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Find video protocols related to scientific articles indexed in Pubmed.
Plasmid-encoding extended-spectrum ?-lactamase CTX-M-55 in a clinical Shigella sonnei strain, China.
Future Microbiol
PUBLISHED: 11-19-2014
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ABSTRACT? Aims: To characterize a clinical Shigella sonnei strain harboring a conjugatable bla CTX-M-55-borne plasmid. Materials & methods: S. sonnei strain #1081 was isolated from a dysentery patient in China. A CTX-M-55-encoding plasmid harbored in this strain was transformed to Escherichia coli, and then its complete nucleotide sequence was determined by next generation sequencing. The MIC values of bacterial strains were tested by using Vitec(®) 2 (Biomerieux, Marcy l'Etoile, France). Results: Strain #1081 conferred the resistance to multiple beta-lactam antibiotics. bla CTX-M-55 was the only known antibiotic resistance gene and located in a 3090-bp ISEcp1-bla CTX-M-55-orf477 transposition unit carried by a conjugatable plasmid p1081-CTXM in #1081. The ISEcp1-mediated transposition provided a sole promoter, which was located adjacently upstream of the inverted repeat right element of ISEcp1, to drive the expression of CTX-M-55. Conclusion: Plasmid p1081-CTXM was a close variant of the IncI2-type plasmid pHN1122-1 that was harbored in a faecal E. coli strain recovered from a dog in China, indicating the potential transfer of CTX-M-55-encoding plasmids from faecal flora E. coli to human pathogen S. sonnei.
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Histone methyltransferase hSETD1A is a Novel Regulator of Metastasis in Breast Cancer.
Mol. Cancer Res.
PUBLISHED: 11-07-2014
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Epigenetic alteration is a hallmark of all cancers. Such alterations lead to modulation of fundamental cancer-related functions, such as proliferation, migration, and invasion. In particular, methylation of Histone H3 Lysine 4 (H3K4), a histone mark generally associated with transcriptional activation, is altered during progression of several human cancers. While the depletion of H3K4 demethylases promotes breast cancer metastasis, the effect of H3K4 methyltransferases on metastasis is not clear. Nevertheless, gene duplications in the human SETD1A (hSETD1A) H3K4 methyltransferase are present in almost half of breast cancers. Herein, expression analysis determined that hSETD1A is up-regulated in multiple metastatic human breast cancer cell lines and clinical tumor specimens. Ablation of hSETD1A in breast cancer cells led to a decrease in migration and invasion in vitro and to a decrease in metastasis in nude mice. Furthermore, a group of matrix metalloproteinases (including MMP2, MMP9, MMP12, MMP13, and MMP17) were identified which were down-regulated upon depletion of hSETD1A and demonstrated a decrease in H3K4me3 at their proximal promoters based on chromatin immunoprecipitation (ChIP) analysis. These results provide evidence for a functional and mechanistic link among hSETD1A, MMPs, and metastasis in breast cancer, thereby supporting an oncogenic role for hSETD1A in cancer.
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1,5-diaminonaphthalene hydrochloride assisted laser desorption/ionization mass spectrometry imaging of small molecules in tissues following focal cerebral ischemia.
Anal. Chem.
PUBLISHED: 10-03-2014
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A sensitive analytical technique for visualizing small endogenous molecules simultaneously is of great significance for clearly elucidating metabolic mechanisms during pathological progression. In the present study, 1,5-naphthalenediamine (1,5-DAN) hydrochloride was prepared for matrix-assisted laser desorption/ionization (MALDI) mass spectrometry imaging (MSI) of small molecules in liver, brain, and kidneys from mice. Furthermore, 1,5-DAN hydrochloride assisted LDI MSI of small molecules in brain tissue of rats subjected to middle cerebral artery occlusion (MCAO) was carried out to investigate the altered metabolic pathways and mechanisms underlying the development of ischemic brain damage. Our results suggested that the newly prepared matrix possessed brilliant features including low cost, strong ultraviolet absorption, high salt tolerance capacity, and fewer background signals especially in the low mass range (typically m/z < 500), which permitted us to visualize the spatial distribution of a broad range of small molecule metabolites including metal ions, amino acids, carboxylic acids, nucleotide derivatives, peptide, and lipids simultaneously. Nineteen endogenous metabolites involved in metabolic networks such as ATP metabolism, tricarboxylic acid (TCA) cycle, glutamate-glutamine cycle, and malate-aspartate shuttle, together with metal ions and phospholipids as well as antioxidants underwent relatively obvious changes after 24 h of MCAO. The results were highly consistent with the data obtained by MRM MS analysis. These findings highlighted the promising potential of the organic salt matrix for application in the field of biomedical research.
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Characteristics of CARMA1-BCL10-MALT1-A20-NF-?B expression in T cell-acute lymphocytic leukemia.
Eur. J. Med. Res.
PUBLISHED: 09-04-2014
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Knowledge of the oncogenic signaling pathways of T-cell acute lymphoblastic leukemia (T-ALL) remains limited. Constitutive aberrant activation of the nuclear factor kappa B (NF-?B) signaling pathway has been detected in various lymphoid malignancies and plays a key role in the development of these carcinomas. The zinc finger-containing protein, A20, is a central regulator of multiple NF-?B-activating signaling cascades. A20 is frequently inactivated by deletions and/or mutations in several B-and T-cell lymphoma subtypes. However, few A20 mutations and polymorphisms have been reported in T-ALL. Thus, it is of interest to analyze the expression characteristics of A20 and its regulating factors, including upstream regulators and the CBM complex, which includes CARMA1, BCL10, and MALT1.
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Nonlinear effects in paul traps operated in the second stability region: analytical analysis and numerical verification.
J. Am. Soc. Mass Spectrom.
PUBLISHED: 09-03-2014
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Paul trap working in the second stability region has long been recognized as a possible approach for achieving high-resolution mass spectrometry (MS), which however is still far away from the experimental implementations because of the narrow working area and inefficient ion trapping. Full understanding of the ion motional behavior is helpful for solving the problem. In this article, the ion motion in a superimposed octopole field, which was characterized by the nonlinear Mathieu equation, was solved analytically using Poincare-Lighthill-Kuo (PLK) method. This method equivalently described the nonlinear disturbance by an effective quadrupole field with perturbed Mathieu parameters, a u (') and q u (') , which would bring huge convenience in the studies of nonlinear ion dynamics and was, therefore, used for rapid evaluation of the nonlinear effects of ion motion. Fourth-order Runge-Kutta method (4th R-K) indicated the error of PLK for characterizing the frequency shift of ion motion was within 15%.
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Plasma-based ambient sampling/ionization/transmission integrated source for mass spectrometry.
Analyst
PUBLISHED: 08-23-2014
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Better sensitivity and interface of ambient sampling/ionization mass spectrometry remain a challenge. Herein, a novel, plasma-based, ambient sampling/ionization/transmission (PASIT) integrated source in a pin-to-funnel configuration was developed for the sensitive analysis of complex samples. With the funnel sleeve directly affected by direct-current discharge plasma, PASIT combines the ability to sample/ionize analyte molecules and then efficiently collect/transport charged mass species under atmospheric pressure and consequently shows an improved sensitivity. The integrated source enhances the signal intensity by more than 2 orders of magnitude compared with the previous pin-to-plate plasma source without significant background addition. A surface limit of detection (LOD) of 130 fmol mm-(2) (S/N = 3) has been achieved for clenbuterol on filter paper with an argon carrier gas. Demonstrated applications include the direct determination of active ingredients from drugs and symbolic compounds from natural plants and cholesterol from mouse brain tissue sections.
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Lysosomal pH rise during heat shock monitored by a lysosome-targeting near-infrared ratiometric fluorescent probe.
Angew. Chem. Int. Ed. Engl.
PUBLISHED: 08-22-2014
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Heat stroke is a life-threatening condition, featuring a high body temperature and malfunction of many organ systems. The relationship between heat shock and lysosomes is poorly understood, mainly because of the lack of a suitable research approach. Herein, by incorporating morpholine into a stable hemicyanine skeleton, we develop a new lysosome-targeting near-infrared ratiometric pH?probe. In combination with fluorescence imaging, we show for the first time that the lysosomal pH?value increases but never decreases during heat shock, which might result from lysosomal membrane permeabilization. We also demonstrate that this lysosomal pH?rise is irreversible in living cells. Moreover, the probe is easy to synthesize, and shows superior overall analytical performance as compared to the existing commercial ones. This enhanced performance may enable it to be widely used in more lysosomal models of living cells and in further revealing the mechanisms underlying heat-related pathology.
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Thermo-responsive release of curcumin from micelles prepared by self-assembly of amphiphilic P(NIPAAm-co-DMAAm)-b-PLLA-b-P(NIPAAm-co-DMAAm) triblock copolymers.
Int J Pharm
PUBLISHED: 07-24-2014
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Thermo-responsive micelles are prepared by self-assembly of amphiphilic triblock copolymers composed of a poly(l-lactide) (PLLA) central block and two poly(N-isopropylacrylamide-co-N,N-dimethylacrylamide) (P(NIPAAm-co-DMAAm)) lateral blocks, using solvent evaporation/film hydration method. The resulting micelles exhibit very low critical micelle concentration (CMC) which slightly increases from 0.0113 to 0.0144mgmL(-1) while the DMAAm content increases from 31.8 to 39.4% in the hydrophilic P(NIPAAm-co-DMAAm) blocks. The lower critical solution temperatures (LCST) of copolymers varies from 44.7°C to 49.4°C in water as determined by UV spectroscopy, and decreases by ca. 3.5°C in phosphate buffered saline (PBS). Curcumin was encapsulated in the core of micelles. High drug loading up to 20% is obtained with high loading efficiency (>94%). The LCST of drug loaded micelles ranges from 37.5 to 38.0°C with drug loading increasing from 6.0 to 20%. The micelles with diameters ranging from 47.5 to 88.2nm remain stable over one month due to the negative surface charge as determined by zeta potential (-12.4 to -18.7mV). Drug release studies were performed under in vitro conditions at 37°C and 40°C, i.e. below and above the LCST, respectively. Initial burst release is observed in all cases, followed by a slower release. The release rate is higher at 40°C than that at 37°C due to thermo-responsive release across the LCST. On the other hand, micelles with lower drug loading exhibit higher release rate than those with higher drug loading, which is assigned to the solubility effect. Peppas' theory was applied to describe the release behaviors. Moreover, the in vitro cytotoxicity of copolymers was evaluated using MTT assay. The results show that the copolymers present good cytocompatibility. Therefore, the nano-scale size, low CMC, high drug loading and stability, as well as good biocompatibility indicate that these thermo-responsive triblock copolymer micelles present a good potential as carrier for targeted delivery of anticancer drugs.
