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991.
Liu XS Jiang M Zhang M Tang D Clemo HF Higgins RS Tseng GN 《American journal of physiology. Heart and circulatory physiology》2007,292(1):H560-H571
The nature of electrical remodeling in a canine model of ischemic cardiomyopathy (ICM; induced by repetitive intracoronary microembolizations) that exhibits spontaneous ventricular tachycardia is not entirely clear. We used the patch-clamp technique to record action potentials and ionic currents of left ventricular myocytes isolated from the region affected by microembolizations. We also used the immunoblot technique to examine channel subunit expression in adjacent affected tissue. Ventricular myocytes and tissue isolated from the corresponding region of normal hearts served as control. ICM myocytes had prolonged action potential duration (APD) and more pronounced APD dispersion. Slow delayed rectifier current (I(Ks)) was reduced at voltages positive to 0 mV, along with a negative shift in its voltage dependence of activation. Immunoblots showed that there was no change in KCNQ1.1 (I(Ks) pore-forming or alpha-subunit), but KCNE1 (I(Ks) auxiliary or beta-subunit) was reduced, and KCNQ1.2 (a truncated KCNQ1 splice variant with a dominant-negative effect on I(Ks)) was increased. Transient outward current (I(to)) was reduced, along with an acceleration of the slow phase of recovery from inactivation. Immunoblots showed that there was no change in Kv4.3 (alpha-subunit of fast-recovering I(to) component), but KChIP2 (beta-subunit of fast-recovering component) and Kv1.4 (alpha-subunit of slow-recovering component) were reduced. Inward rectifier current was reduced. L-type Ca current was unaltered. The immunoblot data provide mechanistic insights into the observed changes in current amplitude and gating kinetics of I(Ks) and I(to). We suggest that these changes, along with the decrease in inward rectifier current, contribute to APD prolongation in ICM hearts. 相似文献
992.
Autoactivation of matriptase in vitro: requirement for biomembrane and LDL receptor domain 总被引:1,自引:0,他引:1
Lee MS Tseng IC Wang Y Kiyomiya K Johnson MD Dickson RB Lin CY 《American journal of physiology. Cell physiology》2007,293(1):C95-105
In live cells, autoactivation of matriptase, a membrane-bound serine protease, can be induced by lysophospholipids, androgens, and the polyanionic compound suramin. These structurally distinct chemicals induce different signaling pathways and cellular events that somehow, in a cell type-specific manner, lead to activation of matriptase immediately followed by inhibition of matriptase by hepatocyte growth factor activator inhibitor 1 (HAI-1). In the current study, we established an analogous matriptase autoactivation system in an in vitro cell-free setting and showed that a burst of matriptase activation and HAI-1-mediated inhibition spontaneously occurred in the insoluble fractions of cell homogenates and that this in vitro activation could be attenuated by a soluble suppressive factor(s) in cytosolic fractions. Immunofluorescence staining and subcellular fractionation studies revealed that matriptase activation occurred in the perinuclear regions. Solubilization of matriptase from cell homogenates by Triton X-100 or sonication of cell homogenates completely inhibited the effect, suggesting that matriptase activation requires proper lipid bilayer microenvironments, potentially allowing appropriate interactions of matriptase zymogens with HAI-1 and other components. Matriptase activation occurred in a narrow pH range (from pH 5.2 to 7.2), with a sharp increase in activation at the transition from pH 5.2 to 5.4, and could be completely suppressed by moderately increased ionic strength. Protease inhibitors only modestly affected activation, whereas 30 nM (5 µg/ml) of anti-matriptase LDL receptor domain 3 monoclonal antibodies completely blocked activation. These atypical biochemical features are consistent with a mechanism for autoactivation of matriptase that requires protein-protein interactions but not active proteases. hepatocyte growth factor activator inhibitor 1; protease activation; low-density lipoprotein 相似文献
993.
994.
Kpp95, isolated on Klebsiella pneumoniae, is a bacteriophage with the morphology of T4-type phages and is capable of rapid lysis of host cells. Its double-stranded genomic DNA (ca. 175 kb, estimated by pulsed-field gel electrophoresis) can be cut only by restriction endonucleases with a cleavage site flanked either by A and T or by T, as tested, suggesting that it contains the modified derivative(s) of G and/or C. Over 26 protein bands were visualized upon sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the virion proteins. N-terminal sequencing indicated that the most abundant band (46 kDa) is the major coat protein (gp23) which has been cleaved from a signal peptide likely with a length similar to that of T4. Phylogenetic analyses based on the sequences of the central region (263 amino acid residues) of gp23 and the full length of gp18 and gp19 placed Kpp95 among the pseudo-T-even subgroup, most closely related to the coliphage JS98. In addition to being able to lyse many extended-spectrum beta-lactamase strains of K. pneumoniae, Kpp95 can lyse Klebsiella oxytoca, Enterobacter agglomerans, and Serratia marcescens cells. Thus, Kpp95 deserves further studies for development as a component of a therapeutic cocktail, owing to its high efficiencies of host lysis plus extended host range. 相似文献
995.
