models adjusted for BMI (odds ratio = 4.37, 95% confidence interval: 1.44, 13.28). These results suggest that childhood OCPs may be associated with IR and lower leptin.
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Palliative care knowledge, attitudes and perceived self-competence of nurses working in Vietnam.
Int J Palliat Nurs
PUBLISHED: 09-25-2014
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To explore palliative care knowledge, attitudes and perceived self-competence of nurses working in oncology settings in Hanoi, Vietnam.
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Antenatal and early infant predictors of postnatal growth in rural Vietnam: a prospective cohort study.
Arch. Dis. Child.
PUBLISHED: 09-24-2014
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To determine which antenatal and early-life factors were associated with infant postnatal growth in a resource-poor setting in Vietnam.
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Scrub typhus in the northern provinces of Vietnam: an observational study of admissions to a national referral hospital.
Trans. R. Soc. Trop. Med. Hyg.
PUBLISHED: 09-23-2014
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Scrub typhus is a common cause of fever in parts of South East and Southern Asia. Little is known about the disease burden in Vietnam.
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In Vitro Study of CaTiO3-Hydroxyapatite Composites for Bone Tissue Engineering.
ASAIO J.
PUBLISHED: 09-20-2014
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A biocomposite composed of hydroxyapatite (HAp) and CaTiO3 was fabricated to study the phase stability, mechanical strength, and biocompatibility for bone tissue engineering. To investigate the optimal concentrations for the biocomposite, different HAp concentrations (0%, 50%, 70%, and 100%) were mixed with CaTiO3 and sintered in a microwave furnace. X-ray diffraction patterns of CaTiO3/HAp composites indicated the phase stability of CaTiO3/HAp. Mechanical properties were characterized by Vickers hardness, Young modulus, fracture toughness, brittleness, and compressive strength. MC3T3-E1 cells were used for in vitro studies to investigate the biocompatibility of CaTiO3/HAp composites, using 3-[4, 5-dimethylthiazol-2-yl]-2, 5-diphenyltetrazolium bromide (MTT) assay and immunofluorescence. The in vitro studies confirmed the highest cell viability on 70HAp at 1, 3, and 7 days. Collagen Type I, osteopontin, and osteocalcin expressions were evaluated by Western blotting and a strong signal of collagen Type I and osteopontin expression was shown by cells grown on 70HAp and 100HAp. Interestingly, osteocalcin signal was found only on 70HAp at day 7. The expression of alkaline phosphatase and osteopontin confirmed that the 70HAp expressed the strongest fluorescent signal as compared with pure materials. Thus considering the biological properties, 70HAp biocomposite was found ideal for bone tissue engineering.
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Platelet-rich plasma encapsulation in hyaluronic acid/gelatin-BCP hydrogel for growth factor delivery in BCP sponge scaffold for bone regeneration.
J Biomater Appl
PUBLISHED: 09-20-2014
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Microporous calcium phosphate based synthetic bone substitutes are used for bone defect healing. Different growth factor loading has been investigated for enhanced bone regeneration. The platelet is a cellular component of blood which naturally contains a pool of necessary growth factors that mediate initiation, continuation, and completion of cellular mechanism of healing. In this work, we have investigated the encapsulation and immobilization of platelet-rich plasma (PRP) with natural polymers like hyaluronic acid (HA) and gelatin (Gel) and loading them in a biphasic calcium phosphate (BCP) scaffold, for a synthetic-allologous hybrid scaffold. Effect of PRP addition in small doses was evaluated for osteogenic potential in vitro and in vivo. BCP (10%) mixed HA-Gel hydrogel with or without PRP, was loaded into a BCP sponge scaffold. We investigated the hydrogel-induced improvement in mechanical property and PRP-mediated enhancement in biocompatibility. In vitro studies for cytotoxicity, cell attachment, and proliferation were carried out using MC3T3-E1 pre-osteoblast cells. In in vitro studies, the cell count, cell proliferation, and cell survival were higher in the scaffold with PRP loading than without PRP. However, in the in vivo studies using a rat model, the PRP scaffold was not superior to the scaffold without PRP. This discrepancy was investigated in terms of the interaction of PRP in the in vivo environment.
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Quantitative ultrasensitive bright-field RNA in situ hybridization with RNAscope.
Methods Mol. Biol.
PUBLISHED: 09-15-2014
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RNA in situ hybridization (ISH) can provide valuable morphological context for molecular markers on one hand and enable morphological analysis in molecular context on the other hand. It has become increasingly important, thanks to increasing interest in new biomarkers and noncoding RNAs in both research and clinical applications. We have developed an ultrasensitive RNA ISH technology, RNAscope, employing a unique probe design strategy that allows target-specific signal amplification while suppressing background noise. This approach enables single RNA molecule detection in formalin-fixed paraffin-embedded (FFPE) specimens under standard bright-field microscopy and is capable of multiplex detection at the single cell level. After staining, target-specific signals appear as punctate dots present in individual cells in well-preserved tissue morphological context, which facilitates both semiquantitative manual scoring and software-assisted quantitative analysis. Here, we present detailed protocols of RNAscope for FFPE tissue sections. The step-by-step protocols describe tissue preparation, pretreatment, probe hybridization, signal amplification, visualization, and analysis. We also highlight the critical steps for ensuring successful staining.
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Dual-color ultrasensitive bright-field RNA in situ hybridization with RNAscope.
Methods Mol. Biol.
PUBLISHED: 09-15-2014
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In situ hybridization (ISH) techniques have been important to the study of gene expression signatures in cells and tissues. The ability to detect multiple targets simultaneously is especially valuable, since it allows dissecting gene expression of distinct cell types with precise cellular and subcellular resolution within morphological context. Recently, we have reported using a novel dual-color ultrasensitive bright-field RNA in situ hybridization for detection of clonally restricted immunoglobulin light chain mRNA expression in B cell lymphomas. Here, we present detailed protocols of RNAscope 2-Plex assays for FFPE tissue sections. The protocols describe the tissue preparation, pretreatment, probe hybridization, signal amplification, visualization, and analysis, as well as emphasize the critical steps for ensuring successful staining.
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Steroidal Constituents from the Edible Sea Urchin Diadema savignyi Michelin Induce Apoptosis in Human Cancer Cells.
J Med Food
PUBLISHED: 09-12-2014
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Abstract Bioassay-directed fractionation and purification were used to isolate 12 steroids (1-12) from a CH2Cl2 extract of the edible Vietnamese sea urchin Diadema savignyi Michelin. The cytotoxic activity of the CH2Cl2 extract and 12 steroids was evaluated in three human cancer cell lines (HL-60, PC-3, and SNU-C5). Relative to the effects of the positive control, mitoxantrone, the CH2Cl2 extract (with an inhibitory concentration of 50% [IC50] values ranging from 1.37±0.15 to 3.11±0.15 ?g/mL) and compounds 2 (with IC50 values ranging from 5.29±0.11 to 6.80±0.67 ?M) and 11 (with IC50 values ranging from 4.95±0.07 to 6.99±0.28 ?M) exhibited potent cytotoxic effects against all three tested human cancer cell lines. In addition, the CH2Cl2 extract and compounds 2 and 11 were found to induce apoptosis. The induction of apoptosis was accompanied by alterations of the apoptosis-related protein expression, inactivation of ERK1/2 mitogen-activated protein kinase signaling, and decreased c-Myc expression. These data suggest that compounds 2 and 11 from the edible sea urchin D. savignyi may have potential for the treatment of colon cancer, leukemia, and prostate cancer as complementary cancer remedies.
