miR-219-5p 通过靶向 HMGA2 抑制前列腺癌细胞生长和转移。
MiR-219-5p inhibits prostate cancer cell growth and metastasis by targeting HMGA2.
机构信息
Department of Urology, The Third Affiliated Hospital of Sun Yat-Sen University, Guangzhou, China.
出版信息
Eur Rev Med Pharmacol Sci. 2020 May;24(9):4710-4718. doi: 10.26355/eurrev_202005_21159.
OBJECTIVE
To investigate the expression of micro ribonucleic acid (miR)-219-5p in prostate cancer (PCa), its influences on the biological functions of PCa, and its mechanism.
PATIENTS AND METHODS
The expression differences of miR-219-5p and high mobility group protein A2 (HMGA2) in 30 pairs of PCa tissues and para-carcinoma tissues were detected via quantitative Real Time-Polymerase Chain Reaction (qRT-PCR), and the difference in miR-219-5p expression in PCa cell lines and normal prostatic epithelial cells was also determined via qRT-PCR. The human PC-3 cells were divided into negative control group and miR-219-5p overexpression group. Methyl thiazolyl tetrazolium (MTT) and colony formation assays were adopted to detect the cell proliferative ability, and flow cytometry was applied to determine the cell apoptosis. The expression of apoptosis-related proteins was measured via Western blotting, and the invasive and migratory abilities of the cells were examined through wound-healing and transwell assays. Bioinformatics prediction software and luciferase reporter assay were employed to verify the targets that might be controlled by miR-219-5p. Rescue experiment was conducted to clarify whether the inhibitory effects of miR-219-5p on the growth and metastasis of PC-3 cells depend on the inhibition of HMGA2.
RESULTS
It was shown in qRT-PCR results that the expression level of miR-219-5p was downregulated remarkably in PCa tissues and cell lines, but overexpressed miR-219-5p could repress the proliferation and promote the apoptosis of PC-3 cells notably. The results of wound-healing and transwell assays indicated that overexpressed miR-219-5p was able to suppress the invasion and metastasis of PC-3 cells. According to Western blotting results, overexpressed miR-219-5p could up-regulate the expressions of pro-apoptotic proteins [Bax, cleaved-caspase-3 and cleaved-poly-ADP-ribose-polymerase (PARP)] and reverse the epithelial-mesenchymal transition (EMT) of PCa cells. It was predicted via the bioinformatics software that HMGA2 gene might be a target gene of miR-219-5p. The Dual-Luciferase reporter assay confirmed that there was a direct regulatory relationship between miR-219-5p and HMGA2. The rescue experiment manifested that overexpressed HMGA2 could reverse the inhibition of miR-219-5p on the growth and metastasis of PC-3 cells.
CONCLUSIONS
MiR-219-5p suppresses the growth and metastasis abilities of prostate cancer cells by directly repressing the expression of HMGA2.
目的
研究微小 RNA（miR）-219-5p 在前列腺癌（PCa）中的表达，及其对 PCa 生物学功能的影响和机制。
患者和方法
采用实时定量聚合酶链反应（qRT-PCR）检测 30 对 PCa 组织和癌旁组织中 miR-219-5p 和高迁移率族蛋白 A2（HMGA2）的表达差异，并通过 qRT-PCR 检测 PCa 细胞系和正常前列腺上皮细胞中 miR-219-5p 的表达差异。将人 PC-3 细胞分为阴性对照组和 miR-219-5p 过表达组。采用噻唑蓝（MTT）和集落形成实验检测细胞增殖能力，采用流式细胞术检测细胞凋亡。采用 Western blot 检测凋亡相关蛋白的表达，采用划痕愈合和 Transwell 实验检测细胞的侵袭和迁移能力。采用生物信息学预测软件和荧光素酶报告实验验证可能受 miR-219-5p 调控的靶基因。通过 rescue 实验明确 miR-219-5p 对 PC-3 细胞生长和转移的抑制作用是否依赖于对 HMGA2 的抑制。
结果
qRT-PCR 结果显示，miR-219-5p 在 PCa 组织和细胞系中表达显著下调，但过表达 miR-219-5p 可显著抑制 PC-3 细胞的增殖并促进其凋亡。划痕愈合和 Transwell 实验结果表明，过表达 miR-219-5p 可抑制 PC-3 细胞的侵袭和转移。Western blot 结果显示，过表达 miR-219-5p 可上调促凋亡蛋白（Bax、cleaved-caspase-3 和 cleaved-poly-ADP-ribose-polymerase [PARP]）的表达并逆转 PCa 细胞的上皮间质转化（EMT）。生物信息学软件预测显示，HMGA2 基因可能是 miR-219-5p 的靶基因。双荧光素酶报告实验证实 miR-219-5p 与 HMGA2 之间存在直接的调控关系。rescue 实验表明，过表达 HMGA2 可逆转 miR-219-5p 对 PC-3 细胞生长和转移的抑制作用。
结论
miR-219-5p 通过直接抑制 HMGA2 的表达抑制前列腺癌细胞的生长和转移能力。
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