A genetic approach toward defining the role of Factor X in development and coagulation

Abstract

Activated Factor X (FX) is a vitamin K-dependent serine protease that plays a crucial role in blood coagulation by converting prothrombin to thrombin. There are no humans with complete deficiency of FX. The overall objective of this study is to investigate the role of FX in development and coagulation by generating FX-deficient mouse models and determining the embryonic pattern of FX expression. First, a FX knock-out mouse model was engineered by eliminating FX exon 8 and its 3’ flanking sequence; this resulted in embryonic or perinatal lethality in FX -/- mice. Approximately 50% of FX -/- embryos died between days 10-12 of development. Fatal bleeding in the remaining FX(-/-) animals occurred soon after birth. The cause of embryonic lethality remains unclear. Next, a viable mouse model of FX deficiency was generated. These mice expressed a FX variant with normal antigen levels but a low level of activity (4-9% in humans carrying the analogous defect, Pro343→Ser, termed FX Friuli). Homozygous mice had FX coagulation activity of ~5.5% of normal, which was sufficient to rescue both embryonic and perinatal lethality. To generate mice with even lower FX activity, FX(F/F) and FX(+/-) mice were crossed. Their offspring, FX(-/F) mice, indeed had even lower FX activity (1-3% of normal) and nonetheless also showed complete rescue of embryonic and perinatal lethality. Therefore, while complete absence of FX is incompatible with murine and human survival, minimum FX activity as low as 1% demonstrates sufficiency in rescue of FX knockout mice from lethality. FX(-/F) mice provided an opportunity to assess the relative contributions of maternal and embryonic FX as well as FX activity and antigen to embryonic survival. FX(-/F) mice also facilitated investigation into a developmental role of FX by allowing for examination of FX(-/-) embryos born to mothers with minimal levels of FX activity. Furthermore, FX expression in multiple embryonic tissues was detected by in situ hybridization and immunostaining. Combined, these results represent an initial investigation into defining the role of FX in development.%%%%Ph.D., Biomedical Science – Drexel University, 2005