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Inhibition of group IVA cytosolic phospholipase A2 by thiazolyl ketones in vitro, ex vivo, and in vivo.
J. Med. Chem.
PUBLISHED: 09-03-2014
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Group IVA cytosolic phospholipase A2 (GIVA cPLA2) is the rate-limiting provider of pro-inflammatory mediators in many tissues and is thus an attractive target for the development of novel anti-inflammatory agents. In this work, we present the synthesis of new thiazolyl ketones and the study of their activities in vitro, in cells, and in vivo. Within this series of compounds, methyl 2-(2-(4-octylphenoxy)acetyl)thiazole-4-carboxylate (GK470) was found to be the most potent inhibitor of GIVA cPLA2, exhibiting an XI(50) value of 0.011 mole fraction in a mixed micelle assay and an IC50 of 300 nM in a vesicle assay. In a cellular assay using SW982 fibroblast-like synoviocytes, it suppressed the release of arachidonic acid with an IC50 value of 0.6 ?M. In a prophylactic collagen-induced arthritis model, it exhibited an anti-inflammatory effect comparable to the reference drug methotrexate, whereas in a therapeutic model, it showed results comparable to those of the reference drug Enbrel. In both models, it significantly reduced plasma PGE2 levels.
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Adenosine promotes vascular barrier function in hyperoxic lung injury.
Physiol Rep
PUBLISHED: 09-01-2014
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Hyperoxic lung injury is characterized by cellular damage from high oxygen concentrations that lead to an inflammatory response in the lung with cellular infiltration and pulmonary edema. Adenosine is a signaling molecule that is generated extracellularly by CD73 in response to injury. Extracellular adenosine signals through cell surface receptors and has been found to be elevated and plays a protective role in acute injury situations. In particular, ADORA2B activation is protective in acute lung injury. However, little is known about the role of adenosine signaling in hyperoxic lung injury. We hypothesized that hyperoxia-induced lung injury leads to CD73-mediated increases in extracellular adenosine, which is protective through ADORA2B signaling pathways. To test this hypothesis, we exposed C57BL6, CD73(-/-), and Adora2B(-/-) mice to 95% oxygen or room air and examined markers of pulmonary inflammation, edema, and monitored lung histology. Hyperoxic exposure caused pulmonary inflammation and edema in association with elevations in lung adenosine levels. Loss of CD73-mediated extracellular adenosine production exacerbated pulmonary edema without affecting inflammatory cell counts. Furthermore, loss of the ADORA2B had similar results with worsening of pulmonary edema following hyperoxia exposure without affecting inflammatory cell infiltration. This loss of barrier function correlated with a decrease in occludin in pulmonary vasculature in CD73(-/-) and Adora2B(-/-) mice following hyperoxia exposure. These results demonstrate that exposure to a hyperoxic environment causes lung injury associated with an increase in adenosine concentration, and elevated adenosine levels protect vascular barrier function in hyperoxic lung injury through the ADORA2B-dependent regulation of occludin.
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Mathematical modeling of bacterial kinetics to predict the impact of antibiotic colonic exposure and treatment duration on the amount of resistant enterobacteria excreted.
PLoS Comput. Biol.
PUBLISHED: 09-01-2014
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Fecal excretion of antibiotics and resistant bacteria in the environment are major public health threats associated with extensive farming and modern medical care. Innovative strategies that can reduce the intestinal antibiotic concentrations during treatments are in development. However, the effect of lower exposure on the amount of resistant enterobacteria excreted has not been quantified, making it difficult to anticipate the impact of these strategies. Here, we introduce a bacterial kinetic model to capture the complex relationships between drug exposure, loss of susceptible enterobacteria and growth of resistant strains in the feces of piglets receiving placebo, 1.5 or 15 mg/kg/day ciprofloxacin, a fluoroquinolone, for 5 days. The model could well describe the kinetics of drug susceptible and resistant enterobacteria observed during treatment, and up to 22 days after treatment cessation. Next, the model was used to predict the expected amount of resistant enterobacteria excreted over an average piglet's lifetime (150 days) when varying drug exposure and treatment duration. For the clinically relevant dose of 15 mg/kg/day for 5 days, the total amount of resistant enterobacteria excreted was predicted to be reduced by 75% and 98% when reducing treatment duration to 3 and 1 day treatment, respectively. Alternatively, for a fixed 5-days treatment, the level of resistance excreted could be reduced by 18%, 33%, 57.5% and 97% if 3, 5, 10 and 30 times lower levels of colonic drug concentrations were achieved, respectively. This characterization on in vivo data of the dynamics of resistance to antibiotics in the colonic flora could provide new insights into the mechanism of dissemination of resistance and can be used to design strategies aiming to reduce it.
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Blockade of IL-6 Trans signaling attenuates pulmonary fibrosis.
J. Immunol.
PUBLISHED: 08-29-2014
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Idiopathic pulmonary fibrosis (IPF) is a lethal lung disease with progressive fibrosis and death within 2-3 y of diagnosis. IPF incidence and prevalence rates are increasing annually with few effective treatments available. Inhibition of IL-6 results in the attenuation of pulmonary fibrosis in mice. It is unclear whether this is due to blockade of classical signaling, mediated by membrane-bound IL-6R?, or trans signaling, mediated by soluble IL-6R? (sIL-6R?). Our study assessed the role of sIL-6R? in IPF. We demonstrated elevations of sIL-6R? in IPF patients and in mice during the onset and progression of fibrosis. We demonstrated that protease-mediated cleavage from lung macrophages was important in production of sIL-6R?. In vivo neutralization of sIL-6R? attenuated pulmonary fibrosis in mice as seen by reductions in myofibroblasts, fibronectin, and collagen in the lung. In vitro activation of IL-6 trans signaling enhanced fibroblast proliferation and extracellular matrix protein production, effects relevant in the progression of pulmonary fibrosis. Taken together, these findings demonstrate that the production of sIL-6R? from macrophages in the diseased lung contributes to IL-6 trans signaling that in turn influences events crucial in pulmonary fibrosis.
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A new phenolic component from Triticum aestivum sprouts and its effects on LPS-stimulated production of nitric oxide and TNF-? in RAW 264.7 cells.
Phytother Res
PUBLISHED: 08-29-2014
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An unusual new phenolic component, triticumoside (1), and eight known compounds, isoorientin (2), isoscoparin (3), (2R)-2-O-?-D-glucopyranosyloxy-4,7-dimethoxy-2H-1,4-benzoxazin-3(4H)-one (4), adenosine (5), ?-sitosterol (6), daucosterol (7), 6?-O-linolenoyl daucosterol (8), ?-tocopherol (9), were isolated fromTriticum aestivum sprouts. The hybrid structure of 1, which is a hybrid between a flavone and a polyoxygenated benzene, is rarely found in natural sources. In addition, the effects of these compounds on LPS-induced NO and TNF-? production in RAW 264.7 cells were evaluated. At a concentration of 2.0 ?M, compounds 2-4 significantly inhibited the production of both NO and TNF-?. Compound 1 exhibited inhibitory activity on the secretion of TNF-? at concentrations as low as 2.0 ?M, but it did not reduce NO levels at any of the tested concentrations.