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Effects of cations on small fragment of DNA polymerase I using a novel FRET assay.
Acta Biochim. Biophys. Sin. (Shanghai)
PUBLISHED: 06-27-2014
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DNA polymerase I (PolI) digested by protease produces a small fragment (SF) containing 5'-3' exonuclease activity. The 5'-3' exonuclease activity of polI cleaves the downstream RNA primer strands during DNA replication in vivo. Previous in vitro studies suggested its capability of cleaving duplex from 5' terminal and a flap-structure-specific endonuclease activity. From the crystal structures of other nucleases and biochemical data, a two-metal-ion mechanism has been proposed but has not been determined. In this study, we cloned, expressed, and purified the SF protein, and established a novel fluorescence resonance energy transfer (FRET) assay to analyze the catalytic activity of the SF protein. The effects of several metal ions on its catalytic capability were analyzed using this FRET assay. Results showed that Mg2+, Mn2+, and Zn2+ were able to activate the cleavage of SF, while Ca2+, Ni2 +, and Co2+ were not suitable for SF catalysis. The effects of K+, Na+, and dNTP were also determined.
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Overexpression of histone deacetylases in cancer cells is controlled by interplay of transcription factors and epigenetic modulators.
FASEB J.
PUBLISHED: 06-19-2014
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Histone deacetylases (HDACs) that deacetylate histone and nonhistone proteins play crucial roles in a variety of cellular processes. The overexpression of HDACs is reported in many cancer types and is directly linked to accelerated cell proliferation and survival. However, little is known about how HDAC expression is regulated in cancer cells. In this study, we found that HDAC1 and HDAC2 promoters are regulated through collaborative binding of transcription factors Sp1/Sp3 and epigenetic modulators, including histone H3K4 methyltransferase SET1 and histone acetyltransferase p300, whose levels are also elevated in colon cancer cell lines and patient samples. Interestingly, Sp1 and Sp3 differentially regulate HDAC1 and HDAC2 promoter activity. In addition, Sp1/Sp3 recruits SET1 and p300 to the promoters. SET1 knockdown (KD) results in a loss of the H3K4 trimethylation mark at the promoters, as well as destabilizes p300 at the promoters. Conversely, p300 also influences SET1 recruitment and H3K4me3 level, indicating a crosstalk between p300 and SET1. Further, SET1 KD reduces Sp1 binding to the HDAC1 promoter through the increase of Sp1 acetylation. These results indicate that interactions among transcription factors and epigenetic modulators orchestrate the activation of HDAC1 and HDAC2 promoter activity in colon cancer cells.
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Organic salt NEDC (N-naphthylethylenediamine dihydrochloride) assisted laser desorption ionization mass spectrometry for identification of metal ions in real samples.
Analyst
PUBLISHED: 05-21-2014
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The significance of metals in life and their epidemiological effects necessitate the development of a direct, efficient, and rapid method of analysis. The matrix assisted laser desorption/ionization technique is on the horns of a dilemma of metal analysis as the conventional matrixes have high background in the low mass range. An organic salt, NEDC (N-naphthylethylenediamine dihydrochloride), is applied as a matrix for identification of metal ions in the negative ion mode in the present work. Sixteen metal ions, Ba(2+), Ca(2+), Cd(2+), Ce(3+), Co(2+), Cu(2+), Fe(3+), Hg(2+), K(+), Mg(2+), Mn(2+), Na(+), Ni(2+), Pb(2+), Sn(2+) and Zn(2+), in the form of their chloride-adducted clusters were systematically tested. Mass spectra can provide unambiguous identification through accurate mass-to-charge ratios and characteristic isotope patterns. Compared to ruthenium ICP standard solution, tris(2,2'-bipyridyl)dichlororuthenium(ii) (C30H24N6Cl2Ru) can form organometallic chloride adducts to discriminate from the inorganic ruthenium by this method. After evaluating the sensitivity for Ca, Cu, Mg, Mn, Pb and Zn and plotting their quantitation curves of signal intensity versus concentration, we determined magnesium concentration in lake water quantitatively to be 5.42 mg L(-1) using the standard addition method. There is no significant difference from the result obtained with ICP-OES, 5.8 mg L(-1). Human urine and blood were also detected to ascertain the multi-metal analysis ability of this strategy in complex samples. At last, we explored its applicability to tissue slice and visualized sodium and potassium distribution by mass spectrometry imaging in the normal Kunming mouse brain.
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A pilot study exploring the mechanisms involved in the longitudinal propagation of acute aortic dissection through computational fluid dynamic analysis.
Cardiology
PUBLISHED: 04-24-2014
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This study sought to elucidate the underlying hemodynamic mechanisms involved in the longitudinal propagation of acute, type-B aortic dissections.
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Involvement of Wnt5a within the cerebrospinal fluid-contacting nucleus in nerve injury-induced neuropathic pain.
Int. J. Neurosci.
PUBLISHED: 04-18-2014
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Studies have demonstrated that the cerebrospinal fluid-contacting nucleus (CSF-CN) is involved in neuropathic pain, but the underlying molecular mechanisms still largely remain obscure. Emerging evidence suggests that spinal Wnt5a plays a crucial role in regulation of chronic pain. However, little is known about the potential role of the supraspinal Wnt5a in the development of chronic pain. To investigate whether Wnt5a exists in the CSF-CN and its role in neuropathic pain, double-labeled immunofluorescence staining was used to identify the expression of Wnt5a in the CSF-CN and western blot analysis of the CSF-CN was employed to verify the alteration of Wnt5a protein in the process of neuropathic pain. In the present study, we demonstrated that Wnt5a is distributed in the CSF-CN and the Wnt5a protein was up-regulated by nerve injury-induced nociceptive stimuli. Furthermore, lateral intracerebroventricular injection of Wnt5a antagonist Box5 attenuated the chronic constriction injury (CCI)-induced neuropathic pain and down-regulated the expression of Wnt5a in the CSF-CN. These data extend our understanding of the role of Wnt5a in supraspinal site and demonstrate that the CSF-CN participates in nerve injury-induced neuropathic pain via the regulation of Wnt5a.
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Algicidal effects of four Chinese herb extracts on bloom-forming Microcystis aeruginosa and Chlorella pyrenoidosa.
Environ Technol
PUBLISHED: 04-08-2014
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Extracts from four Chinese herbs, Phellodendri chinensis cortex, Artemisia annua L., Scutellaria baicalensis G. and Citrus reticulate peel were tested for their algicidal effects on Microcystis aeruginosa and Chlorella pyrenoidosa. The results showed that M. aeruginosa was more susceptible than C. pyrenoidosa. The growth of M. aeruginosa was significantly inhibited (p < 0.05) by the four herb extracts. Among the four herbs, P. chinensis cortex and S. baicalensis had the greatest inhibitory effects on M. aeruginosa, followed by C reticulate peel and A. annua. The 50% effective concentrations (EC50) of S. baicalensis, P chinensis cortex, C. reticulate peel and A. annua were 0.87, 0.88, 5.27 and 1 1.16 gherb L-1, respectively. The growth of C. pyrenoidosa was moderately inhibited by the herb extracts individually. The EC5o concentrations for S. baicalensis, P. chinensis cortex, C. reticulate peel andA. annua were 8.67, 11.67, 12.81 and 12.44 g herb L-1', respectively. Extract from S. baicalensis displayed stronger algicidal effects on C. pyrenoidosa than the other three herbs, although no lethal effect on C. pyrenoidosa was observed during the cultivation period. Compared with corresponding individual extract at the same dosage, the binary mixtures of the four herb extracts enhanced the algicidal effects on M. aeruginosa. The maximum inhibitory rates of all binary mixtures of the four herb extracts were all above 92% during the 10-day incubation. The results demonstrate that Chinese herbs, such as P. chinensis cortex or S. baicalensis and their combinations, could offer an effective alternative for mitigating outbreaks of harmful algal blooms in water bodies.
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Total Endovascular Repair of Thoracoabdominal Aortic Aneurysms With Non-Customized Stent Grafts.
Ann. Thorac. Surg.
PUBLISHED: 03-31-2014
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Total endovascular repair of thoracoabdominal aortic aneurysms with customized branched or fenestrated endografts could be technically challenging outside large-volume centers. This study aims to describe a new endovascular strategy for use of both noncustomized stent grafts and flow-diverting stents in treating complicated thoracoabdominal aortic aneurysms.
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Quantitative assessment of protein adsorption on microparticles with particle mass spectrometry.
Anal. Chem.
PUBLISHED: 03-28-2014
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In this paper, particle mass spectrometry (PMS), which consists of an aerodynamic desorption/ionization (AD) source, a quadrupole ion trap (QIT) mass analyzer, and a charge detector, was exploited to characterize the protein adsorption on microparticles based on the mass variations of microparticles before and after protein adsorption. This method is simple and has low sample cost. Importantly, its mass resolution is good enough to distinguish the microparticles with and without protein. For the adsorption of bovine serum albumin (BSA) on 3 ?m porous poly styrene-divinylbenzene (poly S-DVB), the minimum mass increase that can be resolved by PMS corresponds to 128 fg (1.8 ng/cm(2)) or 1.17 × 10(6) BSA molecules on each poly S-DVB particle. With PMS, the adsorption process of BSA on poly S-DVB spheres was successfully characterized, and the obtained maximum adsorption capacity qm and dissociation constant Kd were consistent with that determined by the conventional depletion method. In addition, the influence of surface modification of silica particles on the enzyme immobilization was evaluated. Compared with C4 (propyldimethylsilane), C8 (octyldimethylsilane), and Ph (phenyldimethylchlorosilane), the CN (cyanoethyldimethylchlorosilane) functionalized silica particles were screened to be most beneficial for the immobilization of both lysozyme and trypsin.
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Cancer stem-like cell: a novel target for nasopharyngeal carcinoma therapy.
Stem Cell Res Ther
PUBLISHED: 03-27-2014
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Nasopharyngeal carcinoma (NPC) is the most common cancer originating in the nasopharynx, and is extremely common in southern regions of China. Although the standard combination of radiotherapy and chemotherapy has improved the efficiency in patients with NPC, relapse and early metastasis are still the common causes of mortality. Cancer stem-like cells (CSCs) or tumor initial cells are hypothesized to be involved in cancer metastasis and recurrence. Over the past decade, increasing numbers of studies have been carried out to identify CSCs from human NPC cells and tissues. The present paper will summarize the investigations on nasopharyngeal CSCs, including isolation, characteristics, and therapeutic approaches. Although there are still numerous challenges to translate basic research into clinical applications, understanding the molecular details of CSCs is essential for developing effective strategies to prevent the recurrence and metastasis of NPC.
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Characteristics of A20 gene polymorphisms in T-cell acute lymphocytic leukemia.
