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author:("Li, liangshan")
1.  Relaxation of Rat Aorta by Farrerol Correlates with Potency to Reduce Intracellular Calcium of VSMCs 
Farrerol, isolated from Rhododendron dauricum L., has been proven to be an important multifunctional physiologically active component, but its vasoactive mechanism is not clear. The present study was performed to observe the vasoactive effects of farrerol on rat aorta and to investigate the possible underlying mechanisms. Isolated aortic rings of rat were mounted in an organ bath system and the myogenic effects stimulated by farrerol were studied. Intracellular Ca2+ ([Ca2+]in) was measured by molecular probe fluo-4-AM and the activities of L-type voltage-gated Ca2+ channels (LVGC) were studied with whole-cell patch clamp in cultured vascular smooth muscle cells (VSMCs). The results showed that farrerol significantly induced dose-dependent relaxation on aortic rings, while this vasorelaxation was not affected by NG-nitro-l-arginine methylester ester or endothelium denudation. In endothelium-denuded aortas, farrerol also reduced Ca2+-induced contraction on the basis of the stable contraction induced by KCl or phenylephrine (PE) in Ca2+-free solution. Moreover, after incubation with verapamil, farrerol can induce relaxation in endothelium-denuded aortas precontracted by PE, and this effect can be enhanced by ruthenium red, but not by heparin. With laser scanning confocal microscopy method, the farrerol-induced decline of [Ca2+]in in cultured VSMCs was observed. Furthermore, we found that farrerol could suppress Ca2+ influx via LVGC by patch clamp technology. These findings suggested that farrerol can regulate the vascular tension and could be developed as a practicable vasorelaxation drug.
doi:10.3390/ijms15046641
PMCID: PMC4013652  PMID: 24747597
farrerol; rat aorta; VSMCs; vasorelaxation
2.  The Adenylyl Cyclase Inhibitor MDL-12,330A Potentiates Insulin Secretion via Blockade of Voltage-Dependent K+ Channels in Pancreatic Beta Cells 
PLoS ONE  2013;8(10):e77934.
Objective
Adenylyl cyclases (ACs) play important role in regulating pancreatic beta cell growth, survival and secretion through the synthesis of cyclic AMP (cAMP). MDL-12,330A and SQ 22536 are two AC inhibitors used widely to establish the role of ACs. The goal of this study was to examine the effects of MDL-12,330A and SQ 22536 on insulin secretion and underlying mechanisms.
Methods
Patch-clamp recording, Ca2+ fluorescence imaging and radioimmunoassay were used to measure outward K+ currents, action potentials (APs), intracellular Ca2+ ([Ca2+]i) and insulin secretion from rat pancreatic beta cells.
Results
MDL-12,330A (10 µmol/l) potentiated insulin secretion to 1.7 times of control in the presence of 8.3 mmol/l glucose, while SQ 22536 did not show significant effect on insulin secretion. MDL-12,330A prolonged AP durations (APDs) by inhibiting voltage-dependent K+ (KV) channels, leading to an increase in [Ca2+]i levels. It appeared that these effects induced by MDL-12,330A did not result from AC inhibition, since SQ 22536 did not show such effects. Furthermore, inhibition of the downstream effectors of AC/cAMP signaling by PKA inhibitor H89 and Epac inhibitor ESI-09, did not affect KV channels and insulin secretion.
Conclusion
The putative AC inhibitor MDL-12,330A enhances [Ca2+]i and insulin secretion via inhibition of KV channels rather than AC antagonism in beta cells, suggesting that the non-specific effects is needed to be considered for the right interpretation of the experimental results using this agent in the analyses of the role of AC in cell function.
doi:10.1371/journal.pone.0077934
PMCID: PMC3812155  PMID: 24205033
3.  Pharmacokinetic Study of Di-Phenyl-Di-(2,4-Difluobenzohydroxamato)Tin(IV): Novel Metal-Based Complex with Promising Antitumor Potential 
Di-phenyl-di-(2,4-difluobenzohydroxamato)tin(IV)(DPDFT), a new metal-based arylhydroxamate antitumor complex, showed high in vivo and in vitro antitumor activity with relative low toxicity, but no data was reported regarding its pharmacokinetics and dependent toxicity. In this paper, a rapid, sensitive, and reproducible HPLC method in vivo using Diamonsil ODS column with a mixture of methanol and phosphoric acid in water (30 : 70, V/V, pH 3.0) as mobile phase was developed and validated for the determination of DPDFT. The plasma was deproteinized with methanol that contained acetanilide as the internal standard (I.S.). The photodiode array detector was set at a wavelength of 228 nm at room temperature and a linear curve over the concentration range 0.1~25 μg·mL−1 (r = 0.9993) was obtained. The method was used to determine the concentration-time profiles for DPDFT in the plasma after single intravenous administration with doses of 5, 10, 15 mg·kg−1 to rats. The pharmacokinetics parameter calculations and modeling were carried out using the 3p97 software. The results showed that the concentration-time curves of DPDFT in rat plasma could be fitted to two-compartment model.
doi:10.1155/2012/210682
PMCID: PMC3287010  PMID: 22400014
4.  Potentiation of Th17 cytokines in aging process contributes to the development of colitis 
Cellular immunology  2010;266(2):208-217.
