摘要
目的 探讨长链非编码RNA(lncR)HOXA-AS3和微小RNA-29a(miR-29a)对前列腺癌(PCa)细胞凋亡、自噬的影响。方法 体外培养人PCa细胞(DU-145、LNCaP、C4-2B)和人正常前列腺细胞RWPE-1,采用RT-qPCR法检测细胞中lncR-HOXA-AS3和miR-29a表达。应用双荧光素酶报告基因实验验证lncR-HOXA-AS3和miR-29a之间的靶向调控关系。取对数生长期LNCaP细胞,分为si-NC组、si-HOXA-AS3组、si-HOXA-AS3+anti-miR-NC组和si-HOXA-AS3+anti-miR-29a组,利用脂质体转染法分别将si-NC、si-HOXA-AS3、si-HOXA-AS3+anti-miR-NC、si-HOXA-AS3+anti-miR-29a转染至各组LNCaP细胞,应用RT-qPCR法检测各组细胞中lncR-HOXA-AS3和miR-29a表达,应用流式细胞术检测各组细胞凋亡情况,应用Western blotting法检测各组细胞中凋亡相关蛋白Caspase-3及自噬相关蛋白LC3II、LC3I、Beclin 1表达。结果 与正常前列腺细胞相比,人PCa细胞中lncR-HOXA-AS3表达均明显升高(P均<0.05),而miR-29a表达均降低(P均<0.05)。双荧光素酶报告基因显示lncR-HOXA-AS3能靶向miR-29a抑制荧光素酶活性(P<0.05)。与si-NC组比较,si-HOXA-AS3组LNCaP细胞lncR-HOXA-AS3表达降低(P<0.05),miR-29a表达升高(P<0.05),细胞凋亡率升高(P均<0.05),Caspase-3、LC3Ⅱ/LC3I及Beclin 1蛋白表达升高(P均<0.05)。与si-HOXA-AS3+anti-miR-NC组比较,si-HOXA-AS3+anti-miR-29a组LNCaP细胞miR-29a表达下降(P<0.05),细胞凋亡率降低(P<0.05),Caspase-3、LC3Ⅱ/LC3I及Beclin1蛋白表达下降(P均<0.05)。结论 抑制lncR-HOXA-AS3表达能靶向调控miR-29a基因促进PCa细胞凋亡和自噬。
Objective To investigate the effects of long chain non-coding RNA(lncR) HOXA-AS3 and microRNA-29a(miR-29a) on apoptosis and autophagy of prostate cancer(PCa) cells.Methods Human PCa cells(DU-145,LNCaP,C4-2B) and human normal prostate cells RWPE-1 were cultured in vitro,and RT-qPCR was used to detect the relative expression of lncR-HOXA-AS3 and miR-29a in the cells.Dual luciferase reporter assay was used to verify the targeted regulatory relationship between lncR-HOXA-AS3 and miR-29a.LNCaP cells in the logarithmic growth phase were divided into the si-NC group,si-HOXA-AS3 group,si-HOXA-AS3+anti-miR-NC group and si-HOXA-AS3+ anti-miR-29a group.Si-NC,si-HOXA-AS3,si-HOXA-AS3+anti-miR-NC,si-HOXA-AS3+ anti-miR-29a were transfected into LNCaP cells of the above groups by liposome transfection.The expression changes of lncR-HOXA-AS3 and miR-29a in each group were detected by RT-qPCR,and the apoptosis of cells in each group was detected by flow cytometry,and the expression levels of apoptosis-related proteins Caspase-3,autophagy related proteins LC3II,LC3I and Beclin 1 in each group were detected by Western blotting.Results Compared with the normal prostate cells,the expression of lncR-HOXA-AS3 significantly increased(P<0.05),while the expression of miR-29a significantly decreased in human PCa cells(P<0.05).Dual luciferase reporter genes showed that lncR-HOXA-AS3 inhibited luciferase activity by targeting miR-29a(P<0.05).Compared with the si-NC group,the lncR-HOXA-AS3 expression of LNCaP cells decreased,and the miR-29a expression increased(both P<0.05),the apoptosis rate significantly increased(P<0.05),and the protein expression levels of Caspase-3,LC3II/LC3I and Beclin 1 significantly increased in the si-HOXA-AS3 group(all P<0.05).Compared with the si-HOXAAS3+anti-miR-NC group,the expression of miR-29a significantly decreased(P<0.05),the apoptosis rate significantly decreased(P<0.05),and the expression levels of Caspase-3,LC3II/LC3I,and Beclin 1 proteins significantly decreased in the si-HOXA-AS3+anti-miR-29a group(all P
作者
李强
刘晶
张国民
王亮
刘志飞
朱研峰
LI Qiang;LIU Jing;ZHANG Guomin;WANG Liang;LIU Zhifei;ZHU Yanfeng(Department of Urology,Kailuan General Hospital Linxi Hospital,Tangshan 063100,China;不详)
出处
《山东医药》
CAS
2024年第10期16-20,共5页
Shandong Medical Journal
基金
河北省医学科学研究课题(20220216)。
