Correlation Study Of The Expression And Methylation Of TWEAK-related Genes In Peripheral Blood Of Patients With Systemic Lupus Erythematosus

Objectives:Tumor necrosis factor-like weak inducer of apoptosis（TWEAK）related genes（TWEAK,Fn14 and LIGHT）promoter methylation,mRNA expression in peripheral blood and TWEAK serum concentration in patients with systemic lupus erythematosus（SLE）were analyzed.Analyze the correlation of above results with disease activity and laboratory indicators,and explore the role and epigenetic mechanism of the three genes in the pathogenesis of SLE.Methods:A total of 178 SLE patients diagnosed in the Department of Rheumatology and Immunology of the First Affiliated Hospital of Kunming Medical University from 2019 to 2021 were recruited as the disease group,and 131 healthy volunteers matched with age and sex during the same period were recruited as the healthy control group.（1）Quantitative real-time polymerase chain reaction（qRT-PCR）was used to detect the mRNA expression of TWEAK、Fn14 and LIGHT in the peripheral blood of all participants in the two groups;（2）40 subjects were randomly selected from each of the two groups,and the serum concentrations of TWEAK in the two groups were detected by enzyme linked immunosorbent assay（ELISA）;（3）50 subjects were randomly selected from each of the two groups,and the DNA methylation of TWEAK、Fn14 and LIGHT promoter region and single CpG site were detected by MassARRAY;（4）Collect laboratory indicators of SLE patients,analyze the correlation of mRNA expression,methylation levels and TWEAK serum concentration with the laboratory indicators of SLE,and explore the expression differences and epigenetic mechanisms of TWEAK-related genes in SLE.Results:1.The expression of TWEAK、Fn14 mRNA in the SLE patients were significantly lower than that of the healthy control group,and the expression of LIGHT mRNA in the SLE group was significantly higher than that of the healthy control group（P<0.05）;2.There was no significant difference in TWEAK serum concentration between SLE group and healthy control group（P>0.05）,but the TWEAK serum concentration in the renal involvement group was significantly lower than that in the non-renal involvement group,and the TWEAK serum concentration in the active group（SLEDAI≥10）was significantly lower than that in the stable group（SLEDAI<10）,the difference is statistically significant.3.The total methylation of TWEAK gene in the SLE group was significantly higher than that of the healthy control group,and the methylation of the renalinvolvement group was higher than that of the non-renal involvement group.There were no significant differences in the total methylation levels of Fn14 and LIGHT genes between the SLE group and the healthy control group,the renal involvement group and the non-renal involvement group,and the active group and the stable group.Furthermore,the methylation of single CpG site was compared.The methylation level of TWEAK gene CpG₁2.13.14,15.16,25.26,39,44.45,55.56,57.58 in the SLE group was significantly higher than that in the healthy control group,the CpG₄4.45,57.58 methylation level in the renal involvement group was significantly higher than that of the non-renal involvement group,the methylation level of CpG₁5.16,43 in active group was significantly higher than that of stable group（P<0.05）.However,there were no significant differences in the methylation level of single CpG site of Fn14 and LIGHT genes between the SLE group and the healthy control group,the renal involvement group and the non-renal involvement group,and the active group and the stable group.4.The study found that TWEAK mRNA expression was negatively correlated with serum uric acid（r=-0.165,P=0.03）;Fn14 mRNA expression was negatively correlated with IL-10（r=-0.211,P=0.028）;LIGHT mRNA expression was positively correlated with IL-10（r=0.299,P=0.02）.The TWEAK serum concentration was negatively correlated with SLED AI（r=-0.419,P=0.007）.The methylation of TWEAK gene was negatively correlated with TWEAK mRNA（r=-0.438,P<0.001）.The methylation of TWEAK gene CpG₁5.16 was positively correlated with the SLEDAI score（r=0.389,P=0.005）,the methylation of CpG₂5.26 was positively correlated with 24-hour microalbumin and uric acid（r=0.324,P=0.039;r=0.294,P=0.042）,the methylation of CpG₄4.45 was positively correlated with IL-6（r=0.387,P=0.026）,the methylation of CpG₅5.56 was positively correlated with uric acid（r=0.334,P=0.020）.Conclusions:TWEAK,Fn14 and LIGHT genes may be involved in the pathogenesis of SLE.The promoter region of TWEAK gene presents a hypermethylated state in the peripheral blood of SLE,and may be one of the regulatory mechanisms of its low expression,thereby participating in the pathogenesis and progression of SLE.The abnormal expression of Fn14 and LIGHT genes in SLE may not be regulated by DNA methylation modification.