The Association Of Ctla-4, Vdr And Mic-a Gene Polymorphism With Latent Autoimmune Diabetes In Adults

【Objective】To investigate the association of cytotoxic T lymphocyte-associated antigen 4 gene,vitamin D receptor gene and MICA gene exon 5 polymorphism with latent autoimmune diabetes in adults.【Methods】The diagnosis and classification for diabetes mellitus was referred to diagnosis standard in ADA. Patients with LADA and type 2 diabetes in clinics, the ward of general department, the Department of Endocrinology of Shenzhen Second People's Hospital were randomly selected and were divided into LADA group and type 2 diabetes group. Health Physical Examination personnel in Medical Examination Center of Hospital were randomly selected and were classified as control group.LADA patients,type 2 diabetes and control personnel were classified according to standards. Fasting blood glucose,insulin,CP and blood sugar,insulin,CP 2 hours after meals were chemical examined by Laboratory Department and the Department of Nuclear Medicine. GADAb, ICA of LADA patients,type 2 diabetes and control personnel were determined by the kit purchased from the Beyondbio company of Canada. DNA was extracted by the kit purchased from the OMEGA company of United States.Primers were designed after searching over the gene bank. The coding region of the gene of CTLA-4,VDR,MIC-A was amplified by PCR. Amplified PCR products were evaluated by 2% agarose gel electrophoresis and ethidium bromide staining,were observed and took photographs with ultra-violet transmission analysator. The DNA of CTLA-4,VDR was identified with restriction enzyme digestion. Digested products were evaluated by 2% agarose gel electrophoresis and ethidium bromide staining,were observed and took photographs with ultra-violet transmission analysator. Amplified PCR products of MIC-A gene were analyzed by denaturing polyacrylamide gel electrophoresis(PAGE) and silver staining,were observed and took photographs with ultra-violet transmission analysator.【Results】DNA was successfully extracted from blood. The coding region of the gene of CTLA-4,VDR,MIC-A was amplified by PCR. The DNA of CTLA-4,VDR was identified with restriction enzyme digestion. Digested products were evaluated by 2% agarose gel electrophoresis and ethidium bromide staining. Amplified PCR products of MIC-A gene were analyzed by denaturing polyacrylamide gel electrophoresis(PAGE) and silver staining.A highly significant increase in the frequency of the G allele was seen in patients with LADA compared with controls(P<0.01).This reflected an increase in the GG genotype in patients(P=1.3×10^-2)and a significant decrease in the AA genotype(P<0.01).The frequency of the G allele was significantly higher in patients with LADA than in type 2 diabetes patients (P<0.01).This reflected an increase in the GG genotype in patients(P=3.4×10^-2 )and a significant decrease in the AA genotype(P<0.01 ).A highly significant increase in the frequency of the B allele was seen in T2DM patients compared with controls(P=2.7×10^-2).This reflected an increase in the Bb genotype in patients(P=1.6×10^-2)and a significant decrease in the bb genotype(P=1.6×10^-2).The Bb or bb frequency was not significantly higher in patients with LADA than in the control group(P>0.05),and so was the B allele.A highly significant increase in the frequency of the A5.1 allele was seen in patients with LADA compared with controls(P<0.05).The A5.1 allele frequency was not significantly higher in patients with T2DM than in the control group(P>0.05)【Conclusion】The polymorphism of MICA and CTLA-4 gene A/G49 confers susceptibility to LADA. The study shows that the vitamin D receptor BsmⅠpolymorphism may be not related to LADA,but may be tightly related to T2DM.