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Analyzing feed-forward loop relationship in aging phenotypes: Physical activity and physical performance.
Mech. Ageing Dev.
PUBLISHED: 08-26-2014
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We present evidence of feed-forward loop relationships and positive association between physical activity and performance levels, which are components of frailty, using measures from 431 high functioning women initially aged 70-79 years followed over 7 visits. Physical activity levels were assessed using a questionnaire. Grip strength was measured using a handheld dynamometer and usual walking speed was measured over 4-m. The results suggest that a reduction in physical activity would not only degrade physical performance, but it would further reduce physical activity through declines in physical performance. As both physical activity and physical performance impact frailty, improvement of physical activity could help reduce frailty directly as well as indirectly via improved physical performance. Our findings support a priori hypothesis that feed-forward loops are present in the phenotype of frailty, which is due to dysregulated energetics. A methodologically broader implication is that we introduce modeling and analysis of feed-forward loop data here. The feed-forward loop, as we define it, is different from the concept of feedback loops used in biochemical systems. Generalizing our model of two-variable feed-forward loop, three, four or multivariable feed-forward loop can be applied to other biological systems.
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Liposomal melatonin rescues methamphetamine-elicited mitochondrial burdens, proapoptosis, and dopaminergic degeneration through the inhibition PKC? gene.
J. Pineal Res.
PUBLISHED: 08-24-2014
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We have demonstrated that mitochondrial oxidative damage and PKC? overexpression contribute to methamphetamine-induced dopaminergic degeneration. Although it is recognized that antioxidant melatonin is effective in preventing neurotoxicity induced by methamphetamine, its precise mechanism remains elusive. C57BL/6J wild type mice exhibited a similar degree of dopaminergic deficit when methamphetamine was administered during light and dark phases. Furthermore, dopaminergic neuroprotection by genetic inhibition of PKC? during the light phase was comparable to that during the dark phase. Thus, we have focused on the light phase in order to examine whether melatonin modulates PKC?-mediated neurotoxic signaling after multiple high doses of methamphetamine. To enhance the bioavailability of melatonin, we applied liposomal melatonin. Treatment with methamphetamine resulted in hyperthermia, mitochondrial translocation of PKC?, oxidative damage (mitochondria > cytosol), mitochondrial dysfunction, pro-apoptotic changes, ultrastructural mitochondrial degeneration, dopaminergic degeneration, and behavioral impairment in wild type mice. Treatment with liposomal melatonin resulted in a dose-dependent attenuation against degenerative changes induced by methamphetamine in wild type mice. Attenuation by liposomal melatonin might be comparable to that by genetic inhibition (using PKC?((-/-)) mice or PKC? antisense oligonucleotide). However, liposomal melatonin did not show any additional protective effects on the attenuation by genetic inhibition of PKC?. Our results suggest that the circadian cycle cannot be a key factor in modulating methamphetamine toxicity under the current experimental condition, and that PKC? is one of the critical target genes for melatonin-mediated protective effects against mitochondrial burdens (dysfunction), oxidative stress, pro-apoptosis, and dopaminergic degeneration induced by methamphetamine. This article is protected by copyright. All rights reserved.
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Strength and Function Response to Clinical Interventions of Older Women Categorized by Weakness and Low Lean Mass Using Classifications From the Foundation for the National Institute of Health Sarcopenia Project.
J. Gerontol. A Biol. Sci. Med. Sci.
PUBLISHED: 08-18-2014
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The Foundation for the National Institutes of Health Sarcopenia Project developed data-driven cut-points for clinically meaningful weakness and low lean body mass. This analysis describes strength and function response to interventions based on these classifications.
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Nuclear translocation of hARD1 contributes to proper cell cycle progression.
PLoS ONE
PUBLISHED: 08-18-2014
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Arrest defective 1 (ARD1) is an acetyltransferase that is highly conserved across organisms, from yeasts to humans. The high homology and widespread expression of ARD1 across multiple species and tissues signify that it serves a fundamental role in cells. Human ARD1 (hARD1) has been suggested to be involved in diverse biological processes, and its role in cell proliferation and cancer development has been recently drawing attention. However, the subcellular localization of ARD1 and its relevance to cellular function remain largely unknown. Here, we have demonstrated that hARD1 is imported to the nuclei of proliferating cells, especially during S phase. Nuclear localization signal (NLS)-deleted hARD1 (hARD1?N), which can no longer access the nucleus, resulted in cell morphology changes and cellular growth impairment. Notably, hARD1?N-expressing cells showed alterations in the cell cycle and the expression levels of cell cycle regulators compared to hARD1 wild-type cells. Furthermore, these effects were rescued when the nuclear import of hARD1 was restored by exogenous NLS. Our results show that hARD1 nuclear translocation mediated by NLS is required for cell cycle progression, thereby contributing to proper cell proliferation.
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Prepubertal organochlorine pesticide concentrations and age of pubertal onset among Russian boys.
Environ Int
PUBLISHED: 08-10-2014
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In animal studies, organochlorine pesticide (OCP) exposure alters pubertal development; however, epidemiological data are limited and inconsistent.
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Late-onset Alzheimer's risk variants in memory decline, incident mild cognitive impairment, and Alzheimer's disease.
Neurobiol. Aging
PUBLISHED: 08-04-2014
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We tested association of nine late-onset Alzheimer's disease (LOAD) risk variants from genome-wide association studies (GWAS) with memory and progression to mild cognitive impairment (MCI) or LOAD (MCI/LOAD) in older Caucasians, cognitively normal at baseline and longitudinally evaluated at Mayo Clinic Rochester and Jacksonville (n>2000). Each variant was tested both individually and collectively using a weighted risk score. APOE-e4 associated with worse baseline memory and increased decline with highly significant overall effect on memory. CLU-rs11136000-G associated with worse baseline memory and incident MCI/LOAD. MS4A6A-rs610932-C associated with increased incident MCI/LOAD and suggestively with lower baseline memory. ABCA7-rs3764650-C and EPHA1-rs11767557-A associated with increased rates of memory decline in subjects with a final diagnosis of MCI/LOAD. PICALM-rs3851179-G had an unexpected protective effect on incident MCI/LOAD. Only APOE-inclusive risk scores associated with worse memory and incident MCI/LOAD. The collective influence of the nine top LOAD GWAS variants on memory decline and progression to MCI/LOAD appears limited. Discovery of biologically functional variants at these loci may uncover stronger effects on memory and incident disease.
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Chemical constituents of Triticum aestivum and their effects on adipogenic differentiation of 3T3-L1 preadipocytes.
Arch. Pharm. Res.
PUBLISHED: 07-25-2014
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In this report, we investigated the anti-obesity effect of wheat sprouts and their component compounds. Twenty compounds (1-20) were isolated from Triticum aestivum. Among them, glycolipids 1-5 were determined for the first time from T. aestivum and its sprouts. The HPLC analysis demonstrated that compounds 1-3, 5, 8, 12, and 14 were major peak in the HPLC chromatogram of the active fraction. The effects of the compounds on lipid accumulation were assessed at concentrations ranging from 1.0 to 100 ?M. At concentration of 10.0 ?M, compounds 1-7, 10-15, and 17-19 significantly decreased lipid accumulation in 3T3-L1 preadipocytes. Glycolipids 1, 2, and phenolic 17 significantly reduced lipid accumulation in the differentiated adipocytes in a concentration-dependent manner. Quantitative analysis based on measurement of the optical density of Oil Red O indicated that, at 100 ?M, compounds 1, 2, and 17 reduced lipid accumulation by 41, 37, and 48 %, respectively, compared with the positive control.
