The Potential Mechanism Of RNA N6-Methyladenosine In PBMCs Of Primary Sj?gren’s Syndrome And Primary Biliary Cholangitis

ObjectiveTo detect and elucidate the N6-methyladenosine-related genes expressed differently between PBMCs in pSS patients,healthy controls and symptom controls,and to determine the potential role of m6A in the development of pSS disease.Methods1.This study included 44 patients with pSS,50 age-and sex-matched healthy controls(HCs),and 11 age-and sex-matched symptom controls(with similar dryness symptoms to pSS patients,but not fulfilling the classification criteria,non-SS sicca patients).RT-qPCR was used to measure mRNA levels of m6A-related genes(including METTL3,WTAP,RBM15,ALKBH5,FTO,YTHDF1,YTHDF2,YTHDF3,YTHDC1,and YTHDC2),ISG15,and USP18 in peripheral blood mononuclear cells from pSS patients,healthy controls,and symptom controls.2.Clinical features and laboratory test results of pSS patients and symptom controls were collected.The correlation between m6A-related genes and laboratory indicators and disease activity in pSS patients was analyzed.3.Binary logistic regression was used to determine whether m6A-related genes are risk factors for pSS.4.The correlation between m6A-related genes and interferon genes in pSS patients was analyzed,which may indicate the possible mechanism of interaction between m6A-related genes and interferon-stimulatory genes in peripheral blood mononuclear cells of patients with pSS in the pathogenesis of pSS.Results1.The mRNA levels of m6A writers(METTL3 and RBM15),erasers(ALKBH5 and FTO),and readers(YTHDF1,YTHDF2,YTHDF3,YTHDC1,and YTHDC2)were all significantly higher in PBMCs from patients with pSS than in HCs.The mRNA levels of m6A writers(METTL3 and WTAP)and readers(YTHDF2,YTHDF3,and YTHDC2)were lower in PBMCs from patients with pSS compared to patients with non-SS sicca.2.The expression of METTL3,RBM15,FTO,YTHDF1,YTHDF2,YTHDC1,and YTHDC2 was positively correlated with the level of C-reactive protein(CRP)of patients with pSS.The mRNA level of YTHDF1 in PBMCs from patients with pSS was negatively correlated with the EULAR Sjogren’s syndrome disease activity index(ESSDAI)score.In patients with pSS,FTO,YTHDC1,and YTHDC2 were also related to white blood cells(WBCs),neutrophils,lymphocytes,and monocytes.Increased mRNA level of ALKBH5 in PBMCs was a risk factor for pSS,as determined by binary logistic regression analysis.3.The mRNA level of ISG15 was positively correlated with that of FTO,YTHDF2,YTHDF3,and YTHDC2 in patients with pSS.Conclusions1.Compared with HCs,the expression of METTL3,RBM15,ALKBH5,FTO,YTHDF1,YTHDF2,YTHDF3,YTHDC1,and YTHDC2 was considerably higher in PBMCs from patients with pSS.In comparison with patients with non-SS sicca,the expression of METTL3,WTAP,YTHDF2,YTHDF3,and YTHDC2 was reduced in PBMCs from patients with pSS.2.The m6A elements correlating with clinical variables may indicate the disease activity and inflammation status of pSS.Elevated expression of ALKBH5 was a risk factor for pSS.3.The dynamic process of m6A modification is active in pSS.m6A elements(FTO,YTHDF2,YTHDF3,or YTHDC2)might target ISG15,stimulate the expression of ISG15,and activate the type Ⅰ IFN signaling pathway,playing an active role in initiating the autoimmunity in pSS.ObjectiveTo detect and elucidate the m6A-related genes expressed differently between PBMCs in PBC patients and healthy controls,and to determine the potential role of m6 A in the development of PBC.Methods1.This study enrolled 32 PBC patients and 32 age-sex-matched healthy controls(HCs).We detected the mRNA levels of METTL3,WTAP,RBM15,FTO,ALKBH5,YTHDF1,YTHDF2,YTHDF3,YTHDC1,and YTHDC2 in peripheral blood mononuclear cells from PBC patients and HCs.2.The clinical features and laboratory findings of PBC patients were also collected.The correlation between m6A-related genes and laboratory indicators in PBC patients was analyzed.3.Binary logistic regression was applied to evaluate whether the m6A elements were the risk factors for PBC.Results1.The mRNA levels of m6A writers(METTL3 and RBM15),eraser ALKBH5,and reader YTHDF1 were all increased in peripheral blood mononuclear cells from PBC patients compared with HCs.2.In peripheral blood mononuclear cells of PBC patients,there was a negative correlation between WTAP and C4.ALKBH5 was positively correlated with TBIL and negatively correlated with neutrophil absolute value and red cell count.FTO is negatively correlated with GGT.YTHDF1 was negatively correlated with IgG and neutrophil absolute values.YTHDF2 was negatively correlated with the absolute values of C3,IgM and neutrophils.YTHDF3 is negatively correlated with GGT and ALT.It was positively correlated with LDH and TBIL.YTHDC1 was negatively correlated with ALP and C3.YTHDC2 showed a moderate positive correlation with ESR.3.The binary logistic regression analysis showed that the increased mRNA levels of METTL3,RBM15,ALKBH5 and YTHDF2 were risk factors for PBC.Conclusions1.The expression of m6A writers,erasers,and readers were all elevated in PBMCs from PBC patients.2.The mRNA levels of m6A-related genes in peripheral blood mononuclear cells of PBC patients were correlated with ALP,GGT,ALT,LDH and TBIL,which may indicate liver parenchymal injury,bilirubin metabolism and cholestasis.WTAP,YTHDF2,YTHDC1,and YTHDC2 were associated with ESR,C3,and C4,and may indicate disease activity and inflammation levels in PBC patients.YTHDF1 and YTHDF2 were correlated with serum IgM and IgG levels,which may reflect PBC immune disorder.3.Increased expression of METTL3,RBM15,ALKBH5 were risk factors for PBC.