Summary
Th17 cells, which produce IL-17 and IL-22, promote autoimmunity in mice and have been implicated in the pathogenesis of autoimmune/inflammatory diseases in humans. However, the Th17 immune response in the aging process is still not clear. In the present study, we found that the induction of IL-17-produing CD4+ T cells was significantly increased in aged individuals compared with young healthy ones. The mRNA expression of IL-17, IL-17F, IL-22, and RORC2 was also significantly increased in aged people. Similar to humans, Th17 cells as well as mRNAs encoding IL-17, IL-22 and RORγt were dramatically elevated in naïve T cells from aged mouse compared to young ones. In addition, CD44 positive IL-17-producing CD4+ T cells were significantly higher in aged mice, suggesting that memory T cells are an important source of IL-17 production. Furthermore, the percentage of IL-17-produing CD4+ T cells generated in co-culture with dendritic cells from either aged or young mice did not show significant differences, suggesting that dendritic cells do not play a primary role in the elevation of Th17 cytokines in aged mouse cells. Importantly, transfer of CD4+CD45Rbhi cells from aged mice induced more severe colitis in RAG−/− mice compared to cells from young mice, Taken together, these results suggest that Th17 immune responses are elevated in aging humans and mice and may contribute to the increased development of inflammatory disorders in the elderly.
doi:10.1016/j.cellimm.2010.10.007
PMCID: PMC3006034  PMID: 21074754
Th17 cell; Aging; IL-17; IL-22; T cell; Dendritic cell
5.  Pharmacokinetics and Tissue Distribution Study of Praeruptorin D from Radix Peucedani in Rats by High-Performance Liquid Chromatography (HPLC) 
Praeruptorin D (PD), a major pyranocoumarin isolated from Radix Peucedani, exhibited antitumor and anti-inflammatory activities. The aim of this study was to investigate the pharmacokinetics and tissue distribution of PD in rats following intravenous (i.v.) administration. The levels of PD in plasma and tissues were measured by a simple and sensitive reversed-phase high-performance liquid chromatography (HPLC) method. The biosamples were treated by liquid-liquid extraction (LLE) with methyl tert-butyl ether (MTBE) and osthole was used as the internal standard (IS). The chromatographic separation was accomplished on a reversed-phase C18 column using methanol-water (75:25, v/v) as mobile phase at a flow rate of 0.8 mL/min and ultraviolet detection wave length was set at 323 nm. The results demonstrate that this method has excellent specificity, linearity, precision, accuracy and recovery. The pharmacokinetic study found that PD fitted well into a two-compartment model with a fast distribution phase and a relative slow elimination phase. Tissue distribution showed that the highest concentration was observed in the lung, followed by heart, liver and kidney. Furthermore, PD can also be detected in the brain, which indicated that PD could cross the blood-brain barrier after i.v. administration.
doi:10.3390/ijms13079129
PMCID: PMC3430287  PMID: 22942756
praeruptorin D; Radix Peucedani; HPLC; pharmacokinetics; tissue distribution
6.  Comparison of the Phenolic Content and Antioxidant Activities of Apocynum venetum L. (Luo-Bu-Ma) and Two of Its Alternative Species 
The leaves of Apocynum venetum L. (AV), a native Chinese plant, have been used as folk medicine in China and Japan. This study evaluated the content of the active antioxidant component and antioxidant activities of AV, and its two alternative species, Poacynum pictum (Schrenk) Baill. (PP) and Poacynum hendersonii (Hook.f.) Woodson (PH). The total phenolic and total flavonoid contents were determined. In addition, the quantitative analysis of two major flavonoid compounds (hyperoside and isoquercitrin) was carried out by HPLC. The antioxidant activities were investigated by the 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging activity method, the reducing power test and the chelating ability of ferrous ions. The highest total phenolic and flavonoid contents were observed in the AV methanolic extract, followed by the PP and PH methanolic extracts. HPLC analysis indicated that isoquercitrin was one of the major components in all three species, however, hyperoside was only detected in AV at high levels. All the antioxidant assays we performed demonstrated that the AV extract was markedly superior to those of the other two species.
doi:10.3390/ijms11114452
PMCID: PMC3000093  PMID: 21151449
Apocynum venetum L.; Poacynum pictum (Schrenk) Baill.; Poacynum hendersonii (Hook.f.) Woodson; antioxidant activity; phenolic; flavonoid; HPLC
7.  Characterization and Functional Studies of a FYVE Domain-Containing Phosphatase in C. elegans 
Journal of cellular biochemistry  2008;104(5):1843-1852.
