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Acta Pharmacologica Sinica 2004年第5期25卷 75-80页

作者： Xue-bo YAN, Fan-jie MENG, Bo SONG, Guang-yi ZHANG2research center for biochemistry and molecular biology, xuzhou medical college, xuzhou 221002, china research center for biochemistry and molecular biology Xuzhou Medical College

AIM: To investigate whether brain ischemia induces serine phosphorylation of neuronal nitric oxide synthase(nNOS) by Ca2+/calmodulin-dependent protein kinase II (CaMKII) and the interaction between CaMKIIa and nNOSin rat hippocampus. METHODS: Brain ischemia was induced by bilateral carotid artery occlusion *** and the interaction of proteins were studied by immunoprecipitation and immunoblotting. Weinvestigated during brain ischemia serine phosphorylation and amount of nNOS in crude membranes fraction (P)and cytosolic fraction (S), interaction between CaMKIIa and nNOS, and the effects of 1-[N,O-bis-(5-isoquinolinesulfonyl)-N-methyl-L-tyrosyl]-4-phenylpiperazine (KN-62, a selective inhibitor of CaMKII) on phos-phorylation and the interaction of proteins in P. RESULTS: Serine phosphorylation of nNOS in P increased persis-tently during brain ischemia, and 15 min ischemia-induced serine phosphorylation of nNOS was attenuated signifi-cantly by KN-62. But there was no serine phosphorylation of nNOS in S. The distributions of nNOS were notaffected by ischemia and KN-62. However, the binding levels of both CaMKIIa with nNOS and Thr286autophosphorylated CaMKIIa with nNOS increased after ischemia, and were diminished by KN-62. CONCLUSION:CaMKII interacted with nNOS and regulated serine phosphorylation of nNOS during brain ischemia.

关键词： serine phosphorylation nitric oxide synthase calmodulin kinase KN 62 brain ischemia hippo-campus

Activation and involvement of JNK1/2 in hydrogen peroxideinduced neurotoxicity in cultured rat cortical neurons

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Acta Pharmacologica Sinica 2004年第5期25卷 88-94页

作者： Wei WANG, Can GAO, Xiao-yu HOU, Yong LIU, Yan-yan ZONG, Guang-yi ZHANG2research center for biochemistry and molecular biology, xuzhou medical college, xuzhou 221002, china research center for biochemistry and molecular biology Xuzhou Medical College

AIM: To investigate the role of c-Jun N-terminal protein kinase 1 and 2 (JNK1/2) and the main signal pathway forits activation in hydrogen peroxide (H2O2) induced apoptotic-like cortical cell death. METHODS: Using the modelof oxidative stress induced by H2O2, the expression and diphosphorylation of JNK1/2 was examined by immunoblottinganalysis, and neuronal apoptotic like cell death was determined by 4’,6-diamidino-2-phenylindole (DAPI) ***: The elevation in diphosphorylation level of JNK1/2 (4.40-/5.61-fold vs sham control) was associatedwith the concentration of H2O2 (0-100 mol/L) and the development of apoptotic-like cell death (11.04 %-81.01 %).There was no alteration of JNK1/2 protein expression following H2O2 treatment and recovery at different timepoints. Administration with JNK1/2 antisense oligonucleotides not only significantly decreased JNK1/2 proteinexpression and activation level, but also significantly reduced cortical cell death induced by H2O2 ***, both JNK1/2 diphosphorylation and apoptotic-like cell death were largely prevented by pretreatmentwith (5S,10R)-()-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-imine hydrogen maleate (MK-801)or omission of Ca2+ in incubation medium with ethylene glycol-bis(2-aminoethylether)-N,N,N’,N’-tetraacetic acid(EGTA). CONCLUSION: JNK1/2 is activated and participates in H2O2-induced apoptotic-like death in cultured ratcortical neurons mainly via N-methyl-D-aspartate (NMDA) receptor-mediated influx of extracellular Ca2+.