Hematology
PUBLISHED: 03-11-2014
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A20 is a repressor of NF-?B and was recently shown to be frequently inactivated by deletions or mutations in several types of lymphomas including T-cell lymphoma. Little is known about the characteristics of A20 mutations in T-cell acute lymphoblastic leukemia (T-ALL). In this study, we analyzed A20 polymorphisms and characterized their features in 11 cases with T-ALL, 30 samples from healthy Chinese individuals, and 3 cells lines including CCRF-CEM, Molt-4, and Toledo cells. Two frequent A20 polymorphisms were found: a CCT deletion at position 12384 and a nucleotide exchange (A to C) at position 13751 (rs2307859 and rs661561). The homozygous form (CC) of rs661561 was detected in all 10 cases with detectable T-ALL, while only 80% (24/30) of the healthy controls had this genotype. We found one T-ALL case without the above frequent single-nucleotide polymorphisms (SNPs) in which a T to G mutation at position 12486 was found, which results in an amino acid exchange (Phe127Cys; rs2230926). Similar results were found in Molt-4 cells, which lack the frequent SNPs but have a heterozygous polymorphism at position 13749 (C > T) (rs5029948). Interestingly, the T-ALL case with the Phe127Cys mutation and Molt-4 cells demonstrated a high A20 copy number as measured by real-time polymerase chain reaction amplification with three primer sets that cover different regions of the A20 gene, corresponding to a high A20 and low NF-?B expression level. In conclusion, we characterized the features of A20 polymorphisms in T-ALL, and found that a low frequency A20 mutation, which was thought to be involved in malignant T-ALL development, might function differently in T cell lymphomas.
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Interface solution isoelectric focusing with in situ MALDI-TOF mass spectrometry.
Electrophoresis
PUBLISHED: 02-17-2014
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This paper describes a simple and reusable microfluidic device combining solution IEF (sIEF) with MALDI-TOF MS for rapid proteomic and metabolic analysis of microliter samples. The device contains two glass plates with nanoliter microwell arrays, which can be assembled to form a fluidic path for sIEF separation, and reconfigured for dividing separated bands. One microliter samples can be loaded and separated by sIEF into static bands in 10?30 min. After a slipping operation, the static IEF bands can be divided into nanoliter droplets in microwells without mobilization, and the device can be opened for in situ MALDI-TOF MS detection without loss of separation resolution. The performance of the device is characterized by separating and identifying intact proteins. The applicability in metabolic analysis is demonstrated by preliminary experiments on profiling small molecular metabolites in cerebrospinal fluid microdialysates from rat brain.
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Dissecting the function of the adult ?-globin downstream promoter region using an artificial zinc finger DNA-binding domain.
Nucleic Acids Res.
PUBLISHED: 02-04-2014
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Developmental stage-specific expression of the ?-type globin genes is regulated by many cis- and trans-acting components. The adult ?-globin gene contains an E-box located 60 bp downstream of the transcription start site that has been shown to bind transcription factor upstream stimulatory factor (USF) and to contribute to efficient in vitro transcription. We expressed an artificial zinc finger DNA-binding domain (ZF-DBD) targeting this site (+60 ZF-DBD) in murine erythroleukemia cells. Expression of the +60 ZF-DBD reduced the recruitment and elongation of RNA polymerase II (Pol II) at the adult ?-globin gene and at the same time increased the binding of Pol II at locus control region (LCR) element HS2, suggesting that Pol II is transferred from the LCR to the globin gene promoters. Expression of the +60 ZF-DBD also reduced the frequency of interactions between the LCR and the adult ?-globin promoter. ChIP-exonuclease-sequencing revealed that the +60ZF-DBD was targeted to the adult ?-globin downstream promoter and that the binding of the ZF-DBD caused alterations in the association of USF2 containing protein complexes. The data demonstrate that targeting a ZF-DBD to the adult ?-globin downstream promoter region interferes with the LCR-mediated recruitment and activity of Pol II.
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The outer membrane protein P2 (OmpP2) of Haemophilus parasuis induces proinflammatory cytokine mRNA expression in porcine alveolar macrophages.
Vet. J.
PUBLISHED: 02-04-2014
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Porins, expressed by Gram-negative bacteria, have several biological effects on host tissues or cells. The outer membrane protein P2 (OmpP2), a member of the porin family, has been identified as a multifunctional protein involved in the pathogenicity of Haemophilus parasuis. In the present study, it was shown that OmpP2 (0.5-10 ?g/mL) from H. parasuis Nagasaki strain up-regulated mRNA expression of interleukin (IL)-1?, IL-1?, IL-6 and IL-8 in porcine alveolar macrophages (PAM, 3D4/31) in vitro, in a dose-dependent manner. Moreover, OmpP2 porin induced a more prolonged cytokine response in PAM than that of the lipooligosaccharide (LOS) from this microorganism. The data demonstrate that H. parasuis OmpP2 can stimulate proinflammatory cytokine mRNA expression, suggesting that this particular porin might play an important role in the pathogenesis of disease caused by H. parasuis.
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Direct renin inhibition with aliskiren protects against myocardial ischemia/reperfusion injury by activating nitric oxide synthase signaling in spontaneously hypertensive rats.
J Am Heart Assoc
PUBLISHED: 01-30-2014
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We tested the hypothesis that direct renin inhibition with aliskiren protects against myocardial ischemia/reperfusion (I/R) injury in spontaneously hypertensive rats (SHR), and examined the mechanism by which this occurs.
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The binding behavior of itraconazole with hemoglobin: studies from multi-spectroscopic techniques.
Spectrochim Acta A Mol Biomol Spectrosc
PUBLISHED: 01-21-2014
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The interactions between hemoglobin (Hb) and itraconazole (ITZ) are investigated in details using UV-vis spectra, circular dichroism spectroscopy, steady state fluorescence, three-dimensional fluorescence spectra, synchronous fluorescence and time-resolved fluorescence spectra at molecular level. The UV-vis studies represent that ITZ can access into heme group and lead to it explored in aqueous medium. CD spectra suggest ITZ could combine with amino acid residues in polypeptide chain and cause a partial unfolding of Hb (reducing of the ?-helix content). Steady state fluorescence/synchronous fluorescence (taking into account inner filter effects) and three-dimensional fluorescence/time-resolved fluorescence spectroscopy results reveal that ITZ alters polarity and conformation around the fluorophore molecule. The interaction processes are static quenching mechanisms. The negative of ?H(0) and ?S(0) indicate that hydrogen bonds and van der Waals are the main force.
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Preparation, blood coagulation and cell compatibility evaluation of chitosan-graft-polylactide copolymers.
Biomed Mater
PUBLISHED: 01-21-2014
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Biodegradable chitosan-graft-polylactide (PLA-CS) copolymers were prepared by the grafting of a poly(L-lactide) (PLLA) or poly(D-lactide) (PDLA) precursor to the backbone of chitosan using 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC ? HCl) and N-hydroxysuccinimide (NHS) as a coupling agent. The blood and cell compatibility of the graft copolymers were investigated in comparison to PLLA and PDLA homopolymers. The coagulation properties of PLA-CS were evaluated by hemolysis, plasma recalcification, dynamic blood clotting and protein absorption assays. PLA-CS copolymers present similar hemolysis ratio and plasma recalcification time as PLA, but slower dynamic blood clotting and lower protein absorption. The cell viability was assessed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT), agar diffusion and lactate dehydrogenase (LDH) experiments. All the samples presented no effect on the viability to cells. Inflammatory cytokine analysis using sandwich ELISAs revealed that PLA-CS would not stimulate inflammatory activity.
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GRK3 is essential for metastatic cells and promotes prostate tumor progression.
Proc. Natl. Acad. Sci. U.S.A.
PUBLISHED: 01-13-2014
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The biochemical mechanisms that regulate the process of cancer metastasis are still poorly understood. Because kinases, and the signaling pathways they comprise, play key roles in regulation of many cellular processes, we used an unbiased RNAi in vitro screen and a focused cDNA in vivo screen against human kinases to identify those with previously undocumented roles in metastasis. We discovered that G-protein-coupled receptor kinase 3 (GRK3; or ?-adrenergic receptor kinase 2) was not only necessary for survival and proliferation of metastatic cells, but also sufficient to promote primary prostate tumor growth and metastasis upon exogenous expression in poorly metastatic cells in mouse xenograft models. Mechanistically, we found that GRK3 stimulated angiogenesis, at least in part through down-regulation of thrombospondin-1 and plasminogen activator inhibitor type 2. Furthermore, GRK3 was found to be overexpressed in human prostate cancers, especially in metastatic tumors. Taken together, these data suggest that GRK3 plays an important role in prostate cancer progression and metastasis.
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Glial cell-derived neurotrophic factor attenuates neuropathic pain in a mouse model of chronic constriction injury: possible involvement of E-cadherin/p120ctn signaling.
J. Mol. Neurosci.
PUBLISHED: 01-12-2014
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Treating neuropathic pain is a major clinical challenge, and several key molecules associated with nociception have been suggested as potential targets for novel analgesics. Many studies have reported the anti-nociceptive effects of glial cell-derived neurotrophic factor (GDNF), but the underlying mechanism remains largely unknown. The present study was performed to assess the effects of GDNF in a mouse model of chronic constriction injury (CCI)-induced neuropathic pain. We also determined the potential role of E-cadherin/p120 catenin (p120ctn) signaling in these effects. Mice received an intrathecal acute injection of PBS, GDNF, and DECMA-1 (an E-cadherin functional blocking antibody) or a combination of DECMA-1 with GDNF on the testing days. Our results demonstrated that CCI caused a rapid decrease in E-cadherin and membrane-associated p120ctn in the spinal dorsal horn. Together, these data demonstrated that E-cadherin-associated p120ctn was upregulated by GDNF and that this upregulation was inhibited by pre-treatment with DECMA-1. Moreover, DECMA-1 significantly inhibited the effect of GDNF on thermal hyperalgesia. These data suggest that GDNF might have a therapeutic potential for the treatment of CCI-induced neuropathic pain and that the E-cadherin/p120ctn might play a role in GDNF-induced attenuation of thermal hyperalgesia.
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NCAM signaling mediates the effects of GDNF on chronic morphine-induced neuroadaptations.