This study investigated the mechanism of oleic acid (OA)-induced disassembly of myofibrils in cardiomyocytes. OA treatment disrupted myofibrils, as revealed by the disorganization of several sarcomeric proteins. Since focal adhesions (FAs) are implicated in myofibril assembly, we examined structural changes in FAs after OA treatment. Immunofluorescence studies with antibodies against FA proteins (vinculin, integrin beta1D, and paxillin) showed that FAs and costameres disintegrated or disappeared after OA treatment and that the changes in FA proteins occurred prior to myofibril disassembly. The effects of OA on FAs and myofibrils were reversed after removal of OA. OA decreased expression of integrin beta1D, paxillin, vinculin, and actin, and induced tyrosine dephosphorylation of FA kinase (FAK) and paxillin. These effects were blocked by pretreatment with sodium orthovanadate, a protein tyrosine phosphatase (PTP) inhibitor. This inhibitor also prevented OA-induced myofibril disassembly, indicating the involvement of PTP in myofibril disassembly. Furthermore, OA increased protein levels of PTP-PEST. The upregulation of this phosphatase correlated with the tyrosine dephosphorylation of paxillin and FAK, which are targets for PTP-PEST. In addition, OA decreased RhoA activity and the phosphorylation of cofilin, a downstream target of RhoA. Cofilin dephosphorylation increased its actin-severing activity and led to the depolymerization of F-actin, which might provide another potential mechanism for OA-induced myofibril disassembly. 相似文献
996.
Wei-Chun Huang Ting-Yuan Tseng Ying-Ting Chen Cheng-Chung Chang Zi-Fu Wang Chiung-Lin Wang Tsu-Ning Hsu Pei-Tzu Li Chin-Tin Chen Jing-Jer Lin Pei-Jen Lou Ta-Chau Chang 《Nucleic acids research》2015,43(21):10102-10113
G-quadruplex (G4) is a promising target for anti-cancer treatment. In this paper, we provide the first evidence supporting the presence of G4 in the mitochondrial DNA (mtDNA) of live cells. The molecular engineering of a fluorescent G4 ligand, 3,6-bis(1-methyl-4-vinylpyridinium) carbazole diiodide (BMVC), can change its major cellular localization from the nucleus to the mitochondria in cancer cells, while remaining primarily in the cytoplasm of normal cells. A number of BMVC derivatives with sufficient mitochondrial uptake can induce cancer cell death without damaging normal cells. Fluorescence studies of these anti-cancer agents in live cells and in isolated mitochondria from HeLa cells have demonstrated that their major target is mtDNA. In this study, we use fluorescence lifetime imaging microscopy to verify the existence of mtDNA G4s in live cells. Bioactivity studies indicate that interactions between these anti-cancer agents and mtDNA G4 can suppress mitochondrial gene expression. This work underlines the importance of fluorescence in the monitoring of drug-target interactions in cells and illustrates the emerging development of drugs in which mtDNA G4 is the primary target. 相似文献
997.
Pei-Yu Wang Wei-Cheng Lin Yi-Cheng Tsai Mei-Ling Cheng Yu-Hung Lin Shu-Heng Tseng Archan Chakraborty Li-Mei Pai 《Genetics》2015,201(4):1511-1523
CTP synthase (CTPsyn) plays an essential role in DNA, RNA, and lipid synthesis. Recent studies in bacteria, yeast, and Drosophila all reveal a polymeric CTPsyn structure, which dynamically regulates its enzymatic activity. However, the molecular mechanism underlying the formation of CTPsyn polymers is not completely understood. In this study, we found that reversible ubiquitination regulates the dynamic assembly of the filamentous structures of Drosophila CTPsyn. We further determined that the proto-oncogene Cbl, an E3 ubiquitin ligase, controls CTPsyn filament formation in endocycles. While the E3 ligase activity of Cbl is required for CTPsyn filament formation, Cbl does not affect the protein levels of CTPsyn. It remains unclear whether the regulation of CTPsyn filaments by Cbl is through direct ubiquitination of CTPsyn. In the absence of Cbl or with knockdown of CTPsyn, the progression of the endocycle-associated S phase was impaired. Furthermore, overexpression of wild-type, but not enzymatically inactive CTPsyn, rescued the endocycle defect in Cbl mutant cells. Together, these results suggest that Cbl influences the nucleotide pool balance and controls CTPsyn filament formation in endocycles. This study links Cbl-mediated ubiquitination to the polymerization of a metabolic enzyme and reveals a role for Cbl in endocycles during Drosophila development. 相似文献
998.
