Introduction: Alloantibodies contribute significantly to the development of chronic rejection, the leading cause of long-term graft failure in organ transplantation. As interest in treating and preventing antibody-mediated rejection continues to grow, there is an increasing need to develop sustainable nonhuman primate models of sensitization for development of effective therapies. Currently, no established consensus for flow cytometric immunophenotyping of plasma cells exists for nonhuman primates. Here, we characterize bone marrow aspirates with plasma cell markers identified in humans, and further analyzed the populations for the expression of possible antigen targets.

Methods: Four adult rhesus macaques underwent skin grafting for sensitization. Donor specific antibody production was confirmed in all animals. Bone marrow aspirates were collected at least 10 weeks after skin grafting, and analyzed with polychromatic flow cytometry. Clones for all antibodies used were confirmed as rhesus-reactive through the NIH nonhuman primate reagent resource. Mann-Whitney U test was used for statistical analysis, with p<0.05 indicating statistical significance.

Results: Without immunosuppression, skin grafts rejected by 10.73 ± 3.86 days. CD38+CD138+ cells represented an average of 1.30% ± 0.39 of viable, non-CD20, CD19+CD27+ cells expressing intracellular IgG+ (Figure A). Of these plasma cells, 89.9% ± 2.8 expressed LFA-1 and 20.5% ± 3.9 expressed VLA-4 (Figure B). Compared to CD20+ B cells, plasma cells had increased constitutive expression of CD80 (MFI = 1098.3 vs 2742.3, p=0.021).

Conclusions: Immune markers identified in normal human plasma cells (CD20loCD19+CD27+IgG+CD38+CD138+) were used to identify a consistent population of plasma cells in rhesus macaques. Flow cytometric analysis may be used to supplement current techniques for monitoring antibody secreting plasma cells, such as ELISPOT. Based on surface expression on these plasma cells, adhesion molecule LFA-1 and costimulation marker CD80 may offer potential therapeutic targets for desensitization.

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