J. Mol. Neurosci.
PUBLISHED: 01-08-2014
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Glial cell line-derived neurotrophic factor (GDNF) is a potent neurotrophic factor for midbrain dopamine (DA) neurons, while the DA neurons in the ventral tegmental area (VTA) is a crucial part of the neural circuits associated with drug addiction. Recently, more and more evidence suggests that GDNF plays an important role in negatively regulating the neuroadaptations induced by chronic exposure to drugs, which was thought to be the neurobiological basis of drug addiction, but the underlying mechanism is still unknown. More recently, the neural cell adhesion molecule (NCAM), which plays an important role in the process of neural plasticity, has been identified as an alternative signaling receptor for GDNF. The purpose of this study was to investigate whether NCAM was involved in the effects of GDNF on the neuroadaptations induced by chronic morphine exposure. Immunostaining results showed that NCAM was widely expressed in the VTA of rats, including all the DA neurons. The results also showed that the phosphorylation of NCAM-associated FAK, but not the total NCAM, was upregulated by GDNF, and this upregulation was inhibited by pre-treatment with the NCAM function-blocking antibody. Moreover, pre-treatment with the antibody could antagonize the effect of GDNF on inhibiting the neuroadaptations induced by chronic morphine exposure, including the decreases of the number and length of neurites and the size of cell bodies of VTA dopamine neurons, as well as the increase of tyrosine hydroxylase in the VTA dopamine neurons. These results suggest that NCAM signaling is involved in the negative regulatory effects of GDNF on chronic morphine-induced neuroadaptations.
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Effect of piracetam on the cognitive performance of patients undergoing coronary bypass surgery: A meta-analysis.
Exp Ther Med
PUBLISHED: 01-08-2014
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Cognitive impairments are observed in numerous patients following coronary bypass surgery, and piracetam are nootropic compounds that modulate cerebral functions by directly enhancing cognitive processes. The present meta-analysis was conducted to evaluate the protective effect of piracetam on the cognitive performance of patients undergoing coronary bypass surgery. The relevant studies were identified by searching Medline, EMBASE, PubMed and the Cochrane Library up to June 2013 and the pertinent bibliographies from the retrieved studies were reviewed. Data were selected from the studies according to predefined criteria. The meta-analysis included two randomized control trials involving 184 patients and including the Syndrom-Kurz test (SKT). Findings of the meta-analysis showed that following treatment the change from baseline observed in five SKT subtest scores, conducted with piracetam patients, indicated a significant advantage over those patients that were in the placebo group. The subtests included immediate pictured object recall, weighted mean difference (WMD)=0.91, 95% confidence interval (CI) 0.51-1.31, P<0.00001; delayed pictured object recall, WMD=0.74, 95% CI 0.19-1.28, P=0.008; delayed picture recognition, WMD=0.82, 95% CI 0.31-1.31, P=0.001; immediate word recall, WMD=0.87, 95% CI 0.47-1.28, P<0.0001; and letter interference, WMD=3.46, 95% CI -5.69 to -1.23, P=0.002. These results indicated that piracetam may have been effective in improving the short-term cognitive performance of patients undergoing coronary bypass surgery. High quality, well-controlled and longer randomized trials are required to corroborate this result.
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The relationship between PthA expression and the pathogenicity of Xanthomonas axonopodis pv. citri.
Mol. Biol. Rep.
PUBLISHED: 01-03-2014
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Citrus canker disease, caused by Xanthomonas axonopodis pv. citri, affects almost all citrus species and cultivars and hascaused severe damage to the citrus industry worldwide. PthA is considered the main pathogenesis effector of the pathogen. This research aimed to temporally and spatially analyze the expression of the PthA protein of the bactrium during its culture, and then try to understand the relationship between the PthA expression levels and the pathogenicity. The relationship between the expression of PthA and the pathogenicity of X. axonopodis pv. citri was fully investigated by using SDS-PAGE, Western blot, ELISA and field inoculation, It was found that bacteria cultured for 36 h had the highest expression of PthA and showed the most virulent pathogenicity. The conservation duration of the pathogen isolates influenced their PthA expression and the pathogenicity, and negative relationship between the duration and the expression of PthA and pathogenicity. When the stored pathogen bacteria were cultured in liquid LB medium, they were able to regain activated, showing higher PthA expression level and enhanced pathogenicity, even though the activity was inferior, in terms of both PthA expression and pathogenicity, than the freshly isolated ones. Seven isolates from different citrus orchards displayed almost identical protein expression profiles. It could conclude that the expressions of PthA was positively related to pathogenicity.
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Histone deacetylase 1 and p300 can directly associate with chromatin and compete for binding in a mutually exclusive manner.
PLoS ONE
PUBLISHED: 01-01-2014
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Lysine acetyltransferases (KATs) and histone deacetylases (HDACs) are important epigenetic modifiers and dynamically cycled on active gene promoters to regulate transcription. Although HDACs are recruited to gene promoters and DNA hypersensitive sites through interactions with DNA binding factors, HDAC activities are also found globally in intergenic regions where DNA binding factors are not present. It is suggested that HDACs are recruited to those regions through other distinct, yet undefined mechanisms. Here we show that HDACs can be directly recruited to chromatin in the absence of other factors through direct interactions with both DNA and core histone subunits. HDACs interact with DNA in a non-sequence specific manner. HDAC1 and p300 directly bind to the overlapping regions of the histone H3 tail and compete for histone binding. Previously we show that p300 can acetylate HDAC1 to attenuate deacetylase activity. Here we have further mapped two distinct regions of HDAC1 that interact with p300. Interestingly, these regions of HDAC1 also associate with histone H3. More importantly, p300 and HDAC1 compete for chromatin binding both in vitro and in vivo. Therefore, the mutually exclusive associations of HDAC1/p300, p300/histone, and HDAC1/histone on chromatin contribute to the dynamic regulation of histone acetylation by balancing HDAC or KAT activity present at histones to reorganize chromatin structure and regulate transcription.
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The TCR ?? Repertoire and Relative Gene Expression Characteristics of T-ALL Cases with Biclonal Malignant V?1 and V?2 T Cells.
DNA Cell Biol.
PUBLISHED: 12-13-2013
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Despite significant improvement in our understanding of T-cell acute lymphoblastic leukemia (T-ALL) biology and pathogenesis, many questions remain unanswered. In previous studies, we found a T-ALL case with two malignant T-cell clones with V?1D?2D?3J?1 and V?2D?3J?2 rearrangements. In this study, we further characterized T-ALL cases with two malignant clones containing V?1D?3J?1 and V?2D?1J?1 rearrangements using fine-tiling array comparative genomic hybridization, ligation-mediated polymerase chain reaction (LM-PCR), sequencing, and reverse transcription polymerase chain reaction (RT-PCR) analysis. We further analyzed the distribution and clonality of the T-cell receptor (TCR) V? and V? subfamily T cells in the two T-ALL cases by RT-PCR and GeneScan. Monoclonal V?1 and V?2 subfamilies were confirmed in both samples, the V?3 through V?7 subfamilies could not be detected in the T-ALL samples, whereas the oligoclonal V?8 subfamily could be identified. Based on the clinical finding that both of the T-ALL cases with two malignant T-cell clones had a poor outcome, we attempted to compare the expression pattern of genes related to T-cell activation and proliferation between cases with the malignant V?1 and V?2 T-cell clones and T-ALL cases with a mono-malignant V? T-cell clone. We selected two T-ALL cases with V?J? rearrangements and analyzed the expression level of Notch1, TAL1, and the CARMA-BCL10-MALT-A20-NF-?B pathway genes by real-time PCR. A20 had significantly higher expression in the biclonal compared with the monoclonal T-ALL group (p=0.0354), and there was a trend toward higher expression for the other genes in the biclonal group with the exception of TAL1, although the differences were not statistically significant. In conclusion, we identified two T-ALL cases with biclonal malignant T-cell clones and described the characteristics of the biclonal T-ALL subtype and its gene expression pattern. Thus, our findings may improve the understanding of biclonal T-ALL.
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hSETD1A regulates Wnt target genes and controls tumor growth of colorectal cancer cells.
Cancer Res.
PUBLISHED: 11-18-2013
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hSETD1A is a member of the trithorax (TrxG) family of histone methyltransferases (HMTs) that methylate H3K4 at promoters of active genes. Although misregulation of MLL family proteins has been associated with acute leukemia, the role of hSETD1A in cancer remains unknown. In this study, we report that hSETD1A and its associated H3K4me3 are up-regulated in human colorectal cancer (CRC) cells and patients. Depletion of hSETD1A inhibited CRC cell growth, colony formation, and tumor engraftment. Genome-wide expression profiling of CRC cells reveals that approximately 50% of Wnt/?-catenin target genes are affected by the hSETD1A knockdown (KD) suggesting that hSETD1A regulates a subset of canonical Wnt-signaling target genes. We further demonstrate that hSETD1A is recruited to promoters of those Wnt-signaling target genes through its interaction with ?-catenin, a master regulator of the Wnt-signaling pathway. The recruitment of the hSETD1A HMT complex confers promoter-associated H3K4me3 that leads to assembly of transcription preinitiation complex (PIC) and transcriptional activation. Furthermore, the expression levels of hSETD1A are positively correlated with H3K4me3 enrichment at the promoters of Wnt/?-catenin target genes and the aberrant activation of these genes in human CRC. These results provide new biologic and mechanistic insights into the cooperative role of hSETD1A and ?-catenin in regulation of Wnt target genes as well as in CRC cell growth in vitro and in vivo.
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Histone Lysine-specific Demethylase 1 (LSD1) Protein Is Involved in Sal-like Protein 4 (SALL4)-mediated Transcriptional Repression in Hematopoietic Stem Cells.
J. Biol. Chem.
PUBLISHED: 10-25-2013
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The stem cell protein SALL4 plays a critical role in hematopoiesis by regulating the cell fate. In primitive hematopoietic precursors, it activates or represses important genes via recruitment of various epigenetic factors such as DNA methyltransferases, and histone deacylases. Here, we demonstrate that LSD1, a histone lysine demethylase, also participates in the trans-repressive effects of SALL4. Based on luciferase assays, the amine oxidase domain of LSD1 is important in suppressing SALL4-mediated reporter transcription. In freshly isolated adult mouse bone marrows, both SALL4 and LSD1 proteins are preferentially expressed in undifferentiated progenitor cells and co-localize in the nuclei. Further sequential chromatin immunoprecipitation assay confirmed that these two factors share the same binding sites at the promoter regions of important hematopoietic regulatory genes including EBF1, GATA1, and TNF. In addition, studies from both gain- and loss-of-function models revealed that SALL4 dynamically controls the binding levels of LSD1, which is accompanied by a reversely changed histone 3 dimethylated lysine 4 at the same promoter regions. Finally, shRNA-mediated knockdown of LSD1 in hematopoietic precursor cells resulted in altered SALL4 downstream gene expression and increased cellular activity. Thus, our data revealed that histone demethylase LSD1 may negatively regulate SALL4-mediated transcription, and the dynamic regulation of SALL4-associated epigenetic factors cooperatively modulates early hematopoietic precursor proliferation.
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[Effect of folic acid for treatment of acute cerebral infarction in rats].
