摘要
目的:探讨亚低温诱导的冷诱导RNA结合蛋白(CIRP)对肝L02细胞缺血再灌注损伤的保护作用及可能的机制。方法:取对数期生长的肝L02细胞株,分为正常对照组(NC)、常温缺血缺氧组(I)、亚低温缺血缺氧组(MH+I)、常温缺血再灌注组(IR)、亚低温缺血再灌注组(MH+IR),分别探讨亚低温对L02细胞缺血缺氧和再灌注损伤的作用。缺血缺氧组置于三气培养箱中培养12 h,再灌注组恢复正常培养条件继续培养2、4、6 h。将亚低温组的培养温度设置为32℃。分别检测各组肝酶(AST、ALT、LDH)的水平。构建pEGFP-C1-CIRP质粒,另取对数期生长的肝L02细胞,分为4组:正常对照组、常温缺血再灌注组(IR)、IR+pEGFP-C1-CIRP组、IR+pEGFP-C1组(阴性对照组)。检测每组AST、ALT、LDH,及各组细胞的活力、超氧化物歧化酶(SOD)活性和丙二醛(MDA)含量。结果:I组与MH+I组ALT、AST、LDH无显著差异。与IR组比较,MH+IR组的AST、ALT、LDH降低。与IR+pEGFP-C1组相比较,IR+pEGFP-C1-CIRP组AST、ALT、LDH均下降,细胞活力明显上升。IR+pEGFPC1-CIRP组SOD和MDA分别为(18.4±1.5) U/mg prot、(0.97±0.10) nmol/mg prot,与IR+pEGFP-C1相比差异均有统计学意义(P<0.05)。结论:亚低温降低L02细胞缺血缺氧阶段的损伤作用不明显,但对缺血后再灌注阶段的损伤有明确的保护作用。上调表达的CIRP蛋白能够显著降低缺血再灌注对L02细胞的氧化应激损伤。
Objective:To investigate the protective effects of the cold-inducible RNA binding protein(CIRP) induced by mild hypothermia from ischemia-reperfusion in liver L02 cells and the possible mechanisms.Methods:L02 cells were divided into normal control group(NC), ischemia group(I),ischemia with mild hypothermia group(MH+I), ischemia-reperfusion group(I/R), and ischemia-reperfusion with mild hypothermia group(MH+IR).The ischemia models of L02 cells were cultured in a tri-gas incubator.The IR models were performed after ischemia for 12 hours by restoring to normal culture conditions for 2, 4, and 6 hours, respectively.The temperature of mild hypothermia groups were set to 32 ℃.Then, L02 cells were divided into four groups:NC Group, I/R Group, I/R with pEGFP-C1-CIRP group(IR+pEGFP-C1-CIRP), and I/R with pEGFP-C1 group(IR+pEGFP-C1,negative control).The levels of serum liver enzymes(ALT, AST,and LDH)were detected.Cell vitality in different groups was measured by CCK-8 method.Superoxide dismutase(SOD)and malondialdehyde(MDA)were detected to determine the oxidative damages in different groups.Results:Compared with those in I group, the levels of AST, ALT, and LDH in MH+I group showed no statistically significant difference.Compared with IR group, AST, ALT, and LDH levels in MH+IR group were lower after 4 hours and 6 hours.Compared with IR+pEGFP-C1, AST, ALT, and LDH in IR+pEGFP-C1-CIRP group decreased and the cell vitality increased.The SOD activity and MDA level were(18.4±1.5) U/mg prot and(0.97±0.10) nmol/mg prot respectively in IR+pEGFP-C1-CIRP group, which showed a statistically significant difference when compared with IR+pEGFP-C1 group(P<0.05).Conclusion:The protective effect of mild hypothermia from ischemia injury in L02 cells is unremarkable, but there is clear protective effect in reperfusion cells, and the induced CIRP may play important roles in this process.
作者
夏志平
王伟
王彦峰
叶啟发
XIA Zhiping;WANG Wei;WANG Yanfeng;YE Qifa(The 3rd Xianya Hospital of Central South University&Research Center of National Health Ministry on Transplantation Medicine Engineering and Technology,Charigsha 410013,Hunan,China;Zhongnan Hospital of Wuhan University,Institute of Hepatobiliary Diseases of Wuhan University,Transplant Center of Wuhan University,&Hubei Key Laboratory of Medical Technology on Transplantation,Wuhan 430071,Hubei,China)
出处
《武汉大学学报（医学版）》
CAS
2021年第4期619-623,643,共6页
Medical Journal of Wuhan University
基金
国家自然科学基金青年基金项目(编号:81600587)。
作者简介
夏志平,男,1984—,医学博士,主治医师,主要从事器官移植的临床与基础研究,E-mail:xiazhiping@whu.edu.cn;通讯作者:叶啟发,男,1954—,医学博士,二级教授,博士生导师,主要从事肝胆外科与腹部器官移植研究,E-mail:yqf_china@163.com。
