半枝莲含药血清对肝癌H22细胞凋亡及线粒体膜电位的影响

Effects of Scutellaria Barbata drug-containing serum on apoptosis and mitochondrial transmembrane potential of hepatoma H22 cells

目的:探讨中药半枝莲提取物(Scutellaria Barbata extract,ESB)含药血清对肝癌H22细胞凋亡率及线粒体膜电位的影响。方法:体外培养小鼠H22肝癌细胞,以ESB高、中、低剂量含药血清培养液作用于H22细胞,并设立空白血清及5-氟尿嘧啶(fluorouracil,5-Fu)组(阳性对照组)。应用四甲基偶氮唑盐法检测药物对细胞增殖的抑制作用;透射电镜下观察肿瘤细胞的超微结构变化;碘化丙啶标记,流式细胞术检测肿瘤细胞周期及细胞凋亡情况;采用荧光探针罗丹明123负载,激光扫描共聚焦显微镜下观察H22细胞的荧光强度,分析H22细胞线粒体膜电位的大小。结果:ESB含药血清有一定的抑制H22细胞增殖的作用,高、中剂量组96 h抑制率分别为(47.5±6.4)%和(36.3±4.5)%。透射电镜下可见部分细胞呈典型凋亡形态学改变。流式细胞仪检测结果显示,ESB中、高剂量组有明显的诱导凋亡作用,其凋亡率分别为(3.15±1.12)%和(7.83±2.25)%,与空白组相比差异有统计学意义(P<0.01)。空白对照组、5-Fu组和ESB低、中、高剂量,H22细胞线粒体膜电位分别为(245.45±67.37)、(127.42±41.35)和(213.68±65.52)、(186.34±56.37)、(142.65±39.44),各组线粒体膜电位与细胞凋亡率呈负相关(r=-0.826,P<0.01)。结论:ESB含药血清可有效诱导肿瘤细胞凋亡,其机制可能与降低肿瘤细胞线粒体膜电位有关。

Objective：To investigate the effects of serum containingScutellaria Barbataextract（ESB） on apoptosis rate and mitochondrial transmembrane potential（MTP） of liver cancer cell line H22from micein vitro.Methods：H22cells were culturedin vitroand divided into5groups： blank control group,low-dose ESB group,medium-dose ESB group,high-dose ESB group and fluorouracil（5-Fu） group.Methyl thiazolyl tetrazolium assay was utilized to determine the proliferation rates of H22cells.Cellular morphology was observed under a transmission electron microscope（EM）.The rhodamine123was used as a fluorescence probe to label the H22cells,and the fluorescence intensities were observed with a laser scanning confocal microscope.The fluorescence intensity of H22cells indicated the MTP of H22cells.Results：The inhibition of serum containing ESB on the proliferation of H22cellsin vitrowas observed in a time-dependent manner.The typical morphological changes of apoptosis were observed after incubation with ESB-containing serum in high dose for48h.The apoptosis rates of blank control group,5-Fu group,low-dose ESB group,medium-dose ESB group and high-dose ESB group were（0.51±0.32）%,（11.26±2.97）%,（1.07±0.46）%,（3.15±1.12）%,（7.83±2.25）% respectively.ESB could reduce the MTP of H22cells from mice as compared with the untreated group.The MTPs of the blank control group,5-Fu group,and low-,medium-and high-dose ESB groups were（245.45±67.37）,（127.42±41.35）,（213.68±65.52）,（186.34 ±56.37） and（142.65±39.44） respectively,which were negatively correlated with the apoptosis rates.Conclusion：ESB-containing serum effectively induces apoptosis,which may be related to the decrease of MTP in H22 cells.