Nan Fang Yi Ke Da Xue Xue Bao
PUBLISHED: 09-27-2013
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To test whether folic acid offers protection of the brain tissue against acute cerebral infarction in rats.
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Modeling and self-assembly behavior of PEG-PLA-PEG triblock copolymers in aqueous solution.
Nanoscale
PUBLISHED: 08-02-2013
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A series of poly(ethylene glycol)-polylactide-poly(ethylene glycol) (PEG-PLA-PEG) triblock copolymers with symmetric or asymmetric chain structures were synthesized by combination of ring-opening polymerization and copper-catalyzed click chemistry. The resulting copolymers were used to prepare self-assembled aggregates by dialysis. Various architectures such as nanotubes, polymersomes and spherical micelles were observed from transmission electron microscopy (TEM), cryo-TEM and atomic force microscopy (AFM) measurements. The formation of diverse aggregates is explained by modeling from the angle of both geometry and thermodynamics. From the angle of geometry, a "blob" model based on the Daoud-Cotton model for star polymers is proposed to describe the aggregate structures and structural changes with copolymer composition and molar mass. In fact, the copolymer chains extend in aqueous medium to form single layer polymersomes to minimize the systems free energy if one of the two PEG blocks is short enough. The curvature of polymersomes is dependent on the chain structure of copolymers, especially on the length of PLA blocks. A constant branch number of aggregates (f) is thus required to preserve the morphology of polymersomes. Meanwhile, the aggregation number (N(agg)) determined from the thermodynamics of self-assembly is roughly proportional to the total length of polymer chains. Comparing f to N(agg), the aggregates take the form of polymersomes if N(agg) ? f, and change to nanotubes if N(agg) > f to conform to the limits from both curvature and aggregation number. The length of nanotubes is mainly determined by the difference between N(agg) and f. However, the hollow structure becomes unstable when both PEG segments are too long, and the aggregates eventually collapse to yield spherical micelles. Therefore, this work gives new insights into the self-assembly behavior of PEG-PLA-PEG triblock copolymers in aqueous solution which present great interest for biomedical and pharmaceutical applications.
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Carbon nanodots as a matrix for the analysis of low-molecular-weight molecules in both positive- and negative-ion matrix-assisted laser desorption/ionization time-of-flight mass spectrometry and quantification of glucose and uric acid in real samples.
Anal. Chem.
PUBLISHED: 07-05-2013
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Carbon nanodots were applied for the first time as a new matrix for the analysis of low-molecular-weight compounds by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) in both positive- and negative-ion modes. A wide range of small molecules including amino acids, peptides, fatty acids, as well as ?-agonists and neutral oligosaccharides were analyzed by MALDI MS with carbon nanodots as the matrix, and the lowest 0.2 fmol limits-of-detection were obtained for octadecanoic acid. Clear sodium and potassium adducts and deprotonated signals were produced in positive- and negative-ion modes. Furthermore, the glucose and uric acid in real samples were quantitatively determined by the internal standard method with the linear range of 0.5-9 mM and 0.1-1.8 mM (R(2) > 0.999), respectively. This work gives new insight into the application of carbon nanodots and provides a general approach for rapid analysis of low-molecular-weight compounds.
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First report of liver abscess caused by Salmonella enterica serovar Dublin.
J. Clin. Microbiol.
PUBLISHED: 06-19-2013
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This is the first reported case of liver abscess attributable to Salmonella serovar Dublin infection and also the fourth case of Salmonella liver abscess complicated with hepatocellular carcinoma reported since 1990. Drainage combined with intravenous antibiotics resulted in improvement, but recovery regressed again. Subsequent hepatic left lobectomy led to full recovery.
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USF1 and hSET1A mediated epigenetic modifications regulate lineage differentiation and HoxB4 transcription.
PLoS Genet.
PUBLISHED: 06-01-2013
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The interplay between polycomb and trithorax complexes has been implicated in embryonic stem cell (ESC) self-renewal and differentiation. It has been shown recently that WRD5 and Dpy-30, specific components of the SET1/MLL protein complexes, play important roles during ESC self-renewal and differentiation of neural lineages. However, not much is known about how and where specific trithorax complexes are targeted to genes involved in self-renewal or lineage-specification. Here, we report that the recruitment of the hSET1A histone H3K4 methyltransferase (HMT) complex by transcription factor USF1 is required for mesoderm specification and lineage differentiation. In undifferentiated ESCs, USF1 maintains hematopoietic stem/progenitor cell (HS/PC) associated bivalent chromatin domains and differentiation potential. Furthermore, USF1 directed recruitment of the hSET1A complex to the HoxB4 promoter governs the transcriptional activation of HoxB4 gene and regulates the formation of early hematopoietic cell populations. Disruption of USF or hSET1A function by overexpression of a dominant-negative AUSF1 mutant or by RNA-interference-mediated knockdown, respectively, led to reduced expression of mesoderm markers and inhibition of lineage differentiation. We show that USF1 and hSET1A together regulate H3K4me3 modifications and transcription preinitiation complex assembly at the hematopoietic-associated HoxB4 gene during differentiation. Finally, ectopic expression of USF1 in ESCs promotes mesoderm differentiation and enforces the endothelial-to-hematopoietic transition by inducing hematopoietic-associated transcription factors, HoxB4 and TAL1. Taken together, our findings reveal that the guided-recruitment of the hSET1A histone methyltransferase complex and its H3K4 methyltransferase activity by transcription regulator USF1 safeguards hematopoietic transcription programs and enhances mesoderm/hematopoietic differentiation.
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Differentiation of Chinese liquors by using ambient glow discharge ionization mass spectrometry.
Analyst
PUBLISHED: 05-22-2013
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Chinese liquors are often a very important part of social event in China. Driven by high profit, some illegal traders often use inferior liquors instead of the products with high quality to cheat the customer. Therefore, it is highly required to authenticate Chinese liquors. In this paper a novel method based on ambient glow discharge ionization mass spectrometry has been developed to differentiate Chinese liquors. Volatile components from liquor samples were ionized by the plasma generated by glow discharge and then detected by a commercial linear ion trap mass spectrometer. Consequently, the fingerprint mass spectra of several Chinese liquors were obtained. Combined with principal component analysis, this new method was successfully applied to differentiate different brands of Chinese liquors without any sample pretreatment. Compared with conventional methods, this novel method has the advantages of easy operation, high speed, and high efficiency, which make it a potential tool in the fields of food safety, atmosphere chemistry.
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Bone marrow-derived Gr1+ cells can generate a metastasis-resistant microenvironment via induced secretion of thrombospondin-1.
Cancer Discov
PUBLISHED: 04-30-2013
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Metastatic tumors have been shown to establish permissive microenvironments for metastases via recruitment of bone marrow-derived cells. Here, we show that metastasis-incompetent tumors are also capable of generating such microenvironments. However, in these situations, the otherwise prometastatic Gr1(+) myeloid cells create a metastasis-refractory microenvironment via the induction of thrombospondin-1 (Tsp-1) by tumor-secreted prosaposin. Bone marrow-specific genetic deletion of Tsp-1 abolished the inhibition of metastasis, which was restored by bone marrow transplant from Tsp-1(+) donors. We also developed a 5-amino acid peptide from prosaposin as a pharmacologic inducer of Tsp-1 in Gr1(+) bone marrow cells, which dramatically suppressed metastasis. These results provide mechanistic insights into why certain tumors are deficient in metastatic potential and implicate recruited Gr1(+) myeloid cells as the main source of Tsp-1. The results underscore the plasticity of Gr1(+) cells, which, depending on the context, promote or inhibit metastasis, and suggest that the peptide could be a potential therapeutic agent against metastatic cancer.
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Novel Poly(L-lactide-co- ? -caprolactone) Matrices Obtained with the Use of Zr[Acac] 4 as Nontoxic Initiator for Long-Term Release of Immunosuppressive Drugs.
Biomed Res Int
PUBLISHED: 04-26-2013
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Slowly degradable copolymers of L-lactide and ? -caprolactone can provide long-term delivery and may be interesting as alternative release systems of cyclosporine A (CyA) and rapamycin (sirolimus), in which available dosage forms cause a lot of side effects. The aim of this study was to obtain slowly degradable matrices containing immunosuppressive drug from PLACL initiated by nontoxic Zr[Acac]4. Three kinds of poly(L-lactide-co- ? -caprolactone) (PLACL) matrices with different copolymer chain microstructure were used to compare the release process of cyclosporine A and rapamycine. The influence of copolymer chain microstructure on drug release rate and profile was also analyzed. The determined parameters could be used to tailor drug release by synthesis of demanded polymeric drug carrier. The studied copolymers were characterized at the beginning and during the degradation process of the polymeric matrices by NMR spectroscopy, GPC (gel permeation chromatography), and DSC (differential scanning calorimetry). Different drug release profiles have been observed from each kind of copolymer. The correlation between drug release process and changes of copolymer microstructure during degradation process was noticed. It was determined that different copolymer composition (e.g., lower amount of caprolactone units) does not have to influence the drug release, but even small changes in copolymer randomness affect this process.
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Induced dual-nanospray: a novel internal calibration method for convenient and accurate mass measurement.
J. Am. Soc. Mass Spectrom.
PUBLISHED: 03-18-2013
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Accurate mass information is of great importance in the determination of unknown compounds. An effective and easy-to-control internal mass calibration method will dramatically benefit accurate mass measurement. Here we reported a simple induced dual-nanospray internal calibration device which has the following three advantages: (1) the two sprayers are in the same alternating current field; thus both reference ions and sample ions can be simultaneously generated and recorded. (2) It is very simple and can be easily assembled. Just two metal tubes, two nanosprayers, and an alternating current power supply are included. (3)With the low-flow-rate character and the versatility of nanoESI, this calibration method is capable of calibrating various samples, even untreated complex samples such as urine and other biological samples with small sample volumes. The calibration errors are around 1 ppm in positive ion mode and 3 ppm in negative ion mode with good repeatability. This new internal calibration method opens up new possibilities in the determination of unknown compounds, and it has great potential for the broad applications in biological and chemical analysis.
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Genetic manipulation in Sulfolobus islandicus and functional analysis of DNA repair genes.
Biochem. Soc. Trans.
PUBLISHED: 01-30-2013
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Recently, a novel gene-deletion method was developed for the crenarchaeal model Sulfolobus islandicus, which is a suitable tool for addressing gene essentiality in depth. Using this technique, we have investigated functions of putative DNA repair genes by constructing deletion mutants and studying their phenotype. We found that this archaeon may not encode a eukarya-type of NER (nucleotide excision repair) pathway because depleting each of the eukaryal NER homologues XPD, XPB and XPF did not impair the DNA repair capacity in their mutants. However, among seven homologous recombination proteins, including RadA, Hel308/Hjm, Rad50, Mre11, HerA, NurA and Hjc, only the Hjc nuclease is dispensable for cell viability. Sulfolobus encodes redundant BER (base excision repair) enzymes such as two uracil DNA glycosylases and two putative apurinic/apyrimidinic lyases, but inactivation of one of the redundant enzymes already impaired cell growth, highlighting their important roles in archaeal DNA repair. Systematically characterizing these mutants and generating mutants lacking two or more DNA repair genes will yield further insights into the genetic mechanisms of DNA repair in this model organism.
