Volume 41, Issue 5, Pages 714-720 (November 2004) Patterns of transgene expression and viral clearance from the transplanted liver following ex vivo adenovirus-mediated gene transfer  Gideon Zamir, Andrew E. Gelman, Kim M. Olthoff, Fotini Debonera, Xavier Aldeguer, Abraham Shaked  Journal of Hepatology  Volume 41, Issue 5, Pages 714-720 (November 2004) DOI: 10.1016/j.jhep.2004.07.008 Copyright © 2004 European Association for the Study of the Liver Terms and Conditions

Fig. 1 (I) The effect of immunosuppression on the level and persistence of transgene (lacZ) expression in the transplanted liver following ex vivo infection of the liver graft with AdlacZ. (A) Syngeneic OLT (Lewis donor to Lewis recipient). Recipients received no immunosuppression, Cyclosporine A, or transplanted with grafts co-infected with AdCTLA-4Ig. (B) Allogeneic OLT (ACI donor to Lewis recipient) or ACI to nude (athymic) recipients. The level of expression was determined by counting the number of hepatocytes showing positive X-gal staining. (II) Disappearance of β-gal expression from the transplanted liver following AdlacZ ex vivo infection. X-gal staining of representative liver sections at different times after allogeneic liver transplantation (ACI to Lewis). Liver grafts were infected ex vivo with AdlacZ and AdCTLA-4Ig. Despite efficient control of the alloimmune response by local expression of CTLA-4Ig, lacZ expression ceases by day 30 after transplantation. Journal of Hepatology 2004 41, 714-720DOI: (10.1016/j.jhep.2004.07.008) Copyright © 2004 European Association for the Study of the Liver Terms and Conditions

Fig. 2 Clearance of adenovirus DNA from the transplanted liver. Agarose gel analysis of different AdlacZ DNA sequences after PCR amplification. Whole liver DNA was analyzed at different times after syngeneic liver transplantation under no immunosuppression, Cyclosporine treatment or AdCTLA-4Ig co-infection. Smaller DNA fragments (E4 0.6kb>1kb lacZ>3kb lacZ) persist longer in the liver with no apparent prolongation by immunosuppression. Journal of Hepatology 2004 41, 714-720DOI: (10.1016/j.jhep.2004.07.008) Copyright © 2004 European Association for the Study of the Liver Terms and Conditions

Fig. 3 Post transplantation liver damage and hepatocyte turnover. (A) BrdU immunohistochemistry. Hepatocyte DNA replication is demonstrated by BrdU immunohistochemistry in normal liver and 10 days after ex vivo gene transfer and transplantation. Brown stained nuclei represent BrdU-positive hepatocytes. In the allogeneic or syngeneic setting, there is only mild hepatocyte turnover (≈10% of hepatocytes) after adenovirus infection and transplantation. (B) AST levels following transplantation of syngeneic or allogeneic liver grafts. In rats receiving unmanipulated syngeneic grafts, there is a mild transient elevation in AST most probably due to ischemia-reperfusion injury. Adenovirus-mediated gene transfer per se does not result in additional liver injury. In allogeneic transplantation there is massive liver damage in untreated recipients which is mostly abrogated by local expression of CTLA-4Ig. Journal of Hepatology 2004 41, 714-720DOI: (10.1016/j.jhep.2004.07.008) Copyright © 2004 European Association for the Study of the Liver Terms and Conditions