中国科学院昆明动物研究所机构知识库

非酒精性脂肪肝（Nonalcoholic fatty liver disease，NAFLD）是一种进行性疾病。它开始表现为肝脏的脂肪变性，进一步发展为非酒精性脂肪性肝炎（Nonalcoholic steatohepatitis，NASH），最终发展成为肝纤维化。但是，对于非酒精性脂肪肝是如何发生发展，我们仍然知之甚少。在本研究中，我们利用蛋白质组学策略去探究NAFLD的发病机制。根据喂养的食物不同，我们随机将树鼩分为以下三组：高脂高胆固醇组 (HFHC)，高脂低胆固醇组 (HFLC) 和对照组 (CON)。我们分别在3周、6周和10周牺牲动物，收集肝脏组织用于比较蛋白质组学分析。通过基于SILAC的定量蛋白质组分析，一共鉴定到4517个蛋白质，其中3589个蛋白质同时被定量。模型组与对应的对照组相比，777个蛋白质发生显著的差异变化。对这些差异表达的蛋白质KEGG 通路的分析表明，这些蛋白参与了多种生物过程，包括蛋白质折叠、内质网应激、细胞骨架、脂肪酸降解、氧化磷酸化等生物过程。其中，脂肪酸降解途径中的多个关键蛋白，在NAFLD表达都下降了，这说明线粒体里的脂肪酸的beta-氧化分解受到影响；而内质网应激应答途径被激活。将这些差异蛋白质进行蛋白质相互作用分析（protein?protein interaction，PPI），我们发现VIM，GRB2，S100A10和S100A11可能成为NAFLD，特别是发展成为肝脏炎症与纤维化的新的生物标志物。这些蛋白质的表达通过定量PCR，蛋白免疫印迹，以及免疫组化的分析，得到了进一步的验证。对于这些重要蛋白质的具体功能和精确的作用机制，我们需要进一步的研究。我们通过腺病毒介导的过表达，研究mS100A11在高脂高胆固醇诱导的NAFLD发展过程中的作用。实验表明，过表达mS100A11使肝脏脂肪积累，加剧高脂高胆固醇诱导的NAFLD的发生。进一步发现，S100A11无论在体内还是体外都能增加DGAT2的mRNA和蛋白表达。 DGAT2是甘油三脂合成的关键酶。因此，我们提出S100A11上调DGAT2的表达，促进甘油三脂的合成，从而加剧非酒精性脂肪肝的形成。总之，本研究不但提供了NAFLD发病发生过程中新的生物标志物（GRB2 、Vimentin和ACSL4），还首次揭示了S100A11在非酒精性脂肪肝发展中的重要作用。

Nonalcoholic fatty liver disease (NAFLD), a progressive disease that begins with simple steatosis, may develop to nonalcoholic steatohepatitis (NASH) and fibrosis. NAFLD is the most commonly chronic liver disease in the worldwide currently. In countries with a Western diet containing a high fat and/or cholesterol content, NASH and hepatic fibrosis are widely prevalent. However, the underlying mechanism of how NAFLD progresses to hepatic fibrosis is poorly understood.We took the advantage of proteomics to explore the underlying mechanism of NAFLD. In this study, based on feeding diets, tree shrews were randomly divided into three groups, high-fat high-cholesterol group (HFHC), high-fat low-cholesterol group (HFLC) and control group. We scarified tree shrews and took their liver tissue to perform proteomics at 3 w, 6 w, and 10 w, respectively. By SILAC (stable isotope labeling by amino acids in cell culture), 4517 proteins were identified totally, and 3589 proteins among them were quantified. Compared with the control group, the expression of 777 proteins was significantly changed in two diet induced groups. Interestingly, direct protein?protein interaction network analyses of these significantly changed proteins showed that vimentin (VIM), growth factor receptor-bound protein 2 (GRB2), S100 calcium binding protein (S100A10 and S100A11) form a complexes in a sub-network. These novel biomarkers are up-regulated with NAFLD progression to hepatic fibrosis. Their interaction networks clearly enhanced the interpretation and understanding of the pathogenesis of hepatic fibrosis. KEGG pathway analyses suggested that these proteins with altered expression involve into multiple vital physiological pathways, including regulation of protein folding, ER stress, actin cytoskeleton, fatty acid degradation, oxidative phosphorylation, and other cellular process. The fatty acid degradation pathway were down-regulated in NAFLD compared to control, while ER stress response pathway was activated. The increased protein levels of vimentin, GRB2, ACSL4, and S100A11 were validated by western blot analysis and immunohistochemical or indirect quantitive PCR.The precise mechanisms of some new candidate proteins are being investigated. We explore function of mS100A11 by adenovirus mediated over-expression in the progression of high-fat high-cholesterol diets induced mice NAFLD. Overexpresion of S100A11 promotes liver steatosis progression induced by HFHC diets. Overexpresion of S100A11 makes liver accumulate more lipids，especially triglyceride (TAG). Further, we found S100A11 up-regulated DGAT2 expression at mRNA level and protein level in vivo and in vitro. DGAT2 is a critical eznzyme in the TAG synthesis. It suggesstsed that S100A11 may up-regulate DGAT2 expression to increase TAG synthesis, to promote liver steatosis progression.In summary, this work not only provides novel biomarkers (GRB2 , Vimentin, S100A11) for diseases transition of NAFLD, but also firstly reveals S100A11 as a new player in the pathogenesis of NAFLD.