Abstract
Objectives In diabetes mellitus, hyperglycemia-mediated nonenzymatic glycosylation of fibrinogen protein plays a crucial role in the pathogenesis of micro- and macrovascular complications especially atherosclerosis via the generation of advanced glycation end products (AGEs). Methylglyoxal (MG) induces glycation of fibrinogen, resulting in structural alterations that lead to autoimmune response via the generation of neoepitopes on protein molecules. The present study was designed to probe the prevalence of autoantibodies against MG-glycated fibrinogen (MG-Fib) in type 2 diabetes mellitus (T2DM), atherosclerosis (ATH), and diabetic atherosclerosis (T2DM-ATH) patients. Design and Methods. The binding affinity of autoantibodies in patients' sera (T2DM, n = 100; ATH, n = 100; and T2DM-ATH, n = 100) and isolated immunoglobulin G (IgG) against native fibrinogen (N-Fib) and MG-Fib to healthy subjects (HS, n = 50) was accessed by direct binding ELISA. The results of direct binding were further validated by competitive/inhibition ELISA. Moreover, AGE detection, ketoamines, protein carbonyls, hydroxymethylfurfural (HMF), thiobarbituric acid reactive substances (TBARS), and carboxymethyllysine (CML) concentrations in patients' sera were also determined. Furthermore, free lysine and free arginine residues were also estimated. Results The high binding affinity was observed in 54% of T2DM, 33% of ATH, and 65% of T2DM-ATH patients' samples with respect to healthy subjects against MG-Fib antigen in comparison to N-Fib (p < 0.05 to p < 0.0001). HS sera showed nonsignificant binding (p > 0.05) with N-Fib and MG-Fib. Other biochemical parameters were also found to be significant (p < 0.05) in the patient groups with respect to the HS group. Conclusions These findings in the future might pave a way to authenticate fibrinogen as a biomarker for the early detection of diabetes-associated micro- and macrovascular complications.
Highlights
In diabetes mellitus, hyperglycemia-mediated nonenzymatic glycosylation or glycation of biological macromolecules plays a critical role in the pathogenesis of secondary complications especially micro- and macrovascular complications [1, 2]
A protein A agarose column was purchased from Sigma, and polystyrene plates were obtained from Nunc (Denmark)
The results revealed that the average saturation concentration of MGFib is 50 μg mL-1 of immunoglobulin G (IgG) from type 2 diabetes mellitus (T2DM) and T2DM-ATH patients
Summary
Hyperglycemia-mediated nonenzymatic glycosylation or glycation of biological macromolecules (proteins, lipids, and DNA) plays a critical role in the pathogenesis of secondary complications especially micro- and macrovascular complications [1, 2]. Schiff’s bases via keto-enol tautomerism and acid-base catalysis convert to irreversible ketoamines or Amadori products, and the latter undergo rearrangements, Oxidative Medicine and Cellular Longevity dehydrations, and cyclizations to form heterogeneous molecular adducts, known as advanced glycation end products (AGEs) [5, 6]. Glycoxidation of glycation-adducts and autoxidation of glucose lead to the generation of reactive oxygen species (ROS) such as metal-catalyzed superoxide radicals (O2) via reaction and hydroxyl radicals transition (OH) via Fenton’s reaction, thereby inducing oxidative stress [7, 8]. Besides ROS generation, both autoxidation and glycoxidation give rise to aldehyde and/or ketone groups possessing highly electronegative dicarbonyls, called reactive carbonyl species (RCS), i.e., glyoxal (GO), methylglyoxal (MG), and 3-. ROS-mediated oxidative stress induces lipid peroxidation and oxidation of amino acids, which result in the generation of RCS, especially
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