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A new phenylpropanoid and an alkylglycoside from Piper retrofractum leaves with their antioxidant and ?-glucosidase inhibitory activity.
Bioorg. Med. Chem. Lett.
PUBLISHED: 07-25-2014
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Two new compounds, piperoside (1) and isoheptanol 2(S)-O-?-D-xylopyranosyl (1?6)-O-?-D-glucopyranoside (11), along with 10 known compounds 3,4-dihydroxyallylbenzene (2), 1,2-di-O-?-D-glucopyranosyl-4-allylbenzene (3), tachioside (4), benzyl-O-?-D-glucopyranoside (5), icariside F2 (6), dihydrovomifoliol-3'-O-?-D-glucopyranoside (7), isopropyl O-?-D-glucopyranoside (8), isopropyl primeveroside (9), n-butyl O-?-D-glucopyranoside (10), isoheptanol 2(S)-O-?-D-apiofuranosyl-(1?6)-O-?-D-glucopyranoside (12), were isolated from the leaves of Piper retrofractum. Their structures were determined from 1D-NMR, 2D-NMR, and HR-ESI-MS spectral, a modified Mosher's method, and comparisons with previous reports. All of the isolated compounds showed modest ?-glucosidase inhibitory (4.60±1.74% to 11.97±3.30%) and antioxidant activities under the tested conditions.
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Triterpene saponins from the sea cucumber Stichopus chloronotus.
Nat Prod Commun
PUBLISHED: 07-17-2014
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Sea cucumbers have been used as a dietary delicacy and important ingredient in Asian traditional medicine and functional foods over many centuries. Using combined chromatographic methods, six triterpene saponins (1-6), including a new compound, stichloroside F (1), were isolated from a methanol extract of the sea cucumber Stichopus chloronotus Brandt. Their structures were determined on the basis of spectroscopic (1H and 13C NMR, HSQC, HMBC, 1H-1lH COSY, ROESY) and FTICR-MS data and by comparison with literature values.
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Burden of care for persons with disabilities in Vietnam.
Health Soc Care Community
PUBLISHED: 07-10-2014
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Vietnam has more than 6 million persons with disabilities (PWD), or 7.8% of the population. To provide better services for them, it is important to investigate the care they currently receive, and to obtain evidence on the health outcomes from that care. This study aimed to estimate the quality of life and functional status of a group of PWD in Vietnam and the cost of care they receive. This was an analytical study exploring the time and cost of informal care, the cost of illness (prevalence-based, patient perspective), quality of life using EuroQoL and functional status using the Barthel Index. The sample was selected from urban and rural areas of Quang Tri province in Central Vietnam, using systematic random sampling. Data were collected by face-to-face interviews, and in a 1-month diary recorded during July-August 2010 for summer and in December 2010 for winter. The costs are presented in 2010 USD values. The data were analysed by descriptive, univariate and multivariate statistics to summarise and explore the relationships among dependent and independent variables. The study sample included 210 PWD, with an average age of 38 years and duration of disability on average 26 years. The health-related quality of life measured in terms of the health utility score (0 = death, 1 = full health) was on average 0.44 and 0.39 in summer and winter respectively. The total cost of illness per year per case was USD 971 (83% of gross domestic product per capita); explanatory variables were the age of the PWD, receiving community-based rehabilitation, receiving government support and the severity of the disability. This illustrates the importance of services and support for reduction of the economic burden on the family. In conclusion, the results of this study provide information on the burden caused by disabilities in rural and urban households in Vietnam.
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Postnatal growth outcomes and influence of maternal gestational weight gain: a prospective cohort study in rural Vietnam.
BMC Pregnancy Childbirth
PUBLISHED: 07-08-2014
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Suboptimal weight gain during pregnancy may result in adverse outcomes for both the mother and child, including increased risk of pre-eclampsia and gestational diabetes, delivery of low birth weight and small-for-gestational age (SGA) infants, and preterm delivery. The objectives of this study were to identify maternal predictors of rate of weight gain in pregnancy, and to evaluate the association of gestational weight gain with infant postnatal growth outcomes.
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Expression, crystallization and preliminary X-ray crystallographic analysis of cystathionine ?-lyase from Acinetobacter baumannii OXA-23.
Acta Crystallogr F Struct Biol Commun
PUBLISHED: 06-17-2014
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Multidrug-resistant Acinetobacter baumannii (Ab) has emerged as a leading nosocomial pathogen because of its resistance to most currently available antibiotics. Cystathionine ?-lyase (CBL), a pyridoxal 5'-phosphate (PLP)-dependent enzyme, catalyzes the second step in the transsulfuration pathway, which is essential for the metabolic interconversion of the sulfur-containing amino acids homocysteine and methionine. The enzymes of the transsulfuration pathway are considered to be attractive drug targets owing to their specificity to microbes and plants. As a potential target for the development of novel antibacterial drugs, the AbCBL protein was expressed, purified and crystallized. An AbCBL crystal diffracted to 1.57?Å resolution and belonged to the trigonal space group P3112, with unit-cell parameters a = b = 102.9, c = 136.5?Å. The asymmetric unit contained two monomers, with a corresponding VM of 2.3?Å(3)?Da(-1) and a solvent content of 46.9%.
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IL-12-producing monocytes and HLA-E control HCMV-driven NKG2C+ NK cell expansion.
J. Clin. Invest.
PUBLISHED: 06-06-2014
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Human cytomegalovirus (HCMV) infection is the most common cause of congenital viral infections and a major source of morbidity and mortality after organ transplantation. NK cells are pivotal effector cells in the innate defense against CMV. Recently, hallmarks of adaptive responses, such as memory-like features, have been recognized in NK cells. HCMV infection elicits the expansion of an NK cell subset carrying an activating receptor heterodimer, comprising CD94 and NKG2C (CD94/NKG2C), a response that resembles the clonal expansion of adaptive immune cells. Here, we determined that expansion of this NKG2C+ subset and general NK cell recovery rely on signals derived from CD14+ monocytes. In a coculture system, a subset of CD14+ cells with inflammatory monocyte features produced IL-12 in response to HCMV-infected fibroblasts, and neutralization of IL-12 in this model substantially reduced CD25 upregulation and NKG2C+ subset expansion. Finally, blockade of CD94/NKG2C on NK cells or silencing of the cognate ligand HLA-E in infected fibroblasts greatly impaired expansion of NKG2C+ NK cells. Together, our results reveal that IL-12, CD14+ cells, and the CD94/NKG2C/HLA-E axis are critical for the expansion of NKG2C+ NK cells in response to HCMV infection. Moreover, strategies targeting the NKG2C+ NK cell subset have the potential to be exploited in NK cell-based intervention strategies against viral infections and cancer.
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CryptoDex: A randomised, double-blind, placebo-controlled phase III trial of adjunctive dexamethasone in HIV-infected adults with cryptococcal meningitis: study protocol for a randomised control trial.