关键词： c-Jun N-terminal protein kinase calcium N-methylaspartate receptors hydrogen peroxide signaltransduction apoptosis cerebral cortex

Competitive binding of postsynaptic density 95 and Ca2+-calmodulin dependent protein kinase II to N-methyl-D-aspartate receptor subunit 2B in rat brain

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Acta Pharmacologica Sinica 2004年第2期25卷 50-54页

作者： Fan-jie MENG , Jun GUO, Bo SONG, Xue-bo YAN, Guang-yi ZHANG2 research center of biochemistry and molecular biology, xuzhou medical college, xuzhou 221002, china research center of biochemistry and molecular biology Xuzhou Medical College Xuzhou 221002 China

AIM: To investigate the interactions among postsynaptic density 95 (PSD-95), Ca2+-calmodulin dependent protein kinase IIα (CaMKIIα), and N-methyl-D-aspartate receptor subunit 2B (NR2B) during ischemia and reperfusion in hippocampus of rats. METHODS: Brain ischemia was induced by four-vessel occlusion procedure in rats. Immu- noprecipitation and immunoblotting were performed to study the interactions and phosphorylation of proteins. The association-dissociation of PSD-95 and CaMKIIα to and from N-methyl-D-aspartate (NMDA) receptor induced by ischemia and reperfusion and the effects of 1-[N,O-bis-(5-isoquinolinesulfonyl)-N-methyl-L-tyrosyl]-4-phenyl-pip- erazine (KN-62, a selective inhibitor of CaMKII) on these protein interactions were investigated. Coimmuno- precipitation and immunoblotting were performed for the studies of interactions among proteins. RESULTS: The alternations of the binding level of PSD-95 and CaMKIIα to NR2B during ischemia and reperfusion demonstrated the negative correlation to each other. Pre-administration of KN62 through both cerebral ventricles inhibited the 10 min ischemia-induced increase of the binding of PSD-95 to NR2B and, on the contrary, promoted the binding of CaMKIIα to NR2B. CONCLUSION: PSD-95 competes with CaMKII to bind to NR2B during ischemia and reperfusion in rat hippocampus.

关键词： N-methyl-D-aspartate receptors Ca2+-calmodulin dependent protein kinase brain ischemia

Neuroprotection of selenite against ischemic brain injury through negatively regulating early activation of ASK1/JNK cascade via activation of PI3K/AKT pathway

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Acta Pharmacologica Sinica 2007年第1期28卷 19-27页

作者： Qing WANG Quan-guang ZHANG Dong-na WU Xiao-hui YIN Guang-yi ZHANG~2 research center for biochemistry and molecular biology,xuzhou medical college,xuzhou 221002,china researchcenterforbiochemistryandmolecularbiology XuzhouMedicalCollegeXuzhou221002China

Aim:To investigate whether selenite,a known antioxidant,could decrease theactivation of apoptosis signal regulating kinase 1/c-jun N-terminal kinase(ASK1/JNK)signaling cascade in cerebral ischemia/reperfusion(I/R)by activating thephosphatidylinositol 3-kinase(PI3K)/AKT pathway in rat hippocampi,and theneuroprotective effect of selenite against ischemic injury after 15 min of transientbrain ***:Transient global brain ischemia was induced by4-vessel occlusion into adult male Sprague-Dawley rats weighing 250-300 *** were pretreated only with selenite(0.3 mg/kg dissolved in 0.9% saline)every24 h for 7 d by means of intravenous injection of the tail or combined with LY294002from d 5 by left cerebral ventricle injection before ***:Selenitesignificantly increased AKT 1 activation and decreased the activation of ASK1/JNK cascade via phosphorylating ASK1 at Ser-83 residue by AKT1 during earlyreperfusion after 15 min transient global brain *** the contrary,com-bined pretreatment of the rats with LY294002(a specific PI3K inhibitor)and selen-ite significantly inhibited the effects solely with ***:The activa-tion of the pro-apoptotic ASK1/JNK cascade,which is closely associated withoxidative stress,could be suppressed by selenite through activating the anti-apoptotic PI3K/AKT pathway during early reperfusion after cerebral ischemia inrat hippocampi.

