Expression of xenogeneic MHC class II molecules in HLA‐DR⁺ and ‐DR^– cells: influence of retrovirus vector design and cellular context

Abstract

We recently established that molecular chimeras of major histocompatibility complex (MHC) class II molecules, created via retroviral transfer of allogeneic class II cDNAs into bone marrow cells (BMCs), alleviated complications associated with mixed BMC chimeras while leading to T cell tolerance to renal grafts sharing the transferred class II. Initially demonstrated for allogeneic transplants in miniature swine, this concept was extended to T-dependent antibody (Ab) responses to xenogeneic antigens (Ags) in the pig --> baboon combination. Successful down-regulation of T cell responses appeared, however, to be contingent on a tight lineage-specific expression of transferred class II molecules. The present studies were, therefore, designed to evaluate the influence of construct design and cellular environment on expression of retrovirally transferred xenogeneic class II cDNAs. Proviral genomes for pig class II SLA-DR expression, differing only at the marker neo(r) or enhanced green fluorescent protein (EGFP) gene, showed increased membrane SLA-DR density on HLA-DR(-) fibroblasts as well as HLA-DR(+), TF-1 erythroleukemia cells. More importantly, HLA-DR(+) human B cell lines, although efficiently transduced with pig DR retroviruses, exhibited unstable surface pig DR. Surface pig DR- B cells, nevertheless, stimulated autologous human T cells pre-sensitized to pig Ags, a proliferation likely occurring through presentation of class II-derived peptides. Collectively, these data suggest that surface expression of transferred class II molecules is not related to the ability of recipient cells to synthesize xenogeneic class II molecules but rather to their Ag processing capacities.

Intracellular MHC Class II Controls Regulatory Tolerance to Allogeneic Transplants

“…Additional data from our group support the MHCII peptide hypothesis. We observed that human B cell lines, transduced with a pig MHCII DR gene, did not express surface pig DR but stimulated autologous human T cell proliferation, which was only blocked by anti-human MHCII mAbs (41). …”

Potential role of major histocompatibility complex class II peptides in regulatory tolerance to vascularized grafts.

“…Unexpectedly, the Tg product was not expressed at the surface of the manipulated BM cells. This finding was confirmed in vitro by showing that prototypic class II committed cell lines, such as dendritic, B, and endothelial cells, were unable to produce membrane-bound Tg class II heterodimers after transduction (23).…”

“…Further mechanistic studies revealed that class II ϩ B-cell lines transduced by foreign Tg class II cDNA, leading to negative expression of surface Tg molecules, were still able to trigger proliferation of self-CD4 ϩ T cells, strongly suggestive of peptide presentation of the transgenic class II product on B-cell class II heterodimers (23). Antibody-blocking experiments confirmed that the Tg class II-derived peptides were preferentially presented on surface class II molecules of the B cells.…”

Regulation of T-cell functions by MHC class II self-presentation

“…Nonetheless, foreign class II chains sequestered in transduced APCs appeared to be processed as peptides docked selectively on APC surface class II heterodimers. Indeed, the proliferation of T helper (Th) cells to self-class II þ APCs, transduced with foreign class II, could be blocked by monoclonal antibodies (mAbs) to APC class II, but not to transgenic class II [23]. Because bone-marrow class II þ APCs are instrumental in the induction of T-cell tolerance [24], they might additionally be key participants in regulatory tolerance induction following class II gene therapy.…”