胞外环境Ca~(2 )变化诱导人肿瘤细胞凋亡

目的 :观察胞外环境Ca2 变化对人肿瘤细胞的直接诱导凋亡作用。方法 :通过EGTA螯合胞外Ca2 或添加CaCl2 改变胞外环境Ca2 。用PI和Hoechst3334 2荧光双染观察细胞核形态 ,用流式细胞仪检测凋亡百分率。分别借助荧光探针Fluo 3和Rh12 3检测胞内Ca2 和线粒体膜电位 (ΔΨm)变化。用环孢菌素A(CsA)阻断PT孔 ,研究其在凋亡中的作用。结果 :胞外环境Ca2 升高或降低均诱导人胃癌MGC 80 3和喉癌Hep细胞凋亡 ,恢复胞外Ca2 阻断凋亡。随胞外Ca2 变化胞内Ca2 相应升高或降低 ,但线粒体ΔΨm均表现为急剧下降。CsA对MGC 80 3细胞凋亡无明显抑制作用 ,轻微促进Hep细胞凋亡。结论 :胞外环境Ca2 变化引起胞内Ca2 相应变化和线粒体ΔΨm下降并通过PT孔非依赖性途径诱导肿瘤细胞凋亡。

Alterations of extracellular calcium induce apoptosis in human tumor cell lines

Purpose：To study the direct apoptosis-inducing effect of extracellular calcium alterations on human tumor cells. Methods：Cells were incubated with calcium chelator EGTA to lower extracellular Ca2+ or CaCl2 to raise it. Apoptosis was detected by fluorescence microscope and flow cytometry. Intracellular Ca2+ and mitochondria transmembrane potential (ΔΨm) were measured by fluorescent probes Fluo-3 and Rh123 respectively. The role of mitochondria permeability transition pore (MTP) was also investigated by using the specific blocker of MTP, CsA. Results：Both EGTA and excess CaCl2 induced apoptosis in the human gastric cancer cell line MGC-803 and the human laryngocarcinoma cell line Hep, which was blocked by restoring extracellular calcium. The changes of intracellular calcium corresponded to the decrease or increase of extracellular calcium, while ΔΨm was dramatically decreased in both cases. CsA did not protect MGC-803 cells against apoptosis and slightly aggravated it in Hep cells.Conclusions：Alterations of extracellular calcium was accompanied by changes of intracellular calcium and decrease of ΔΨm and induced MTP-independent apoptosis in human tumor cell lines MGC-803 and Hep.