Jie Liu Lijuan Wang Feifei Zhao Serena Tseng Cyndhavi Narayanan Lei Shura Stephen Willingham Maureen Howard Susan Prohaska Jens Volkmer Mark Chao Irving L. Weissman Ravindra Majeti 《PloS one》2015,10(9)
CD47 is a widely expressed cell surface protein that functions as a regulator of phagocytosis mediated by cells of the innate immune system, such as macrophages and dendritic cells. CD47 serves as the ligand for a receptor on these innate immune cells, SIRP-alpha, which in turn delivers an inhibitory signal for phagocytosis. We previously found increased expression of CD47 on primary human acute myeloid leukemia (AML) stem cells, and demonstrated that blocking monoclonal antibodies directed against CD47 enabled the phagocytosis and elimination of AML, non-Hodgkin’s lymphoma (NHL), and many solid tumors in xenograft models. Here, we report the development of a humanized anti-CD47 antibody with potent efficacy and favorable toxicokinetic properties as a candidate therapeutic. A novel monoclonal anti-human CD47 antibody, 5F9, was generated, and antibody humanization was carried out by grafting its complementarity determining regions (CDRs) onto a human IgG4 format. The resulting humanized 5F9 antibody (Hu5F9-G4) bound monomeric human CD47 with an 8 nM affinity. Hu5F9-G4 induced potent macrophage-mediated phagocytosis of primary human AML cells in vitro and completely eradicated human AML in vivo, leading to long-term disease-free survival of patient-derived xenografts. Moreover, Hu5F9-G4 synergized with rituximab to eliminate NHL engraftment and cure xenografted mice. Finally, toxicokinetic studies in non-human primates showed that Hu5F9-G4 could be safely administered intravenously at doses able to achieve potentially therapeutic serum levels. Thus, Hu5F9-G4 is actively being developed for and has been entered into clinical trials in patients with AML and solid tumors (ClinicalTrials.gov identifier: NCT02216409). 相似文献
999.
Shih-Feng Liu Ho-Chang Kuo Ching-Wan Tseng Hung-Tu Huang Yung-Che Chen Chia-Cheng Tseng Meng-Chih Lin 《PloS one》2015,10(9)
Background
Oxidative stress is known to be involved in the pathogenesis of chronic obstructive pulmonary disease (COPD). Evidence suggests that leukocytes mitochondria DNA (mtDNA) is susceptible to undergo mutations, insertions, or depletion in response to reactive oxidative stress (ROS). We hypothesize that mtDNA copy number is associated with the development of COPD.Methodology/Principal Findings
Relative mtDNA copy number was measured by a quantitative real-time PCR assay using DNA extracted from peripheral leukocytes. MtDNA copy number of peripheral leukocytes in the COPD group (n = 86) is significantly decreased compared with non-smoker group (n = 77) (250.3± 21.5 VS. 464.2± 49.9, P<0.001). MtDNA copy number in the COPD group was less than that in the healthy smoking group, but P value nearly achieved significance (250.3± 21.5 VS. 404.0± 76.7, P = 0.08) MtDNA copy number has no significance with age, gender, body mass index, current smoking, and pack-years in COPD group, healthy smoker group and no smoker group, respectively. Serum glutathione level in the COPD group is significantly decreased compared with healthy smoker and non-smoker groups (4.5± 1.3 VS. 6.2± 1.9 and 4.5± 1.3 VS. 7.1±1.1 mU/mL; P<0.001 respectively). Pearson correlation test shows a significant liner correlation between mtDNA copy number and serum glutathione level (R = 0.2, P = 0.009).Conclusions/Significance
COPD is associated with decreased leukocyte mtDNA copy number and serum glutathione. COPD is a regulatory disorder of leukocytes mitochondria. However, further studies are needed to determine the real mechanisms about the gene and the function of mitochondria. 相似文献1000.
Yu-Mu Chen Chien-Hao Lai Huang-Chih Chang Tung-Ying Chao Chia-Cheng Tseng Wen-Feng Fang Chin-Chou Wang Yu-Hsiu Chung Yi-Hsi Wang Mao-Chang Su Kuo-Tung Huang Hung-Chen Chen Ya-Chun Chang Meng-Chih Lin 《PloS one》2015,10(8)