The Study Of The Correlation Between MMp-9 Gene Promoter Regions Methylation Status And Diabetic Nephropathy

Objective: The aim of the present study is to investigate the methylation level of the MMP-9 gene promoter and the level of MMP-9 expression in patients with different periods of diabetic nephropathy, and to analyze their possible correlation with DN.Methods: According to the diagnosis and classification of diabetes standard(WHO 1999), a total of 120 patients continuously selected from the hospital endocrinology clinic visits, including sugar regulation damaged and type 2 diabetes patients. There are another 30 cases of healthy volunteers with the same period in our hospital check-up. All the study objects are divided into five groups: group A(control group),group B(sugar regulation damaged group), group C(simple diabetes group), group D(early-stage diabetic nephropathy group), and group E(clinical-stage diabetic nephropathy group), to collect general information and biochemical examination indexes respectively with 30 people in each group. Genomic DNA was extracted from the peripheral blood leukocytes and then it was treated with sodium bisulfate. The processed DNA was amplified by methylation-specific PCR(MSP) to detect the MMP-9 gene regions methylation status. The serum levels of MMP-9 protein were determinated by enzyme-linked immunosorbent assay(ELISA). The results were carried out by SPSS 22.0 to statistical analysis. By comparing the difference between different groups, it can be concluded that whether MMP-9 gene promoter regions methylation status is relevant to DN.Results: The MMP-9 expression levels of control group, sugar regulation damaged group, simple diabetes group, the early-stage diabetic nephropathy group and the clinical-stage diabetic nephropathy group were listed as follows respectively: 100.87±4.88ng/ml, 101.93±4.00ng/ml, 132.67±4.82ng/ml, 212.03±5.75ng/ml and 246.27 ±5.02ng/ml. Compared with control group, there was no significant difference in the MMP-9 concentration of sugar regulation damaged group(P>0.05). The MMP-9 protein levels in the early-stage DN and clinical-stage DN groups were significantly higher than those in the control group and Type 2 DM group(P<0.05),with the highest level in the clinical-stage diabetic nephropathy group(P<0.05). The MMP-9 gene promoter regions methylation positive rate of control group, sugar regulation damaged group, simple diabetes group, the early-stage diabetic nephropathy group and the clinical-stage diabetic nephropathy group were respectively 93.33%, 90.00%, 46.67%, 23.33% and 6.67%. Compared with control group, there was no obvious difference in sugar regulation damaged group(P>0.05). The MMP-9 methylation levels in the early-stage DN and clinical-stage DN groups were dramatically lower than those in the control group and Type 2 DM group(P<0.05), with the lowest methylation level in clinical-stage diabetic nephropathy group(P<0.05). Correlation analysis showed that the demethylation level of the promoter was positively correlated with levels of MMP-9 expression, urinary albumin-to-creatinine ratio(UACR), blood urea nitrogen, creatinine, while it was negatively correlated with glomerular filtration rate(GFR).Conclusion: The MMP-9 gene promoter methylation levels gradually reduce with the progress of DN. MMP-9 protein expression levels gradually rise with the progress of DN. MMP-9 gene promoter regions demethylation is possibly involved in the occurrence or development of diabetic nephropathy through regulating the MMP-9 protein expression.