摘要
探讨过氧化物诱导人类肝细胞（ＬＯ２细胞）、内皮细胞（ＥＶＣ３０４细胞）凋亡时Ｃａ２＋流变化不同特点和意义及丹参提取物Ｒｘａ通过Ｃａ２＋信号系统介导的细胞保护效应。方法用膜联蛋白荧光素（Ａｎｎｅｘｉｎ－Ｖ－Ｆｌｕｏｓ）与碘化丙啶（ＰＩ）双标记流式细胞术（ＦＣＭ）分析Ｈ２Ｏ２作用不同时限Ｒｘａ处理组与未处理组凋亡细胞指数分布；用全细胞膜片钳放大系统显微荧光检测术同步分析同期Ｃａ２＋流变化。结果Ｈ２Ｏ２诱导细胞凋亡时ＬＯ２细胞［Ｃａ２＋］ｉ、Ｃａ２＋跃升，Ａｎｎｅｘｉｎ－Ｖ＋细胞指数均较ＥＶＣ３０４细胞为高；［Ｃａ２＋」ｉ＞４００ｎｍｏｌ／Ｌ后１～２小时可观察到细胞凋亡早期膜翻转现象。Ｒｘａ组［Ｃａ２＋］ｉ、Ａｎｎｅｘｉｎ．Ｖ＋和ＰＩ＋细胞指数均较未处理组为低。结论Ｈ２Ｏ２可诱导ＬＯ２、ＥＶＣ３０４细胞Ｃａ２＋依赖性细胞凋亡；［Ｃａ２＋］ｉ＞４００ｎｍｏｌ／Ｌ可能为细胞凋亡启动的早期信号；ＬＯ２细胞较ＥＶＣ３０４细胞抗凋亡能力为差；Ｒｘａ能有效降低［Ｃａ２＋］ｉ，减轻细胞损伤程度。
Objective To investigate the different characteristics of intracellular free Ca2+ concentration ([Ca2+1i) during the periods of hydrogen peroxide (H2O2)-mediated apoptosis of human hepatocytes(LO2 cells) and endothelial cells (EVC304 cells), and the effective cytoprotection of Rxa via Ca2+ signalpathway. Method L02 cells and EVC304 cells were respectively divided into 3 groups: (1) Control group;(2) H2O2 injury group (H2O2 10 μmol/L); and (3) Rxa-treatment group: (Rxa 2 nmol/L). Apoptotic cellswere analyzed quantitatively by flow cytometry with Annexin-V-flous. The [Ca2+] i was analyzed by wholecell pacth clamp combined with single cell micro-fluorescence [Ca2+] i measurement. Result The levels of[Ca2+ ]i, Ca2+ spiking and index of apoptotic cells of L02 cells were higher than that of EVC304 cells; the index of apoptotic cells was increased significantly when [Ca2+] i>400 mmol/L. The datum in Rxa-treatmentgroups were lower than that of no-treatment groups. Conclusion H2O2 induced apoptosis depending on Ca2+in L02 and EV304 cells: it may be one of the early apoptotic signal mediators that [Ca2+] i reached 400 nmol/L or more; Rxa had effective cytoprotection, which probably related to its calcium blocking effect, especiallyin L02 cells.
出处
《中华实验外科杂志》
CSCD
北大核心
1999年第2期147-149,共3页
Chinese Journal of Experimental Surgery
基金
国家自然科学基金!No.39770938
