IDF21-0074 Role of methylglyoxal modified low density lipoprotein in type 2 diabetes mellitus and its role in periodontal disease

Abstract

Background: Glycation, a physiological and pathological process mainly affects proteins, nucleic acids and lipids. Methylglyoxal (MGO) along with free radicals is known to contribute to the pathogenesis of several diseases, in disorders particularly having enhanced production of acetone and amino-acetone such as diabetes mellitus (DM). The accumulation of MGO induces β-cells dysfunction, insulin resistance as well as impairment of endothelial functions in DM patients. Also, the increased level of MGO intensifies ROS generation and AGEs formation which further accelerates the co-morbidities associated with type 2 diabetes mellitus (T2DM). There is an increased susceptibility and severity of periodontal diseases in case of patients with diabetes and vice-versa. Aim: The likely role of native low density lipoprotein (LDL) and glycated LDL in type 2 diabetes mellitus (T2DM) and periodontitis (PD) as well as in T2DM+PD patients has been evaluated. Method: Sera from T2DM+PD T2DM and PD patients (n=80), were screened for the presence of antibodies against MGO-LDL and native LDL. Besides, the conformational changes in LDL isolated from T2DM+PD, T2DM and PD patients (possibly due to glycoxidaton) were also studied by UV absorption, fluorescence and CD spectroscopy. ThT, Comet assay, SEM and TEM were also done. Cellular toxicity of the isolated LDL from both T2DM+PD, T2DM and PD patients was demonstrated by comet assay. Biochemical parameters like increase in carbonyl content, TBARS, TNF α and IL1β indicate LDL oxidation and increase in pro-inflammatory cytokines. Results: MGO-LDL (in comparison to parallel control) proved to be a potent antigen showing very high binding with T2DM+PD, T2DM and PD sera as observed in direct binding ELISA. The stronger binding of T2DM+PD, T2DM and PD auto-antibodies to MGO-LDL suggests the importance of this modification in the onset and pathogenesis of these diseases. Inhibition ELISA results showed very high percent inhibition of antibodies from T2DM+PD, T2DM and PD patients by MGO-LDL in comparison to native LDL pointing towards the possible role of glycoxidation of LDL in these patients. The modified LDL might have further generated high levels of circulating auto-antibodies in the sera of these patients. Further, to ascertain the specificity of auto-antibodies from T2DM+PD, T2DM and PD patients, competitive inhibition ELISA studies were done on IgG isolated from the sera of these patients. The samples of isolated IgG from T2DM+PD (T2DM+PD-IgG), T2DM (T2DM-IgG) and PD (PD-IgG) showed very strong recognition of MGO-LDL in comparison to the native counterpart. The specificity and affinity of T2DM+PD, T2DM and PD auto-antibodies towards MGO-LDL was further confirmed by gel retardation assay. The gel mobility results confirmed specificity of MGO-LDL towards auto-antibodies from T2DM and PD patients. Furthermore, high recognition of experimentally induced anti-MGO-LDL antibodies by T2DM+PD-LDL, T2DM-LDL and PD-LDL compared to NHS-LDL confirmed that the observed structural damage in T2DM+PD-LDL, T2DM-LDL and PD-LDL is due to glycoxidation. Discussion: The neo-epitopes on LDL created by its exposure to MGO appear to be responsible for induction and elevated levels of these antibodies, and thus MGO-LDL may be considered as potential antigenic candidate for auto-immune response in diabetes and periodontitis patients with a potential for biomarker development.