蛋白质组学技术筛选大鼠肝脏大部切除后肝再生相关差异表达蛋白

Proteomics technique in screening of differential proteins for liver regeneration after partial hepatectomy in rats

目的:应用蛋白质组学技术观察大鼠肝脏大部分切除术后蛋白表达情况,筛选肝脏切除后肝再生相关的差异表达蛋白。方法:大鼠随机分为肝大部切除组(n=35)和假手术对照组(n=5)。肝大部切除组大鼠无菌条件下切除肝左外叶、左中叶和中叶共约70%肝脏;假手术组大鼠仅开腹,不行肝大部切除。肝大部切除术后不同时间点(2、12、24、36、48、72、168h)各处死5只大鼠,取右叶肝组织,提取大鼠肝脏总蛋白,以假手术组为对照,进行二维电泳和质谱分析,筛选差异表达蛋白,并对其中差异显著蛋白进行Western印迹鉴定。结果:电泳图谱显示肝脏70%切除术后2h差异表达蛋白点开始增多,36h达到高峰;共筛选出78个差异表达蛋白点,质谱技术鉴定出其中35个有意义的蛋白点。35个差异蛋白根据动态变化趋势可分为5类:3类丰度上调蛋白和2类丰度下调蛋白(各自间上调或下调时间和幅度均有所差异);这些蛋白主要涉及以下代谢途径:氧化应激反应、急性期反应蛋白、脂类代谢蛋白、能量代谢的酶类、信号转导、神经递质降解等,部分蛋白功能未知。Western印迹鉴定证实其中的prohibitin蛋白在肝大部切除术后2h开始上调,36h达高峰,48h后趋于正常水平。结论:多种信号和代谢途径参与肝脏大部切除后的肝脏再生过程。

Objective：To screen for the differentially expressed proteins during rat liver regeneration after partial hepatectomy（PH） by proteomics technique. Methods： Healthy male SD rats were randomly divided into 2 groups： hepatectomy group and sham operated group. The hepatectomy model was produced by 70% PH（n= 35） and the sham operated rats（n= 5）underwent the same surgical protocol without hepatectomy. Rats were executed at 2, 12, 24, 36, 48, 72 and 168 h after partial hepatectomy （each time 5 rats） and the right lobes were harvested. The total protein was extracted and analyzed by two-dimensional gel electrophoresis and mass spectrometric analysis. The differential proteins were then analyzed by Western blotting. Results： The spots of differential protein began to rise at 2 h after PH and peaked at 36 h after PH. A total of 78 protein spots were identified and 35 significant protein spots were found by mass spectrometric analysis. The 35 protein sports fell into 5 types according to their dynamic changes： 3 up-regulated and 2 down-regulated （with different regulation time periods and amplitudes） ; and their functions involved oxidative stress response, acute reaction, lipid and energy metabolism, intracellular signaling transduction, cell proliferation, etc. , with some having unknown functions. Western blotting analysis showed that the prohibitin protein began to increase 2 h after PH and decreased to the normal level after 48 h. Conclusion.. It is indicated that many proteins and signal transduction pathways participate in the liver regeneration after partial hepatectomy.