The hyperglycemic byproduct methylglyoxal impairs anticoagulant activity through covalent adduction of antithrombin III

Abstract

Introduction The blood coagulation system is a tightly regulated balance of procoagulant and anticoagulant factors, disruption of which can cause clinical complications. Diabetics are known to have a hypercoagulable phenotype, along with increased circulating levels of methylglyoxal (MGO) and decreased activity of the anticoagulant plasma protein antithrombin III (ATIII). MGO has been shown to inhibit ATIII activity in vitro, however the mechanism of inhibition is incompletely understood. As such, we designed this study to investigate the kinetics and mechanism of MGO-mediated ATIII inhibition. Methods MGO-mediated ATIII inhibition was confirmed using inverse experiments detecting activity of the ATIII targets thrombin and factor Xa. Fluorogenic assays were performed in both PBS and plasma after incubation of ATIII with MGO, at molar ratios comparable to those observed in the plasma of diabetic patients. LC-coupled tandem mass spectrometry was utilized to investigate the exact mechanism of MGO-mediated ATIII inhibition. Results and conclusions MGO concentration-dependently attenuated inhibition of thrombin and factor Xa by ATIII in PBS-based assays, both in the presence and absence of heparin. In addition, MGO concentration-dependently inhibited ATIII activity in a plasma-based system, to the level of plasma completely deficient in ATIII, again both in the presence and absence of heparin. Results from LC-MS/MS experiments revealed that MGO covalently adducts the active site Arg 393 of ATIII through two distinct glyoxalation mechanisms. We posit that active site adduction is the mechanism of MGO-mediated inhibition of ATIII, and thus contributes to the underlying pathophysiology of the diabetic hypercoagulable state and complications thereof.

Thrombin generation and platelet activation in cytoreductive surgery combined with hyperthermic intraperitoneal chemotherapy - A prospective cohort study

“…In this study, glucose concentration increased during surgery, peaked during chemotherapy (related to the dextrose 5% solution used for chemotherapy instillation) but continued to be increased on day 3 and 7 postoperative [ 14 ]. In diabetic patients, the hyperglycemic byproduct methylglyoxal impairs anticoagulant activity through covalent adduction of antithrombin III [ 57 ]. Acute hyperglycemia has been related with a transient increase in thrombin generation [ 58 ].…”

Coagulatory Defects in Type-1 and Type-2 Diabetes

“…Fibrinogen is not the only protein affected by poor glycaemic control; plasminogen, the precursor of plasmin, undergoes increased glycation in individuals with T1DM, thereby leading to a reduced fibrinolytic-activity of plasmin [51]. Furthermore, the activity of antithrombin is inhibited by methylglyoxal, a by-product of hyperglycaemia [56]. In addition, in healthy individuals subjected to combined hyperglycaemia and hyperinsulinemia, the tissue factor pathway has been shown to be increasingly activated (compared to an euglycemia-hyperinsulinemia group), as reflected by elevated concentrations of activated factor VII and tissue factor pathway inhibitor, as well as in an increase in factor VII activity [57].…”

Type 1 Diabetes Mellitus in the SARS-CoV-2 Pandemic: Oxidative Stress as a Major Pathophysiological Mechanism Linked to Adverse Clinical Outcomes

“…Simultaneously, the concentrations of anti-coagulant proteins, including protein C and protein S, are reduced [ 136 , 137 ]. The activity of antithrombin is also impaired by methylglyoxal, a by-product of hyperglycemia [ 138 ]. Regarding the anti-fibrinolytic proteins, the concentrations of thrombin-activatable fibrinolysis inhibitor and 2-macroglobulin are elevated in patients with T1DM [ 139 , 140 ].…”