亚低温在抑制小鼠脓毒症相关急性肾损伤中铁死亡的作用探究

Effect of mild hypothermia on ferroptosis in mice with sepsis-related acute kidney injury

目的:探讨亚低温是否通过激活核因子-E2相关因子2(Nrf2)抑制小鼠脓毒症相关急性肾损伤(S-AKI)中的铁死亡,从而发挥保护作用。方法:选择15只SPF级C57BL/6小鼠随机分为假手术组(S组)、脓毒症常温组(N组)、脓毒症亚低温组(H组),每组5只。盲肠结扎穿孔术建立小鼠脓毒症模型,分别对3组进行相应处理。苏木精—伊红(HE)染色观察小鼠肾组织损伤情况,酶标仪法测定血清尿素氮(BUN)及肌酐(Cr)水平、肾组织还原型谷胱甘肽(GSH)、丙二醛(MDA)含量,蛋白免疫印迹法(western blotting)和实时荧光定量PCR(RT-qPCR)测定肾组织酰基辅酶A合成酶长链家族成员4(ACSL4)、谷胱甘肽过氧化酶4(GPX4)、Nrf2及其下游因子血红素氧合酶1(HO-1)的蛋白及mRNA表达水平。结果:与S组比较,N组和H组肾组织病理有不同程度损伤,BUN、Cr水平升高,GSH含量减少,MDA含量增加,ACSL4蛋白及mRNA表达升高、GPX4蛋白及mRNA表达降低、Nrf2和HO-1蛋白及mRNA表达显著升高(均P<0.05);H组与N组比较,病理损伤减轻,BUN、Cr水平下降,GSH含量增加,MDA含量减少,ACSL4蛋白及mRNA水平表达降低,GPX4蛋白及mRNA表达升高,Nrf2和HO-1蛋白及mRNA表达显著升高(均P<0.05)。结论:亚低温可通过激活Nrf2抑制铁死亡,在S-AKI中发挥保护作用。

Objective:To investigate whether mild hypothermia could inhibit ferroptosis in mice with sepsis-associated acute kidney injury(S-AKI)by activating NF-E2-related factor 2(Nrf2),thus playing a protective role.Methods:15 SPF C57BL/6 mice were randomly divided into sham operation group(group S),sepsis normal temperature group(group N)and sepsis mild hypothermia group(group H),with 5 mice in each group.Cecal ligation and puncture was used to establish a mouse model of sepsis and corresponding treatment was carried out in the three groups.Hematoxylin-eosin(HE)staining was used to observe the damage of kidney tissue.The levels of urea nitrogen(BUN)and creatinine(Cr)in serum,the contents of reduced glutathione(GSH)and malondialdehyde(MDA)in kidney tissue were determined by enzyme-labeled assay.Western blotting and real-time fluorescence quantitative polymerase chain reaction(RT-qPCR)were used to determine the protein and mRNA expression levels of acyl CoA synthetase long chain family member 4(ACSL4),glutathione peroxidase 4(GPX4),Nrf2 and its downstream factor heme oxygenase 1(HO-1)in kidney tissue.Results:Compared with group S,there were different degrees in the pathological changes of kidney tissue in N and H groups,the levels of BUN and Cr in serum increased,GSH content decreased,MDA content increased,the expression of ACSL4 protein and mRNA increased,the expression of GPX4 protein and mRNA decreased,and the expressions of NRF2 and HO-1 protein and mRNA significantly increased(all P<0.05).Compared with group N,group H showed reduced pathological damage,decreased levels of BUN and Cr,increased GSH content,decreased MDA content,decreased expression of ACSL4 protein and mRNA,increased expression of GPX4 protein and mRNA,and significantly increased expressions of Nrf2 and HO-1 protein and mRNA(all P<0.05).Conclusion:Mild hypothermia can inhibit ferroptosis by activating Nrf2 and play a protective role in S-AKI.