[Mechanisms of human glutamic acid decarboxylase 65 DNA vaccine preventing diabetes in non-obese diabetic mice].

Objective To investigate the mechanisms of human GAD65 DNA vaccine preventing insulitis and diabetes in NOD mice.Methods Female NOD mice at 4 weeks of age were randomly divided into PBS ( n =21), pcDNA( n =20), and hGAD65( n =21) groups. Mice in each group received two intramusclar injections of 0 05 ml PBS alone , 50 μg pcDNA3 1 and 50 μg DNA vaccine emulsified in 0 05 ml PBS 7 days apart respectivly. The accumulative diabetes incidence was followed up to 30weeks of age in each group of NOD mice. Pancreas was removed from NOD mice at 12 weeks of age in each group ( n =10) to score insulitis severity by routine H E staining. The apoptotic β cells in islets were observed with double labeling technique of TUNEL in situ combined standard sensitive avdin biotin complex (sABC) immunohistochemical method. Their spleens were for cell culture and total RNA extraction. Spleen IL 4, IFN γ, NF ATc and NF ATp mRNA levels were tested by RT PCR. IL 4 and IFN γ levels in sera and supernatants of spleen cells were measured by ELISA. Results (1) At 30 weeks of age, the diabetes incidence was 95 2%, 80 0% and 61 9% in PBS, pcDNA and hGAD65 group respectively. The diabetes incidence in the PBS group was higher than that in hGAD65 group ( P =0 008). (2) At 12 weeks of age , the insulitis scores in hGAD65 group was lower than that in PBS group ( P =0 001) and pcDNA group ( P =0 027) respectively. (3) The apoptotic β cell rates in hGAD65 group was lower than that in PBS group( P =0 014) and pcDNA group ( P =0 023). (4) IL 4 levels in sera, spleen IL 4 and NF ATc mRNA level in hGAD65 group were higher than those in PBS group(all P 0 05) and pcDNA group (all P 0 05) respectively, NF ATp mRNA level in hGAD65 group was lower than that in PBS group( P 0 05).Conclusion Human GAD65 DNA vaccine via downregulating NF ATp and upregulating NF ATc and IL 4, makes Th cells deviate to Th2, and sequently prevents insulitis, beta cell apoptosis and diabetes onset in NOD mice.