Alcohol consumption is a critical risk factor for hepatic pathogenesis, including alcoholic liver diseases (ALD), but implications of alcohol-induced dysregulation of microRNA (miRNA) in ALD pathogenesis are not completely understood. In the present study, C57BL/6J male mice were treated with saline (CON; oral gavage; n = 8) or alcohol (EtOH; 3 g/kg body weight; oral gavage; n = 8) for 7 days. A total of 599 miRNAs and 158 key mRNAs related to fatty liver and hepatotoxicity pathways were assessed in mice liver tissues. The mRNA expression datasets were then utilized to predict interactions with miRNAs that were changed by alcohol consumption. Predicted miRNA-mRNA interactions were validated using in vitro miRNA transfection experiments. The results showed that let-7a was significantly decreased in the EtOH group and Rb1 mRNA was predicted as a target gene. This was further supported by an inverse correlation of RB1 and let-7a expression in mice liver tissue. Additionally, key protein expressions involved in RB1-apoptosis axis [i.e., p73, cleaved CASP-3 (cCASP-3), and cCASP-7] showed a trend of increase in the EtOH mice; this was also confirmed by capase-3 enzyme activity and terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling assay in livers of mice that had consumed alcohol. In line with our in vivo observations, alcohol treatment suppressed the let-7a expression and subsequently upregulated p73, cCASP-3, and cCASP-7 protein expressions in mice hepatocytes. Additional proteins in the apoptosis regulatory pathway (i.e., MDM2-p53 axis) were significantly changed in response to let-7a suppression in the cells. Taken together, the current study provides mechanistic evidence that alcohol consumption-induced let-7a suppression results in the upregulation of RB1, thereby promoting hepatic apoptosis through induction of pro-apoptotic proteins (e.g., p73), and by, at least in part, preventing MDM2-mediated p53 degradation.

中文翻译：

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急性饮酒诱导的let-7a抑制通过调节小鼠中的Rb1加剧肝细胞凋亡。

饮酒是包括酒精性肝病（ALD）在内的肝病发病机制的关键危险因素，但是酒精引起的微小RNA（miRNA）失调在ALD发病机制中的意义尚不完全清楚。在本研究中，C57BL / 6J雄性小鼠用生理盐水（CON；口服管； n = 8）或酒精（EtOH； 3 g / kg体重；口服管； n = 8）治疗7天。在小鼠肝组织中评估了与脂肪肝和肝毒性途径相关的总共599个miRNA和158个关键mRNA。然后将mRNA表达数据集用于预测与因饮酒而改变的miRNA的相互作用。使用体外miRNA转染实验验证了预测的miRNA-mRNA相互作用。结果表明，EtOH组中let-7a显着减少，Rb1 mRNA被预测为靶基因。小鼠肝脏组织中RB1和let-7a表达的负相关进一步支持了这一点。此外，参与RB1细胞凋亡轴的关键蛋白表达（即p73，CASP-3（cCASP-3）和cCASP-7裂解）在EtOH小鼠中有增加的趋势。食用酒精的小鼠肝脏中的capase-3酶活性和末端脱氧核苷酸转移酶介导的dUTP缺口末端标记试验也证实了这一点。与我们的体内观察一致，酒精处理抑制了小鼠肝细胞中let-7a的表达，并随后上调了p73，cCASP-3和cCASP-7的蛋白表达。凋亡调节途径中的其他蛋白质（即，MDM2-p53轴）响应let-7a抑制在细胞中发生了显着变化。综上所述，当前的研究提供了机制性证据，表明饮酒诱导的let-7a抑制导致RB1的上调，从而通过诱导促凋亡蛋白（例如p73）促进肝细胞凋亡，并且至少部分地通过诱导凋亡。防止MDM2介导的p53降解。