miR-144-3p靶向调控E2F3抑制膀胱癌T24细胞的增殖与侵袭

miR-144-3p inhibits proliferation and invasion of bladder cancer T24 cells through targeted regulation of E2F3

目的:探讨miR144-3p在膀胱癌组织及细胞中的表达及其对T24细胞增殖与侵袭的影响。方法:选用2018年2月至2018年12月空军军医大学唐都医院手术切除的36例膀胱癌组织及10例正常膀胱上皮组织标本,以及人膀胱癌细胞株T24和正常尿路上皮细胞株SV-HUC-1,用qPCR法检测膀胱癌组织和细胞中miR144-3p的表达水平。用脂质体转染技术分别将miR-144-3p mimics、miR-NC等转染进T24细胞,通过MTT法、流式细胞术和Transwell小室法分别检测T24细胞的增殖、细胞周期和侵袭能力。利用TargetScan软件预测miR-144-3p与E2F转录因子3(E2F transcription factor 3,E2F3)的结合位点,用双荧光素报告基因实验验证miR-144-3p与E2F3的靶向关系,WB实验检测细胞中miR-144-3p与E2F3的表达水平。结果:miR-144-3p在膀胱癌组织和细胞中低表达(均P<0.01),其中肌层浸润性膀胱癌组织中的表达水平低于非肌层浸润膀胱癌组织(P<0.05)。双荧光素报告基因实验证实,miR-144-3p与E2F3表达存在靶向调节关系。miR-144-3p过表达可抑制T24细胞的增殖和侵袭能力(均P<0.01),同时下调E2F3的表达水平(P<0.01);当上调E2F3表达时,对细胞侵袭和增殖的抑制作用则被逆转。结论:miR-144-3p在膀胱癌组织中低表达,其通过靶向调控E2F3表达从而抑制膀胱癌细胞的增殖与侵袭。

Objective: To investigate the expression of miR144-3p in bladder cancer tissues and cells and its effect on the proliferation and invasion of T24 cells. Methods: A total of 36 cases of bladder cancer tissue specimens and 10 cases of normal bladder epithelial tissue specimens were collected from Tangdu Hospital of Air Force Medical University during February 2018 and December 2018. In addition, bladder cancer T24 cell line and normal urothelial cell line SV-HUC-1 were also collected for this study. The levels of miR144-3p in bladder cancer tissues and cells were detected by q PCR methods. The miR-144-3p mimics and miR-NC were transfected into T24 cells by LipofectamineTM 2000, respectively. The proliferation, cell cycle distribution and invasion abilities were detected by MTT, Flow cytometry and Transwell chamber methods, respectively. TargetScan software was used to predict the binding site between miR-144-3p and E2F3(E2 F transcription factor 3);Dual luciferase reporter gene assay was used to verify the relationship between miR-144-3p and E2F3;and WB was used to detect the expression levels of miR-144-3p and E2F3 in cells. Results: The expression of miR-144-3p was downregulated in bladder cancer tissues and cells(all P<0.01). In addition, the expression level of miR-144-3p in muscular invasive bladder cancer tissues was significantly lower than that in non-muscular invasive bladder cancer tissues(P<0.05).Dual luciferase reporter gene assay confirmed that there was a targeted relationship between miR-144-3p and E2F3. Overexpression of miR-144-3p inhibited the proliferation and invasion of T24 cells(all P<0.01) and downregulated the expression of E2F3(P<0.01);upregulation of E2F3 could reverse the inhibitory effect of miR-144-3p overexpression on proliferation and invasion of T24 cells.Conclusion: miR-144-3p has low expression level in bladder cancer tissues. It inhibits proliferation and invasion of bladder cancer cells by downregulating E2F3.