CLASS I MHC PROTEIN TOLEROGENS FOR RAT CARDIAC ALLOGRAFTS ACROSS MAJOR HISTOCOMPATIBILITY BARRIERS.

Abstract

241 Soluble rat heavy chain class I MHC proteins were produced with an E. coli-based protein expression system. Site-directed mutagenesis using PCR-based SOEing was utilized to produce allochimeric class I major histocompatibility complex (MHC) proteins. The allochimeric protein α1hn-RT1.Ac contains donor Brown Norway(BN) amino acid (a.a.) residues of the highly polymorphic donor alpha-1-helical region on the background of the recipient PVG (RT1.Ac) class I MHC a.a. sequence. In a tolerogenic protocol, 50 μg wild-type RT1.An injected by portal vein (PV) at the time of transplantation to PVG rats plus a 7-day course of oral cyclosporine (CsA, 4 mg/kg/day) prolonged the survival of BN cardiac allografts to a mean survival time (MST) of>200days (n=7, p=0.03). Administration of CsA alone produced a MST of 11.0±1.0 days, (n=25). Administration of 10 μg of α1hn-RT1.Ac via PV at the time of transplantation to PVG rats with CsA prolonged the survival of BN heart allograft to a MST>200days, (n=6, p=0.006). In an oral tolerogenic protocol, PVG rats were pre-treated with 100 μg wild-type RT1.An daily for 5 days(days −4 to 0) before transplant of a BN cardiac allograft plus peri-operative CsA yielded a MST>180 days (n=3, p=0.001). Using a 3H-thymidine uptake assay, analysis of the pan-T-cells from long-term allograft survivors demonstrated markedly decreased proliferation upon challenge with irradiated BN splenocytes in vitro as compared to T-cells from rats with rejected allografts and naïve untreated rats. The data demonstrate that the MHC class I tolerogens are effective for the induction of specific tolerance across MHC barriers.