miR-199b-3p通过靶向PLCε抑制前列腺癌细胞的恶性增殖

miR-199b-3p Suppresses Prostate Cancer Cells Malignant Proliferation by Targeting PLCε

该研究的目的是确定miR-199b-3p在前列腺癌(PCa)中的表达及其对PCa细胞增殖的影响及作用机制。通过实时定量聚合酶链反应(RT-qPCR)检测miR-199b-3p在PCa组织、良性前列腺增生(BPH)组织、PCa细胞及人正常前列腺上皮细胞(RWPE-1)中的表达,并分析其表达与PCa临床病理特征的关系。蛋白质印迹法(Western blot)用于检测磷脂酰肌醇特异性磷脂酶C epsilon(PLCε)的表达。采用CCK-8法和克隆形成实验对其体外增殖进行评估。Edu测定法用于检测细胞对Edu的吸收。荧光素酶报告实验被用来验证miR-199b-3p和PLCε的靶向结合情况。结果显示,在PCa组织中miR-199b-3p的表达水平明显低于BPH组织,且与临床病理特征中的组织学分期相关。上调miR-199b-3p可以显著抑制PCa细胞的增殖和Edu的摄取能力,PLCε被鉴定为miR-199b-3p的下游靶基因,且其表达量与miR-199b-3p的表达量呈负相关。此外,补救实验结果显示,上调PLCε能够逆转miR-199b-3p在PCa细胞增殖中的抑制作用。总之,miR-199b-3p可通过靶向PLCε3′非编码区(3′-UTR)负性调控PLCε进而抑制PCa细胞的恶性增殖。

The purpose of this study was to determine the expression of miR-199b-3p in PCa (prostate cancer) and its effect on PCa cells proliferation.The expression of miR-199b-3p in PCa tissues,BPH (benign prostatic hyperplasia) tissues,PCa cells and RWPE-1 were detected by RT-qPCR (real-time quantitative polymerase chain reaction).In addition,we examined the relationship between miR-199b-3p and the clinicopathological features of PCa in PCa tissues.Western blot analysis was used to detect the expression of PLCε (phospholipiase C epsilon).CCK-8 assay and colony-forming assay were applied to assess the proliferation in vitro.Edu assay was used to detect cell uptake of Edu.Luciferase reporter assay was used to confirm the binding of miR-199b-3p and PLCε.The results showed that the expression level of miR-199b-3p in PCa tissues was significantly lower than that in BPH tissues,and correlated with the histological stage in various clinicopathological characteristics.Upregulation of miR-199b-3p can significantly inhibit the proliferation of PCa cells and the uptake capacity of Edu.PLCε was identified as the downstream target gene of miR-199b-3p.In addition,the expression of PLCε was negatively correlated with that of miR-199b-3p.Furthermore,rescue experiments showed that the overregulation of PLCε could rescue inhibitory effect of miR-199b-3p on cell proliferation.In a word,miR-199b-3p negatively regulates PLCε by targeting its 3?-UTR (3?-untranslated region),then inhibiting the malignant proliferation of PCa cells.