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目的 探讨过氧化物诱导人类肝细胞（Ｌ０２ 细胞） 凋亡时重要细胞信使Ｃａ２＋ 和同期细胞膜超微结构变化特点和意义，以及丹参提取物有效成分之一Ｒｘａ 通过Ｃａ２＋ 信号系统介导的细胞保护效应．方法 实验以正常Ｌ０２ 细胞为对照（ Ｌ１ 组） ，分Ｒｘａ 处理组（ＬａＨ 组， Ｒｘａ ２ ｍ ｍｏｌ／ Ｌ） 和未处理组（ ＬＨ 组） ， 在 Ｈ２Ｏ２（１０ μｍ ｏｌ／ Ｌ） 作用于Ｌ０２ 细胞０ ，０－５ ，１ ，２ ，４ ，６ ，８ ｈ ，用膜联蛋白荧光素（ ＡｎｎｅｘｉｎＶＦｌｕｏｓ） 、碘化丙啶（ＰＩ） 双标记流式细胞术（ＦＣＭ） 与荧光显微镜联合检测术分析各检测点正常细胞、早期凋亡（ ＡｎｎｅｘｉｎＶ＋） 细胞、死亡（ ＰＩ＋） 细胞指数，用Ｆｕｒａ２／ ＡＭ标记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，用扫描电镜技术观察同期细胞膜超微结构变化．结果 ＬＨ 组Ｌ０２ 细胞在Ｈ２Ｏ２ 作用后０－５ ｈ 出现Ｃａ２＋ 跃升，１ ｈ［Ｃａ２ ＋］ｉ ＞ ４００ ｎ ｍ ｏｌ／ Ｌ后出现Ｃａ２＋ 振荡现象；２ ｈ ～４ ｈ 荧光显微镜下可观察到细胞凋亡早期膜翻转现象，电镜下可见少量膜小泡出现；４ ｈ ～６ ｈ 后ＦＣＭ 检测到ＡｎｎｅｘｉｎＶ＋ ?
Objective To investigate the characteristics and significance of ultrastructural changes of important cell messenger Ca2 + and cell membrane during peroxides-induced apoptosis in human hepatocytes (L02 cells) and the protective effect of Rxa mediated by Ca2 + signaling system . Methods L02 cells were treated with L02 cells (L1 group), treated with Rxa (2 mmol / L) and untreated group (Rx) (FCM) with AnnexinVFluos, Propidium Iodide (PI) double staining and fluorescence microscope at 0, 0-5, 1, 2, 4, The detection of normal cells, early apoptosis (AnnexinV +) cells and the index of death (PI +) cells in each test site were detected by the combined detection technique. The whole cell patch clamp system was amplified by Fura2 / AM. Changes, the use of scanning electron microscopy observation of cell membrane ultrastructure during the same period. Results Ca2 + oscillations were observed in LH group at 0-5 hours after H2O2 treatment. Ca2 + oscillations were observed after 1 hour [Ca2 +] i> 400 nmol / L. In 2 hours and 4 hours, apoptosis was observed under fluorescence microscope The phenomenon of membrane turnover in the early stage of death, a small amount of membrane vesicles can be seen under electron microscopy; 4 h ~ 6 h after FCM detected Annexin V +?