Molecular Hepatology - Alcohol Associated Diseases, II. Medical Clinic, Medical Faculty Mannheim, Heidelberg University, Theodor-Kutzer-Ufer 1-3, 68167 Mannheim, Germany.
J Hepatol. 2011 Aug;55(2):369-78. doi: 10.1016/j.jhep.2010.11.027. Epub 2010 Dec 22.
BACKGROUND & AIMS: In injured liver, TGF-β affects all hepatic cell types and participates in wound healing and fibrogenesis. TGF-β downstream signaling is highly complex and cell type dependent, involving Smad and non-Smad signaling cascades thus requiring tight regulation. Endocytosis has gained relevance as important mechanism to control signaling initiation and termination. In this study, we investigated endocytic mechanisms for TGF-β mediated Smad and non-Smad signaling in hepatocytes.
Endocytosis in hepatocytes was elucidated using chemical inhibitors, RNAi, viral gene transfer and caveolin-1-/- mice. TGF-β signaling was monitored by Western blot, reporter assays and gene expression analysis.
In hepatocytes, Smad activation is to a large degree accomplished AP-2 complex dependent on the hepatocyte surface without the necessity of clathrin coated pit formation or an endocytic step. In contrast, non-Smad/AKT pathway activation required functional dynamin mediated endocytosis and the presence of caveolin-1, an essential protein for caveolae formation. Furthermore, these two TGF-β signaling initiation platforms discriminate distinct signaling routes that integrate at the transcriptional level as shown for TGF-β target genes, Id1, Smad7, and CTGF. Endocytosis inhibition increased canonical Smad signaling and culminated in a superinduction of Id1 and Smad7 expression, whereas caveolin-1 mediated AKT pathway activation was required for maximal CTGF induction.
Endocytosis is critical for TGF-β signaling regulation in hepatocytes and determines gene expression signature and (patho)physiological outcome.
在受损的肝脏中，TGF-β 影响所有的肝细胞类型，并参与伤口愈合和纤维化形成。TGF-β 的下游信号转导非常复杂，依赖于细胞类型，涉及 Smad 和非 Smad 信号级联，因此需要严格的调节。内吞作用作为控制信号起始和终止的重要机制已经得到了重视。在本研究中，我们研究了肝细胞中 TGF-β 介导的 Smad 和非 Smad 信号的内吞作用机制。
使用化学抑制剂、RNAi、病毒基因转移和 caveolin-1-/- 小鼠来阐明肝细胞中的内吞作用。通过 Western blot、报告基因分析和基因表达分析监测 TGF-β 信号转导。
在肝细胞中，Smad 激活在很大程度上是由 AP-2 复合物完成的，而不需要网格蛋白包被凹陷的形成或内吞步骤。相反，非 Smad/AKT 途径的激活需要功能性 dynamin 介导的内吞作用和 caveolin-1 的存在，caveolin-1 是 caveolae 形成的必需蛋白。此外，这两个 TGF-β 信号起始平台区分了不同的信号通路，这些信号通路在转录水平上整合，如 TGF-β 靶基因 Id1、Smad7 和 CTGF 所示。内吞作用抑制增加了经典的 Smad 信号，并最终导致 Id1 和 Smad7 表达的超诱导，而 caveolin-1 介导的 AKT 途径的激活是最大程度诱导 CTGF 所必需的。
内吞作用对于肝细胞中 TGF-β 信号转导的调节至关重要，决定了基因表达特征和（病理）生理结果。
