糖氧剥夺/再灌注损伤对人正常肝L02细胞自噬水平的影响

目的 探讨糖氧剥夺/再灌注损伤(OGD/R)对人正常肝L02细胞自噬水平的影响.方法 体外培养人正常肝L02细胞,制备OGD/R模型,模拟临床的肝脏缺血再灌注损伤.分为5组:正常对照组、OGD 6 h/R 1、3、6、12 h组.随后在倒置显微镜下观察L02细胞形态;采用MTT比色法检测L02细胞的相对增殖情况;Western blot法检测自噬相关蛋白LC3、Beclin-1和p62的表达变化.结果 与正常对照组相比,随着再灌注时间的延长,OGD/R组处理L02细胞的形态损伤逐渐加剧,增殖活性逐渐降低,呈时间依赖性.自噬相关蛋白LC3和Beclin-1在OGD 6 h/R 1 h即有所增高,随着再灌注时间的延长,LC3的表达量逐渐增高并于再灌注6 h时大量增加,于再灌注12 h时达到最大值(P<0.01);Beclin-1蛋白的表达量随再灌注时间的延长逐渐增加于6 h和12 h急剧增加(P<0.01);p62蛋白于OGD 6 h/R 1 h时和3 h无明显变化,而在再灌注6 h时开始急剧增加,于12 h达到最大值(P<0.01).结论 OGD/R可诱导人正常肝L02细胞自噬水平上调,并且导致细胞的自噬性死亡.

The level of autophagy induced by oxygen glucose deprivation/reperfusion injury in L02 cell

Objective To explore the level of autophagy induced by oxygen glucose deprivation/reperfusion(OGD/R) injury in L02 cell.Methods L02 cells were cultured to establish the model of OGD/R injury and simulate clinical hepatic ischemia-reperfusion injury.The L02 cells were randomly divided into 5 groups : normal control group, oxygen-glucose deprivation 6 h/reperfusion 1,3,6,12 h group (OGD 6 h/R 1,3,6,12 h).Then observe the form changes of the L02 cells by optical microscope.The appreciation of the company's relative L02 cells was detected by MTT.The expression of autophagy related proteins such as Beclin-1, LC3 and p62 were evaluated by Western blot.Results Compared with the normal control group, the form damaged and the cells proliferation activity of L02 cells in the OGD/R group were gradually increased in a time-dependent manner.Compared with the normal control group, autophagy related proteins LC3 , Beclin-1 were increased at OGD 6 h/R 1 h.The expression of LC3 was gradually increased as the time went on and was increased gradually at OGD 6 h/R 6 h, reached a peak at OGD 6 h/R 12 h(P<0.01).The expression of Beclin-1 was gradually increased as the time went on and was increased gradually at OGD 6 h/R 6 h and OGD 6 h/R 12 h (P<0.01).The expression of p62 had no obvious change at OGD 6 h/R 1 h and OGD 6 h/R 3 h, began to increase sharply at OGD 6 h/R 6 h and reached a peak at OGD 6 h/R 12 h(P<0.01).Conclusion Our data suggests that oxygen-glucose deprivation/reperfusion may increase the level of autophagy and lead to autophagic cell death in L02 cell.