Membrane-bound or soluble truncated RT1.A^a rat class I major histocompatibility antigens induce specific alloimmunity

Transfectants that express membrane-bound (MB) or secrete soluble truncated (TR) rat class I RT1.A^a major histocompatibility (MHC) antigens induce alloimmunity in vivo. The MB-RT1.A^a was produced by transfecting the full-length RT1.A^a cDNA, including the α₁, α₂, and α₃, transmembrane and intracellular domains. The TR-RT1.A^a cDNA insert included only the extracellular α₁, α₂, and α₃ domains; a stop codon was placed in front of the transmembrane domain. Following full-length sequencing, MB-RT1.A^a and TR-RT1.A^a cDNAs were translated in vitro into glycosylated MB-RT1.A^a (45 kDa) and TR-RT1.A^a (36 kDa) proteins, respectively. Each cDNA construct was individually subcloned into the pSG5 vector before transfection into Buffalo (BUF; RT1^b) hepatoma cells. FACscan analysis with anti-RT1.A^a-specific R2/15S monoclonal antibody (MAb) confirmed surface expression of RT1.A^a molecules on the MB-RT1.A^a, but not on the TR-RT1.A^a, transfectants. In contrast, enzyme-linked immunoadsorbent assays documented the presence of soluble RT1.A^a molecules in supernates from cells transfected with the TR- RT1.A^a, but not from cells transfected with the MB-RT1.A^a, cDNA. Subcutaneous injection of MB-RT1.A^a or TR-RT1.A^a transfectants to BUF or Wistar Furth (WF; RT1(u)) rats induced accelerated rejection of ACI (RT1^a) but not third-party Brown Norway (RT1(n)) heart allografts. Furthermore, supernates of TR-RT1.A^a, but not of MB-RT1.A^a, transfectants immunized WF hosts toward ACI hearts. Thus, both intact MB-RT1.A^a and soluble TR-RT1.A^a class I alloantigensinduce potent sensitization against alloantigens.