黄芩苷抑制PDK1介导糖代谢重编程干预RA滑膜炎症

Baicalin inhibits PDK1 to mediate glucose metabolism reprogramming and intervene rheumatoid arthritis synovial inflammation

本研究从唇形科植物黄芩主要活性成分黄芩苷(baicalin,BC)对胶原诱导性关节炎(collagen-induced arthritis,CIA)大鼠作用出发,以类风湿关节炎(rheumatoid arthritis,RA)滑膜炎症关键效应细胞—成纤维样滑膜细胞(fibroblast like synoviocytes,FLSs)糖代谢重编程角度展开机制探讨。建立CIA大鼠模型和肿瘤坏死因子-α(tumor necrosis factor-α,TNF-α)诱导的类风湿关节炎滑膜成纤维细胞(rheumatoid arthritis synovial fibroblasts,RASFs)体内外模型,观测BC给药后CIA大鼠关节炎指数(arthritis index,AI)评分及组织病理学变化,并ELISA法检测血清和细胞上清中炎症因子水平,免疫组织化学及Western blot法检测G蛋白偶联受体81(G-protein-coupled receptor 81,GPR81)、丙酮酸脱氢酶激酶1(pyruvate dehydrogenase kinase 1,PDK1)蛋白表达。试剂盒测定糖代谢重编程中关键产物水平及酶活性。结果显示,BC(50、100和200 mg·kg-1)剂量依赖性减轻CIA大鼠关节炎症状,抑制滑膜增生,减轻炎性细胞浸润,下调促炎因子TNF-α、白细胞介素(interleukin,IL)-1β,上调抑炎因子IL^(-1)0水平,同时降低乳酸、丙酮酸、乙酰辅酶A、柠檬酸分泌水平和乳酸脱氢酶B(lactate dehydrogenase B,LDH-B)活性,并下调GRP81、PDK1表达,提示BC调节糖代谢重编程过程。而GPR81抑制剂3-OBA抑制乳酸摄取后,LDH-B活性呈现显著升高,提示BC抑制从糖酵解到氧化磷酸化重编程代谢中的关键酶PDK1表达。本研究所有动物实验操作均按照安徽中医药大学实验动物护理中心伦理标准进行(批准号:AHUCM-rats-2021049)。以上研究揭示黄芩苷通过抑制PDK1蛋白表达,介导RASFs从糖酵解向氧化磷酸化的代谢重编程,缓解CIA大鼠关节炎症。

This study started from the effect of baicalin(BC),the main active component of the labiaceae plant Scutellaria baicalensis,on collagen-induced arthritis(CIA)in rats,to explore the mechanism of glucose metabolism reprogramming in fibroblast like synoviocytes(FLSs),a key effector cell of synovial inflammation in rheumatoid arthritis(RA).First of all,CIA rats and tumor necrosis factor-α(TNF-α)-induced RASFs in vitro and in vivo models were established,the arthritis index(AI)score and histopathological changes of CIA rats after BC administration were observed,and the levels of inflammatory factors in serum and cell supernatant were quantified by ELISA,immunocytochemistry and Western blot were used to detect the expression of G-protein-coupled receptor 81(GPR81)and pyruvate dehydrogenase kinase 1(PDK1)proteins.In addition,the kit was used to measure the levels of key products and enzyme activities in glucose metabolism reprogramming.The results showed that BC(50,100 and 200 mg·kg-1)could alleviate the symptoms of arthritis in CIA rats in a dose-dependent manner,inhibit synovial hyperplasia,alleviate the infiltration of inflammatory cells,down-regulate the levels of pro-inflammatory factors TNF-αand interleukin(IL)-1β,and up-regulate the levels of anti-inflammatory factor IL-10 in CIA rats.At the same time,the secretion levels of lactate,pyruvate,acetyl-CoA,citrate and the activity of lactate dehydrogenase B(LDH-B)were decreased,and the expressions of GRP81 and PDK1 were down-regulated,suggesting that BC mediated the reprogramming process of glucose metabolism.However,when GPR81 inhibitor 3-OBA inhibited lactate uptake,the activity of LDH-B was significantly increased,suggesting that BC inhibited the expression of PDK1,a key enzyme in the reprogramming metabolism from glycolysis to oxidative phosphorylation.All animal experiments in this study were conducted in accordance with the ethical standards of the Laboratory Animal Care Center of Anhui University of Chinese Medicine(approval number:AHUCM-rats-2021049).These studies revealed that baicalin mediated metabolic reprogramming of RASFs from glycolysis to oxidative phosphorylation by inhibiting PDK1 protein expression,and alleviated joint inflammation in CIA rats.