Inhibition Of FGF21 On OxLDL-Induced HUVEs Pyroptosis Through The TET2-UQCRC1 Pathway

Chronic inflammation runs through the process of atherosclerosis(AS)development and is the main pathophysiological basis of atherosclerosis.Fibroblast growth factor 21(FGF21)is a multi-effect protein factor secreted by liver and adipose tissue.It can inhibit the development of atherosclerosis through anti-inflammatory,lipid-lowering,anti-oxidative stress and other mechanisms.The study found that after vascular endothelial cells(VECs)were stimulated by oxidized low-density lipoprotein(oxLDL),mitochondrial function was abnormal,and a large number of inflammatory mediators were released,causing an inflammatory response and promoting vascular endothelial cell Pyrocytosis.Pyrocytosis,also known as programmed necrosis,has been shown to play an important role in the development of As.However,whether FGF21 can inhibit oxLDL-induced endothelial cell Pyrocytosis and its mechanism of action is unclear,so this study intends to explore the inhibitory effect of FGF21 on oxLDL-induced endothelial cell Pyrocytosis and its molecular mechanism.[Objective]: To Inhibitory effect of FGF21 on ox-LDL-induced pyroptosis of vascular endothelial cells and its mechanism[Methods]:1.Study the effect of FGF21 on oxLDL-induced endothelial cell pyroptosis at the cellular level.Immunofluorescence and Western blot were used to detect the expression levels of Caspase1,NLRP3,and IL-1β related to pyroptosis;flow cytometry,ATP assay kit,and ROS fluorescent probe were used to detect mitochondrial function.2.After treating human umbilical vein endothelial cells(HUVECs)with 100 mg /L oxLDL,gene chip technology was used to analyze the changes of mitochondrial gene expression profile.Immunofluorescence and Western blot were used to detect the changes of mitochondrial ubiquinone cytochrome c reductase core protein 1(UQCRC1)protein level.3.siRNA transfection technology knocks down the expression of UQCRC1,immunofluorescence and Western blot to detect the expression level of UQCRC1 protein,and analyze the transfection efficiency.Western blot detection UQCRC1 siRNA transfection of HUVECs pyroptosis-related protein Caspase1,NLRP3,IL-1β and DNA hydroxymethylase TET2 protein expression levels.Flow cytometry,ATP content detection kit,and ROS fluorescent probe were used to detect the changes of mitochondrial function after FGF21 interfered with UQCRC1 siRNA.4.Bioinformatics analysis of the CG island of the UQCRC1 gene promoter,construction of TET2 low and high expression lentiviral vectors,and transfection ofHUVECs cells respectively.Western blot was used to detect changes in the TET2-UQCRC1 pathway and the effect of FGF21 on the pathway.[Results] : 1.Western blot results show that FGF21 can inhibit the expression of Caspase-1,NLRP3 and IL-1β related to oxLDL-induced pyroptosis in HUVECs.Immunofluorescence analysis showed that after oxLDL treatment,caspase-1 protein expression increased,but was partially reversed by FGF21.2.Compared with the control group,the mitochondrial membrane potential of the oxLDL treatment group decreased,the ATP level was significantly reduced,and the ROS level was significantly increased,but they could be partially reversed by FGF21.3.The mitochondrial gene chip results showed that UQCRC1 was significantly down-regulated by oxLDL.oxLDL significantly down-regulated the expression of UQCRC1,but could be partially reversed by FGF21.4.After the interference of UQCRC1 siRNA,the pyroptosis expression levels of Caspase-1,NLRP3 and IL-1β were significantly up-regulated,but could be partially reversed by FGF21.5.After UQCRC1 siRNA interference,the mitochondrial membrane potential of HUVECs decreased,ATP levels decreased significantly,and ROS production levels increased significantly,but they were all inhibited by FGF21.6.Bioinformatics analysis found that there are two CpG islands on the UQCRC1 gene promoter,so FGF21 may mediate the up-regulation of UQCRC1 expression through TET2.Western blot resultsshowed that oxLDL significantly down-regulated the expression of TET2,but could be reversed by FGF21.7.The results of transfection of highand low-expression TET2 lentivirus showed that UQCRC1 was down-regulated when TET2 was low and UQCRC1 was up-regulated when TET2 was overexpressed.[Conclusion]: FGF21 inhibits oxLDL-induced pyroptosis in human umbilical vein endothelial cells through the TET2-UQCRC1-ROS pathway.