Preston, R. J. | bioRxiv Science

bioRxiv2023

GLYCOLYTIC REPROGRAMMING FUELS MYELOID CELL-DRIVEN HYPERCOAGULABILITY

BackgroundMyeloid cell metabolic reprogramming is a hallmark of inflammatory disease, however, its role in inflammation-induced hypercoagulability is poorly understood. Objective/MethodsUsing novel myeloid cell-based global haemostasis assays and murine models of immunometabolic disease, we evaluated the role of inflammation-associated metabolic reprogramming in regulating blood coagulation. ResultsGlycolysis was essential for enhanced activated myeloid cell tissue factor expression and decryption, driving increased cell-dependent thrombin generation in response to inflammatory challenge. Similarly, inhibition of glycolysis enhanced activated macrophage fibrinolytic activity via reduced plasminogen activator inhibitor 1 (PAI-1)-activity. Macrophage polarisation or activation markedly increased endothelial protein C receptor (EPCR) expression on monocytes and macrophages, leading to increased myeloid cell-dependent protein C activation. Importantly, inflammation-dependent EPCR expression on tissue-resident macrophages was also observed in vivo. Adipose tissue macrophages from obese mice fed a high-fat diet exhibited significantly enhanced EPCR expression and APC generation compared to macrophages isolated from the adipose tissue of healthy mice. Similarly, the induction of colitis in mice prompted infiltration of EPCR+ innate myeloid cells within inflamed colonic tissue that were absent from the intestinal tissue of healthy mice. ConclusionCollectively, this study identifies immunometabolic regulation of myeloid cell hypercoagulability, opening new therapeutic possibilities for targeted mitigation of thrombo-inflammatory disease. ESSENTIALSO_LIInflammation-mediated glycolytic reprogramming enables myeloid cell-induced hypercoagulability and antifibrinolytic activity. C_LIO_LI2-Deoxy-D-glucose (2-DG) inhibits the expression of transcription factors necessary for inflammation-induced procoagulant gene expression. C_LIO_LIMyeloid cell membrane regulation of tissue factor procoagulant activity is glycolysis-dependent. C_LIO_LIActivation of myeloid innate immunity dysregulates activated protein C anticoagulant pathway activity. C_LI

immunology↗

bioRxiv2021

Complement Regulator Factor H is a Cofactor for Thrombin in both Pro- and Anticoagulant Roles

BackgroundComplement FH (FH) is a key regulator of complement activity whereas thrombin (FIIa) is central to hemostasis with both pro- and anticoagulant functions. Both have separately been shown to have auxiliary activities across the two systems. The purpose of this study was to determine the effect of FH on pro- and anti-coagulant functions and investigate the interaction between FH and thrombin. MethodsTail bleeding time and hemolysis were measured in FH-deficient mice (CFH-/-). Activated partial thromboplastin time (aPTT) was determined in FH-depleted human plasma. FH effect on fibrin clot generation was investigated in turbidity assays and on activated protein C (APC) generation. Binding affinity of thrombin with FH was determined using surface plasmon resonance (SPR). ResultsTail bleeding time in CFH-/- mice was significantly prolonged compared to wild type mice. The aPTT in FH-depleted human plasma was elevated compared to normal plasma and restored by adding back FH to depleted plasma. Accordingly, FH enhanced thrombin-mediated fibrin clot generation by shortening lag time, increasing rate of clot formation and maximum turbidity, and affected clot structure. Despite this, FH also increased the rate of thrombin-mediated protein C (PC) activation, both in the presence and absence of soluble recombinant thrombomodulin (TM). Nanomolar affinity binding of FH with thrombin, but not prothrombin, was confirmed. ConclusionComplement FH binds thrombin with strong affinity and acts as a novel cofactor that enhances both pro- and anticoagulant actions of thrombin. These data highlight an important role for FH in hemostasis. Key pointsO_LIAbsence of FH prolongs tail bleeding time in CFH-/- mice and activated partial thromboplastin time (aPTT) is elevated in human FH-depleted plasma. C_LIO_LIFH acts a cofactor for thrombin by enhancing fibrin generation, altering fibrin clot structure and enhancing TM-thrombin mediated protein C activation C_LI

immunology↗