NOD8对H2O2诱导的人肝细胞凋亡的影响  

Effects of NOD8 on H_2O_2-induced apoptosis in human hepatocytes

作　　者：郑媛媛[1] 杨文欣[1] 周晗[1] 胡巢凤[1] 

机构地区：[1]暨南大学医学院病理生理学系国家中医药管理局病理生理实验室,广东广州510632

出　　处：《中国病理生理杂志》2014年第11期2033-2037,共5页Chinese Journal of Pathophysiology

基　　金：广东省自然科学基金资助项目(No.S2012010008161);暨南大学“211工程”三期预研项目

摘　　要：目的：探讨NOD8对H2O2诱导的人肝细胞L02凋亡的影响。方法：p EGFP-C2及p EGFP-NOD8重组质粒经Jet PRIME介导转染L02细胞;用H2O2诱导细胞凋亡。实验分为p EGFP-C2组、p EGFP-C2＋H2O2组和p EGFP-NOD8＋H2O2组。采用MTT法检测细胞活性,Western blotting检测细胞NOD8的蛋白表达,Hoechst 33342染色检测细胞凋亡情况,流式细胞术检测细胞凋亡率,比色法检测细胞caspase-3活性。结果：通过MTT检测不同浓度（0.2-2 mmol/L）H2O2刺激6 h后的细胞活性,确定1 mmol/L H2O2为诱导细胞凋亡的剂量。Western blotting检测结果显示,转染p EGFP-NOD8质粒的细胞NOD8蛋白表达明显增加。Hoechst 33342染色法观察发现,p EGFPC2＋H2O2组有较多细胞出现强蓝色荧光细胞核,细胞凋亡较多,而p EGFP-NOD8＋H2O2组细胞凋亡明显减少。流式细胞术分析显示,p EGFP-C2＋H2O2组的细胞凋亡率明显升高,p EGFP-NOD8＋H2O2组的细胞凋亡率则显著下降。p EGFP-C2＋H2O2组细胞的caspase-3活性明显升高,而p EGFP-NOD8＋H2O2组细胞的caspase-3活性显著下降。结论：NOD8可抑制H2O2诱导的L02细胞凋亡,其作用机制可能与NOD8抑制细胞的caspase-3活性有关。AIM： To observe the effects of NOD8 on H2O2-induced apoptosis in human L02 hepatocytes.METHODS： p EGFP-C2 and p EGFP-NOD8 plasmids were transfected into L02 cells by Jet PRIME,respectively. The apoptosis of these transfected cells was induced by H2O2. The cells were divided into p EGFP-C2 group,p EGFP-C2 ＋ H2O2 group and p EGFP-NOD8 ＋ H2O2 group. MTT assay was used to detect the cell viability. NOD8 protein expression was determined by Western blotting. The cell apoptosis was observed by Hoechst 33342 staining and apoptotic rate was evaluated by flow cytometry. The caspase-3 activity was analyzed by a colorimetric method. RESULTS： L02 cells were stimulated by H2O2 at concentrations of 0. 2 - 2 mmol / L for 6 h,and H2O2 at concentration of 1 mmol / L was chosen to induce apoptosis determined by MTT assay. The protein expression of NOD8 significantly increased in the cells transfected with p EGFPNOD8 plasmid. More cellular nucleus with strong blue fluorescence by Hoechst 33342 staining in p EGFP-C2 ＋ H2O2 group were observed,indicating that apoptosis was increased,while the apoptosis in p EGFP-NOD8 ＋ H2O2 group significantly reduced. The apoptotic rate in p EGFP-C2 ＋ H2O2 group was obviously increased,whereas that in p EGFP-NOD8 ＋ H2O2 group was significantly decreased. The caspase-3 activity in p EGFP-C2 ＋ H2O2 group was remarkably increased. By contrast,the activity of caspase-3 was significantly reduced in p EGFP-NOD8 ＋ H2O2 group. CONCLUSION： NOD8 inhibits H2O2-induced apoptosis in L02 cells and the mechanism may be related to inhibition of caspase-3 activity.

关 键 词：L02细胞 过氧化氢 NOD8 半胱氨酸蛋白酶-3 

分 类 号：R363[医药卫生—病理学]