Ning Beh, Wang Shao-An, Young Ming-Jer, Chen Yung-Ching, Hung Yun, Huong Tran Thu, Chang Wen-Chang, Wang Yi-Ching, Yu Ming-Lung, Hsu Kai-Cheng, Hung Jan-Jong
Department of Biotechnology and Bioindustry Sciences, National Cheng Kung University, Tainan, 701, Taiwan.
Graduate Institute of Medical Sciences, College of Medicine, Taipei Medical University, Taipei, Taiwan.
J Biomed Sci. 2025 May 30;32(1):54. doi: 10.1186/s12929-025-01148-4.
Ubiquitin-specific peptidase 24 (USP24), a deubiquitinating enzyme, regulates protein stability by removing ubiquitin. This study investigates the role of UPS24 in lipid metabolism, inflammation, and fibrosis. It also explores the effect of targeting USP24 on metabolic disorders, focusing on high-fat diet (HFD)-induced obesity and liver diseases.
This study utilized CRISPR/Cas9 to create functional knockout mice (USP24) and treated HFD-fed mice with USP24 inhibitor (USP24-i-101). The effects of USP24 inhibition or knockout on 3T3-L1 derived adipocytes, primary hepatocytes, hepatic stellate cells, and murine hepatocyte cell line AML12 (alpha mouse liver 12) cells were assessed with RNA-sequencing. Molecular mechanisms and the interaction between USP24 and PKA-Cα were studied with co-immunoprecipitation. Downstream signaling pathways involving CREB, SREBP1, PPARγ, and C/EBPβ, as well as USP24 role in liver inflammation and fibrosis, were studied using western blot and real-time PCR. Clinical and animal tissue samples were examined with immunohistochemistry to identify the correlations between USP24 and metabolic-associated liver diseases.
Knockout or inhibition of USP24 reduced body weight, lipid accumulation, inflammation, and fibrosis in HFD-fed mice. The expression of genes related to lipogenesis, inflammation, and fibrosis was downregulated in USP24 mice and those treated with USP24 inhibitor (USP24-i-101). USP24 inhibition decreased lipid droplet accumulation in adipocytes and hepatocytes, suppressed inflammation in hepatocytes and AML12 cells, and reduced fibrosis in hepatic stellate cells. Mechanistically, USP24 expression was upregulated by PKA activation during adipocyte differentiation, leading to increased PKA-Cα stability and CREB phosphorylation, which promoted lipogenic gene expression. Free fatty acids (FFA) increased USP24 expression, activating NF-κB and TGFβ pathways to induce inflammation (Cox2) and fibrosis (α-SMA). USP24 was highly expressed in patients with metabolic dysfunction-associated steatohepatitis (MASH) and correlated with Cox2 and α-SMA levels.
泛素特异性肽酶24（USP24）是一种去泛素化酶，通过去除泛素来调节蛋白质稳定性。本研究调查了USP24在脂质代谢、炎症和纤维化中的作用。它还探讨了靶向USP24对代谢紊乱的影响，重点关注高脂饮食（HFD）诱导的肥胖和肝脏疾病。
本研究利用CRISPR/Cas9技术创建功能性基因敲除小鼠（USP24），并用USP24抑制剂（USP24-i-101）处理高脂饮食喂养的小鼠。通过RNA测序评估USP24抑制或敲除对3T3-L1衍生脂肪细胞、原代肝细胞、肝星状细胞和小鼠肝细胞系AML12（α小鼠肝脏12）细胞的影响。通过免疫共沉淀研究USP24与PKA-Cα之间的分子机制和相互作用。使用蛋白质免疫印迹和实时聚合酶链反应研究涉及CREB、SREBP1、PPARγ和C/EBPβ的下游信号通路，以及USP24在肝脏炎症和纤维化中的作用。通过免疫组织化学检查临床和动物组织样本，以确定USP24与代谢相关肝病之间的相关性。
敲除或抑制USP24可降低高脂饮食喂养小鼠的体重、脂质积累、炎症和纤维化。在USP24基因敲除小鼠和用USP24抑制剂（USP24-i-101）处理的小鼠中，与脂肪生成、炎症和纤维化相关的基因表达下调。USP24抑制减少了脂肪细胞和肝细胞中的脂滴积累，抑制了肝细胞和AML12细胞中的炎症，并减少了肝星状细胞中的纤维化。从机制上讲，在脂肪细胞分化过程中，PKA激活上调了USP24的表达，导致PKA-Cα稳定性增加和CREB磷酸化增加，从而促进脂肪生成基因的表达。游离脂肪酸（FFA）增加了USP24的表达，激活NF-κB和TGFβ通路以诱导炎症（Cox2）和纤维化（α-SMA）。USP24在代谢功能障碍相关脂肪性肝炎（MASH）患者中高表达，并与Cox2和α-SMA水平相关。
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