175: A New Paradigm for Tissue Regeneration: Preparative Irradiation for Cell-Based Therapies as an Alternative to Organ Transplantation

Abstract

Orthotopic liver transplantation (OLT) is the only curative therapy for patients with terminal liver diseases. A significant number of patients fail to get OLT due to shortage of donor livers. Hepatocyte transplantation (HT) is being explored as a therapeutic alternative to OLT for the treatment of end-stage liver diseases. However, the number of cells that can be safely transplanted into the liver and the inability of the transplanted hepatocytes to proliferate in the host liver currently limit the clinical application of this procedure. We hypothesized that a preparative regimen of hepatic irradiation (HIR), in combination with hepatic mitotic stimuli, would suppress host hepatocellular proliferation and induce post-mitotic death, thus ”making room“ for transplanted, nonirradiated hepatocytes to proliferate preferentially and repopulate the irradiated host liver. C57Bl/6 mice received HIR (0-50 Gy) to the anterior liver lobes after exposing the liver by laparotomy and shielding the posterior liver lobes and other abdominal organs, followed by an injection of a recombinant adenoviral vector expressing human hepatocyte growth factor (Ad-HGF, 5-10 x 1010 particles). One day after HIR, the mice received intrasplenic HT with 0.8-1×106 primary hepatocytes, freshly isolated from congenic, beta-galactosidase-expressing transgenic (ROSA)26 C57Bl/6 mice. Control mice received Ad-HGF alone, or HIR + Ad-LacZ with HT. Animals were sacrificed at 7 days, 1, 2, 4 and 6 months after HT and the liver sections were examined for beta-galactosidase-expressing donor cells after histochemical and hematoxylin-eosin staining. Compared to unirradiated controls, HIR augmented the engraftment of transplanted hepatocytes, with an increase in the number of integrated donor hepatocytes into liver cords by 7 days (p<0.01). There was progressive repopulation of the irradiated anterior liver lobes by transplanted hepatocytes with near-total (80-95%) replacement of the host hepatocytes by the donor cells within 6 months. In contrast, the unirradiated posterior and caudate liver lobes exhibited occasional presence of beta-galactosidase-expressing donor hepatocytes, indicating engraftment without repopulation. H&E staining demonstrated normal liver architecture without any evidence of hepatic radiation injury. Liver function tests of the transplanted animals were normal. Preparative irradiation is routinely used for bone marrow transplantation, but we are the first to use HIR to facilitate the engraftment and host liver repopulation by donor cells after HT in rodent models. HIR significantly enhanced the integration of transplanted cells in the host liver. Furthermore, partial liver irradiation enabled selective repopulation of a liver lobe by the donor cells, thereby increasing the safety of our regimen. Clinically, preparative HIR could be safely administered using radiosurgery or 3-D conformal RT and combined with systemic administration of hepatic growth factors to facilitate HT for 1) the treatment of inherited liver diseases, 2) liver failure, 3) ex vivo hepatic gene therapy, 4) rescuing patients with liver cancer, following chemo-radiation therapy, and 5) expanding human hepatocytes in animal liver for generating animal models for human-specific infections.