The myotubularin (MTM) enzymes are phosphotidylinositol-3-phosphate and phosphatidylinositol 3,5-bisphosphate phosphatases. Mutation of MTM1, the founder member of this family, is responsible for X-linked myotubular myopathy in humans. Here, we have isolated and characterized a Caenorhabditis elegans homology of the enzymes designated ceMTM3. ceMTM3 preferably dephosphorylates phosphotidylinositol-3-phosphate and contains a FYVE lipid-binding domain at its C-terminus which binds phosphotidylinositol-3-phosphate. Immunoblotting analyses revealed that the enzyme is expressed during the early development and adulthood of the animal. Immunofluorescent staining revealed predominant expression of the enzyme in eggs and muscles. Knockdown of the enzyme by using feeding-based RNA interference resulted in an increased level of phosphotidylinositol-3-phosphate and caused severe impairment of body movement of the worms at their post-reproductive ages and significantly shortened their lifespan. This study thus reveals an important role of the MTM phosphatases in maintaining muscle function, which may have clinical implications in prevention and treatment of sarcopenia.
doi:10.1002/jcb.21752
PMCID: PMC2562989  PMID: 18393358
Tyrosine phosphatase; myotubularin; phosphoinositide; aging; locomotion
8.  Identification of an Acquired JAK2 Mutation in Polycythemia Vera 
The Journal of biological chemistry  2005;280(24):22788-22792.
Polycythemia vera (PV) is a human clonal hematological disorder. The molecular etiology of the disease has not been identified. PV hematopoietic progenitor cells exhibit hypersensitivity to growth factors and cytokines, suggesting possible abnormalities in protein tyrosine kinases and phosphatases. By sequencing the entire coding regions of cDNAs of candidate enzymes, we identified a G:C-to-T:A point mutation of the JAK2 tyrosine kinase in 20 out of 24 PV blood samples but none in 12 normal samples. The mutation has varying degrees of heterozygosity and is apparently acquired. It changes conserved Val617 to Phe in the pseudokinase domain of JAK2 that is known to have an inhibitory role. The mutant JAK2 has enhanced kinase activity, and when overexpressed together with the erythropoietin receptor in cells, it caused hyperactivation of erythropoietin-induced cell signaling. This gain-of-function mutation of JAK may explain the hyper sensitivity of PV progenitor cells to growth factors and cytokines. Our study thus defines a molecular defect of PV.
doi:10.1074/jbc.C500138200
PMCID: PMC1201515  PMID: 15863514
PV, polycythemia vera; PTK, protein tyrosine kinase; PTP, protein tyrosine phosphatase; EPO, erythropoietin; EPOR, erythropoietin receptor
9.  Erlotinib Effectively Inhibits JAK2V617F Activity and Polycythemia Vera Cell Growth 
The Journal of biological chemistry  2006;282(6):3428-3432.
JAK2V617F, a mutant of tyrosine kinase JAK2, is found in most patients with polycythemia vera (PV) and a substantial proportion of patients with idiopathic myelofibrosis or essential thrombocythemia. The JAK2 mutant displays a much increased kinase activity and generates a PV-like phenotype in mouse bone marrow transplant models. This study shows that the anti-cancer drug erlotinib (Traceva™) is a potent inhibitor of JAK2V617F activity. In vitro colony culture assays revealed that erlotinib at micro-molar concentrations effectively suppresses the growth and expansion of PV hematopoietic progenitor cells while having little effect on normal cells. Furthermore, JAK2V617F-positive cells from PV patients show greater susceptibility to the inhibitor than their negative counterparts. Similar inhibitory effects were found with the JAK2V617F-positive human erythroleukemia HEL cell line. These data suggest that erlotinib may be used for treatment of JAK2V617F-positive PV and other myeloproliferative disorders.
doi:10.1074/jbc.C600277200
PMCID: PMC2096634  PMID: 17178722
10.  Transcription factor IRF8 directs a silencing programme for TH17 cell differentiation 
Nature Communications  2011;2:314-.
TH17 cells are recognized as a unique subset of T helper cells that have critical roles in the pathogenesis of autoimmunity and tissue inflammation. Although RORγt is necessary for the generation of TH17 cells, the molecular mechanisms underlying the functional diversity of TH17 cells are not fully understood. Here we show that a member of interferon regulatory factor (IRF) family of transcription factors, IRF8, has a critical role in silencing TH17-cell differentiation. Mice with a conventional knockout, as well as a T cell-specific deletion, of the Irf8 gene exhibited more efficient TH17 cells. Indeed, studies of an experimental model of colitis showed that IRF8 deficiency resulted in more severe inflammation with an enhanced TH17 phenotype. IRF8 was induced steadily and inhibited TH17-cell differentiation during TH17 lineage commitment at least in part through its physical interaction with RORγt. These findings define IRF8 as a novel intrinsic transcriptional inhibitor of TH17-cell differentiation.
The molecular mechanisms that regulate TH17 cell diversity are poorly understood. Ouyang et al. show that the transcription factor interferon regulatory factor-8 is required for TH17-cell differentiation and that its absence increases the severity of an experimental model of colitis.
doi:10.1038/ncomms1311
PMCID: PMC3112536  PMID: 21587231

Results 1-10 (10)