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The hydrogen sulfide donor, GYY4137, exhibits anti-atherosclerotic activity in high fat fed apolipoprotein E(-/-) mice.
Br. J. Pharmacol.
PUBLISHED: 01-11-2013
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Atherosclerosis is associated with reduced vascular hydrogen sulfide (H2 S) biosynthesis. GYY4137 is a novel slow-releasing H2 S compound that may effectively mimic the time course of H2 S release in vivo. However, it is not known whether GYY4137 affects atherosclerosis.
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Changing trends and serotype distribution of Shigella species in Beijing from 1994 to 2010.
Gut Pathog
PUBLISHED: 01-01-2013
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Shigella species are a common cause of acute diarrheal disease in China. In this study, we characterized the changing trends and serotype distribution of Shigella species in Beijing from 1994 to 2010. A total of 5999 Shigella strains were isolated and serotyped from the 302nd Hospital in Beijing. The annual number of Shigella isolates reached a peak (n = 1192; 19.84%) in 1996 and then decreased annually, reaching the lowest point (n = 24; 0.41%) in 2010. S. flexneri 2a and S. sonnei were the most frequently isolated Shigella, with their respective isolates making up 53.3% and 27.6% of the total. Isolates of S. flexneri 4c, 4a, and x made up 3% respectively of the total isolates. Significant decreases in percentage of S. flexneri over time were observed. S. sonnei surpassed S. flexneri 2a as the predominant serotype in 2000. Most isolates were recovered from July to September; 13.6% of the isolates were recovered from children aged 0 to 5 years, and 16% were recovered from those aged 21 to 25 years. S. flexneri 2a and 5 were recovered mostly from males (33.41%, p < 0.001; and 0.46%, p < 0.001%; respectively), whereas S. flexneri 2b and 6, and S. sonnei were most often isolated from females. Continuous monitoring of Shigella showed that all 4 species and 27 serotypes were present in Beijing, China, during the study period. The emergence of S. sonnei and the overall decreasing isolation rate of Shigella in Beijing can potentially aid in the development of vaccine and control strategies for shigellosis in the city.
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Celastrol prevents atherosclerosis via inhibiting LOX-1 and oxidative stress.
PLoS ONE
PUBLISHED: 01-01-2013
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Celastrol is a triterpenoid compound extracted from the Chinese herb Tripterygium wilfordii Hook F. Previous research has revealed its anti-oxidant, anti-inflammatory, anti-cancer and immunosuppressive properties. Here, we investigated whether celastrol inhibits oxidized low-density lipoprotein (oxLDL) induced oxidative stress in RAW 264.7 cells. In addition, the effect of celastrol on atherosclerosis in vivo was assessed in apolipoprotein E knockout (apoE(-/-)) mouse fed a high-fat/high-cholesterol diet (HFC). We found that celastrol significantly attenuated oxLDL-induced excessive expression of lectin-like oxidized low density lipoprotein receptor-1(LOX-1) and generation of reactive oxygen species (ROS) in cultured RAW264.7 macrophages. Celastrol also decreased I?B phosphorylation and degradation and reduced production of inducible nitric oxide synthase (iNOS), nitric oxide (NO) and proinflammatory cytokines such as tumor necrosis factor (TNF)-? and IL-6. Celastrol reduced atherosclerotic plaque size in apoE(-/-) mice. The expression of LOX-1 within the atherosclerotic lesions and generation of superoxide in mouse aorta were also significantly reduced by celastrol while the lipid profile was not improved. In conclusion, our results show that celastrol inhibits atherosclerotic plaque developing in apoE(-/-) mice via inhibiting LOX-1 and oxidative stress.
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Evaluation of a novel paclitaxel-eluting stent with a bioabsorbable polymeric surface coating in the porcine artery injury model.
Acta Cardiol
PUBLISHED: 10-17-2011
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Drug-eluting stents (DES) are unique in allowing sustained release after a single short intervention. The challenge with DES still remaining is the optimal combination of a biocompatible drug-eluting matrix including an antiproliferative drug. We studied the role of a novel paclitaxel-eluting stent with a bioabsorbable polymer coating in preventing vascular restenosis in the porcine artery injury model.
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Comparative proteomic analysis of a Haemophilus parasuis SC096 mutant deficient in the outer membrane protein P5.
Microb. Pathog.
PUBLISHED: 08-17-2011
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Outer membrane protein A (OmpA) is a major structural component of the outer membranes and functions as a multifaceted molecular with many diverse roles in Gram-negative bacteria. In Haemophilus parasuis, OmpA has been recognized and named as OmpP5 in genomic literature. In this study, to determine the precise functions of OmpP5, an ompP5 deficient mutant (?ompP5) of a H. parasuis serovar 4 filed strain SC096 was constructed using a natural transformation method. Compared to the wild-type SC096 strain, the ?ompP5 mutant displayed a detectable delay in growth. However, the wild-type and mutant strains were indistinguishable with respect to the other phenotypes including resistance to killing by porcine and rabbit sera, adhesion to and invasion of porcine umbilicus veins endothelial cells (PUVEC) and porcine kidney epithelial cells (PK-15). To analyze the differences of proteome expression between wild-type and mutant strains, a 2-dimensional gel electrophoresis (2-DE)-based proteomics comparison was performed. There were 24 differentially expressed proteins which were mainly involved in carbohydrate, lipid, nucleotide and amino acid metabolism, or served as transcription and translation factors and chaperone proteins. Collectively, loss of OmpP5 expression in the H. parasuis SC096 strain resulted in global protein expression changes which might be responsible for novel phenotypes occurred in ?ompP5 mutant.
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Regulation of nucleosome landscape and transcription factor targeting at tissue-specific enhancers by BRG1.
Genome Res.
PUBLISHED: 07-27-2011
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Enhancers of transcription activate transcription via binding of sequence-specific transcription factors to their target sites in chromatin. In this report, we identify GATA1-bound distal sites genome-wide and find a global reorganization of the nucleosomes at these potential enhancers during differentiation of hematopoietic stem cells (HSCs) to erythrocytes. We show that the catalytic subunit BRG1 of BAF complexes localizes to these distal sites during differentiation and generates a longer nucleosome linker region surrounding the GATA1 sites by shifting the flanking nucleosomes away. Intriguingly, we find that the nucleosome shifting specifically facilitates binding of TAL1 but not GATA1 and is linked to subsequent transcriptional regulation of target genes.
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Serum resistance in Haemophilus parasuis SC096 strain requires outer membrane protein P2 expression.
FEMS Microbiol. Lett.
PUBLISHED: 07-25-2011
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Haemophilus parasuis outer membrane protein P2 (OmpP2), the most abundant protein in the outer membrane, has been identified as an antigenic protein and a potential virulence factor. To study the precise function of OmpP2, an ompP2-deficient mutant (?ompP2) of a H. parasuis serovar 4 clinical strain SC096 was constructed by a modified natural transformation system. Compared with the wild-type SC096 strain, the ?ompP2 mutant showed a pronounced growth defect and exhibited significantly greater sensitivity to the bactericidal action of porcine and rabbit sera, whereas the complemented strain could restore the growth and serum resistance phenotypes. The results indicated that H. parasuis OmpP2 from SC096 strain is an important surface protein involved in serum resistance.
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Enzyme-Catalyzed Degradation of Biodegradable Polymers Derived from Trimethylene Carbonate and Glycolide by Lipases from Candida Antarctica and Hog Pancreas.
J Biomater Sci Polym Ed
PUBLISHED: 07-05-2011
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Enzyme-catalyzed degradation of poly(trimethylene carbonate) homo-polymer (PTMC) and poly(trimethy- lene carbonate-co-glycolide) co-polymer (PTGA) was investigated in the presence of lipases from Candida antarctica and Hog pancreas. Degradation was monitored by gravimetry, size-exclusion chromatography (SEC), nuclear magnetic resonance (NMR), tensiometry and environmental scanning electron microscopy (ESEM). PTMC can be rapidly degraded by Candida antarctica lipase with 98% mass loss after 9 days, while degradation by Hog pancreas lipase leads to 27% mass loss. Introduction of 16% glycolide units in PTMC chains strongly affects the enzymatic degradation. Hog pancreas lipase becomes more effective to PTGA co-polymer with a mass loss of 58% after 9 days, while Candida antarctica lipase seems not able to degrade PTGA. Bimodal molecular weight distributions are observed during enzymatic degradation of both PTMC and PTGA, which can be assigned to the fact that the surface is largely degraded while the internal part remains intact. The composition of the PTGA co-polymer remains constant, and ESEM shows that the polymers are homogeneously eroded during enzymatic degradation. Contact angle measurements confirm the enzymatic degradation mechanism, i.e., enzyme adsorption on the polymer surface followed by enzyme-catalyzed chain cleavage.
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Spectroscopic response of ferrocene derivatives bearing a BODIPY moiety to water: a new dissociation reaction.
Chemistry
PUBLISHED: 06-09-2011
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Ferrocence and its derivatives have long been known to be a class of stable organometallic compounds, and their dissociation usually occurs under harsh conditions. Here we report a new type of ferrocene derivatives, 4,4-difluoro-8-ferrocenyl-1,3,5,7-tetramethyl-4-bora-3a,4a-diaza-s-indacene and 4,4-difluoro-2,6-diethyl-8-ferrocenyl-1,3,5,7-tetramethyl-4-bora-3a,4a-diaza-s-indacene, which surprisingly can hydrolyze under mild conditions. These two derivatives, initially developed as donor-acceptor probes for reactive oxygen species by incorporating the electron donor of ferrocene as a quencher into the fluorophore of BODIPY (boron dipyrromethene difluoride), barely emit fluorescence. Upon reaction with H(2)O under the irradiation of natural light at room temperature, however, both of the probes display a dramatic color change and fluorescence retrievement. Detailed experimental results reveal that the reaction of the probes with H(2)O leads to the removal of a cyclopentadiene unit and iron(II), yielding a BODIPY derivative that retains the other cyclopentadiene unit and shows a large fluorescence enhancement (over 100-fold). Moreover, the increase in fluorescence intensity is directly proportional to microamount of water, and the presence of both light and H(2)O is indispensable in the reaction, which makes the present system of great potential not only for determining water but also for forming a AND logic gate. Most importantly, the present mild dissociation reaction may give a new insight into the stability of ferrocene and its derivatives.
