摘要
目的 探讨细胞外信号调节激酶 (ERK)在精 甘 天冬 丝氨酸 (RGDS)四肽诱导肝星状细胞 (HSC)凋亡中的作用。方法 将培养的HSC细胞分为 6组 :①空白对照组 ,②纤维连接蛋白 (FN)组 ,③FN +RGDS(2 5mg/L)组 ,④FN +RGDs(5 0mg/L)组 ,⑤FN +RGDS(10 0mg/L)组 ,⑥FN +精 甘 谷 丝氨酸 (RGES ,10 0mgL)组。采用3 H 胸腺嘧啶核苷 (3 H TdR)掺入法测定HSC增生 ;透射电镜和末端脱氧核苷酸转移酶介导的脱氧三磷酸尿苷缺口末端标记 (TUNEL)方法测定HSC凋亡 ;甲苯胺兰染色法测定细胞粘附率 ;分别应用Western印迹和逆转录 聚合酶链反应 (RT PCR)技术测定ERK蛋白及其mRNA表达。结果 ① 2 5、5 0、10 0mg/L浓度的RGDS四肽呈剂量依赖性抑制HSC增生并诱导HSC凋亡 (P <0 .0 5 )。②RGDS四肽作用于HSC 2h ,HSC粘附率下降 (P <0 .0 1)。③在RGDS四肽干预组 ,HSC的ERK及其mRNA表达下调。结论 RGDS四肽可抑制HSCs增生并诱导其凋亡 ;RGDS四肽诱导HSCs凋亡的效应与其抗粘附作用、抑制ERK蛋白和ERK1mRNA表达有关。
Objective To explore the role of extracellular signal-regulated kinase(ERK)in the apoptosis of hepatic stellate cells(HSC) induced by Arg-Gly-Asp-Ser (RGDS) tetrapeptide. Methods The cultrued HSC cells were divided into six groups including untreated control, fibronertin (FN), FN+RGDS (25 mg/L), FN+RGDS (50 mg/L), FN+RGDS(100 mg/L) and FN+Arg-Gly-Glu-Ser(RGES, 100 mg/L) groups. 3H-thymidine incorporation, transmission electron microscopy and TUNEL were employed to estimate the influence of RGDS on proliferation and apoptosis of HSC. The HSC adhesion rates were observed by toluidine blue colorimetric assay. The expressions of ERK mRNA and ERK protein in HSC were detected by RT-PCR and Western blotting. Results Compared with control and fibronectin groups, RGDS tetrapeptide at concentrations of 25 mg/L, 50 mg/L and 100 mg/L inhibited the proliferation of HSC(P<0.01). After exposure to RGDs tetrapeptide, the chromatins of HSC condensed, shrank and aggregated along the inside of nuclear membranes existing in the forms of ball, petal and crescent. RGDS tetrapeptide induced the HSC apoptosis in a dose-dependent manner (P<0.05). Compared with those in RGES group, HSC adhesion inhibition rates were decreased after being added RGDS tetrapeptide for 2 h (P<0.01). ERK protein and ERK mRNA were down-regulated by RGDS tetrapeptide. Conclusions RGDS tetrapeptide can induce the apoptosis of HSC in both dose-dependent and time-dependent manners in vitro, which may be related to the disruption of cell matrix adhesion and the down-regulation of ERK expression.
出处
《中华消化杂志》
CAS
CSCD
北大核心
2004年第3期162-166,共5页
Chinese Journal of Digestion
基金
河北省自然科学基金 ( 3 0 13 61)
