METHODS: NASH rat model was established through long-term feeding HFD, and the steatosis cell model was stimulated via palmitate acid (PA). The therapeutic effect of THC was evaluated in terms of liver function, lipid metabolism, liver pathophysiology, inflammation and oxidative stress in vivo, and lipid accumulation in vitro. The alteration in lipophagy was identified by using western blot and immunofluorescence. mTORC1-TFEB signaling pathway was measured by qRT-PCR, western blot and protein-ligand docking. In addition, chloroquine and MHY1485 were further introduced to validate the effect of THC on lipophagy and mTORC1-TFEB signaling pathway, respectively.
RESULTS: THC effectively improved hepatic steatosis, inflammation and oxidative stress in NASH rats, and reduced lipid accumulation in steatosis L02 cells and Hep G2 cells. THC promoted lipophagy with increasing LC3B-II as well as decreasing P62 expression via lysosomal biogenesis upregulation, which was greatly weakened after chloroquine intervention. mTORC1-TFEB is a critical pathway for regulating lysosome in autophagy, THC treatment induced TFEB nucleus translocation via inhibiting mTORC1 to upregulate lysosomal biogenesis. However, these effects were partly eliminated by mTORC1 activator MHY1485.
CONCLUSIONS: THC restored lipophagy to reduce lipid accumulation by regulating mTORC1-TFEB pathway in NASH rats and steatosis hepatocytes. These findings suggested that THC represents a therapeutic candidate for NASH treatment.
方法:通过长期喂养HFD建立NASH大鼠模型,通过棕榈酸(PA)刺激脂肪变性细胞模型。从肝功能方面评估THC的治疗效果,脂质代谢,肝脏病理生理学,体内炎症和氧化应激,和体外脂质积累。通过使用蛋白质印迹和免疫荧光鉴定脂质吞噬的改变。通过qRT-PCR检测mTORC1-TFEB信号通路,蛋白质印迹和蛋白质-配体对接。此外,进一步引入氯喹和MHY1485,以验证THC对吸脂性和mTORC1-TFEB信号通路的影响,分别。
结果:THC有效改善了肝脏脂肪变性,NASH大鼠的炎症和氧化应激,和减少脂肪变性L02细胞和HepG2细胞中的脂质积累。THC通过溶酶体生物发生上调促进LC3B-II和P62表达降低,氯喹干预后大大减弱。mTORC1-TFEB是调节自噬中溶酶体的关键途径,THC处理通过抑制mTORC1上调溶酶体生物发生诱导TFEB核易位。然而,mTORC1激活剂MHY1485部分消除了这些影响.
结论:THC通过调节NASH大鼠和脂肪变性肝细胞中的mTORC1-TFEB通路来恢复脂质吞噬以减少脂质积累。这些发现提示THC代表NASH治疗的治疗候选物。