Normally factor (F) VIII is not expressed in megakaryocytes, but when human FVIII was transgenically expressed in murine megakaryocytes, it was stored in platelet ␣-granules and released at sites of injury. This platelet FVIII (pFVIII) is effective in correcting hemostasis, even in the presence of circulating inhibitors, so it offers a potential gene therapy strategy for hemophilia A. To understand clot development by pFVIII, we have examined clot response to laser injury in both cremaster arterioles and venules in FVIII null mice either infused with FVIII or transgenic for pFVIII. In both sets of vessels, pFVIII is at least as effective as infused FVIII. However, there are temporal and spatial differences in fibrin and platelet accumulation within clots depending on how FVIII is delivered. These differences may be related to the temporal and spatial distribution of the ␣-granular-released FVIII within the developing clot, and may explain the increased frequency and size of embolic events seen with pFVIII. These observations may not only have implications for the use of pFVIII in gene therapy for hemophilia A, but may also have physiologic consequences, explaining why many procoagulant factors are delivered both in the plasma and in platelet ␣-granules. (Blood. 2008;112:1101-1108)
IntroductionWe and others have shown that human B-domainless factor (F) VIII can be expressed in murine megakaryocytes, stored in platelet ␣-granules independent of von Willebrand factor (VWF), and released at sites of injury in hemophilia A FVIII null mice. [1][2][3] Correction of bleeding occurred in FVIII null mice transgenic for platelet-specific FVIII (pFVIII) expression, in spite of there being no detectable plasma FVIII level. In addition, pFVIII is protected from circulating inhibitors, 3,4 a significant problem in the hemophilia A population. 5 Several editorials have, therefore, suggested that this pFVIII gene therapy strategy may be of benefit in hemophilia A patients with problematic inhibitors who do not respond to present-day therapies to eliminate such inhibitors. 6,7 Aside from a megakaryocyte-specific gene therapy, all other FVIII-based gene therapy strategies for hemophilia A correct plasma FVIII levels. 8 We believe that the details of how FVIII released from activated platelets affects clot development must be examined to be assured that there are no untoward side effects before clinical application. Several clotting models have been examined in FVIII null mice, which suggest that pFVIII is at least as efficacious as plasma FVIII. The tail exsanguination model, which involves both arterials and venules of various sizes and which extends over 16 hours, suggested very high relative efficacy for pFVIII, but we have shown that this model may be too sensitive to pFVIII, likely due to hypovolemia with subsequent blood stasis in the tail. 1 A cuticular bleed model, which also involves both arterial and venule injury and which extends over 8 hours, suggested less relative efficacy. 1 Since both the tail exsanguination and c...
