摘要
目的探讨铁死亡在大鼠脂肪肝缺血再灌注损伤中的表达及作用。方法随机数字表法将36只7~8周龄清洁级雄性SD大鼠分为6组:普通饲料(ND)和高脂饲料(HFD)分别喂养的假手术(Sham)、肝脏缺血再灌注损伤(HIRI)和HIRI联合铁死亡抑制剂(Fer-1)治疗(HIRI+Fer-1)组,每组6只。油红O和苏木精-伊红(HE)染色观察肝脏脂变和病理损伤。生化分析仪检测谷丙转氨酶(ALT)和谷草转氨酶(AST)水平。比色法检测肝脏丙二醛(MDA)、还原型谷胱甘肽(GSH)、Fe2+含量。实时定量反转录聚合酶链反应(RT-qPCR)检测前列腺素内过氧化物合酶2(Ptgs2)信使RNA(mRNA)表达水平。蛋白质印迹法检测谷胱甘肽过氧化物酶4(GPX4)、铁蛋白重链(FTH1)和转铁蛋白受体1(TFR1)的蛋白表达水平。两组间比较采用t检验。结果高脂饲料连续喂养20周成功建立重度(>60%)混合大泡型脂肪肝。肝功能及HE检测结果显示,ND和HFD大鼠的HIRI组损伤均重于各自的Sham和HIRI+Fer-1组,同时在HIRI组中,HFD大鼠ALT[(1723.00±336.70)U/L]和AST[(1536.00±361.50)U/L]均高于ND[ALT:(804.10±113.40)U/L、AST:(1001.00±118.20)U/L,t=6.339、3.447,P<0.05]。铁死亡指标显示,ND和HFD大鼠的HIRI组铁死亡发生均高于各自的Sham和HIRI+Fer-1组,且在HIRI组中,HFD大鼠肝脏Fe^(2+)[(200.50±13.04)μmol/L]和MDA[(346.30±56.00)nmol/g]均高于ND[Fe^(2+):(161.50±16.41)μmol/L和MDA:(200.80±55.70)nmol/g,t=4.557、4.512,P<0.05],GSH低于ND(1.52±0.09)μg/mg比(2.64±0.41)μg/mg,t=6.512,P<0.05;Ptgs2的mRNA相对表达(14.65±1.22)高于ND(9.18±1.27,t=7.575,P<0.05);GPX4(0.36±0.03)和FTH1(0.38±0.04)蛋白表达均低于ND(GPX4:0.63±0.02和FTH1:0.62±0.03,t=11.600、8.507,P<0.05),TFR1表达(1.76±0.08)高于ND(1.47±0.07,t=4.354,P<0.05),差异均有统计学意义。结论HFD大鼠较ND大鼠对HIRI耐受性显著降低,其机制可能与脂肪肝铁代谢异常、铁离子增多致铁死亡发生有关。
Objective To investigate the expression and effect of ferroptosis on ischemia-reperfusion injury of fatty liver in rats.Methods Totally,367-8-week-old male Sprague Dawley rats were divided into 6 groups using random number table method:sham operation(Sham),hepatic ischemic reperfusion injury(HIRI)and HIRI+Ferrostatin-1(HIRI+Fer-1)group for normal diet(ND)and high-fat diet(HFD)rats,respectively.Oil Red O staining and hematoxylin-eosin(HE)staining were used to observe the lipidosis and pathological damage of liver.Automatic biochemical analyzer wass used to determine alanine aminotransferase(ALT)and aspartate aminotransferase(AST)levels.Colorimetric method was used to measure the content of malondialdehyde(MDA),reduced glutathione(GSH)and Fe2+in liver.Real-time quantitative reverse transcription polymerase chain reaction(RT-qPCR)was used to detect the mRNA expression of prostaglandin-endoperoxide synthase 2(Ptgs2)in liver.Standard Western blotting was used to detect the protein expression of glutathione peroxidase(GPX4),ferritin heavy chain(FTH1)and transferrin receptor 1(TFR1)in liver.The t test was used for comparison between two groups.Results HE and oil red O staining showed that severe(>60%)mixed macrovesicular hepatic steatosis was successfully established after 20 weeks of continuous HFD feeding.Liver function and HE showed that the injury of HIRI group in ND and HFD rats was significantly worse than that in Sham and HIRI+Fer-1 groups,and in HIRI group,ALT[(1723.00±336.70)U/L]and AST[(1536.00±361.50)U/L]in HFD rats were higher than in ND rats[ALT:(804.10±113.40)U/L and AST:(1001.00±118.20)U/L,t=6.339,3.447,P<0.05].The ferroptosis index showed that ferroptosis in the HIRI group of ND and HFD rats was more significantly than that in Sham and HIRI+Fer-1 groups,and in the HIRI group,liver Fe^(2+)[(200.50±13.04)μmol/L]and MDA[(346.30±56.00)nmol/g]in HFD rats were higher than in ND rats[Fe^(2+):(161.50±16.41)μmol/L and MDA:(200.80±55.70)nmol/g,t=4.557,4.512,P<0.05],and GSH[(1.52±0.09)μg/mg]lower[(2.64�
作者
李想
吴龙龙
田轩
曹欢
袁梦淑
宋红丽
Li Xiang;Wu Longlong;Tian Xuan;Cao Huan;Yuan Mengshu;Song Hongli(The First Central Clinical College of Tianjin Medical University,Tianjin 300192,China;The Medical College,Nankai University,Tianjin 300074,China;Department of Organ Transplantation,Tianjin First Central Hospital,Tianjin 300192,China)
出处
《中华实验外科杂志》
CAS
北大核心
2022年第4期649-652,共4页
Chinese Journal of Experimental Surgery
基金
国家自然科学基金(82070639、81670574)。
关键词
铁死亡
脂肪肝
肝脏缺血再灌注损伤
Ferroptosis
Fatty liver
Hepatic ischemia reperfusion injury
