Acute liver failure (ALF) is one of the most common life-threatening diseases in adults without previous liver disease. Glycogen synthase kinase 3β (GSK3β) is a serine/threonine protein kinase that is widely distributed in the cells. Inhibition of its activity can inhibit cell death and promote autophagy through various pathways, thus providing a protective effect. In this study, we aimed to investigate the effect on ALF after inhibition of GSK3β and its potential mechanisms. D- galactosamine(D-Gal) in combination with lipopolysaccharide(LPS) was used to induce ALF in vitro and in vivo. And then GSK3β inhibitor TDZD-8 was used to explore the protective effect against ALF. After TDZD-8 treatment TUNEL staining and flow techniques were used to detect the proportion of apoptosis in liver tissues and cells respectively, while western blotting and immunofluorescence assays were performed to detect the expression levels of apoptosis, pyroptosis and necroptosis-related proteins in tissues and cells. In addition, western blotting was performed to explore the specific mechanism of hepatoprotective effect after GSK3β inhibition to detect the expression levels of TAK1, TRAF6 and HDAC3 after TRAF6 and HDAC3 inhibition alone. The co-localization of TRAF6 and HDAC3 in vitro was detected by immunofluorescence, while the interaction between TRAF6 and HDAC3 was detected by immunoprecipitation assay. Both in vivo and in vitro experiments, GSK3β inhibitor TDZD-8 can significantly alleviate the progression of ALF. Inhibition of GSK3β activity could significantly reduce the level of hepatocyte apoptosis, pyroptosis, necroptosis and improve liver dysfunction and tissue damage. Furthermore, we found that hepatocyte TAK1 and TRAF6 levels decreased and HDAC3 levels increased in ALF, whereas inhibition of GSK3β upregulated TAK1 and TRAF6 levels and decreased HDAC3 expression. GSK3β inhibitor TDZD-8 can prevent the progression of ALF, and its action may involve the TRAF6/HDAC3/TAK1 pathway.

中文翻译：

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抑制 GSK3β 活性可通过抑制多种程序性细胞死亡来缓解急性肝衰竭

急性肝衰竭（ALF）是无肝病史的成年人最常见的危及生命的疾病之一。糖原合成酶激酶 3β (GSK3β) 是一种广泛分布于细胞中的丝氨酸/苏氨酸蛋白激酶。抑制其活性可以通过多种途径抑制细胞死亡并促进自噬，从而提供保护作用。在本研究中，我们旨在探讨抑制GSK3β后对ALF的影响及其潜在机制。D-氨基半乳糖(D-Gal)与脂多糖(LPS)联合用于体外和体内诱导ALF。然后利用GSK3β抑制剂TDZD-8探讨其对ALF的保护作用。TDZD-8处理后，采用TUNEL染色和流式技术分别检测肝组织和细胞凋亡比例，免疫印迹和免疫荧光检测组织和细胞中凋亡、细胞焦亡及坏死性凋亡相关蛋白的表达水平。此外，通过Western blotting检测单独抑制TRAF6和HDAC3后TAK1、TRAF6和HDAC3的表达水平，探讨GSK3β抑制后保肝作用的具体机制。通过免疫荧光检测TRAF6和HDAC3的体外共定位，并通过免疫沉淀法检测TRAF6和HDAC3之间的相互作用。无论是体内还是体外实验，GSK3β抑制剂TDZD-8均能显着缓解ALF的进展。抑制GSK3β活性可显着降低肝细胞凋亡、焦亡、坏死性凋亡水平，改善肝功能障碍和组织损伤。此外，我们发现 ALF 中肝细胞 TAK1 和 TRAF6 水平降低，HDAC3 水平升高，而抑制 GSK3β 上调 TAK1 和 TRAF6 水平并降低 HDAC3 表达。GSK3β抑制剂TDZD-8可以阻止ALF的进展，其作用可能涉及TRAF6/HDAC3/TAK1通路。