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为了探讨三氧化二砷(As2O3)、地塞米松(dexamethasone,Dex)和沙利度胺(thalidomide,Thal)诱导骨髓瘤细胞株U266细胞凋亡时对胞浆内Ca2+浓度([Ca2+]i)的影响,用含15%FBS的RPMI1640培养液培养U266细胞并按5×105/(ml.well)接种于24孔板中,分别用不同浓度的As2O3、Dex和Thal干预8小时后收集U266细胞,用荧光显微镜观察细胞凋亡,以AnnexinV-FITC/PI双参数流式细胞术(FCM)检测细胞凋亡,负载Fluo-3/AM荧光素FCM检测[Ca2+]i。结果显示:①随As2O3、Dex和Thal浓度增加,荧光显微镜下带有荧光信号的凋亡细胞逐渐增多;②As2O3、Dex和Thal作用U266细胞后FCM测得凋亡细胞比例从对照组的0.56%分别升至31.54%、28.35%、21.97%;③As2O3、Dex诱导U266细胞凋亡时[Ca2+]i升高,升高的程度与药物浓度成正比关系;④Thal诱导细胞凋亡时未观察到[Ca2+]i有明显改变。结论:[Ca2+]i的升高是As2O3、Dex诱导U266细胞凋亡的机制之一,Thal诱导U266细胞凋亡与[Ca2+]i的变化无明显关系。
To investigate the effects of As2O3, dexamethasone (Dex) and thalidomide (Thal) on the cytosolic Ca2 + concentration ([Ca2 +] i) in myeloma cell line U266, U266 cells were cultured in RPMI1640 medium containing 15% FBS and inoculated into 24-well plates at 5 × 10 5 / (ml · well). U266 cells were harvested after 8 hours of intervention with different concentrations of As 2 O 3, Dex and Thal respectively. Apoptosis was observed. Apoptosis was detected by Annexin V-FITC / PI dual-parameter flow cytometry (FCM) and [Ca 2+] i was detected by Fluo-3 / AM fluorescein FCM. The results showed that: (1) apoptotic cells with fluorescence signal under fluorescence microscope increased gradually with the increase of As2O3, Dex and Thal concentration; (2) The proportion of apoptotic cells detected by FCM after treated with As2O3, Dex and Thal was 0.56% Up to 31.54%, 28.35% and 21.97%, respectively. (3) The increase of [Ca2 +] i in U266 cells induced by As2O3 and Dex was directly proportional to the concentration of drug; ④ No [Ca2 + i have changed significantly. CONCLUSION: Elevated [Ca2 +] i is one of the mechanisms of As2O3 and Dex-induced apoptosis in U266 cells. Thal-induced U266 cell apoptosis has no significant relationship with [Ca2 +] i.