关键词： cerebral ischemia PI3K/AKT ASK1/JNK LY294002 selenite

Inhibition of Proliferation and Induction of Apoptosis in Human Renal Carcinoma Cells by Anti-telomerase Small Interfering RNAs

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Acta Biochimica et Biophysica Sinica 2006年第7期38卷 500-506页

作者： Jun-Nian ZHENG~(1,3#*)Ya-Feng SUN~(2#)Dong-Sheng PEI~2 Jun-Jie LIU~1 Jia-Cun CHEN~1 Wang LI~1 Xiao-Qing SUN~1 Qi-Duo SHI~3 Rui-Fa HAN~3 Teng-Xiang MA~3 1 Laboratory of Urology,Affiliated Hospital of xuzhou medical college,xuzhou 221002,china 2 research center for biochemistry and molecular biology,xuzhou medical college,xuzhou 221002,china 3 Institute of Urology,Tianjin medical University,Tianjin 300070,china LaboratoryofUrology AffiliatedHospitalofXuzhouMedicalCollegeXuzhou221002China researchcenterforBiochemist'ryandmolecularbiology XuzhouMedicalCollegeXuzhou221002China

Telomerase is an attractive molecular target for cancer therapy because it is present in mostmalignant cells but is undetectable in most normal somatic *** telomerase consists of two subunits,an RNA component(hTR)and a human telomerase reverse transcriptase component(hTERT).Smallinterfering RNA(siRNA),one kind of RNA interferences,has been demonstrated to be an effective methodto inhibit target gene expression in human *** investigated the effects of siRNA targeting at both hTRand hTERT mRNA on the inhibition of telomerase activity in human renal carcinoma cells(HRCCs).Theproliferation and apoptosis of HRCCs were *** treatment of HRCCs using hTR and hTERTsiRNAs resulted in significant decrease of hTR mRNA,hTERT mRNA and hTERT *** siRNA canalso inhibit the telomerase activity and the proliferation of ***,they can induce apoptotic celldeath in a dose-dependent *** these findings,we propose that the inhibition of telomerase activityusing siRNA targeting hTR and hTERT might be a rational approach in renal cancer therapy.

关键词： hTERT hTR telomerase siRNA renal cell carcinoma proliferation apoptosis

Treatment with Vector-expressed Small Hairpin RNAs against Ki67 RNA-induced Cell Growth Inhibition and Apoptosis in Human Renal Carcinoma Cells

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Acta Biochimica et Biophysica Sinica 2006年第4期38卷 254-261页

作者： Jun-Nian ZHENG~(1,3*) Ya-Feng SUN~(2#) Dong-Sheng PEI~2 Jun-Jie LIU~1 Teng-Xiang MA~3 Rui-Fa HAN~3 Wang LI~1 Dian-Bao ZHENG~1 Jia-Cun CHEN~1 Xiao-Qing SUN~1 1 Laboratory of Urology,Affiliated Hospital of xuzhou medical college,xuzhou 221002,china 2 research center for biochemistry and molecular biology,xuzhou medical college,xuzhou 221002,china 3 Institute of Urology,medical University of Tianjin,Tianjin 300211,china LaboratoryofUrology AffiliatedHospitalofXuzhouMedicalCollegeXuzhou221002China researchcenterforbiochemistryandmolecularbiology XuzhouMedicalCollegeXuzhou221002China InstituteofUrology MedicalUniversityofTianjin

Short hairpin RNAs (shRNAs) transcribed *** polymerase Ⅲ promoters can triggersequence-selective gene silencing in mammalian *** virtue of their excellent function in knocking downexpression of cancer-associated genes,shRNAs could be used as new therapeutic agents for *** of Ki67 in renal cancer has been correlated to a more aggressive tumor phenotype,inhibitionof Ki67 protein expression by means of shRNAs seems to be a promising approach for the therapy of *** this study,we constructed an expression plasmid encoding shRNAs against the Ki67 gene,namedpSilencerKi67,and transfected it into human renal carcinoma *** pSilencerKi67 was shown to signifi-cantly knock down the expression of the Ki67 gene in human renal carcinoma cells,resulting in inhibitingproliferation and inducing apoptotic cell death that can be maintained for at least *** findings offer thepromise of using vector-based shRNAs against Ki67 in renal cancer gene therapy.

