| Co-Investigator(Kenkyū-buntansha) |
MIYAZATO Hiroshi University of the Ryukyus, The First Dept. of Surgery, Instructor, 医学部, 助手 (90284976)
SHIRAISHI Masayuki University of the Ryukyus, The First Dept. of Surgery, Assistant Professor, 医学部・附属病院, 講師 (00264482)
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| Research Abstract |
In this project, we aim to establish an adenovirus mediated gene transfer to the liver graft during cold ischemia, using in-situ perfusion of the liver as a model for orthotopic liver transplantation (OLT). The liver graft was transfected ex-vivo with adenovirus vector, carrying marker LacZ gene (AxCALacZ). Ex-vivo setting should allow target liver to be exposed to a high titer of adenovirus vector (m.o.i > 1x10ィイD13ィエD1) for a long time (> 30 min), in which the adenovirus vector will not be diluted in circulating blood or excluded by a host immunological system. Efficient gene transfer to the target liver was achieved by the ex-vivo transfection both in the rats and pigs. Additional systemic inmmunosuppression of the host is also required for successful gene transfer to the porcine liver.
In the porcine-to-human model of xenogeneic liver transplantation, human complement activation can not be inhibited by porcine complement regulator proteins (pCRP), due to the molecular incompatibilit
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ies beyond species. To overcome such incompatibilities, we have tested the adenovirus mediated gene transfer of triple human complement regulator proteins (hCRP; CD46, CD55, CD59) to the porcine liver, using the same ex-vivo procedure as described above. The transfected porcine livers were harvested 24 hours after gene transfer and then were subjected to xenogeneic perfusion with fresh human blood. The gene transfer of CRPs was recognized both in the vascular endothelium and in the sinusoidal endothelium in the porcine liver. Triple CRPs, compared to a single CRP, effectively inhibited human complement activation in the xenoperfused porcine liver, thus resulting in a higher complement levels in the perfusate. Although the eventual destruction of the xenograft porcine liver could not be avoided, the adenovirus mediated gene transter of human CRPs was thought to be useful in inhibiting xenograft rejection in combination with other therapeutic modalities, such as natural antibody depletion and/ or anti-homeostasis. Less
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