Trials
PUBLISHED: 05-16-2014
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Cryptococcal meningitis (CM) is a severe AIDS-defining illness with 90-day case mortality as high as 70% in sub-Saharan Africa, despite treatment. It is the leading cause of death in HIV patients in Asia and Africa.No major advance has been made in the treatment of CM since the 1970s. The mainstays of induction therapy are amphotericin B and flucytosine, but these are often poorly available where the disease burden is highest. Adjunctive treatments, such as dexamethasone, have had dramatic effects on mortality in other neurologic infections, but are untested in CM. Given the high death rates in patients receiving current optimal treatment, and the lack of new agents on the horizon, adjuvant treatments, which offer the potential to reduce mortality in CM, should be tested.The principal research question posed by this study is as follows: does adding dexamethasone to standard antifungal therapy for CM reduce mortality? Dexamethasone is a cheap, readily available, and practicable intervention.
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Fracture risk in diabetic elderly men: the MrOS study.
Diabetologia
PUBLISHED: 05-09-2014
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Diabetes mellitus is associated with increased fracture risk in women but few studies are available in men. To evaluate the relationship between diabetes and prospective non-vertebral fractures in elderly men, we used data from the Osteoporotic Fractures in Men (MrOS) study.
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Phenylalanine-Based Inactivator of AKT Kinase: Design, Synthesis, and Biological Evaluation.
ACS Med Chem Lett
PUBLISHED: 05-08-2014
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Strategies to inhibit kinases by targeting the substrate binding site offer many advantages, including naturally evolved selectivity filters, but normally suffer from poor potency. In this work we propose a strategy to design and prepare covalent substrate-competitive kinase inhibitors as a method to improve potency. We have chosen AKT as the model kinase for this work. Using the AKT-GSK3? cocrystal structure and a reactive cysteine near the substrate binding site, we have identified phenylalanine (Phe) as an appropriate scaffold for the covalent inactivator portion of these inhibitors. By synthesizing compounds that incorporate cysteine-reactive electrophiles into phenylalanine and testing these compounds as AKT inhibitors, we have identified Boc-Phe-vinyl ketone as a submicromolar inactivator of AKT. We also show that Boc-Phe-vinyl ketone (1) potently inhibits AKT1 and inhibits cell growth in HCT116 and H460 cells nearly as well as AKT inhibitors GSK690693 and MK-2206, (2) is selective for kinases that possess an activation loop cysteine such as AKT, (3) requires the vinyl ketone for inactivation, (4) has inactivation that is time-dependent, and (5) alkylates Cys310 of AKT as shown by mass spectrometry. Identification of Boc-Phe-vinyl ketone as a covalent inactivator of AKT will allow the development of peptide and small-molecule substrate-competitive covalent kinase inhibitors that incorporate additional substrate binding elements to increase selectivity and potency. This proof-of-principle study also provides a basis to apply this strategy to other kinases of the AGC and CAMK families.
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Autoacetylation regulates differentially the roles of ARD1 variants in tumorigenesis.
Int. J. Oncol.
PUBLISHED: 05-06-2014
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ARD1 is an acetyltransferase with several variants derived from alternative splicing. Among ARD1 variants, mouse ARD1225 (mARD1225), mouse ARD1235 (mARD1235), and human ARD1235 (hARD1235) have been the most extensively characterized and are known to have different biological functions. In the present study, we demonstrated that mARD1225, mARD1235, and hARD1235 have conserved autoacetylation activities, and that they selectively regulate distinct roles of ARD1 variants in tumorigenesis. Using purified recombinants for ARD1 variants, we found that mARD1225, mARD1235, and hARD1235 undergo similar autoacetylation with the target site conserved at the Lys136 residue. Moreover, functional investigations revealed that the role of mARD1225 autoacetylation is completely distinguishable from that of mARD1235 and hARD1235. Under hypoxic conditions, mARD1225 autoacetylation inhibited tumor angiogenesis by decreasing the stability of hypoxia-inducible factor-1? (HIF-1?). Autoacetylation stimulated the catalytic activity of mARD1225 to acetylate Lys532 of the oxygen-dependent degradation (ODD) domain of HIF-1?, leading to the proteosomal degradation of HIF-1?. In contrast, autoacetylation of mARD1235 and hARD1235 contributed to cellular growth under normoxic conditions by increasing the expression of cyclin D1. Taken together, these data suggest that autoacetylation of ARD1 variants differentially regulates angiogenesis and cell proliferation in an isoform-specific manner.
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An unusual case of primary hepatic lymphoma mimicking sarcoidosis in MRI.
Acta Radiol Short Rep
PUBLISHED: 05-01-2014
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Sarcoidosis is a granulomatous disease, in which liver affection is common, contrary to a primary hepatic lymphoma that is very rarely seen. On MRI both present with almost the same imaging features: hypointense in T1-weighted and hyperintense in T2-weighted sequences. Our patient with a histologically confirmed sarcoidosis in the lungs showed liver lesions that were similar to sarcoidosis manifestations of the liver. Due to size, progression and overlapping features with secondary malignant liver lesions within an interval of 5 months, a biopsy was conducted and confirmed a primary hepatic lymphoma with diffuse large b-cells. Thus, we would recommend performing a biopsy in ambiguous lesions with indistinguishable characteristics and progression within a short follow-up interval.
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Human cytomegalovirus Fc? binding proteins gp34 and gp68 antagonize Fc? receptors I, II and III.
PLoS Pathog.
PUBLISHED: 05-01-2014
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Human cytomegalovirus (HCMV) establishes lifelong infection with recurrent episodes of virus production and shedding despite the presence of adaptive immunological memory responses including HCMV immune immunoglobulin G (IgG). Very little is known how HCMV evades from humoral and cellular IgG-dependent immune responses, the latter being executed by cells expressing surface receptors for the Fc domain of IgG (Fc?Rs). Remarkably, HCMV expresses the RL11-encoded gp34 and UL119-118-encoded gp68 type I transmembrane glycoproteins which bind Fc? with nanomolar affinity. Using a newly developed Fc?R activation assay, we tested if the HCMV-encoded Fc? binding proteins (HCMV Fc?Rs) interfere with individual host Fc?Rs. In absence of gp34 or/and gp68, HCMV elicited a much stronger activation of Fc?RIIIA/CD16, Fc?RIIA/CD32A and Fc?RI/CD64 by polyclonal HCMV-immune IgG as compared to wildtype HCMV. gp34 and gp68 co-expression culminates in the late phase of HCMV replication coinciding with the emergence of surface HCMV antigens triggering Fc?RIII/CD16 responses by polyclonal HCMV-immune IgG. The gp34- and gp68-dependent inhibition of HCMV immune IgG was fully reproduced when testing the activation of primary human NK cells. Their broad antagonistic function towards Fc?RIIIA, Fc?RIIA and Fc?RI activation was also recapitulated in a gain-of-function approach based on humanized monoclonal antibodies (trastuzumab, rituximab) and isotypes of different IgG subclasses. Surface immune-precipitation showed that both HCMV-encoded Fc? binding proteins have the capacity to bind trastuzumab antibody-HER2 antigen complexes demonstrating simultaneous linkage of immune IgG with antigen and the HCMV inhibitors on the plasma membrane. Our studies reveal a novel strategy by which viral Fc?Rs can compete for immune complexes against various Fc receptors on immune cells, dampening their activation and antiviral immunity.
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Development of an appropriate PCR system for the reclassification of Streptococcus suis.