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Chromatin boundaries require functional collaboration between the hSET1 and NURF complexes.
Blood
PUBLISHED: 06-08-2011
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Chromatin insulators protect erythroid genes from being silenced during erythropoiesis, and the disruption of barrier insulator function in erythroid membrane gene loci results in mild or severe anemia. We showed previously that the USF1/2-bound 5HS4 insulator mediates chromatin barrier activity in the erythroid-specific chicken ?-globin locus. It is currently not known how insulators establish such a barrier. To understand the function of USF1, we purified USF1-associated protein complexes and found that USF1 forms a multiprotein complex with hSET1 and NURF, thus exhibiting histone H3K4 methyltransferase- and ATP-dependent nucleosome remodeling activities, respectively. Both SET1 and NURF are recruited to the 5HS4 insulator by USF1 to retain the active chromatin structure in erythrocytes. Knock-down of NURF resulted in a rapid loss of barrier activity accompanied by an alteration of nucleosome positioning, increased occupancy of the nucleosome-free linker region at the insulator site, and increased repressive H3K27me3 levels in the vicinity of the HS4 insulator. Furthermore, suppression of SET1 reduced barrier activity, decreased H3K4me2 and acH3K9/K14, and diminished the recruitment of BPTF at several erythroid-specific barrier insulator sites. Therefore, our data reveal a synergistic role of hSET1 and NURF in regulating the USF-bound barrier insulator to prevent erythroid genes from encroachment of heterochromatin.
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Anisotropic self-assembling micelles prepared by the direct dissolution of PLA/PEG block copolymers with a high PEG fraction.
Langmuir
PUBLISHED: 06-03-2011
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A series of polylactide-poly(ethylene glycol) (PLA-PEG) block copolymers with a high PEG fraction were synthesized by the ring-opening polymerization of L- or D-lactide in the presence of mono- or dihydroxyl PEG using nontoxic zinc lactate as a catalyst. Micelles were then prepared by direct dissolution of the obtained water-soluble copolymers in an aqueous medium without heating or using any organic solvents. Large anisotropic micelles instead of conventional spherical ones were observed from a transmission electron microscopy examination. Various parameters influencing the structure of the novel micelles were considered, such as the copolymer chain structure, molar mass, PEG fraction, copolymer concentration, and stereocomplexation between L- and D-PLA blocks. Anisotropic micelles were obtained for both diblock and triblock copolymers but vanished with increasing molar mass of the copolymers. The morphology of micelles strongly depends on the PEG fraction. Anisotropic micelles were found only in an intermediate EO/LA ratio range in which a higher PEG fraction leads to a higher length/width ratio of micelles. Stereocomplexation between L- and D-PLA or a lower concentration disfavors the formation of anisotropic micelles. Under appropriate concentrations, spherical and anisotropic micelles coexist in the same micellar solution. Moreover, it was found that anisotropic micelles are susceptible to further self-assemble into more organized complex aggregates. Similar results were obtained from light scattering and aqueous gel permeation chromatography measurements. A novel model is proposed to explain the formation of anisotropic micelles and the effects of various parameters on the structure of micelles in an aqueous medium.
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In vivo study on the histocompatibility and degradation behavior of biodegradable poly(trimethylene carbonate-co-D,L-lactide).
Acta Biochim. Biophys. Sin. (Shanghai)
PUBLISHED: 05-12-2011
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The aim of this study was to explore the in vivo behavior and histocompatibility of poly(trimethylene carbonate-co-D,L-lactide) (PDLLA/TMC) and its feasibility of manufacturing cardiovascular stents. Copolymers with 50/50 molar ratio were synthesized by ring-opening polymerization with TMC and D, L-LA, or TMC and L-LA. Poly(L-lactide) (PLLA) was synthesized as a control. The films of the three polymers were implanted into 144 Wistar rats. At different time points of implantation, polymer films were explanted for the evaluation of degradation characteristics and histocompatibility using size exclusion chromatography , nuclear magnetic resonance , environmental scanning electron microscope , and optical microscope. Results showed that there were differences in the percentage of mass loss, molecular weight, shape and appearance changes, and inflammation cell counts between different polymers. With the time extended, the films superficial structure transformed variously, which was rather obvious in the polymer of PDLLA/TMC. In addition, there were relatively lower inflammation cell counts in the PDLLA/TMC and poly(trimethylene carbonate-co-L-lactide) (PLLA/TMC) groups at different time points in comparison with those in the PLLA group. The differences were of statistical significance (P< 0.05) in the group of PDLLA/TMC vs. PLLA, and the group of PLLA/TMC vs. PLLA, but not within the PDLLA/TMC and PLLA/TMC groups (P> 0.05). These results suggested that the polymer of PDLLA/TMC (50/50) with favorable degradation performance and histocompatibility is fully biodegradable and suitable for manufacturing implanted cardiovascular stents.
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A barrier-only boundary element delimits the formation of facultative heterochromatin in Drosophila melanogaster and vertebrates.
Mol. Cell. Biol.
PUBLISHED: 04-25-2011
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Formation of facultative heterochromatin at specific genomic loci is fundamentally important in defining cellular properties such as differentiation potential and responsiveness to developmental, physiological, and environmental stimuli. By the nature of their formation, heterochromatin and repressive histone marks propagate until the chain reaction is broken. While certain active promoters can block propagation of heterochromatin, there are also specialized DNA elements, referred to as chromatin barriers, that serve to demarcate the boundary of facultative heterochromatin formation. In this study, we identified a chromatin barrier that specifically limits the formation of repressive chromatin to a distal enhancer region so that repressive histone modifications cannot reach the promoter and promoter-proximal enhancer regions of reaper. Unlike all of the known boundary elements identified for Drosophila melanogaster, this IRER (irradiation-responsive enhancer region) left barrier (ILB) does not exhibit enhancer-blocking activity. Not only has the ILB been conserved in different Drosophila species, it can also function as an effective chromatin barrier in vertebrate cells. This suggests that the mechanism by which it functions to spatially restrict the formation of repressive chromatin marked by trimethylated H3K27 has also been conserved widely during evolution.
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Lipoxin A4 inhibits the production of proinflammatory cytokines induced by ?-amyloid in vitro and in vivo.
Biochem. Biophys. Res. Commun.
PUBLISHED: 03-31-2011
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Studies increasingly indicate that inflammation induced by ?-amyloid (A?) contributes to the progression of Alzheimers disease (AD). How to inhibit the enhanced production of proinflammatory cytokines stimulated by A? is an important research subject for the treatment of AD. In this study, we investigated the inhibitory effect and the molecular mechanism of the lipoxin A(4) (LXA(4)) on the production of interleukin-1? (IL-1?) and tumor necrosis factor? (TNF?) induced by ?-amyloid in the cortex and hippocampus of mice, and in A?-stimulated BV2 cells, a mouse microglial cell line. LXA(4) down-regulated the protein expression of IL-1? and TNF?, attenuated the gene expressions of IL-1? and TNF?, inhibited the degradation of I?B?, inhibited translocation of NF-?B p65 subunit into the nucleus induced by ?-amyloid in the cortex and hippocampus of mice, and in A?-stimulated BV2 cells, and the inhibitory effects were dose dependently elevated. Our findings suggest that LXA(4) inhibits the production of IL-1? and TNF? induced by ?-amyloid in the cortex and hippocampus of mice, and in BV2 microglial cells via the NF-?B signal pathway.
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Controlled poly(l-lactide-co-trimethylene carbonate) delivery system of cyclosporine A and rapamycine--the effect of copolymer chain microstructure on drug release rate.
Int J Pharm
PUBLISHED: 02-10-2011
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The effect of poly(L-lactide-co-TMC) chain microstructure (and its changes during degradation) on immunosuppressive drugs release process was analyzed. Three kinds of poly(L-lactide-co-TMC) (PLATMC)--two semiblock and one random were used to prepare matrices containing cyclosporine A or rapamycine and drug free matrices. All of them degraded slowly enough to provide long term delivery of immunosuppressive agents. Moreover, copolymer chain microstructure determined the effect of drug loading on the degradation process. It was observed that matrices without drug obtained from semiblock copolymer degraded differently than matrices containing cyclosporine A or rapamycine, whereas all kinds of matrices obtained from random PLATMC degraded in similar way. This is the evidence that the only in case of semiblock copolymer factors concerning the presence of drug and the kind of drug influenced degradation process. Based on the obtained results, correlations between copolymer degradation and drug release process are proposed. According to our outcomes, regular drug release process may be obtained from highly randomized copolymers (R ? 1) that remain amorphous during degradation process. Determination of this factor may help in development of biodegradable systems, in which drug release rate and profile can be tailored by synthesis of polymer with appropriate chain microstructure.
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Role of helix-loop-helix proteins during differentiation of erythroid cells.
Mol. Cell. Biol.
PUBLISHED: 01-31-2011
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Helix-loop-helix (HLH) proteins play a profound role in the process of development and cellular differentiation. Among the HLH proteins expressed in differentiating erythroid cells are the ubiquitous proteins Myc, USF1, USF2, and TFII-I, as well as the hematopoiesis-specific transcription factor Tal1/SCL. All of these HLH proteins exhibit distinct functions during the differentiation of erythroid cells. For example, Myc stimulates the proliferation of erythroid progenitor cells, while the USF proteins and Tal1 regulate genes that specify the differentiated phenotype. This minireview summarizes the known activities of Myc, USF, TFII-I, and Tal11/SCL and discusses how they may function sequentially, cooperatively, or antagonistically in regulating expression programs during the differentiation of erythroid cells.
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[Biocompatibility evaluation of lactide--trimethylene carbonate copolymers].
Sheng Wu Yi Xue Gong Cheng Xue Za Zhi
PUBLISHED: 07-24-2010
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Biocompatibility is the essential property of biomaterials, which is the essence of biomaterial evaluation as well as the foundation of the design and improvement of biomaterials. Several methods were carried out to evaluate the biocompatibility of poly(L-Lactide)-b-poly(trimethylene carbonate (PLLA-b-PTMC) and poly(D,L-Lactide)-b-poly(trimethylene carbonate) (PDLLA-b-PTMC) with poly(L-Lactide) (PLLA) and poly(trimethylene carbonate) (PTMC) as control, including extract liquid experiment, directly contact experiment of materials and cells, hemolytic ratio analysis and platelet adhesion investigation. The results revealed that all the materials exhibited an acceptable cytotoxicity, and proliferation of cells on the modified materials was less than that on the PLLA but more than that on PTMC. The results of hemocompatibility experiments showed that no significant hemolysis was detected when all the materials were in use; in addition, the numbers of platelets adhered on the surface of copolymers were smaller than that on the surface of PLLA, and the degree of platelet deformation was slighter. So, the biocompatibility of copolymers is similar to that of PLLA, the biocompatibility of PLLA is not remarkably changed by modification with PTMC, but rather is improved.
