干预ERRα的表达对体外培养的多发性骨髓瘤细胞MM.1S的凋亡诱导作用

Intervention of ERRα Expression on Apoptosis Induction of Multiple Myeloma MM.1S Cells Cultured in Vitro

目的:探讨两种不同的干预ERRα基因的表达和活性策略对人多发性骨髓瘤细胞株MM.1S凋亡的影响。方法:第一种策略是设计并构建靶向人ERRα基因的sh RNA慢病毒载体,利用293T细胞包装慢病毒后感染骨髓瘤细胞株MM.1S,经嘌呤霉素筛选得到敲低ERRα的稳转细胞株。另一种策略是使用ERRα的特异性反向激动剂XCT790处理细胞。流式细胞术分析敲低ERRα对MM.1S细胞凋亡的影响,Western blot检测敲低ERRα后凋亡相关蛋白的表达情况。结果:成功构建敲低ERRα的sh RNA表达载体,获得ERRα敲低的稳转细胞株;靶向ERRα的sh RNA病毒感染和特异性反向激动剂均能显著下调ERRα在骨髓瘤细胞MM.1S中的表达水平;敲低ERRα能诱导MM.1S细胞发生显著凋亡并且使凋亡相关蛋白cleaved PARP表达水平升高;ERRα的特异性反向激动剂XCT790处理骨髓瘤细胞MM.1S与敲低ERRα导致的效应一致。结论:通过sh RNA靶向敲低ERRα或使用特异性反向激动剂XCT790抑制ERRα的表达均能诱导MM.1S细胞发生显著凋亡,提示ERRα对骨髓瘤细胞的生存有重要影响。

Objective: To investigate the effect of two different approaches ERRα strategy on the apoptosis in multiple myeloma cell line MM.1 S. Methods: For the one strategy, sh RNA was mediated by lentivirus. Stable cell clones were established by transfecting the lentivirus into MM.1 S cells and screened by puromycin. For the other strategy, XCT790, a specific reverse agonist of ERRα, was used to treat MM.1 S cells. The apoptosis of the cells was analyzed by flow cytometry after ERRα was down-regulated. Western blot assay was used to detect the apoptosis of related proteins. Results: The knocked down ERRα was achieved, lentivirus with sh ERRα were successfully infected into MM.1 S and ERRα was reduced significantly. Knockdown of ERRα could induce MM.1 S cell apoptosis dramatically. Meanwhile, the expression of cleaved PARP(a kind of apoptosis related markers) was significantly increased following depletion of ERRα in MM.1 S cells. XCT790 could significantly down-regulate the expression of ERRα protein in MM.1 S cells, which was consistent with the effect caused by sh RNA. Conclusion: Interference the expression of ERRα by sh RNA or XCT790 can induce apparent apoptosis in MM.1 S cells, which indicating that ERRα is crucial for the survival of myeloma cells.