Hemophilia A(HA)is an inherited X-linked recessive bleeding disorder caused by coagulant factor Ⅷ(FⅧ)deficiency.Previous studies showed that introduction of mesenchymal stem cells(MSCs)modified by FⅧ-expressing retrovirus may result in phenotypic correction of HA animals.This study aimed at the investigation of an alternative gene therapy strategy that may lead to sustained FⅧ transgene expression in HA mice.B-domain-deleted human FⅧ(hFⅧBD)vector was microinjected into single-cell embryos of wild-type mice to generate a transgenic mouse line,from which hFⅧBD-MSCs were isolated,followed by transplantation into HA mice.RT-PCR and real-time PCR analysis demonstrated the expression of hFⅧBD in multi-organs of recipient HA mice.Immunohistochemistry showed the presence of hFⅧBD positive staining in multi-organs of recipient HA mice.ELISA indicated that plasma hFⅧBD level in recipient mice reached its peak(77 ng/mL)at the 3rd week after implantation,and achieved sustained expression during the 5-week observation period.Plasma FⅧ activities of recipient HA mice increased from 0%to 32%after hFⅧBD-MSCs transplantation.APTT(activated partial thromboplastin time)value decreased in hFⅧBD-MSCs transplanted HA mice compared with untreated HA mice(45.5 s vs.91.3 s).Our study demonstrated an effective phenotypic correction in HA mice using genetically modified MSCs from hFⅧBD transgenic mice.