J. Microbiol. Methods
PUBLISHED: 04-27-2014
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Thirty-five serotypes of Streptococcus suis (serotypes 1-34 and serotype 1/2) have so far been described on the basis of their polysaccharide capsular antigens. However, in the last decade, some serotype reference strains have been reexamined for their taxonomic status, and the reference strains of serotypes 20, 22, 26, 32, 33, and 34 may be different from taxon S. suis. In the present study, we developed a novel PCR method targeting the recombination/repair protein (recN) gene of S. suis, designated recN PCR, which corresponds to the current reclassification of this bacterium. We compared its specificity with other PCR methods for S. suis, and the results obtained confirmed its specificity. In addition, the detection limits of recN PCR were similar among all the reference strains of authentic S. suis, indicating that the recN PCR gave reliable results against bacterial strains and isolates used in this study. Therefore, recN PCR described in the present study will be a useful tool for the identification of authentic S. suis, and can also be used in epidemiological studies on this bacterium.
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Criteria for clinically relevant weakness and low lean mass and their longitudinal association with incident mobility impairment and mortality: the foundation for the National Institutes of Health (FNIH) sarcopenia project.
J. Gerontol. A Biol. Sci. Med. Sci.
PUBLISHED: 04-17-2014
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This analysis sought to determine the associations of the Foundation for the National Institutes of Health Sarcopenia Project criteria for weakness and low lean mass with likelihood for mobility impairment (gait speed ? 0.8 m/s) and mortality. Providing validity for these criteria is essential for research and clinical evaluation.
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Cutpoints for low appendicular lean mass that identify older adults with clinically significant weakness.
J. Gerontol. A Biol. Sci. Med. Sci.
PUBLISHED: 04-17-2014
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Low lean mass is potentially clinically important in older persons, but criteria have not been empirically validated. As part of the FNIH (Foundation for the National Institutes of Health) Sarcopenia Project, this analysis sought to identify cutpoints in lean mass by dual-energy x-ray absorptiometry that discriminate the presence or absence of weakness (defined in a previous report in the series as grip strength <26kg in men and <16kg in women).
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Grip strength cutpoints for the identification of clinically relevant weakness.
J. Gerontol. A Biol. Sci. Med. Sci.
PUBLISHED: 04-17-2014
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Weakness is common and contributes to disability, but no consensus exists regarding a strength cutpoint to identify persons at high risk. This analysis, conducted as part of the Foundation for the National Institutes of Health Sarcopenia Project, sought to identify cutpoints that distinguish weakness associated with mobility impairment, defined as gait speed less than 0.8 m/s.
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The FNIH sarcopenia project: rationale, study description, conference recommendations, and final estimates.
J. Gerontol. A Biol. Sci. Med. Sci.
PUBLISHED: 04-17-2014
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Low muscle mass and weakness are common and potentially disabling in older adults, but in order to become recognized as a clinical condition, criteria for diagnosis should be based on clinically relevant thresholds and independently validated. The Foundation for the National Institutes of Health Biomarkers Consortium Sarcopenia Project used an evidence-based approach to develop these criteria. Initial findings were presented at a conference in May 2012, which generated recommendations that guided additional analyses to determine final recommended criteria. Details of the Project and its findings are presented in four accompanying manuscripts.
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Personalized drug combinations to overcome trastuzumab resistance in HER2-positive breast cancer.
Biochim. Biophys. Acta
PUBLISHED: 04-08-2014
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HER2-positive (HER2+) breast cancer accounts for 18%-20% of all breast cancer cases and has the second poorest prognosis among breast cancer subtypes. Trastuzumab, the first Food and Drug Administration-approved targeted therapy for breast cancer, established the era of personalized treatment for HER2+ metastatic disease. It is well tolerated and improves overall survival and time-to-disease progression; with chemotherapy, it is part of the standard of care for patients with HER2+ metastatic disease. However, many patients do not benefit from it because of resistance. Substantial research has been performed to understand the mechanism of trastuzumab resistance and develop combination strategies to overcome the resistance. In this review, we provide insight into the current pipeline of drugs used in combination with trastuzumab and the degree to which these combinations have been evaluated, especially in patients who have experienced disease progression on trastuzumab. We conclude with a discussion of the current challenges and future therapeutic approaches to trastuzumab-based combination therapy.
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Versatile in vitro system to study translocation and functional integration of bacterial outer membrane proteins.
Nat Commun
PUBLISHED: 04-07-2014
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Gram-negative bacteria use the type-V secretion pathway to expose proteins at their cell surface, many of which have virulence functions. Translocation of those proteins across the outer membrane occurs either by means of dedicated translocator proteins (two-partner secretion) or covalently fused translocator domains (autotransporters). Translocator proteins and translocator domains are ?-barrels requiring the ?-barrel assembly machinery (BAM) for membrane integration. However, the molecular details of their passage across the envelope and insertion into the outer membrane remain enigmatic, owing in part to the fact that in vitro systems are not available. Here we describe a versatile in vitro reconstitution system that faithfully reproduces both branches of the type-V secretion pathway and the assembly of ?-barrel outer membrane proteins. This system will allow an in-depth analysis of protein secretion across and integration into outer membranes.
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Secondary metabolites from Vietnamese marine invertebrates with activity against Trypanosoma brucei and T. cruzi.
Molecules
PUBLISHED: 04-04-2014
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Marine-derived natural products from invertebrates comprise an extremely diverse and promising source of the compounds from a wide variety of structural classes. This study describes the discovery of five marine natural products with activity against Trypanosoma species by natural product library screening using whole cell in vitro assays. We investigated the anti-trypanosomal activity of the extracts from the soft corals and echinoderms living in Vietnamese seas. Of the samples screened, the methanolic extracts of several marine organisms exhibited potent activities against cultures of Trypanosoma brucei and T. cruzi (EC50 < 5.0 ?g/mL). Among the compounds isolated from these extracts, laevigatol B (1) from Lobophytum crassum and L. laevigatum, (24S)-ergost-4-ene-3-one (2) from Sinularia dissecta, astropectenol A (3) from Astropecten polyacanthus, and cholest-8-ene-3?,5?,6?,7?-tetraol (4) from Diadema savignyi showed inhibitory activity against T. brucei with EC50 values ranging from 1.57 ± 0.14 to 14.6 ± 1.36 ?M, relative to the positive control, pentamidine (EC50 = 0.015 ± 0.003 ?M). Laevigatol B (1) and 5?-cholest-8(14)-ene-3?,7?-diol (5) exhibited also significant inhibitory effects on T. cruzi. The cytotoxic activity of the pure compounds on mammalian cells was also assessed and found to be insignificant in all cases. This is the first report on the inhibitory effects of marine organisms collected in Vietnamese seas against Trypanosoma species responsible for neglected tropical diseases.
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CCR2 deficiency promotes exacerbated chronic erosive neutrophil-dominated chikungunya virus arthritis.
