Zhou Xingang, Wang Peng, Ma Zhiyuan, Li Man, Teng Xiaoying, Sun Lei, Wan Gang, Li Yang, Guo Limei, Liu Honggang
Department of Pathology, Beijing Tongren Hospital, Capital Medical University, Beijing Key Laboratory of Head and Neck Molecular Diagnostic Pathology.
Departments of Pathology.
Appl Immunohistochem Mol Morphol. 2020 Feb;28(2):154-160. doi: 10.1097/PAI.0000000000000724.
Nonalcoholic steatohepatitis (NASH) has the potential to progress to fibrosis, cirrhosis, and hepatocellular carcinoma. Upregulation of sonic hedgehog (Shh) has been documented in development of NASH through sustained cell stress. At the same time, transforming growth factor-β1 (TGF-β1), which is a central element in fibrogenic reactions in various diseases and sites, has been reported to be associated with hepatic inflammation and fibrotic reaction. To explore crosstalk between Shh and TGF-β1 in the development and progression of NASH, we investigated the expression of both these proteins in 135 human specimens of NASH, 35 fatty liver specimens, 35 specimens of alcoholic steatohepatitis with immunohistochemistry. Shh protein was expressed in the cytoplasm of ballooned hepatocytes with an ubiquitin-like pattern. In addition, a few scattered apoptotic hepatocytes in the inflammatory foci showed homogeneous cytoplasmic Shh expression. TGF-β1 protein was observed mainly in the activated hepatic stellate cells (HSCs) which were located in the inflammatory foci surrounding ballooned hepatocytes. Moreover, the mRNA levels of both Shh and TGF-β1 in the liver biopsy specimens from NASH patients was significantly increased compared with those in fatty liver patients. Statistically, there was a significant association of the expressions of Shh and TGF-β1 proteins in NASH (r=0.6, P<0.05). In addition, increased expression of Shh protein significantly parallels the severity of hepatocellular ballooning, lobular, and portal inflammatory responses and progression of fibrosis in NASH patients. Moreover, we found that much HSCs transformed into myofibroblast-like phenotype and migrated downward to HepG2 hepatocellular carcinoma cells with overexpression of Shh by transwell assay. We also observed overexpression of proteins of Shh and TGF-β1 in cultured activated HSCs with confocal microscopy. These findings strongly suggest there is interplay between Shh and TGF-β1 in hepatic inflammatory reactions. Shh secreted through damaged hepatocytes may result in activation of TGF-β1 and subsequent transformation of HSCs, which together modulate the progression of human NASH.
非酒精性脂肪性肝炎（NASH）有可能进展为肝纤维化、肝硬化和肝细胞癌。已有文献记载，在NASH的发展过程中，通过持续的细胞应激，音猬因子（Shh）会上调。同时，转化生长因子-β1（TGF-β1）是各种疾病和部位纤维化反应的核心要素，据报道与肝脏炎症和纤维化反应有关。为了探究Shh和TGF-β1在NASH发生发展过程中的相互作用，我们采用免疫组织化学方法研究了这两种蛋白在135例人类NASH标本、35例脂肪肝标本和35例酒精性脂肪性肝炎标本中的表达情况。Shh蛋白以泛素样模式表达于气球样变肝细胞的细胞质中。此外，炎症灶中一些散在的凋亡肝细胞显示细胞质Shh均匀表达。TGF-β1蛋白主要在位于气球样变肝细胞周围炎症灶中的活化肝星状细胞（HSC）中观察到。此外，与脂肪肝患者相比，NASH患者肝活检标本中Shh和TGF-β1的mRNA水平均显著升高。统计学分析显示，NASH中Shh和TGF-β1蛋白的表达存在显著相关性（r=0.6，P<0.05）。此外，Shh蛋白表达的增加与NASH患者肝细胞气球样变、小叶和门管区炎症反应的严重程度以及纤维化进展显著平行。此外，我们通过Transwell实验发现，过表达Shh时，大量HSC转化为肌成纤维细胞样表型并向下迁移至HepG2肝癌细胞。我们还通过共聚焦显微镜观察到培养的活化HSC中Shh和TGF-β1蛋白的过表达。这些发现有力地表明，Shh和TGF-β1在肝脏炎症反应中存在相互作用。受损肝细胞分泌的Shh可能导致TGF-β1活化以及随后HSC的转化，二者共同调节人类NASH的进展。
