Research On The Correlation Between IgA Nephropathy And The Levels Of HLA-DQB1 And HLA-DRB1 Methylation And Antigen Expression

Background: Genome-wide association studies(GWAS)found that HLA-DQB1 and HLA-DRB1 are related to the disease risk of IgA nephropathy(IgAN),but GWASs can only explain 4～7% of disease heritability.And the pathogenesis and progression mechanisms of HLA-DQB1,HLA-DRB1 in IgA nephropathy are still unclear.Our previous experimental results showed downregulation in the m RNA expression of HLADQB1 and HLA-DRB1 in IgAN patients.It is speculated whether the decrease in m RNA expression is related to the level of methylation in HLA-DQB1 and HLA-DRB1.Thus,this study measured the methylation of HLA-DQB1,HLA-DRB1 promoter and the expression of HLA-DQ,HLA-DR antigen in IgAN patients;the relationships between the level of methylation and antigen expression and the clinical and pathological characteristics were analyzed to further clarify the possible mechanisms of HLA-DQB1 and HLA-DRB1 in the pathogenesis and progression of IgA nephropathy.Methods: The peripheral blood of 47 IgAN patients and 39 healthy people was collected;flow cytometry was used to detect the expression of HLA-DQ and HLA-DR antigen on the peripheral blood lymphocytes(PBLs);the serum galactose-deficient IgA1,(Gd-IgA1)levels were assessed by an ELISA kit.The peripheral blood of 28 IgAN patients and normal individuals was collected;the B cells were isolated from the PBLs by the immunomagnetic beads.According to the results of methylation preliminary experiment,the methylation levels of the selected Cp G sites in HLA-DQB1 promoter of B cells were assessed by bisulfite pyrosequencing.The clinical and pathological data of patients with IgAN were collected at the time of renal biopsy.The correlation among the HLA-DQ,HLA-DR antigen levels and clinical and pathological data were analyzed;the correlation between HLA-DQB1 methylation levels and clinical and pathological data were analyzed.Results: 1.Compared with healthy people,the expression of HLA-DQ antigen on total nucleated cells,B cells,NK cells and granulocytes in the IgAN patients was decreased;2.Compared with healthy people,the expression of HLA-DR antigen on NK cells was decreased in IgAN patients and the expression of HLA-DR antigen on the granulocytes was increased in IgAN patients;3.The serum levels of Gd-IgA1 in the IgAN patients were higher than in healthy people;4.The expression of HLA-DQ antigen on the lymphocytes of IgAN patients was positively correlated with serum Gd-IgA1 levels,the expression of HLA-DQ antigen on NK cells and granulocytes was positively correlated with 24-hour urine protein quantification;5.The expression of HLA-DR antigen on NK cells in IgAN patients was positively correlated with 24-hour urine protein quantification,the expression of HLA-DR antigen on T cells is negatively correlated with estimated Glomerular Filtration Rate(GFR)levels;6.Compared with IgAN patients with 24-hour urine protein quantification≤1g/24 h,the HLA-DQ antigen on granulocytes and HLADR antigen on NK cells were increased in IgAN patients with 24-hour urine protein quantification>1g/24h;7.Compared with IgAN patients without crescentic lesions,the expression of HLA-DR antigen on T cells and NK cells was increased in IgAN patients with crescentic lesions;8.Compared with healthy people,the methylation level of CPG8 in HLA-DQB1 promoter was increased in IgAN patients;there was no significant difference in the methylation level of the HLA-DRB1 promoter between IgAN patients and healthy people;9.There was no correlation between the methylation level of Cp G8 in HLA-DQB1 promoter and clinical and pathological data of IgAN patients.Conclusion: The expression of HLA-DQ and HLA-DR antigens in IgAN patients is positively correlated with 24-hour urine protein quantification and negatively correlated with e GFR levels and highly expressed in IgAN patients with crescent lesions,suggesting a positive correlation with the severity of the disease;The high methylation level of Cp G8 in HLA-DQB1 in B cells is related to the susceptibility of IgAN.