Induction of specific allograft immunity by soluble class I MHC heavy chain protein produced in a baculovirus expression system

Spodoptera Frugiperda (Sf9) insect cells secreted a class I MHC RT1.A(a) heavy chain protein when infected with baculovirus that bore a construct that contained a honeybee melittin secretion (ms) signal attached to RT1.A(a) cDNA. The RT1.A(a) heavy chain protein in the culture supernatant and cell lysate immunoprecipitated in the presence of 5 individual anti-RT1.A(a)-specific mAb. As was revealed by densitometric analysis, the ms signal increased the production (7- to 17-fold) and secretion (20- to 47-fold) of RT1.A(a) protein by Sf9 cells (compared with RT1A(a)- Sf9 cells without the ms signal). Subcutaneous immunization with secreted RT1.A(a) heavy chain proteins of Wistar-Furth (WF;RT1(u)) rats (day -4) accelerated the rejection of ACI (RT1(a)), but not third-party Brown Norway (BN;RT1(n)), heart allografts from 5.9+/-0.5 days in controls to 4.0+/-0.0 days (P<0.001); cell lysate from RT1.A(a)-Sf9 or ms/RT1.A(a)-Sf9 cells reduced ACI heart allograft survival to 3.8+/-0.4 days or 3.7+/-O.5 days, respectively (P<0.001). Indirect presentation of RT1.A(a) heavy chain proteins by syngeneic macrophages shortened the survival of RT1.A(a)-disparate PVG.RS (RT1.A(a)D(u)B(u)C(u)) heart allografts in PVG.1U (RT1(u)) hosts from 6.3+/-0.5 days in controls to 4.0+/-0.0 days (P<0.01). Finally, RT1.A(a) heavy chain proteins injected into the thymus or into the portal vein (day -14) in combination with anti-T cell receptor mAb (days -14 and -13) induced indefinite survival of ACI liver allografts in Lewis (RT1(1)) recipients (>250 days). Thus, indirect presentation of soluble class I MHC heavy chain proteins (produced in a baculovirus/Sf9 cell system) may either sensitize or induce tolerance in the same fashion as native class I MHC alloantigens expressed on donor tissues.