三氧化二砷经内质网应激反应诱导K562/ADM耐药细胞凋亡

Arsenic trioxide induces endoplasmic reticulum stress-related apoptosis in drug-resistant K562/ADM cells

目的探讨三氧化二砷(As2O3)能否通过内质网应激反应性凋亡途径诱发耐药白血病细胞凋亡。方法采用细胞形态学和AnnexinV/PI双染色法检测细胞凋亡,电镜观察凋亡细胞内质网和线粒体形态结构变化;流式细胞术(FCM)测定线粒体跨膜电位(Δψm)、细胞内Ca2+和细胞色素c(Cyt c)含量及caspase-3活性;Western blot法检测GRP78/Bip蛋白表达。结果2、5μmol/L As2O3诱导K562/ADM细胞发生凋亡过程中,内质网明显扩张和脱颗粒,线粒体内外膜融合,嵴紊乱,肿胀,内膜扩张呈空泡样变。线粒体Δψm降低,细胞质Ca2+和Cyt c水平明显升高,caspase-3活性显著增强,GRP78蛋白表达增高。结论As2O3可诱导K562/ADM细胞发生内质网应激反应,通过内质网-线粒体途径诱导耐药白血病K562/ADM细胞凋亡。

Objective To explore whether arsenic trioxide （As2O3 ）-induced apopotosis in drug-resistant leukemia K562/ADM cells may induce in through endoplasmic reticulum stress leukemia cell apopotosis. Methods The apoptosis of K562/ADM cells was identified by double staining of FITC-Annexin V and propidium iodide （PI）, the ultrastructure of the cells, endoplasmic reticulum and mitochondria were observed by transmission electron microscopy. Flow cytometry （FCM） was employed to assess mitochondrial inner membrane potential（ AtOm）, intracellular calcium concentration, cytochrome c （ Cyt c） release and caspase-3 activity. The expression of GRP78 protein was analyzed by Western blot. Results During the apoptotic process of K562/ADM ceils induced with 2μmoL/L and 5μmoL/L As2O3, the endoplasmic reticulum exhibited obvious expansion and degranulation, and the mitoehondria illustrated inner and outer membranes fusion, reduced and confused cristae, swelling and vacuolization. The mitochondrial AtOm decreased, the intracellular calcium concentration and releasing of cytochrome c from mitoehondria increased, and caspase-3 was activated. Western blot result indicated upregulation of GRP78 protein at endoplasmic reticulum in apopototic K562/ADM cells. Conclusion As2O3 can initiate the endoplasmic reticulum stress in K562/ADM cells, and induces to apoptosis of the drug-resistant cell via endoplasmic reticulummitochondrial pathway.