miR-1243通过靶向hnRNPA2B1调控肝癌HepG2细胞增殖和迁移

目的：探讨miR-1243通过靶向调控核不均一核糖核蛋白A2/B1（hnRNPA2B1）表达对肝癌HepG2细胞增殖、迁移的影响及其分子机制。方法：用qPCR和WB法检测40例肝癌组织及其癌旁组织（2019年1月至2021年8月在武汉市第三医院首义院区手术切除标本）和正常人肝细胞 QSG-7701 与肝癌细胞 HepG2、Hep3b、HuH-7 中 miR-1243、hnRNPA2B1 mRNA 水平及hnRNPA2B1、cyclin D1、MMP-2蛋白水平；双荧光素酶报告基因实验验证miR-1243和hnRNPA2B1的靶向关系。HepG2细胞分为对照组（不转染）、miR-NC 组（转染 miR-NC）、miR-1243 mimic 组（转染 miR-1243 mimic）、miR-1243 mimic+pcDNA3.1 组（转染miR-1243 mimic 和 pcDNA3.1）、miR-1243 mimic+pc-hnRNPA2B1 组（转染 miR-1243 mimic 和 pc-hnRNPA2B1）后进行相应转染；WB 法检测肝癌组织及细胞和转染后各组细胞的 hnRNPA2B1、cyclin D1、MMP-2 蛋白表达水平；CCK-8 法检测转染后各组HepG2细胞的增殖能力；划痕愈合实验检测转染后各组HepG2细胞的迁移能力。结果：与癌旁组织或正常人肝细胞QSG-7701相比，肝癌组织和肝癌细胞中miR-1243呈低表达、hnRNPA2B1 mRNA及其蛋白呈高表达（均P<0.05）。双荧光素酶报告基因实验结果证实miR-1243与hnRNPA2B1间存在靶向关系，且miR-1243通过靶向hnRNPA2B1负调控其表达。转染miR-1243 mimic后HepG2细胞中hnRNPA2B1蛋白表达、细胞增殖能力、划痕愈合率及cyclin D1、MMP-2蛋白表达均显著降低（均P<0.05）；而同时过表达 hnRNPA2B1 和 miR-1243 可逆转过表达 miR-1243 对 HepG2 细胞增殖、迁移的抑制作用。结论：miR-1243 通过靶向hnRNPA2B1表达调控肝癌HepG2细胞的增殖和迁移。

The effects of miR-1243 on the proliferation and migration of hepatocellular carcinoma HepG2 cells through targeted regulation of hnRNPA2B1 expression

Objective: To investigate the effects of miR-1243 on the proliferation and migration of hepatocellular carcinoma HepG2 cells by targeting and regulating the expression of heterogeneous nuclear ribonucleoprotein A2/B1 (hnRNPA2B1) and its molecular mechanisms. Methods: qPCR and WB methods were used to detect the levels of miR-1243 and hnRNPA2B1 mRNA and the expression of hnRNPA2B1, cyclin D1 and matrix metalloproteinase-2 (MMP-2) protein in 40 HCC tissues and adjacent tissues (surgically removed in Shouyi Hospital of Wuhan Third Hospital from January 2019 to August 2021), human normal liver QSG-7701 cells and hepatocellular carcinoma HepG2, Hep3b and Huh-7 cells. Dual-luciferase reporter assay was performed to verify the targeting relationship between miR-1243 and hnRNPA2B1. HepG2 cells were randomly divided into control group (not transfected), miR-NC group (transfected with miR-NC), miR-1243 mimic group (transfected with miR-1243 mimic), miR-1243 mimic+pcDNA3.1 group (transfected with miR-1243 mimic and pcDNA3.1), and miR-1243 mimic+pc-hnRNPA2B1 group (transfected with miR-1243 mimic and pc-hnRNPA2B1) and corresponding transfections were then carried out. CCK-8 method was performed to evaluate the proliferation ability of HepG2 cells in each group after transfection. Scratch healing test was performed to evaluate the migration ability of HepG2 cells, and Western blotting was performed to determine the protein expression levels of hnRNPA2B1, cyclin D1 and matrix metalloproteinase-2 (MMP-2). Results: Compared with adjacent tissues or human normal liver cells QSG-7701, the expression level of miR-1243 in liver cancer tissues and cells was significantly reduced, and the expression levels of hnRNPA2B1 mRNA and protein were significantly increased (all P<0.05). The results of Dual-luciferase experiments showed that miR-1243 and hnRNPA2B1 had a targeting relationship, and miR-1243 targeting negatively regulated the expression of hnRNPA2B1 protein. After miR-1243 mimic transfection,the expression of hnRNPA2B1 protein in HepG2 cells, proliferation ability of cells, scratch healing rate, and the expression of cyclin D1 and MMP-2 proteins were all significantly decreased (all P<0.05); the overexpression of both hnRNPA2B1 and miR-1243 could reverse the inhibitory effects of miR-1243 overexpression on the proliferation and migration of HepG2 cells. Conclusion: miR-1243 inhibits the proliferation and migration of liver cancer cells through targeting the expression of hnRNPA2B1.