title: Neo-Epitopes Generated on Hydroxyl Radical Modified GlycatedIgG Have Role in Immunopathology of Diabetes Type 2.
abstract: Glycoxidation plays a crucial role in diabetes and its associated complications. Among the glycoxidation agents, methylglyoxal (MG) is known to have very highglycationpotential witha concomitant generation of reactive oxygen species (ROS) during its synthesis and degradation. The presentstudy probes the MG and ROSinduced structural damage to immunoglobulin G (IgG) and alterations in its immunogenicity in diabetes type 2 patients (T2DM). Human IgG was first glycated with MG followed by hydroxyl radical (OH•) modification. Glycoxidation mediated effects on IgG were evaluated by various physicochemical techniques likeultraviolet (UV) and fluorescence spectroscopy, 8-anilinonaphthalene-1-sulfonic acid (ANS) binding studies, carbonyl andfree sulfhydryl groups assay, matrix assisted laser desorption ionization mass spectrometry-time of flight (MALDI-TOF), red blood cell (RBC) haemolysis assay, Congored (CR) staining analysis and scanning electron microscopy (SEM). The results revealed hyperchromicityin UV, advanced glycation end product (AGE)specific and ANS fluorescence, quenching in tyrosine and tryptophan fluorescence intensity,enhanced carbonyl content,reduction in free sulfhydryl groups,pronounced shift in m/z value of IgGand decrease in antioxidant activity in RBC induced haemolysis assayupon glycoxidation. SEM and CRstaining assay showed highly altered surface morphology in glycoxidised sample as compared to the native. Enzyme linked immunosorbent assay (ELISA) and band shift assay were performed to assess the changes in immunogenicity of IgG upon glyoxidation and its role in T2DM. The serum antibodies derived from T2DM patients demonstrated strong affinity towards OH• treated MG glycatedIgG (OH•-MG-IgG) when compared to native IgG (N-IgG) or IgGs treated with MG alone (MG-IgG) or OH• alone (OH•-IgG). This study shows the cumulating effect of OH• on the glycation potential of MG. The results point towards the modification of IgG in diabetes patients under the effect of glycoxidative stress, leading to the generation of neo-epitopes on theIgG molecule and rendering it immunogenic.
authors.affiliation: Department of Biochemistry, J.N. Medical College, Faculty of Medicine, Aligarh Muslim University, Aligarh, Uttar Pradesh, India.
authors.forename: Sidra
authors.initials: S
authors.lastname: Islam
authors.affiliation: Department of Biotechnology, Government Degree College Baramulla, University of Kashmir, Jammu and Kashmir, India.
authors.forename: Abdul Rouf
authors.initials: AR
authors.lastname: Mir
authors.affiliation: Rajiv Gandhi Centre for Diabetes and Endocrinology, Aligarh Muslim University, Aligarh, Uttar Pradesh, India.
authors.forename: Alok
authors.initials: A
authors.lastname: Raghav
authors.affiliation: Department of Biochemistry, J.N. Medical College, Faculty of Medicine, Aligarh Muslim University, Aligarh, Uttar Pradesh, India.
authors.forename: Farzana
authors.initials: F
authors.lastname: Khan
authors.affiliation: Department of Biochemistry, J.N. Medical College, Faculty of Medicine, Aligarh Muslim University, Aligarh, Uttar Pradesh, India.
authors.forename: Khursheed
authors.initials: K
authors.lastname: Alam
authors.affiliation: Department of Biochemistry, J.N. Medical College, Faculty of Medicine, Aligarh Muslim University, Aligarh, Uttar Pradesh, India.
authors.forename: Asif
authors.initials: A
authors.lastname: Ali
authors.affiliation: Department of Biochemistry, J.N. Medical College, Faculty of Medicine, Aligarh Muslim University, Aligarh, Uttar Pradesh, India.
authors.forename: Moin
authors.initials: M
authors.lastname: Uddin
mesh_terms: D000975:Antioxidants / Q000378:metabolism; D001323:Autoantibodies / Q000276:immunology; D016022:Case-Control Studies; D003924:Diabetes Mellitus, Type 2 / Q000276:immunology*; D000939:Epitopes / Q000276:immunology*; D004912:Erythrocytes / Q000378:metabolism; D017127:Glycation End Products, Advanced / Q000378:metabolism; D006031:Glycosylation; D006461:Hemolysis; D006801:Humans; D017665:Hydroxyl Radical / Q000737:chemistry*; D007074:Immunoglobulin G / Q000737:chemistry*; D010084:Oxidation-Reduction; D011765:Pyruvaldehyde / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism
chemical_list: D000975:Antioxidants; D001323:Autoantibodies; D000939:Epitopes; D017127:Glycation End Products, Advanced; D007074:Immunoglobulin G; D017382:Reactive Oxygen Species; D017665:Hydroxyl Radical; D011765:Pyruvaldehyde