Production and control of coagulation proteins for factor X activation in human endothelial cells and fibroblasts

Abstract

Human endothelial cells (ECs) synthesize, store, and secrete von Willebrand factor multimeric strings and coagulation factor (f) Viii. it is not currently known if ecs produce other coagulation factors for active participation in coagulation. We found that 3 different types of human ECs in primary culture produce clotting factors necessary for fX activation via the intrinsic (fViii-fiX) and extrinsic (tissue factor [TF]-FVII) coagulation pathways, as well as prothrombin. Human dermal fibroblasts were used as comparator cells. TF, FVII, FIX, FX, and prothrombin were detected in ECs, and TF, FVII, FIX, and FX were detected in fibroblasts. In addition, FVII, FIX, FX, and prothrombin were detected by fluorescent microscopy in ec cytoplasm (associated with endoplasmic reticulum and Golgi proteins). fX activation occurred on human umbilical vein EC surfaces without the addition of external coagulation proteins, proteolytic enzymes, or phospholipids. tumour necrosis factor, which suppresses the generation of activated protein C and increases TF, augmented FX activation. Fibroblasts also produced TF, but (in contrast to ECs) were incapable of activating FX without the exogenous addition of FX and had a marked increase in FX activation following the addition of both FX and FVII. We conclude that human ecs produce their own coagulation factors that can activate cell surface fX without the addition of exogenous proteins or phospholipids. Human endothelial cells (ECs) help maintain blood flow and prevent extra-vascular blood loss. The precise molecular contributions of ECs to haemostasis are unknown. We conducted experiments to determine if human ECs can produce coagulation proteins. Comparative experiments were performed with human fibroblasts, sub-EC vascular wall components that are exposed to blood upon vascular wall injury. For many years hepatocytes were considered the predominant (or exclusive) site of coagulation factor production 1-9. In recent murine and human studies, however, liver sinusoidal ECs (LSECs), rather than hepatocytes, were demonstrated to be the primary cellular source of factor (F) VIII 10-13. Other extra-hepatic vascular ECs, including human umbilical vein ECs (HUVECs) and human glomerular microvascular ECs (GMVECs), have also been shown to produce FVIII 14-18. FVIII is stored in EC Weibel-Palade bodies (WPBs), and secreted along with ultra-large (UL) von Willebrand factor (VWF) multimers 18. The FVIII data suggest that ECs may have a more active role in coagulation factor production than previously appreciated. ECs produce surface regulatory proteins that prevent excessive coagulation. These include thrombomodulin (TM), endothelial protein C receptor (EPCR), tissue factor pathway inhibitor (TFPI), and protein C (PC) 19-22. TM-bound thrombin converts PC that is bound to EPCR into activated protein C (APC). APC (and protein S [PS]) inactivates activated FVIII and FV 23-25 and, therefore, limits the functions of FVIII-FIX (intrinsic tenase complex) and FX-FV (prothrombinase complexes). TFPI inhibits t...

COVID-19-associated vasculitis and vasculopathy

“…Because tissue factor, expressed locally at sites of vessel wall injury and from circulating monocytes [21] plays a pivotal role in the thrombotic phenotype of coronary atherosclerosis, an important surface protein antagonist, tissue factor pathway inhibitor (TFPI), has gained considerable attention. A variety of other surface proteins, including protein C, protein S, protein Z, nitric oxide, glycosaminoglycans, β2-glycoprotein 1, tissue plasminogen activator, urokinase-like plasminogen activator and thrombomodulin among others play critical roles [22][23][24].…”

Human Endothelial Cells And Fibroblasts Express And Produce The Coagulation Proteins Necessary For Thrombin Generation

“…Fibroblast supernatant has higher concentrations of PF1.2 (Fig. 2b) and FX than HUVECs, 6 whereas fibroblast lysates have lower amounts of PF1.2 and FX than HUVECs. Furthermore, similar levels of FVII and FIX were measured in fibroblast and HUVEC supernatants despite HUVEC lysate containing 10-fold more FVII and 5-fold more FIX than in fibroblast lysate.…”

“…and FX can be activated on human umbilical vein EC (HUVEC) surfaces without the addition of exogenous proteins, proteolytic enzymes, or phospholipids. 6 Furthermore, TF, FVII, FIX, and FX were detected in human fibroblasts, that are, along with ECs, components of human vascular walls. The cellular source of FV has previously been enigmatic.…”

The Association of Syndecan-1, Hypercoagulable State and Thrombosis and in Patients With Nephrotic Syndrome

“…Endothelial damage caused the release a wide variety of pro-coagulant molecules, such as tissue factors (TF) 13 and Factor V (FV). 14 In recent studies, SDC-1 also served as a promising marker for identifying endothelial dysfunction in NS patients. 8 Highly increased level of SDC-1 was considered as an independent predicator for hypercoagulable state in our study.…”