J. Virol.
PUBLISHED: 04-02-2014
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Chikungunya virus (CHIKV) is a member of a globally distributed group of arthritogenic alphaviruses that cause weeks to months of debilitating polyarthritis/arthralgia, which is often poorly managed with current treatments. Arthritic disease is usually characterized by high levels of the chemokine CCL2 and a prodigious monocyte/macrophage infiltrate. Several inhibitors of CCL2 and its receptor CCR2 are in development and may find application for treatment of certain inflammatory conditions, including autoimmune and viral arthritides. Here we used CCR2(-/-) mice to determine the effect of CCR2 deficiency on CHIKV infection and arthritis. Although there were no significant changes in viral load or RNA persistence and only marginal changes in antiviral immunity, arthritic disease was substantially increased and prolonged in CCR2(-/-) mice compared to wild-type mice. The monocyte/macrophage infiltrate was replaced in CCR2(-/-) mice by a severe neutrophil (followed by an eosinophil) infiltrate and was associated with changes in the expression levels of multiple inflammatory mediators (including CXCL1, CXCL2, granulocyte colony-stimulating factor [G-CSF], interleukin-1? [IL-1?], and IL-10). The loss of anti-inflammatory macrophages and their activities (e.g., efferocytosis) was also implicated in exacerbated inflammation. Clear evidence of cartilage damage was also seen in CHIKV-infected CCR2(-/-) mice, a feature not normally associated with alphaviral arthritides. Although recruitment of CCR2(+) monocytes/macrophages can contribute to inflammation, it also appears to be critical for preventing excessive pathology and resolving inflammation following alphavirus infection. Caution might thus be warranted when considering therapeutic targeting of CCR2/CCL2 for the treatment of alphaviral arthritides.
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Late-onset Alzheimer disease genetic variants in posterior cortical atrophy and posterior AD.
Neurology
PUBLISHED: 03-26-2014
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To investigate association of genetic risk factors for late-onset Alzheimer disease (LOAD) with risk of posterior cortical atrophy (PCA), a syndrome of visual impairment with predominant Alzheimer disease (AD) pathology in posterior cortical regions, and with risk of "posterior AD" neuropathology.
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Enhanced compatibility and initial stability of Ti6Al4V alloy orthodontic miniscrews subjected to anodization, cyclic precalcification, and heat treatment.
Korean J Orthod
PUBLISHED: 03-20-2014
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To evaluate the bioactivity, and the biomechanical and bone-regenerative properties of Ti6Al4V miniscrews subjected to anodization, cyclic precalcification, and heat treatment (APH treatment) and their potential clinical use.
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RNAscope for in situ detection of transcriptionally active human papillomavirus in head and neck squamous cell carcinoma.
J Vis Exp
PUBLISHED: 03-19-2014
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The 'gold standard' for oncogenic HPV detection is the demonstration of transcriptionally active high-risk HPV in tumor tissue. However, detection of E6/E7 mRNA by quantitative reverse transcription polymerase chain reaction (qRT-PCR) requires RNA extraction which destroys the tumor tissue context critical for morphological correlation and has been difficult to be adopted in routine clinical practice. Our recently developed RNA in situ hybridization technology, RNAscope, permits direct visualization of RNA in formalin-fixed, paraffin-embedded (FFPE) tissue with single molecule sensitivity and single cell resolution, which enables highly sensitive and specific in situ analysis of any RNA biomarker in routine clinical specimens. The RNAscope HPV assay was designed to detect the E6/E7 mRNA of seven high-risk HPV genotypes (HPV16, 18, 31, 33, 35, 52, and 58) using a pool of genotype-specific probes. It has demonstrated excellent sensitivity and specificity against the current 'gold standard' method of detecting E6/E7 mRNA by qRT-PCR. HPV status determined by RNAscope is strongly prognostic of clinical outcome in oropharyngeal cancer patients.
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NF-?B inhibitory activity of polyoxygenated steroids from the Vietnamese soft coral Sarcophyton pauciplicatum.
Bioorg. Med. Chem. Lett.
PUBLISHED: 02-10-2014
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Chromatographic purification of the methanolic extract from the soft coral Sarcophyton pauciplicatum led to the isolation of three polyhydroxylated steroids 1-3, including a new compound, sarcopanol A (1). Their structures were elucidated by spectroscopic analysis and by comparison of the spectroscopic data with those of similar compounds previously reported in literature. The anti-inflammatory effects of isolated compounds were evaluated using nuclear factor kappa B (NF-?B) luciferase and reverse transcription polymerase chain reaction (RT-PCR). The effect of isolated compounds on cell growth was evaluated by MTS assays. Compounds 1 and 2 significantly inhibited TNF?/INF?-induced NF-?B transcriptional activity in human keratinocyte (HaCaT) cells in a dose-dependent manner, with EC50 values of 8.27±3.28 and 26.07±5.59 ?M, respectively. Furthermore, the transcriptional inhibition of these compounds was confirmed by a decrease in cyclooxygenase-2 (COX-2), inducible nitric oxide synthase (iNOS), and intercellular adhesion molecule-1 (ICAM-1) gene expression levels in HaCaT cells.
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Triterpenoid saponins from the roots of Rosa rugosa Thunb. as rat intestinal sucrase inhibitors.
Arch. Pharm. Res.
PUBLISHED: 02-07-2014
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Medicinal plants constitute an important source of potential therapeutic agents for diabetes. The purpose of present study is to investigate the effect of root extract of Rosa rugosa Thunb. on inhibition of sucrase related to diabetes mellitus (DM). Bioassay-guided fractionation of the methanol extract led to the identification of 13 triterpenoid saponins (1-13). Their structures were elucidated on the basis of extensive spectroscopic analysis, including 1D, 2D NMR, and MS. The n-butanol fraction showed potent rat intestinal sucrase inhibitory activity with value of 87.62 ± 5.84 % inhibition compared to the positive control acarbose (50.96 ± 2.97 % inhibition at 0.02 mM). Subsequently, compounds 11-13 (1.0 mM) exhibited significant sucrase inhibitory activity, with inhibition percentage values of 41.17 ± 3.52, 46.80 ± 4.00, and 39.39 ± 4.19 %, respectively. Whereas, compounds 2-6, 8, and 10 showed moderate sucrase inhibitory activity (ranging from 13.26 ± 7.00 to 32.08 ± 6.04 % inhibition) at a same concentration. The data provide a starting point for creating new sucrase inhibitors, which may be useful for the development of effective therapies for the treatment of DM.
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Asterosaponins from the Starfish Astropecten monacanthus suppress growth and induce apoptosis in HL-60, PC-3, and SNU-C5 human cancer cell lines.
Biol. Pharm. Bull.
PUBLISHED: 02-05-2014
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Using various chromatographic experiments, six asterosaponins (1-6) were isolated from the MeOH extract of the Vietnamese starfish Astropecten monacanthus. The cytotoxic activities of the MeOH extract and six asterosaponins were evaluated on three human cancer cell lines, HL-60 (promyelocytic leukemia), PC-3 (prostate cancer), and SNU-C5 (colorectal cancer). Relative to the effects of the postitive control mitoxantrone, the MeOH extract (with IC50 values ranging from 0.84±0.03 to 3.96±0.14?µg/mL) and astrosterioside D (5) (with IC50 values ranging from 4.31±0.07 to 5.21±0.15?µM) exhibited potent cytotoxic effects against all three tested human cancer cell lines. In addition, the MeOH extract and astrosterioside D (5) have an effect on leading to apoptosis. Interestingly, the apoptosis of induction was accompanied by down-regulation of phosphatidyl inositol 3-kinase (PI3K)/AKT signaling and extracellular signal-regulated kinase (ERK) 1/2 mitogen-activated protein kinase (MAPK) signaling, and decrease of c-myc expression. Further studies are required to establish use of the asterosaponins from A. monacanthus as remedial and/or nutraceutical purposes.
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Cembranoid diterpenes from the soft coral Lobophytum crassum and their anti-inflammatory activities.
Chem. Pharm. Bull.
