探讨miR-4458及其靶向蛋白BZW2在肝细胞癌（肝癌）中的表达及其对肝癌细胞增殖、迁移和侵袭的影响。

本研究标本来源于2019年1月至2020年12月中国人民解放军联勤保障部队第九〇〇医院收治的247例肝癌患者空腹血清、肝癌组织标本（肝癌组），并收集同期200例健康志愿者空腹血清标本作为对照组。其中肝癌组男153例，女94例；年龄35~71岁，中位年龄49岁。对照组男120例，女80例；年龄30~70岁，中位年龄41岁。所有入组人员均签署知情同意书，符合医学伦理学规定。采用实时荧光定量PCR检测两组患者miR-4458和BZW2 mRNA表达量；Western blot检测肝癌组织和癌旁组织中BZW2蛋白表达情况，分析肝癌组织中BZW2 mRNA表达量与患者临床病理特征的关系；生物信息学预测miR-4458和BZW2的作用关系，并采用双荧光素酶检测验证。培养和转染人肝癌细胞株HepG2,采用CCK-8法、Transwell实验检测调控miR-4458表达对肝癌细胞增殖、迁移和侵袭能力的影响。

肝癌组患者血清miR-4458平均相对表达量为0.85±0.23，明显低于健康对照组的1.06±0.30（t=-7.886，P<0.05）。肝癌组织miR-4458相对表达量为0.77±0.27，明显低于癌旁组织的1.23±0.35（t=-12.646，P<0.05）。肝癌组血清BZW2 mRNA相对表达量为1.67±0.44，明显高于健康对照组的1.13±0.29（t=13.674，P<0.05）。肝癌组织BZW2 mRNA相对表达量为1.98±0.31，明显高于癌旁组织的1.27±0.30（t=16.537，P<0.05）。肝癌组织BZW2蛋白相对表达量为1.57±0.44，明显高于癌旁组织的0.83±0.38（t=16.483，P<0.05）。肝癌组织BZW2 mRNA相对表达量与肝癌患者肿瘤直径、肿瘤包膜、微血管侵犯和肿瘤分化相关（t=-8.208，-13.248，-7.981，7.145；P<0.05）。生物信息平台预测并荧光素酶验证miR-4458和BZW2互为靶向关系。HepG2细胞转染miR-4458抑制剂后BZW2蛋白相对表达量为1.95±0.24，明显高于转染miR-4458模拟物的0.43±0.11（LSD-t=3.384，P<0.05）。HepG2细胞转染miR-4458抑制剂后24、48、72 h增殖、迁移和侵袭能力上调。

肝癌患者中miR-4458低表达，BZW2高表达。miR-4458靶向结合BZW2后下调miR-4458表达可促进肝癌细胞增殖、迁移和侵袭，可能与肝癌患者不良预后相关。

To detect the expression levels of miR-4458 and its target protein BZW2 in hepatocellular carcinoma (HCC) and to evaluate its effects on the proliferation, migration and invasion of HCC cells.

The serum samples under fasting and HCC tissue samples (HCC group) were collected from 247 HCC patients admitted to No.900 Hospital of Joint Logistic Support Forces of PLA from January 2019 to December 2020, and serum samples under fasting from 200 healthy volunteers were collected in the control group. In the HCC group, 153 patients were male and 94 female, aged from 35 to 71 years, with a median age of 49 years. In the control group, 120 patients were male and 80 female, aged from 30 to 70 years, with a median age of 41 years. The informed consents of all patients were obtained and the local ethical committee approval was received. The expression levels of miR-4458 and BZW2 mRNA in two groups were detected by real-time fluorescent quantitative PCR. The expression level of BZW2 protein in HCC and adjacent tissues was measured by Western blot. The association between BZW2 mRNA expression in HCC tissues and clinicopathological features of HCC patients was analyzed. The interaction between miR-4458and BZW2 was predicted by bioinformatics analysis and validated by dual luciferase reporter assay. Human hepatoma cell line HepG2 was cultured and transfected. The effect of regulating miR-4458 expression on the proliferation, migration and invasion of HepG2 cells was evaluated by CCK-8 and Transwell assay.

The average relative expression level of serum miR-4458 in HCC patients was 0.85±0.23, significantly lower than 1.06±0.30 in the control group (t=-7.886, P<0.05). The relative expression level of miR-4458 in HCC tissues was 0.77±0.27, significantly lower than 1.23±0.35 in the adjacent tissues (t=-12.646, P<0.05). The relative expression of serum BZW2 mRNA in the HCC group was 1.67±0.44, significantly higher than 1.13±0.29 in the control group (t=13.674, P<0.05). The relative expression level of BZW2 mRNA in HCC tissues was 1.98±0.31, significantly higher than 1.27±0.30 in adjacent tissues (t=16.537, P<0.05). The relative expression of BZW2 protein in HCC tissues was 1.57±0.44, significantly higher than 0.83±0.38 in adjacent tissues (t=16.483, P<0.05). The relative expression of BZW2 mRNA in HCC tissues was correlated with tumor diameter, tumor encapsulation, microvascular invasion and tumor differentiation (t=-8.208, -13.248, -7.981, 7.145; P<0.05). The mutual targeting relationship between miR-4458 and BZW2 was predicted by bioinformatics analysis and validated by dual luciferase reporter assay. The relative expression of BZW2 protein was 1.95±0.24 in HepG2 cells transfected with miR-4458 inhibitor, significantly higher than 0.43±0.11 in those transfected with miR-4458 mimics (LSD-t=3.384, P<0.05). The proliferation, migration and invasion capabilities of HepG2 cells were up-regulated at 24 h, 48 h and 72 h after transfection with miR-4458 inhibitor.

The miR-4458 is lowly expressed, whereas BZW2 is highly expressed in HCC patients. Down-regulating miR-4458 expression through miR-4458 targeting BZW2 can promote the proliferation, migration and invasion of HCC cells, and it may be associated with poor prognosis of HCC patients.