Curcumin通过调控自噬影响肝细胞上皮-间质转化的分子机制研究

Myofibroblast (MFB) proliferation and activation is the key event in hepatic fibrosis (HF). A large number of studies have confirmed that hepatocytes can be transformed into MFB through epithelial-mesenchymal transformation (EMT). Our previous studies showed that Curcumin (Cur) can effectively inhibit hepatocyte EMT, as well as inhibit HSC activation and proliferation by affecting the Hedgehog (Hh) signaling pathway and autophagy. We herein innovatively put forward that Cur inhibition of the hepatocyte EMT is due to the regulation of cell autophagy. The involved mechanism may be affecting the Hh signal transduction pathways. To test this hypothesis, we will first use HF models induced by CCL4, BDL and MMH cell model interference autophagy genes, to investigate the relationship between autophagy and Cur inhibition of hepatocyte EMT; Then the effect of Cur regulating autophagy and EMT of hepatocyte will be examined under the condition of activation or inhibition of Hh signaling pathways in vivo and in vitro. The purpose of these studies are to clarify the mechanism of Cur regulation of cell autophagy and suppression of hepatocyte EMT by influencing the Hh signaling pathways. Finally we will use Cre; ATG7fl/fl mice to further verify the effect of Cur inhibition of hepatocyte EMT by regulating autophagy. Through those studies, we can interpret the mechanism of Cur reduction of HF from a new perspective, and the results of this study will provide new strategies for pharmacotherapy of hepatic fibrosis in clinical context.

肌成纤维细胞（MFB）的大量生成与活化是肝纤维化（HF）发生的核心环节，大量研究已证实肝细胞经上皮间质转化（EMT）可转变为MFB。我们前期研究发现姜黄素（Cur）能有效抑制肝细胞EMT进程，并能影响Hedgehog（Hh）信号通路与细胞自噬抑制HSC的活化。本项目创新性提出Cur抑制肝细胞EMT的作用与影响细胞的自噬水平相关，具体机制可能涉及到对Hh信号通路的调控。为此，本研究拟利用CCL4、BDL致HF与自噬基因干扰细胞模型，首先明确自噬与Cur抑制肝细胞EMT之间的关系；然后通过体内外激动或抑制Hh信号通路，研究Cur对肝细胞自噬与EMT的影响，以阐明Cur通过影响Hh信号通路调控自噬进而抑制肝细胞EMT的机制；最后利用自噬基因干扰小鼠进一步验证Cur调控自噬抑制肝细胞EMT的作用。本项目从新的视角阐释了Cur抗HF的作用机制，预期研究结果将为抗HF药物的临床治疗提供新的策略。

肌成纤维细胞（MFB）的大量生成与活化是肝脏纤维化发展过程中的核心事件。大量研究证实肝细胞是MFB的重要来源，在肝纤维发生时可通过上皮间质转化（EMT）转变为MFB。我们的研究前期证实姜黄素可抑制肝细胞的EMT进程及向MFB的分化，有研究显示自噬在调控细胞EMT进程中发挥重要作用。因此，我们假设姜黄素可通过调控自噬抑制肝细胞向肌成纤维细胞的转变及肝纤维化的发生与发展。本研究中我们发现姜黄素通过活化肝细胞自噬功能来抑制TGF-β/Smad信号传递进而抑制肝细胞EMT。姜黄素增强肝细胞自噬后可能是通过抑制TTC3 蛋白(Tetratricopeptide repeat domain 3)及诱导Smad泛素化调节因子2（Smad ubiquitination regulatory factor 2）的泛素化和蛋白酶体降解，导致了SMAD2和SMAD3蛋白的多泛素化降解。姜黄素通过激活PPARα抑制肝细胞内ROS水平，调控自噬上游信号通路AMPK、PI3K/AKT/mTOR信号通路，导致肝细胞内自噬流的的增加。本研究证实Curcumin通过激活自噬能有效地减少肝细胞EMT的发生，抑制细胞外基质的产生，为肝纤维治疗提供了潜在的新治疗策略。