摘要
目的:探讨丹参单体IH764-3对H2O2刺激的肝星状细胞(HSCs)增殖、凋亡等细胞行为的影响及细胞外信号调节激酶1(ERK1)在其中的调节作用。方法:应用体外细胞培养技术,采用直接细胞计数法、3H-胸腺嘧啶核苷(3H-TdR)掺入法测定HSCs增殖;透射电镜、膜联蛋白(Annexin-V)/碘化丙啶(PI)双标记流式细胞术测定HSCs凋亡;分别应用Western blot和逆转录-聚合酶链反应(RT-PCR)技术测定ERK1蛋白及其mRNA的表达。结果:①H2O2具有刺激HSCs增殖的作用;②丹参单体IH764-3剂量依赖性抑制H2O2刺激的HSCs增殖;③Annexin-V/PI检测显示,10 mg/L,20 mg/L,30 mg/L及40 mg/L IH764-3干预48 h后各组凋亡率分别为6.35%、9.28%、15.10%、19.69%,而H2O2组为2.30%;30 mg/L IH764-3干预HSCs不同时间(12 h、24 h、48 h)的凋亡率分别是6.73%、10.34%、15.10%,呈时间依赖性;④丹参单体IH764-3干预组,HSCs的ERK1蛋白及其mRNA表达下调。结论:丹参单体IH764-3可以抑制HSCs增殖并诱导其凋亡;这种作用与其抑制ERK1蛋白和ERK1mRNA表达有关。
Objective： To explore the effect of Salvia miltiorrhiza monomer IH764-3 on apoptosis in hydrogen peroxide （H202） -stimulated hepatic stellate cells （HSCs）. Methods： HSCs were cultured in medium with different IH764-3 doses （10 mg/L, 20 mg/L, 30 mg/L, 40 mg/L） and without IH764-3. Direct cell count,3H-thymidine incorporation, Annexin-V/Propidium Iodide double-labeled flow cytometry, TUNEL and transmission electron microscopy were employed to estimate the influence of IH764-3 on proliferation and apoptosis of HSCs. The expression of extracellular signal-regulated kinase 1 （ERK1 ） mRNA and protein in HSCs were detected using RT-PCR and Western blot respectively. Results： It was showed that H2O2 could promote HSC proliferation. In contrast, IH764-3 at concentrations of 10 mg/L, 20 mg/L, 30 mg/L and 40 mg/L inhibited its proliferation. The inhibition rates were 7.13%, 28.36%, 53.80% and 73.10% （ P 〈 0.01）. And the inhibition rates of IH764-3 at concentrations of 30 mg/L at 12 h, 24 h and 48 h were 22.24%, 40.51% and 61.65%. Furthermore, IH764-3 could also induce the HSC apoptosis in dose-dependent and time-dependent manners（ P 〈 0.01）. In addition, after exposed of HSCs to IH764- 3 for 24 h, ERK production decreased and ERK1 mRNA was down-regulated earlier about 2 h after exposure to IH764-3. Conclusion： IH764- 3 may inhibit the proliferation and induce apoptosis of HSCs in both dose-dependent and time-dependent manners, which may be related to down-regulation of ERK expression.
出处
《中国应用生理学杂志》
CAS
CSCD
北大核心
2011年第4期402-406,共5页
Chinese Journal of Applied Physiology
基金
河北省中医药管理局科研课题(2007061)
河北省科技厅资助项目(09966107D)
关键词
肝星状细胞
细胞外信号调节激酶
凋亡
丹参单体
IH764-3
过氧化氢
hepatic stellate cell
extracellular signal-regulated kinase
apoptosis
salvia miltiorrhiza monomer
IH764-3
hydrogen peroxide
