Human endothelial cells and fibroblasts express and produce the coagulation proteins necessary for thrombin generation

Clay T Cohen,Nancy A Turner,Joel L Moake

doi:10.1038/s41598-021-01360-w

Clay T Cohen, Nancy A Turner + Show 1 more

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https://doi.org/10.1038/s41598-021-01360-w

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In a previous study, we reported that human endothelial cells (ECs) express and produce their own coagulation factors (F) that can activate cell surface FX without the additions of external proteins or phospholipids. We now describe experiments that detail the expression and production in ECs and fibroblasts of the clotting proteins necessary for formation of active prothrombinase (FV–FX) complexes to produce thrombin on EC and fibroblast surfaces. EC and fibroblast thrombin generation was identified by measuring: thrombin activity; thrombin–antithrombin complexes; and the prothrombin fragment 1.2 (PF1.2), which is produced by the prothrombinase cleavage of prothrombin (FII) to thrombin. In ECs, the prothrombinase complex uses surface-attached FV and γ-carboxyl-glutamate residues of FX and FII to attach to EC surfaces. FV is also on fibroblast surfaces; however, lower fibroblast expression of the gene for γ-glutamyl carboxylase (GGCX) results in production of vitamin K-dependent coagulation proteins (FII and FX) with reduced surface binding. This is evident by the minimal surface binding of PF1.2, following FII activation, of fibroblasts compared to ECs. We conclude that human ECs and fibroblasts both generate thrombin without exogenous addition of coagulation proteins or phospholipids. The two cell types assemble distinct forms of prothrombinase to generate thrombin.

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In a previous study, we reported that human endothelial cells (ECs) express and produce their own coagulation factors (F) that can activate cell surface FX without the additions of external proteins or phospholipids
In order to investigate the formation of prothrombinase complexes on both EC and fibroblast surfaces, we determined whether or not these cells express and produced FV and FII
In our previous study we found that both ECs and fibroblasts produced tissue factor (TF), FVII, FIX, and FX; FII was only detected in E Cs6

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We reported that human endothelial cells (ECs) express and produce their own coagulation factors (F) that can activate cell surface FX without the additions of external proteins or phospholipids. FV is on fibroblast surfaces; lower fibroblast expression of the gene for γ-glutamyl carboxylase (GGCX) results in production of vitamin K-dependent coagulation proteins (FII and FX) with reduced surface binding. This is evident by the minimal surface binding of PF1.2, following FII activation, of fibroblasts compared to ECs. We conclude that human ECs and fibroblasts both generate thrombin without exogenous addition of coagulation proteins or phospholipids. We report experiments, performed without the exogenous additions of proteins or phospholipids, demonstrating that active prothrombinase (FX–FV–FII) complexes can form, and produce thrombin, on human ECs and fibroblasts. Our analysis of thrombin activity, the FII cleavage product (PF1.2), and TAT, indicate that human ECs and fibroblasts participate actively in hemostasis by promoting thrombin generation, albeit via different processes

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