In vitro immunogenicity of cadaver donor bone marrow cells used for the induction of allograft acceptance in clinical transplantation

Background. The cascade of immunological effects brought about by donor bon e marrow cell (DBMC) infusions to induce allograft acceptance in clinical t ransplantation is not fully understood. Aside from acting as immune respond ing and regulatory cells, the infused DBMC also may sensitize the recipient to the donor antigens. Methods. To analyze this stimulatory activity of DBMC, in vitro mixed lymph ocyte cultures (MLC) and cell-mediated lymphocytotoxicity (CML) culture sys tems analogous to the transplant model with DBMC infusion were used. Results. When responding peripheral blood lymphocytes (PBL) from normal vol unteers were placed in culture with suspensions of Ficoll-purified, T cell- depleted, un-irradiated allogeneic DBMC (NT-DBMC), a reaction was seen in b oth MLC and CML. However, when compared to allogeneic spleen cells as stimu lating cells, the responses to NT-DBMC were of markedly lower magnitude and were not seen when the NT-DBMC was irradiated (3000 R), When responding PB L were stimulated with either NT-DBPMC that had been previously cultured wi th irradiated cells from the responders for 1 meek (activated NT-DBMC), NT- DBMC further depleted of CD15(+) and glycophorin A-positive cells (NT-LP/DB MC), or purified CD34(+) and CD2(+) DBMC subsets, stronger lymphoproliferat ive and cytotoxic responses were observed. Moreover, these responses were n ot abrogated by irradiation of the stimulating DBMC subpopulations. Depleti on of antigen-presenting cells by positive selection of CD3(+) cells from t he responding PBL abrogated MLC and CML reactivity, even when purified NT-L P/DBMC, the most stimulatory cells, were used. This latter observation was in contrast to the responses seen with cultures containing allogeneic stimu lating spleen cell populations. This indicated the requirement for indirect alloantigen presentation, i.e., the failure of these DBMC to stimulate by direct alloantigen presentation. NT-DBMC was able to stimulate responding P BL in secondary MLC and CML responses with an equivalent magnitude, irrespe ctive of whether the stimulators were spleen cells or NT-DBMC. Finally, the MLC and CR-IL responses were inhibited by tacrolimus (FR506), mycophenolic acid (MPA), and cyclosporine (CsA) in a dose dependent manner, in contrast to previously observed refractoriness of DBMC preparations to these agents if DBMC was tested as responder cells or in modulatory assays. Conclusions. These results indicate that DBMC are able to function as effec tive in vitro stimulators, but only by indirect antigen presentation, and t hat the immune responses mediated by them can be downregulated by their own inherent suppressive nature, an effect that can be enhanced by the presenc e of immunosuppressive drugs.