Extra-hepatic sources of FVIII potentially contribute to the coagulation cascade correcting the bleeding phenotype of Hemophilia A mice

A large fraction of FVIII in blood originates from liver sinusoidal endothelial cells although extrahepatic sources also contribute to plasma FVIII levels. Identification of cell-types other than endothelial cells with capacity to synthesize and release FVIII will be helpful for therapeutic approaches in hemophilia A. Recent cell therapy and bone marrow transplantation studies indicated that Kupffer cells, monocytes or mesenchymal stromal cells could synthesize FVIII in sufficient amounts to ameliorate bleeding phenotype in hemophilic mice. To further establish the role of blood cells in expressing FVIII, we studied various mouse and human hematopoietic cell types. We identified FVIII in cells isolated from peripheral and cord blood, as well as bone marrow. Costaining for cell type-specific markers verified that FVIII was expressed in monocytes, macrophages and megakaryocytes. We additionally verified that FVIII was expressed in liver sinusoidal endothelial cells and endothelial cells elsewhere, e.g., in spleen, lungs and kidneys. FVIII was well expressed in sinusoidal endothelial cells and Kupffer cells isolated from human liver, whereas by comparison isolated human hepatocytes expressed FVIII at very low levels. After transplantation of CD34+ human cord blood cells into NOD/SCIDγNull-hemophilia A mice, FACS of peripheral blood showed >40% donor cells engrafted in the majority of mice. In these animals, plasma FVIII activity 12 weeks after cell transplantation was up to 5% and 9 of 12 mice survived tail clip-assay. In conclusion, hematopoietic cells expressed and secrete FVIII in addition to endothelial cells, which should offers further opportunities for understanding mechanisms in FVIII synthesis and replenishment.