5h后再给予100nmol/L FⅦa刺激12h后细胞ProM-MP-7蛋白水平。(2)观察用100nmol/L FⅦa刺激时丝裂原激活蛋白激酶(MAPKs)各亚通路信号蛋白(包括细胞外信号调节激酶ERK1/2,丝裂原激活蛋白激酶P38及c-Jun-N末端蛋白激酶JNK)磷酸化水平变化。(3)在FⅦa刺激前0.5h分别加入ERK1/2,P38和JNK信号通路特异阻断剂PD98059,SB203580和SP600125,培养12h后检测细胞ProMMP-7蛋白的变化。结果:(1)FⅦa可刺激LoVo系细胞表达ProMMP-7并呈时间依赖性和剂量依赖性,在刺激的12h左右ProMMP-7达峰值,是正常对照组的5.5±0.6倍(P=0.006);用TF抗体预处理的LoVo细胞系,其FⅦa上调ProMMP-7表达的作用被完全阻断。(2)用100nmol/L

FⅦa刺激LoVo细胞5min时,MAPKs磷酸化活性蛋白p-ERK1/2和p-P38表达水平开始增加,至10min时达峰值(分别为对照组的2.2±0.3倍和3.9±0.5倍,P值分别为0.02和0.01),存在时间效应关系,而p-JNK活性无改变(为对照组的1.1±0.1倍)。(3)ERK1/2通路阻断剂PD98059及P38通路阻断剂SB203580可部分减少FⅦa上调LoVo细胞系表达ProMMP-7的效应,分别降低了32%±5%(P=0.01)和61%±10%(P=0.009),JNK通路特异性阻断剂SP600125对ProMMP-7的表达无明显影响。结论:FⅦa通过与LoVo细胞表面的TF结合诱导ProMMP-7的表达,并呈时间依赖性和剂量依赖性;ERK1/2和P38MAPKs亚通路参与LoVo细胞TF信号转导通路并与TF/FⅦa调控ProMMP-7表达有关。
目的研究组蛋白去乙酰化酶(HDAC)抑制剂MS-275诱导人肝癌细胞株HepG2细胞周期阻滞作用,并对其信号传导机制进行初步探讨。方法体外培养人肝癌细胞株HepG2;应用MTT比色法观察MS-275对HepG2的生长抑制作用,应用流式细胞仪检测MS-275对细胞周期的影响;Western印迹分析细胞周期蛋白(Cyclin)D1、p21、p-p38MAPK三种蛋白表达的差异。结果HDAC抑制剂MS-275能抑制肝癌细胞生长,具有时间和剂量的依赖性。可以引起细胞周期G0-G1期阻滞。MS-275可以使HepG2细胞p21蛋白表达提高,CyclinD1蛋白表达降低,在SB203580组两蛋白的表达又恢复到对照组水平。p-p38MAPK蛋白在两组表达均提高。结论MS-275能诱导肝癌细胞株的细胞周期阻滞,这种作用可能是通过p38MAPK信号传导途径介导的,与下调CyclinD1的表达、上调p21的表达有关。
目的:探讨p38丝裂素活化蛋白激酶(MAPK)信号传导对低氧预处理(HPC)的细胞保护作用以及HPC后血红素氧化酶-1(HO-1)mRNA表达的影响。方法:选用新生Wistar大鼠心脏进行成纤维细胞培养,给予HPC及低氧/复氧处理(HR)。药物处理组在HPC和HR处理过程中向培养液中加入SB203580。测定细胞培养上清液中乳酸脱氢酶(LDH)浓度和细胞HO-1

Selleck AUY 922 Saracatinib mRNA表达量。结果:低氧/复氧后,HPC/SB组LDH浓度显著高于HPC/NSB组(P<0.01,P
Aim:To

investigate the mechanism of silibinin-protected isoproterenol-inducedapoptosis 无 in rat cardiac myocytes.Methods:The viability of rat cardiac myocyteswas measured by MTT method.The apoptotic ratio was measured by terminaldeoxynucleotidyl transferase-mediated dUTP nick end-labeling.Protein kinase C(PKC) activity assay was carried out according to the instructions of the PepTagnon-radioactive protein kinase C assay kit.Western blot analysis was used toevaluate the level of Ras,Raf-1 and mitogen-activated protein kinase (MAPK)expression.Results:The protective effects of silibinin were significantly sup-pressed by inhibitors,including genistein,manumycin A and GW5074 [inhibitorsfor protein tyrosine kinases (PTK),Ras and Raf-1,respectively].The exposure ofrat cardiac myocytes to isoproterenol alone caused decreased PKC activity,whichwas prevented by pretreatment with silibinin dose-dependently.Simultaneously,the increased expression of Ras and Raf-1 activated by silibinin were blocked bythe PKC inhibitor,stauroporine.