Institute of Liver Diseases, Shuguang Hospital, Shanghai University of Traditional Chinese Medicine, Shanghai 201203, China ; Taizhou Municipal Hospital, Taizhou, Zhejiang 318000, China.
Evid Based Complement Alternat Med. 2013;2013:972863. doi: 10.1155/2013/972863. Epub 2013 Jan 29.
Hepatic fibrosis is a common pathological process of chronic liver diseases and would lead to cirrhosis, and Fuzheng Huayu (FZHY) is an effective Chinese herbal product against liver fibrosis. This study observes FZHY influence on proteome of fibrotic liver with differential proteomic approach and aims to understand FZHY multiple action mechanisms on liver fibrosis. The liver fibrosis models were induced with intraperitoneal injection of dimethylnitrosamine for 4 weeks in rats and divided into model control (model) and FZHY-treated (FZHY) groups, while normal rats were used as normal control (normal). After model establishment, rats in FZHY groups were administered 4 g/kg wt of FZHY for 4 weeks, and normal and model groups were given the same volume of saline. The liver proteins in the above 3 groups were separated by two-dimensional gel electrophoresis (2-DE), the differentially expressed spots were analyzed and compared between normal and model or model and FZHY groups, and then the proteins were identified with mass spectrum analysis and validated partially with western blot and real-time PCR. 1000~1200 spots were displayed on each 2D gel, and a total of 61 protein spots were found with significant intensity difference between normal control or FZHY and model control. 23 most obviously differential spots were excised, and in-gel digestion and 21 peptide mass fingerprints (PMF) were obtained with MALDI-TOF MS analysis, and 14 proteins were identified through protein database searching. Among 14 differentially expressed proteins, 8 proteins in normal and FZHY groups had the same tendency of differential expression compared with the ones in model group. And one of them, vimentin, was validated by western blot and real-time PCR analyses. Our study reveals 12 proteins responsible for fibrogenesis induced by DMN in rats, and among them, 8 proteins in fibrotic liver were regulated by FZHY, including aldehyde dehydrogenase, vimentin isoform (CRA_b), gamma-actin, vimentin, fructose-bisphosphate aldolase B, aldo-keto reductase, S-adenosylhomocysteine hydrolase isoform, and HSP90. It indicates that the action mechanism of FZHY antiliver fibrosis may be associated with modulation of proteins associated with metabolism and stress response, as well as myofibroblast activation. The study provides new insights and data for exploring the liver fibrogenesis pathophysiology and FZHY action mechanism against liver fibrosis.
肝纤维化是慢性肝病的常见病理过程，可导致肝硬化，扶正化瘀（FZHY）是一种有效的抗肝纤维化中药。本研究采用差异蛋白质组学方法观察 FZHY 对纤维性肝蛋白质组的影响，旨在了解 FZHY 对肝纤维化的多种作用机制。采用腹腔注射二甲基亚硝胺 4 周诱导大鼠肝纤维化模型，分为模型对照组（模型）和 FZHY 治疗组（FZHY），正常大鼠为正常对照组（正常）。模型建立后，FZHY 组给予 FZHY4g/kg 体重 4 周，正常组和模型组给予等量生理盐水。采用二维凝胶电泳（2-DE）分离上述 3 组肝蛋白，比较正常组与模型组或模型组与 FZHY 组间差异表达点，然后采用质谱分析进行鉴定，部分用 Western blot 和实时 PCR 进行验证。每张 2-DE 显示 1000~1200 个斑点，正常对照组或 FZHY 与模型对照组之间共发现 61 个蛋白斑点差异明显。选择 23 个差异最明显的斑点进行胶内酶切，MALDI-TOF-MS 分析得到 21 个肽质量指纹图谱（PMF），通过蛋白数据库搜索鉴定出 14 种蛋白。在 14 种差异表达蛋白中，有 8 种蛋白在正常组和 FZHY 组与模型组相比具有相同的差异表达趋势。其中，波形蛋白通过 Western blot 和实时 PCR 分析得到验证。本研究揭示了 12 种 DMN 诱导大鼠肝纤维化形成的蛋白，其中纤维化肝中有 8 种蛋白受 FZHY 调节，包括醛脱氢酶、波形蛋白同工型（CRA_b）、γ-肌动蛋白、波形蛋白、果糖-1,6-二磷酸醛缩酶 B、醛酮还原酶、S-腺苷同型半胱氨酸水解酶同工型和 HSP90。表明 FZHY 抗肝纤维化的作用机制可能与代谢和应激反应相关蛋白以及肌成纤维细胞激活的调节有关。本研究为探索肝纤维化发病机制和 FZHY 抗肝纤维化作用机制提供了新的见解和数据。
