Autoimmune response to AGE modified human DNA: Implications in type 1 diabetes mellitus.

AIMS

Non-enzymatic glycation of DNA both and results in generation of free radicals, known as glycoxidation. Glycoxidation leads to structural perturbation of DNA resulting in generation of neo-antigenic epitopes having implication in autoimmune disorders like diabetes mellitus. In this study human placental DNA was glycated with methylglyoxal (MG) and lysine (Lys) in the presence of Cu and its auto-antibody binding was probed in Type 1 diabetes patients.

METHODS

Glycation was carried out by incubating DNA with MG, Lys and Cu for 24 h at 37 °C. Carboxyethyl deoxyguanosine (CEdG) formed in glycation reaction was studied by LC-MS and the pathway for Amadori formation was studied by ESI-MS techniques. Furthermore, binding characteristics of auto-antibodies in diabetes patients were assessed by direct binding, competitive ELISA and band shift assay.

RESULTS

DNA glycation with MG, Lys and Cu results in the formation of CEdG (marker of DNA glycation) which was confirmed by LC-MS. The intermediate stages of glycation were confirmed by ESI-MS technique. Serum from diabetes patients exhibited enhanced binding and specificity for glycated DNA as compared to native form.

CONCLUSIONS

Glycation of DNA has resulted in structural perturbation causing generation of neo-antigenic epitopes thus recognizing auto-antibodies in diabetes.