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Identification of a subpopulation of reactive large granular mononuclear cells in allogeneic heart transplantation.

This study is designed to test the hypothesis that specific morphologic attributes of peripheral blood mononuclear cells, measurable by flow cytometry, are correlated with the timing and the intensity of allograft injury during the development of heart rejection. A pig model of major histocompatibility complex-mismatched heterotopic heart transplantation with (n = 5) and without (n = 5) cyclosporine administration was monitored serially be telemetered electrocardiography and endomyocardial biopsies. Flow cytometric analysis of peripheral blood mononuclear cells revealed the emergence of a discrete subpopulation of peripheral blood mononuclear cells (7.8% +/- 1.0% and 8.5% +/- 0.9% before transplantation to 16.5% +/- 1.3% and 19.4% +/- 3.0% after transplantation in the untreated and the cyclosporine-treated groups, respectively, p < 0.05), exhibiting characteristic changes in forward and 90-degree light scatter, indicative of increased cell size and granularity, and possibly representing monocytes or large granular lymphocytes. Lymphocyte cell surface-marker studies indicated that 62% of these cells are DH59B+ (monocyte/granulocyte). Because intracellular free calcium is an important second messenger in lymphocyte activation we measured intracellular free calcium by flow cytometry using fluo-3. This subpopulation of cells was found to have similar intracellular free calcium when compared to normal-sized lymphocytes (104 +/- 7 nmol/L versus 101 +/- 5 nmol/L, respectively). We conclude that this lymphocyte subset detected by flow cytometry represents specifically reactive cells that are associated with incipient allograft rejection.