Adenovirus-mediated gene transfer in the transplant setting: early events after orthotopic transplantation of liver allografts expressing TGF-beta1.

ABSTRACT

We hypothesized that adenovirus mediated gene transfer of TGF-beta1 into liver grafts would enhanced local expression of this recombinant protein and down-regulate inflammatory and alloreactive immune response. A full length DNA encoding the murine TGF-beta1 was used to replaced the E1 region of adenovirus type 5 (AdmTGF-beta1). Expression and protein production of biologically active murine TGF-beta1 was tested in AdmTGF-beta1-transduced Hep G2 cells and TGF-beta-sensitive MV1 cells. In the transplant setting, the replication-defective vector was used to perfused cold preserved ACI liver allograft prior to transplantation into Lewis recipients. Control livers were similarly perfused with cold lactated Ringer's solution and were followed without immunosuppression. Animals were sacrificed at 1, 3, and 5 days after transplantation. Intragraft cytokine levels of TNFalpha, and IFNgamma were determined using ELISA and quantitative PCR. TGF-beta1 ELISA of culture supernatants from AdmTGF-beta1 transduced hepatocyte cell line Hep G2 excreted TGF-beta1 in quantities directly correlated with multiplicity of infection (MOI, vectorhepatic cell ratio). The biological activity of the excreted recombinant protein was confirmed by growth inhibition of MV1 TGF-beta-sensitive cells. Enhanced production of TGF-beta1 in transduced allografts was associated with decreased levels of TNFalpha and IFNgamma when compared with nonimmunosuppressed controls. Adenovirus-mediated gene transfer of murine TGF-beta1 into hepatic cells results in the expression of biologically active protein. Transduction of allografts with TGF-beta1 down-regulates TNFalpha and IFNgamma production early after orthotopic transplantation. Graft transduction with TGF-beta1 offers a novel approach to study the effects of single immune modulator on alloreactive immune response, T cell function, and cytokine cascade.