Pig endothelial protein C receptor is functionally compatible with the human protein C pathway.
Xenotransplantation
; 27(2): e12557, 2020 03.
Article
en En
| MEDLINE
| ID: mdl-31556182
BACKGROUND: Endothelial protein C receptor (EPCR) plays an anticoagulant and anti-inflammatory role by promoting the activation of protein C by thrombin bound to thrombomodulin (TBM). Incompatibility between pig TBM and human/primate thrombin is thought to contribute to dysregulated coagulation in pig-to-primate organ xenografts, and expression of human TBM (hTBM) in pigs has shown benefit in preclinical models. However, it is not known whether there are incompatibilities-or molecular barriers-between endogenous pig EPCR (pEPCR) and transgenically expressed human TBM. AIM: To clone and express pEPCR, and determine its function in the human protein C pathway in vitro. METHODS: Pig endothelial protein C receptor cDNA was generated from pig lung RNA by RT-PCR. Primate COS-7 transfectants expressing various combinations of human and pig TBM and EPCR were incubated with human thrombin and human protein C, and tested for TBM cofactor activity. RESULTS: The predicted protein sequence of pEPCR shared 72.3% amino acid sequence identity with hEPCR, and residues critical for protein C binding were conserved. COS-7 cells transfected with hEPCR, pEPCR or vector showed minimal TBM cofactor activity (0.13 ± 0.04, 0.13 ± 0.02 and 0.14 ± 0.06 U, respectively). The cofactor activity of hTBM-transfected cells (1.18 ± 0.29 U) was 8-fold higher than vector-transfected cells (P = .004) and further increased 4-fold and 3-fold by co-transfection with hEPCR (5.01 ± 1.12 U, P = .004) or pEPCR (3.73 ± 0.65 U, P = .003), respectively. CONCLUSIONS: Our data show that pEPCR is largely compatible with the human TBM/thrombin complex, when expressed on COS-7 cells in vitro, promoting the activation of human protein C. These findings suggest that endogenous pEPCR will enhance the activity of transgenic hTBM in the xenograft setting.
Palabras clave
Texto completo:
1
Colección:
01-internacional
Base de datos:
MEDLINE
Asunto principal:
Proteína C
/
Animales Modificados Genéticamente
/
Células Endoteliales
/
Receptor de Proteína C Endotelial
Tipo de estudio:
Prognostic_studies
Límite:
Animals
Idioma:
En
Revista:
Xenotransplantation
Asunto de la revista:
TRANSPLANTE
Año:
2020
Tipo del documento:
Article
País de afiliación:
Australia
Pais de publicación:
Dinamarca