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Epilepsy surgery in tuberous sclerosis complex: emphasis on surgical candidate and neuropsychology.
Epilepsia
PUBLISHED: 07-17-2010
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To discuss neuropsychological outcome and candidate of epilepsy surgery for tuberous sclerosis complex (TSC).
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In vitro degradation behavior of poly(lactide)-poly(ethylene glycol) block copolymer micelles in aqueous solution.
Int J Pharm
PUBLISHED: 05-16-2010
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Self-assembling micelles were prepared from polylactide-poly(ethylene glycol) (PLA-PEG) block copolymer by using two different methods: direct dissolution and dialysis. The in vitro degradation properties of the micelles were investigated at 37°C and monitored by using various techniques. During the investigated degradation time, the size of micelles by dialysis remains stable, while that of micelles by direct dissolution first increases, followed by a collapse of micellar structure. The composition of PLA-PEG copolymers greatly affects the degradation of micelles. Micelles with longer hydrophobic PLA blocks exhibit less size changes due to more compact structure. On the other hand, the structural integrity of L/D mixed micelles is preserved for longer time than that of single micelles, in agreement with the stereocomplexation effect between L-PLA and D-PLA blocks. As degradation proceeds, the average molar mass of copolymer decreases and the distribution becomes wider, especially for micelles by dialysis and L/D mixed micelles with a more compact structure. Additionally, the PEG content in the copolymer chains increases during degradation, leading to a decrease of glass transition and crystallization temperature of the copolymers. However, the residual LA fragments produced by degradation disfavors the crystallization of PEG blocks, thus resulting in the decrease of melting temperature and melting enthalpy.
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Rhodamine B piperazinoacetohydrazine: a water-soluble spectroscopic reagent for pyruvic acid labeling.
Chemistry
PUBLISHED: 04-23-2010
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A new water-soluble reagent, rhodamine B piperazinoacetohydrazine (RBPH), with improved spectroscopic and reaction properties, has been developed and characterized for pyruvic acid labeling. The reagent RBPH is designed and synthesized by using rhodamine B as a spectroscopic unit, and hydrazine as a carbonyl-specific labeling unit; the two units are connected by a well-chosen linker of piperazine, which prohibits the formation of the nonfluorescent spirocyclic structure of rhodamine B, thereby keeping the spectroscopic response of the reagent in a stable state. Such a design enables RBPH not only to maintain its excellent spectroscopic properties over a wide pH range, but also to exist as a stable cation with high water solubility. Moreover, the hydrazino group of RBPH is expected to react selectively with carbonyl compounds under mild conditions through the rapid formation of hydrazones. These important features make RBPH of great potential use in the labeling of aldehydes or ketones in various biosystems, and such an application of RBPH as a precolumn derivatizing reagent has been successfully demonstrated on the analysis of pyruvic acid in human serum by high-performance liquid chromatography with common UV/Vis detection.
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Role of leukaemia inhibitory factor in the induction of pluripotent stem cells in mice.
Cell Biol. Int.
PUBLISHED: 04-16-2010
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iPS (induced pluripotent stem) cells can be induced from somatic cells in mice by genetic manipulation. Most previously established mouse iPS cell lines have been derived using feeder layers supplemented with exogenous LIF (leukaemia inhibitory factor). Although a feeder-free induction system has been developed in recent studies, LIF is still required for reprogramming, but its role in the generation of mouse iPS cells has remained elusive. In this study, we investigated its contribution to the induction of pluripotency. Our results showed that LIF activates AP (alkaline phosphatase) through a c-Myc-dependent mechanism. Moreover, it acts as a protective factor during the transition from AP-positive colonies to Oct3/4-positive cells. These findings illustrate a mechanism by which LIF may integrate signalling into reprogramming.
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USF and NF-E2 cooperate to regulate the recruitment and activity of RNA polymerase II in the beta-globin gene locus.
J. Biol. Chem.
PUBLISHED: 03-17-2010
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The human beta-globin gene is expressed at high levels in erythroid cells and regulated by proximal and distal cis-acting DNA elements, including promoter, enhancer, and a locus control region (LCR). Transcription complexes are recruited not only to the globin gene promoters but also to the LCR. Previous studies have implicated the ubiquitously expressed transcription factor USF and the tissue-restricted activator NF-E2 in the recruitment of transcription complexes to the beta-globin gene locus. Here we demonstrate that although USF is required for the efficient association of RNA polymerase II (Pol II) with immobilized LCR templates, USF and NF-E2 together regulate the association of Pol II with the adult beta-globin gene promoter. Recruitment of Pol II to the LCR occurs in undifferentiated murine erythroleukemia cells, but phosphorylation of LCR-associated Pol II at serine 5 of the C-terminal domain is mediated by erythroid differentiation and requires the activity of NF-E2. Furthermore, we provide evidence showing that USF interacts with NF-E2 in erythroid cells. The data provide mechanistic insight into how ubiquitous and tissue-restricted transcription factors cooperate to regulate the recruitment and activity of transcription complexes in a tissue-specific chromatin domain.
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A near-infrared fluorescent probe for monitoring tyrosinase activity.
Chem. Commun. (Camb.)
PUBLISHED: 03-11-2010
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A new cyanine-based near-infrared fluorescent probe has been developed for monitoring tyrosinase activity.
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Haemo- and cytocompatibility of bioresorbable homo- and copolymers prepared from 1,3-trimethylene carbonate, lactides, and epsilon-caprolactone.
J Biomed Mater Res A
PUBLISHED: 02-27-2010
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A series of bioresorbable polymers were prepared by ring-opening polymerization of L-lactide (LLA), DL-lactide (DLLA), epsilon-caprolactone (CL) and 1,3-trimethylene carbonate (TMC), using low toxic zinc lactate as catalyst. The various PLLA, PTMC, PCL homopolymers, and PLLA-TMC, PDLLA-TMC, PCL-TMC copolymers with 50/50 molar ratios were characterized by using analytical techniques such as proton nuclear magnetic resonance, gel permeation chromatography, tensiometer, and differential scanning calorimetry. The haemo- and cyto-compatibility were investigated in order to evaluate the potential of the polymers as coating material in drug eluting stents. Haemolysis tests show that all the homo- and copolymers present very low haemolytic ratios, indicating good haemolytic properties. Adhesion and activation of platelets were observed on the surface of PLLA, PCL, PLLA-TMC, and PDLLA-TMC films, while less platelets and lower activation were found on PTMC. The most interesting results were obtained with PCL-TMC which exhibited the lowest degree of activation with few adhered platelets, in agreement with its outstanding anticoagulant properties. Both indirect and direct cytocompatibility studies were performed on the polymers. The relative growth ratio data obtained from the liquid extracts during the 6-day cell culture period indicate that all the polymers present very low cytotoxicity. Microscopic observations demonstrate adhesion, spreading and proliferation of human umbilical vein endothelial cells ECV304. Therefore, it is concluded that these bioresorbable polymers, in particular PCL-TMC, are promising candidate materials as drug loading coating material in drug eluting stents.
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Aggregation behavior of self-assembling polylactide/poly(ethylene glycol) micelles for sustained drug delivery.
Int J Pharm
PUBLISHED: 02-18-2010
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A series of diblock copolymers were synthesized by ring-opening polymerization of l- or d-lactide in the presence of monomethoxy poly(ethylene glycol) (mPEG) with molar masses of 2000 and 5000. The aggregation behavior of the resulting water soluble PLA/PEG diblock copolymers in aqueous medium was studied with dynamic and static light scattering (DLS and SLS), in combination with aqueous gel permeation chromatography (GPC). The average hydrodynamic radius (R(h)) of l-PLA/PEG and d-PLA/PEG mixed micelles is lower than that of l-PLA/PEG single micelles due to the stereocomplexation effect between l-PLA and d-PLA blocks. It is also confirmed that the micelle size increases with increasing temperature and hydrophobic block length, but decreases after salt addition. Aqueous GPC and SLS were used to evaluate the molecular weight (M(w)) and aggregation number (N(agg)) of the micelles. Mixed micelles present lower N(agg) than single copolymer micelles due to stereocomplexation. N(agg) decreases with increasing hydrophobic block length, and decreases upon addition of NaCl, in agreement with a more compact structure. In contrast, N(agg) increases with elevating temperature. The average radius of gyration (R(g)) and R(g)/R(h) ratio data show that both increase with increasing temperature, suggesting that micelles exhibit a compact hard-sphere structure at 15 degrees C and a swollen structure at 35 degrees C. As the temperature increases from 15 degrees C to 35 degrees C, the second virial coefficient (A(2)) of PLA/PEG copolymers turns from negative to positive, which means that water changes from poor solvent to good solvent. The fact that the average density (rho) of PLA/PEG micelles decreases with increasing temperature confirms that micelles exhibit a looser structure at higher temperatures due to water swelling effect. The higher rho value of mixed micelles as compared to single micelles also confirms a more compact structure of the former. In addition, due to the much higher N(agg) and lower R(h), PLA/PEG2000 micelles present higher rho value than PLA/PEG5000 ones.
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Analysis and evaluation of the source of heavy metals in water of the River Changjiang.
Environ Monit Assess
PUBLISHED: 02-11-2010
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In order to analyze and evaluate different trace metals on surface water of the Changjiang River, concentrations of dissolved trace metals (Cu, Ni, Fe, Co, Sc, Al, Zn, Pb, Cd, Se, As, Cr, and Hg), major elements (Ca and Mg), and nutrient (NO3- were measured. Samples were taken at 76 positions along Changjiang River in flood and dry seasons during 2007-2008. Spatial distributions identified two main large zones mainly influenced by mineral erosion (sites 1-22) and anthropogenic action (sites 23-76), respectively. Principal component analysis (PCA) and hierarchical cluster analysis were used to identify the variance distinguishing the origin of water. Four significant components were extracted by PCA, explaining 74.91% of total variable. Cu, Ni, Fe, Co, Sc, Al, Ca, and Mg were mainly associated with the weathering and erosion of various rocks and minerals, while an anthropogenic source was identified for Cd and As. Although erosion was one source of Pb and Zn, they were also input by atmospheric deposition and industrial pollutions. NO3- and Se were mainly associated with agriculture activities. However, Hg and Cr showed different sources. CA confirmed and completed the results obtained by PCA, classifying the data into two large groups representing different areas. Group 1 referred to the upper reaches which represented samples mainly corresponding to natural background areas. Group 2 referred to the middle and lower reaches including samples under anthropogenic influence. Meanwhile, group 2 was subdivided into three new groups, representing agricultural, industrial, and various artificial pollution sources, respectively.
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