Xiao Qi, Ye Qi Fa, Wang Wei, Fu Bi Qi, Xia Zhi Ping, Liu Zhong Zhong, Zhang Xing Jian, Wang Yan Feng
Department of Transplant Surgery, The Third Xiangya Hospital, Central South University, Changsha, 410013, PR China.
Department of Transplant Surgery, The Third Xiangya Hospital, Central South University, Changsha, 410013, PR China; Hubei Key Laboratory of Medical Technology on Transplantation, Zhongnan Hospital of Wuhan University, Institute of Hepatobiliary Diseases of Wuhan University, Transplant Center of Wuhan University, Wuhan, 430071, PR China.
Cryobiology. 2017 Apr;75:100-105. doi: 10.1016/j.cryobiol.2017.01.005. Epub 2017 Jan 14.
Mild hypothermia has been well known as an effective way to reduce ischemia reperfusion injury (IRI), while the mechanisms are still unclear. More and more evidences have indicated that miRNAs should been involved in the regulation of IRI and expecially some miRNAs have shown temp-responsiveness for temperature variation. Therefore, the role of miR-122 in mild hypothermia pretreatment after IRI was investigated.
We established a LO2 cell anoxia-reoxygenation injury model to simulate liver IRI. Five groups of differently pretreated L02 cells were studied. ALT, AST and LDH as well as cell viability were measured. Flow cytometric analysis was used to evaluate the apoptosis. The expression of miR-122 was quantified by qRT-PCR. Insulin-like growth factor 1 receptor (IGF-1R), protein kinase B (p-AKT), AKT, forkhead box O3a (p-FOXO3a) and Caspase3 were examined using western blot analysis.
We found that mild hypothermia pretreatment could reduce the hepatocellular injury and induce a significant down-regulation in miR-122 expression after IRI. However, those effects of protection were attenuated by overexpressed miR-122 blockade. We further demonstrated that down-regulation of miR-122 promoted IGF-1R translation and AKT activity, suppressed FOXO3a activity and Caspase3 expression after mild hypothermia pretreatment, which was abrogated by miR-122 mimic.
Our data clearly demonstrate that mild hypothermia pretreatment can down-regulate miR-122 to protect hepatocytes against IRI through activation IGF-1R/AKT signaling pathway and inhibit cells apoptosis.
轻度低温作为减轻缺血再灌注损伤（IRI）的一种有效方法已广为人知，但其机制仍不清楚。越来越多的证据表明，微小RNA（miRNA）参与了IRI的调控，尤其是一些miRNA对温度变化具有温度响应性。因此，本研究探讨了miR-122在IRI后轻度低温预处理中的作用。
我们建立了LO2细胞缺氧复氧损伤模型以模拟肝脏IRI。研究了五组不同预处理的L02细胞。检测了谷丙转氨酶（ALT）、谷草转氨酶（AST）、乳酸脱氢酶（LDH）以及细胞活力。采用流式细胞术分析评估细胞凋亡。通过qRT-PCR定量检测miR-122的表达。使用蛋白质印迹分析检测胰岛素样生长因子1受体（IGF-1R）、蛋白激酶B（p-AKT）、AKT、叉头框O3a（p-FOXO3a）和半胱天冬酶3（Caspase3）。
我们发现轻度低温预处理可减轻肝细胞损伤，并导致IRI后miR-122表达显著下调。然而，过表达的miR-122阻断减弱了这些保护作用。我们进一步证明，轻度低温预处理后miR-122的下调促进了IGF-1R的翻译和AKT活性，抑制了FOXO3a活性和Caspase3表达，而miR-122模拟物可消除这些作用。
我们的数据清楚地表明，轻度低温预处理可通过激活IGF-1R/AKT信号通路和抑制细胞凋亡来下调miR-122，从而保护肝细胞免受IRI损伤。
