Shen Ding-zhu, Tao Qing, Du Jin-xing, Ding Sai-dan, Chen Gao-feng, Hu Yi-yang, Liu Ping
Institute of Liver Diseases, Shuguang Hospital, Shanghai University of Traditional Chinese Medicine, Shanghai 201203, China.
Zhong Xi Yi Jie He Xue Bao. 2010 Feb;8(2):158-67. doi: 10.3736/jcim20100211.
To investigate the effects of Yiguanjian Decoction, a compound traditional Chinese herbal medicine, on rats with cirrhosis based on the method of differential proteomics.
Wistar male rats (n=48) were randomly divided into normal control group (n=12) and model-making group (n=36). Rat cirrhosis model was established by intraperitoneal injection of 50% carbon tetrachloride (CCl4) plus olive oil solution (1 mL/kg, twice weekly for 9 weeks). After 3- and 6-week injection, 6 rats each time were sacrificed for dynamic observation before medicine intervention, and the 24 remained rats were randomly divided into untreated group (n=12) and Yiguanjian Decoction group (n=12) at the first day of the 7th week. All animals were sacrificed by the end of the 9th week, and total protein of liver tissue was isolated by two-dimensional gel electrophoresis (2-DE); some differentially expressed protein spots were analyzed and identified by matrix-assisted laser desorption/ionization two-stage time-of-flight mass spectrometry (MALDI-TOF/TOF-MS) and database querying. Protein expressions of Cu/Zn superoxide dismutase (Cu/Zn SOD) and DJ-1 were validated by Western blot and immunohistochemical methods.
2-DE maps with high resolution and good repeatability were obtained. In all 50 protein spots identified by MALDI-TOF/TOF-MS and database querying, there were 5 protein spots related to oxidative stress named Cu/Zn SOD, DJ-1, glutathione synthetase, glutathione S-transferase Yb-1 subunit and aldo-keto reductase family 7, A2 respectively. Compared with the normal control group, expressions of Cu/Zn SOD, DJ-1, glutathione S-transferase Yb-1 subunit and aldo-keto reductase family 7, A2 in the untreated group were decreased significantly. Expressions of Cu/Zn SOD and aldo-keto reductase family 7, A2 were decreased time-dependently. Compared with the untreated group in 9th week, protein expressions of Cu/Zn SOD, DJ-1, glutathione S-transferase Yb-1 subunit and aldo-keto reductase family 7, A2 in the Yiguanjian Decoction groups were increased significantly while expression of glutathione synthetase was decreased notably. Western blot and immunohistochemical results of Cu/Zn SOD and DJ-1 expressions coincided with proteomics results.
Anti-oxidative depression is a key pathological change of cirrhosis induced by CCl4 in rats, and increasing expression of proteins related to anti-oxidative stress may be a major mechanism of Yiguanjian Decoction in treating cirrhosis induced by CCl4 effectively.
基于差异蛋白质组学方法，探讨中药复方一贯煎对肝硬化大鼠的影响。
将48只雄性Wistar大鼠随机分为正常对照组（n = 12）和造模组（n = 36）。通过腹腔注射50%四氯化碳（CCl4）加橄榄油溶液（1 mL/kg，每周2次，共9周）建立大鼠肝硬化模型。注射3周和6周后，每次处死6只大鼠用于药物干预前的动态观察，剩余24只大鼠在第7周第一天随机分为未治疗组（n = 12）和一贯煎组（n = 12）。第9周结束时处死所有动物，采用双向凝胶电泳（2-DE）分离肝组织总蛋白；部分差异表达蛋白点通过基质辅助激光解吸/电离二级飞行时间质谱（MALDI-TOF/TOF-MS）和数据库查询进行分析鉴定。采用蛋白质免疫印迹法和免疫组织化学方法验证铜/锌超氧化物歧化酶（Cu/Zn SOD）和DJ-1的蛋白表达。
获得了分辨率高、重复性好的2-DE图谱。通过MALDI-TOF/TOF-MS和数据库查询共鉴定出50个蛋白点，其中与氧化应激相关的蛋白点有5个，分别为Cu/Zn SOD、DJ-1、谷胱甘肽合成酶、谷胱甘肽S-转移酶Yb-1亚基和醛糖酮还原酶家族7成员A2。与正常对照组相比，未治疗组中Cu/Zn SOD、DJ-1、谷胱甘肽S-转移酶Yb-1亚基和醛糖酮还原酶家族7成员A2的表达显著降低。Cu/Zn SOD和醛糖酮还原酶家族7成员A2的表达呈时间依赖性降低。与第9周未治疗组相比，一贯煎组中Cu/Zn SOD、DJ-
