Abstract：
Acute cholestatic liver injury (ACLI) is a disease associated with bile duct obstruction that causes liver inflammation and apoptosis. Although G protein-coupled bile acid receptor1 (Gpbar-1) has diverse metabolic roles, its involvement in ACLI-associated immune activation remains unclear. Liver tissues and blood samples from 20 patients with ACLI and 20 healthy individuals were analyzed using biochemical tests, H&E staining, western blotting, and immunohistochemistry to verify liver damage and expression of Gpbar-1. The expression of Gpbar-1, cAMP/PKA signaling, and the NLRP3 inflammasome was tested in wild-type (WT) and Gpbar-1 knockdown (si-Gpbar-1) mice with ACLI induced by bile duct ligation (BDL) and in primary Kupffer cells (KCs) with or without Gpbar-1-siRNA. The results showed that total bile acids and Gpbar-1 expressions were elevated in patients with ACLI. Gpbar-1 knockdown significantly worsened BDL-induced acute hepatic damage, inflammation, and liver apoptosis in vivo. Knockdown of Gpbar-1 heightened KC sensitivity to lipopolysaccharide (LPS) stimulation. Gpbar-1 activation inhibited LPS-induced pro-inflammatory responses in normal KCs but not in Gpbar-1-knockdown KCs. Notably, NLRP3-ASC inflammasome expression was effectively enhanced by Gpbar-1 deficiency. Additionally, Gpbar-1 directly increased intracellular cAMP levels and PKA phosphorylation, thus disrupting the NLRP3-ASC inflammasome. The pro-inflammatory characteristic of Gpbar-1 deficiency was almost neutralized by the NLRP3 inhibitor CY-09. In vitro, M1 polarization was accelerated in LPS-stimulated Gpbar-1-knockdown KCs. Therapeutically, Gpbar-1 deficiency exacerbated BDL-induced ACLI, which could be rescued by inhibition of the NLRP3-ASC inflammasome. Our study reveal that Gpbar-1 may act as a novel immune-mediated regulator of ACLI by inhibiting the NLRP3-ASC inflammasome.
摘要：
急性胆汁淤积性肝损伤（ACLI）是一种与胆管阻塞相关的疾病，可引起肝脏炎症和细胞凋亡。尽管G蛋白偶联的胆汁酸受体1（Gpbar-1）具有不同的代谢作用，其是否参与ACLI相关免疫激活仍不清楚.对20例ACLI患者和20例健康个体的肝组织和血液样本进行生化检测,H&E染色,西方印迹,和免疫组化验证肝损伤和Gpbar-1的表达。Gpbar-1、cAMP/PKA信号的表达,并在胆管结扎（BDL）诱导的ACLI的野生型（WT）和Gpbar-1敲低（si-Gpbar-1）小鼠以及有或没有Gpbar-1-siRNA的原代Kupffer细胞（KCs）中测试了NLRP3炎性体-1-siRNA。结果表明，ACLI患者总胆汁酸和Gpbar-1表达升高。Gpbar-1敲低可显著加重BDL诱导的急性肝损伤,炎症,和体内肝细胞凋亡。敲除Gpbar-1提高了KC对脂多糖(LPS)刺激的敏感性。Gpbar-1激活在正常KC中抑制LPS诱导的促炎反应，但在Gpbar-1敲低的KC中不抑制。值得注意的是,Gpbar-1缺乏可有效增强NLRP3-ASC炎性体表达。此外,gpbar-1直接增加细胞内cAMP水平和PKA磷酸化,从而破坏NLRP3-ASC炎性体。Gpbar-1缺乏的促炎特性几乎被NLRP3抑制剂CY-09中和。体外，在LPS刺激的Gpbar-1敲低的KC中M1极化加速。治疗学上,Gpbar-1缺乏加剧了BDL诱导的ACLI，可以通过抑制NLRP3-ASC炎性体来挽救。我们的研究表明，Gpbar-1可能通过抑制NLRP3-ASC炎性体而充当ACLI的新型免疫介导调节剂。
