1. Coagulation process proceeds on cultured human mesangial cells via expression of factor V
- Author
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Eri Muso, Ning Liu, Takahiko Ono, Hitoshi Kusano, Katsuo Suyama, Shigetake Sasayama, Tadashi Kamata, Fumiaki Nogaki, and Kenji Kasuno
- Subjects
medicine.medical_specialty ,Antibodies ,Fibrin ,chemistry.chemical_compound ,Tissue factor ,Thrombin ,Internal medicine ,medicine ,Humans ,RNA, Messenger ,mesangioproliferative glomerulonephritis ,Cells, Cultured ,biology ,Mesangial cell ,Tumor Necrosis Factor-alpha ,business.industry ,Factor X ,Factor V ,IgA nephropathy ,Immunohistochemistry ,Molecular biology ,Glomerular Mesangium ,intraglomerular coagulation ,Endocrinology ,Models, Chemical ,chemistry ,Coagulation ,Nephrology ,Cell culture ,Factor Xa ,biology.protein ,cross-linked fibrin ,tumor necrosis factor-α ,factor X ,business ,medicine.drug - Abstract
Coagulation process proceeds on cultured human mesangial cells via expression of factor V.BackgroundIn a previous clinicopathological study, we observed mesangial factor V expression accompanied by the intact form of cross-linked fibrin deposition in the active type of IgA nephropathy. The conversion of prothrombin to thrombin by factor Xa is potently accelerated more than 104-fold by the presence of factor V, which is a membrane-bound cofactor. Another membrane-bound cofactor, tissue factor, is known to play an initiating role in the coagulation cascade and to be synthesized in mesangial cells (MCs) by the stimulation of tumor necrosis factor-α (TNF-α). However, the synthesis of factor V, which plays on the terminating stage of prothrombin activation, has not been reported previously in MCs by in vitro study. Our current study tested the coagulation process via expression of factor V by the stimulation of proinflammatory cytokine, TNF-α, in cultured human MCs.MethodsTo evaluate factor V protein expression, immunoperoxidase staining with densitometric evaluation and Western blot analysis were conducted after stimulation of TNF-α. To test factor V activity, stimulated MCs were incubated in combination with factor Xa, prothrombin, fibrinogen and factor XIII, and fibrin production on MCs was assessed after immunoperoxidase staining on the cell surface. In a blocking test using an antibody against factor V, suppression of fibrin production was evaluated to clarify the role of factor V activity. For the evaluation of factor V mRNA expression in cultured human MCs, in situ hybridization and Northern blot analysis were performed.ResultsFactor V protein expression in MCs after TNF-α stimulation increased both time- and dose-dependently. As a marker of factor V activity with exogenous factor Xa, fibrin production on TNF-α–stimulated MCs was increased in a time-dependent manner and was inhibited by the addition of anti-factor V antibody. Factor V mRNA was identified in MCs by in situ hybridization and showed an increase after stimulation with TNF-α on Northern blot analysis.ConclusionsOur data suggest that the coagulation process proceeds on MCs as the result of increased expression of endogenous factor V activity on its cell surface in cooperation with exogenous factor Xa.
- Published
- 2001