LncRNA PICSAR敲低对卵巢癌细胞增殖、迁移和凋亡的影响及其作用机制

Effect and mechanism of LncRNA PICSAR knockdown on proliferation,migration and apoptosis of ovarian cancer cells

目的探讨长链非编码RNA(LncRNA)PICSAR在卵巢癌中的表达,探究LncRNA PICSAR对卵巢癌细胞增殖、迁移、侵袭和凋亡的影响以及其可能的作用机制。方法使用荧光定量PCR法检测卵巢癌组织和细胞系A2780、OVCAR-3、HO-8910以及正常卵巢组织和细胞系IOSE386中LncRNA PICSAR的表达水平。将LncRNA PICSAR表达最高的卵巢癌细胞系分为对照组和实验组,通过脂质体转染技术分别转染阴性对照小干扰RNA(siRNA-NC)或PICSAR小干扰RNA(siRNA-PICSAR)。细胞计数实验(CCK-8)、克隆形成实验、划痕实验和Transwell实验以及流式细胞术分别分析LncRNA PICSAR敲低对卵巢癌细胞增殖、迁移、侵袭和凋亡的影响。Western blot法测定各组细胞的凋亡蛋白Bcl-2、Bax和自噬蛋白LC3B、ATG7、Beclin-1的表达水平。使用雷帕霉素和羟氯喹分别作为自噬激活剂和抑制剂处理卵巢癌细胞,并通过Western blot实验检测细胞凋亡水平。结果与正常卵巢组织相比,卵巢癌组织中LncRNA PICSAR表达水平升高。与IOSE386细胞系比较,卵巢癌细胞系A2780、OVCAR-3、HO-8910中LncRNA PICSAR表达水平均升高。与si-NC组比较,si-PICSAR组卵巢癌细胞的增殖、迁移和侵袭能力均降低,细胞凋亡率增高。si-PICSAR组卵巢癌细胞的自噬水平较si-NC组降低,siRNA-PICSAR转染后,用雷帕霉素激活自噬降低了细胞凋亡水平,而用羟氯喹抑制自噬促进了细胞凋亡。结论LncRNA PICSAR在卵巢癌组织和细胞系中高表达,LncRNA PICSAR敲低能够抑制卵巢癌细胞的恶性生物学行为。敲低LncRNA PICSAR可能是通过调节自噬促进卵巢癌细胞凋亡。

Objective To investigate the expression of long non-coding RNA(LncRNA)PICSAR in ovarian cancer,and explore the effects of LncRNA PICSAR on the proliferation,migration,invasion and apoptosis of ovarian cancer cells as well as its possible mechanism of action.Methods The expression levels of LncRNA PICSAR in ovarian cancer tissue and cell line A2780,OVCAR-3,HO-8910 and normal ovarian tissue and cell line IOSE386 were detected by fluorescence quantitative PCR.Ovarian cancer cell lines with the highest expression of LncRNA PICSAR were divided into control group and experimental group,and transfected with negative control small interfering RNA(siRNA-NC)or PICSAR knockout small interfering RNA(siRNA-PICSAR)by liposome transfection technique,respectively.The effects of LncRNA PICSAR knockdown on the invasion,migration,proliferation and apoptosis of ovarian cancer cells were analyzed by cell counting assay(CCK-8),clonogenic assay,scratch assay,transwell assay and flow cytometry and so on.The expression levels of autophagy related proteins and apoptosis-related proteins in each group were determined by Western blot.Ovarian cancer cells were treated with rapamycin and hydroxychloroquine as autophagy activator and inhibitor,and Western blot assay was used to detect apoptosis.Results The expression level of LncRNA PICSAR in ovarian cancer tissues was higher than that in normal ovarian tissues.Compared with IOSE386 cell line,LncRNA PICSAR expression levels in ovarian cancer cell lines HO-8910,OVCAR-3 and A2780 increased.Compared with the si-NC group,the proliferation,invasion and migration ability of ovarian cancer cells in si-PICSAR group decreased,and the apoptosis rate increased.The autophagy level of ovarian cancer cells in si-PICSAR group was lower than that in si-NC group.After transfection with siRNA-PICSAR,rapamycin activated autophagy to reduce apoptosis,while hydroxychloroquine inhibited autophagy to promote apoptosis.Conclusion LncRNA PICSAR is highly expressed in ovarian cancer tissues and cell lines,and the malignant biological behavior of ovarian cancer cells can be inhibited by knockout of LncRNA PICSAR.The knockdown of LncRNA PICSAR may promote the apoptosis of ovarian cancer cells by regulating autophagy.