关键词： Ki67 gene shRNAs renal cell carcinoma proliferation apoptosis

Integrin was involved in the signaling events of GDNF

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Integrin was involved in the signaling events of GDNF

Ohmefentanyl stereoisomers induce changes of CREB phosphorylation in hippocampus of mice in conditioned place preference paradigm

中国神经科学学会第四次会员代表大会暨第七届全国学术会议

作者： Jun-Ping CAO~1,Jing-Kao YU~1,Chong LI~2,Yu SUN~1,Hong-Hua YUAN~1,Hong-Jun WANG~1,Dian-Shuai GAO~(1*) 1 research center for Neurobiology,2 research center for biochemistry and molecular biology,xuzhou medical college, xuzhou 221002,china

Glial cell line-derived neurotrophic factor(GDNF) is a potent neurotrophic factor for midbrain dopamine(DA) neurons. GDNF exerts its biologic effects through a multi-component receptor system includin

关键词： glial cell line-derived neurotrophic factor integrin dopamine neurons

来源： cnki会议评论

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Cell research 2003年第1期13卷 29-34页

作者： CAN GAO, Li WEI CHEN, JIE CHEN, XUE JUN XU, ZHI QIANG CHISecond Department of Pharmacology, Shanghai Institute of Materia Medica, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200031, chinaresearch center for biochemistry and molecular biology, xuzhou medical college, xuzhou, Jiangsu 221002, china Second Department of Pharmacology Shanghai Institute of Materia Medica Shanghai Institutes for Biological Sciences Chinese Academy of Sciences Shanghai 200031 China

The present study was designed to determine the changes of phosphorylation of cAMP- response element binding protein (CREB) in hippocampus induced by ohmefentanyl stereoisomers (F9202 and F9204) in conditioned place preference (CPP) paradigm. The results showed that mice receiving F9202 and F9204 displayed obvious CPP. They could all significantly stimulate CREB phosphorylation and maintained for a long time without affecting total CREB protein levels. The effect of F9204 was similar to morphine which effect was more potent and longer than F9202. We also examined the effects of ketamine, a noncompetitive N-mthyl-D-aspartate receptor (NR) antagonist, on morphine-, F9202- and F9204- induced CPP and phosphorylation of CREB in hippocampus. Ketamine could suppress not only the place preference but also the phosphorylation of CREB produced by morphine, F9202 and F9204. These findings suggest that alterations in the phosphorylation of CREB be relevant to opiates signaling and the development of opiates depende

关键词： cAMP-response element binding protein (CREB) ohmefentanyl stereo isomers opiates hippocampus conditioned place preference (CPP).

人白细胞介素18在大肠杆菌中的表达、纯化和复性

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生物化学与生物物理学报 2000年第4期32卷 397-400页

作者：裴冬生胡书群赵惠仁徐州医学院生物化学与分子生物学研究中心!徐州221002

用RT PCR从PHA刺激的人外周血单个核细胞 (PBMC)中扩增出人白细胞介素 18(IL 18)的cDNA ,克隆到表达载体pJW 2中 ,经热诱导后在大肠杆菌中得到高效表达。表达的重组IL 18占菌体总蛋白质的 2 0 % ,表达的蛋白质分子量约为 18kD ,与预期... 详细信息

用RT PCR从PHA刺激的人外周血单个核细胞 (PBMC)中扩增出人白细胞介素 18(IL 18)的cDNA ,克隆到表达载体pJW 2中 ,经热诱导后在大肠杆菌中得到高效表达。表达的重组IL 18占菌体总蛋白质的 2 0 % ,表达的蛋白质分子量约为 18kD ,与预期分子量相符。表达产物主要以包涵体的形式存在。包涵体经超声破碎、洗涤后以 8mol/L尿素溶解 ,经SephadexG 10 0柱纯化后 ,纯度可达 90 %以上。纯化的蛋白质在GSH/GSSH的存在下复性 ,得到具有生物学活性的白细胞介素 18。在 0 .5mg/LConA存在的条件下 ,复性产物对人外周血单个核细胞具明显的IFN γ诱生作用。hIL 18的热诱导表达及复性和纯化方法的建立为大量生产rhIL 18打下了基础。

关键词：白细胞介素18 表达纯化复性

Overexpression of PDZl domain inhibits excitotoxicity induced by KA and ischaemic brain injury

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Overexpression of PDZl domain inhibits excitotoxicity induce...