Objective: To investigate the effect of two different approaches ERRα strategy on the apoptosis in multiple myeloma cell line MM.1S.
Methods: For the one strategy, shRNA was mediated by lentivirus. Stable cell clones were established by transfecting the lentivirus into MM.1S cells and screened by puromycin. For the other strategy, XCT790, a specific reverse agonist of ERRα, was used to treat MM.1S 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 shERRα were successfully infected into MM.1S and ERRα was reduced significantly. Knockdown of ERRα could induce MM.1S cell apoptosis dramatically. Meanwhile, the expression of cleaved PARP (a kind of apoptosis related markers) was significantly increased following depletion of ERRα in MM.1S cells. XCT790 could significantly down-regulate the expression of ERRα protein in MM.1S cells, which was consistent with the effect caused by shRNA.
Conclusion: Interference the expression of ERRα by shRNA or XCT790 can induce apparent apoptosis in MM.1S cells, which indicating that ERRα is crucial for the survival of myeloma cells.
题目: 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.
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shRNA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shRNA表达载体，获得ERRα 敲低的稳转细胞株；靶向ERRα的shRNA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shRNA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.
Keywords: ERRα; MM.1S; multiple myeloma.