Hemophilia A is caused by the lack of functional blood-clotting factor VIII. We have used retrovirus-mediated gene transfer to generate various cell lines, rodent as well as human, that secrete the human factor VIII protein. To study whether transplantation of genetically modified fibroblasts is a feasible approach for gene therapy of hemophilia A, we implanted the factor VIII-secreting cells into immune-deficient mice. Implantation of factor VIII-secreting primary human skin fibroblasts resulted in long-term persistence of the transplanted cells; cells recovered from the implants up to 2 months post-implantation still had the capacity to secrete factor VIII when regrown in tissue culture. However, we were unable to detect any human factor VIII in plasma samples of the recipient mice. The absence of human factor VIII in the recipients' plasma is shown to be due neither to (epigenetic) inactivation of the retroviral vector in vivo, nor to inability of the stationary cells to secrete factor VIII protein. However, we did note a rapid clearing of the human factor VIII: CAg from plasma upon intravenous injection of plasma-derived human factor VIII in mice (t1/2 < 60 min vs. 10 hr in humans). This phenomenon can fully explain the apparent absence of human factor VIII in the recipients' plasma.