三氧化二砷、地塞米松和沙利度胺对骨髓瘤细胞U266凋亡及胞浆内Ca2+浓度的影响

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?S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。

Effects of As2O3, Dexamethasone and Thalidomide on Apoptosis and Cytoplasmic [ Ca2+ ] of Myeloma Cell Line U266

To investigate the influence of As2O3, dexamethasone (Dex) and thalidomide (Thal) on apoptosis-induced myeloma cell line U266 cytoplasmic calcium concentrations (Ca2+]i), U266 cells were incubated in the culture of RPMI 1640 with 15% FBS in 24-well plate and exposed to different concentrations of As2O3, Dex and Thal for 8 hours, respectively, then cell apoptosis was analyzed by fluorescence microscopy and flow cytometry (FCM) with Annexin V-FITC/PI double staining, and cytoplasmic free calcium were detected on FCM through Fluo-3/AM loading. The results indicated that (1) apoptotic cells were gradually increased with enhancement of As2O3, Dex and Thal concentrations; (2) apoptotic cell rates increased from 0.56% in control to 31.54%, 28.35% and 21.97% respectively after treatment with As2O3, Dex and Thal; (3) As2O3, Dex induced U266 cell apoptosis accompanied with raise of Ca2+]i; (4) Ca2+]i had no statistically significant changes in Thal-induced apoptotic U266 cells. It is concluded that the raise of Ca2+]i is one of the mechanisms for As2O3 and Dex-induced U266 cells apoptosis, whereas Thal-induced U266 apoptosis has no significant relation to Ca2+]i changes.