PUBLISHED: 02-05-2014
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Nine cembranoid diterpenes 1-9, including four new compounds, crassumols D-G (1-4), were isolated from the methanol extract of the Vietnamese soft coral Lobophytum crassum. Spectroscopic methods were used to elucidate the structures of these compounds. Compound 5 exhibited a potent inhibitory effect on tumor necrosis factor-alpha (TNF?)-induced nuclear factor-kappa B (NF-?B) transcriptional activation in HepG2 cells and significantly inhibited the mRNA expression of cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS) in a dose-dependent manner.
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Identification and characterization of a Trypanosoma congolense 46 kDa protein as a candidate serodiagnostic antigen.
J. Vet. Med. Sci.
PUBLISHED: 02-03-2014
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Trypanosoma congolense is a major livestock pathogen in Africa, causing large economic losses with serious effects on animal health. Reliable serodiagnostic tests are therefore urgently needed to control T. congolense infection. In this study, we have identified one T. congolense protein as a new candidate serodiagnostic antigen. The 46.4 kDa protein (TcP46, Gene ID: TcIL3000.0.25950) is expressed 5.36 times higher in metacyclic forms than epimastigote forms. The complete nucleotide sequences of TcP46 contained an open reading frame of 1,218 bp. Southern blot analysis indicated that at least two copies of the TcP46 gene were tandemly-arranged in the T. congolense genome. The recombinant TcP46 (rTcP46) was expressed in Escherichia coli as a glutathione S-transferase (GST) fusion protein. Western blot analysis and confocal laser scanning microscopy revealed that the native TcP46 protein is expressed in the cytoplasm during all life-cycle stages of the parasite. Moreover, an enzyme-linked immunosorbent assay (ELISA) based on rTcP46 detected the specific antibodies as early as 8 days post-infection from mice experimentally infected with T. congolense. No cross-reactivity was observed in the rTcP46-based ELISA against serum samples from cattle experimentally infected with Babesia bigemina, B. bovis and Anaplasma marginale. These results suggest that rTcP46 could be used as a serodiagnostic antigen for T. congolense infection.
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Functionalized acridin-9-yl phenylamines protected neuronal HT22 cells from glutamate-induced cell death by reducing intracellular levels of free radical species.
Bioorg. Med. Chem. Lett.
PUBLISHED: 02-01-2014
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The in vitro neuronal cell death model based on the HT22 mouse hippocampal cell model is a convenient means of identifying compounds that protect against oxidative glutamate toxicity which plays a role in the development of certain neurodegenerative diseases. Functionalized acridin-9-yl-phenylamines were found to protect HT22 cells from glutamate challenge at submicromolar concentrations. The Aryl(1)-NH-Aryl(2) scaffold that is embedded in these compounds was the minimal pharmacophore for activity. Mechanistically, protection against the endogenous oxidative stress generated by glutamate did not involve up-regulation of glutathione levels but attenuation of the late stage increases in mitochondrial ROS and intracellular calcium levels. The NH residue in the pharmacophore played a crucial role in this regard as seen from the loss of neuroprotection when it was structurally modified or replaced. That the same NH was essential for radical scavenging in cell-free and cell-based systems pointed to an antioxidant basis for the neuroprotective activities of these compounds.
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Pharmacokinetic and Pharmacodynamic Relationship of AMG 811, An Anti-IFN-? IgG1 Monoclonal Antibody, in Patients with Systemic Lupus Erythematosus.
Pharm. Res.
PUBLISHED: 01-30-2014
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To investigate the relationships between AMG 811 exposure, concentration changes in serum IFN-?, and IFN-?-induced protein 10 (CXCL10), and to identify important contributions of baseline covariates to these relationships.
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In vitro evaluation of the antioxidant and cytotoxic activities of constituents of the mangrove Lumnitzera racemosa Willd.
Arch. Pharm. Res.
PUBLISHED: 01-28-2014
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This study performed phytochemical and bioactive assessments of the mangrove Lumnitzera racemosa Willd. leaves. Bioassay-guided fractionation of the methanolic extracts led to the identification of thirty-six compounds (1-36), their structures were elucidated using detailed NMR spectroscopic and MS analysis. The extracts, fractions, and the isolated compounds were screened for potential antioxidant and cytotoxic activities. Antioxidant assays were performed using peroxyl radical-scavenging and reducing assays, whereas cytotoxicity was measured using MTT assays in HL-60 and Hel-299 cell lines. The methanolic extract, CH2Cl2 and n-BuOH fractions (10.0 ?g/mL) exhibited potent antioxidant activity, with Trolox equivalent (TE) values of 24.94 ± 0.59, 28.34 ± 0.20, and 27.09 ± 0.37 (?M), respectively. In addition, the isolated compounds exerted cytotoxic effects in a dose-dependent manner; compounds 1 and 14 exhibited the most potent cytotoxicity in HL-60 cells, with IC50 values of 0.15 ± 0.29 and 0.60 ± 0.16 ?M, respectively. To clarify the mechanism(s) behind these cytotoxic effects, we measured the time-dependent changes in apoptotic markers including the condensation and fragmentation of nuclear chromatin, and the downregulation of p-ERK1/2, p-AKT, and c-Myc levels.
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Expression, crystallization and preliminary X-ray crystallographic analysis of DNA-directed RNA polymerase subunit L from Thermococcus onnurineus NA1.
Acta Crystallogr F Struct Biol Commun
PUBLISHED: 01-28-2014
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RNA polymerase (RNAP) plays a crucial role in gene expression in all organisms. It is a multiprotein complex that produces primary transcript RNA. Generally, the basal transcription apparatus in archaea is simpler than the eukaryotic RNA polymerase II counterpart. To understand the structure and function of archaeal RNAP, the TON-0309 gene encoding DNA-directed RNA polymerase subunit L (ToRNAP_L) from Thermococcus onnurineus NA1 was cloned and the protein was overexpressed in Escherichia coli, purified and crystallized. The purified protein was crystallized using the hanging-drop vapour-diffusion method and the crystal diffracted to 2.10 Å resolution. The crystal belonged to the hexagonal space group P6122, with unit-cell parameters a = b = 42.3, c = 211.2 Å. One molecule was present in the asymmetric unit, with a corresponding VM of 2.5 Å(3) Da(-1) and a solvent content of 50.0%.
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Peroxisome proliferator-activated receptor transactivational effects in HepG2 cells of cembranoids from the soft coral Lobophytum crassum Von Marenzeller.
Arch. Pharm. Res.
PUBLISHED: 01-28-2014
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Peroxisome proliferator-activated receptors (PPARs) are ligand-activated transcription factors that regulate the expression of multiple genes involved in metabolic, anti-inflammatory, and developmental processes. This study evaluated the PPARs transactivational effects of thirteen cembranoid diterpenoids 1-13 from the soft coral Lobophytum crassum, using PPAR-responsive elements-luciferase reporter and GAL4-PPAR chimera assays. All isolated compounds activated the transcription of PPARs in a dose-dependent manner, with EC50 values ranging from 2.07 ± 1.73 to 130.20 ± 1.85 ?M. Moreover, compounds 6-9 affected the transactivation of all three PPAR types, PPAR?, ?, ?(?), in a dose-dependent manner, with EC50 values in a ranging from 11.92 ± 1.23 to 122.50 ± 2.12 ?M. These results provide a scientific rationale for further studies on the soft coral L. crassum and its diterpenoid constituents to develop medicinal products against inflammatory and metabolic diseases.
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