PRETREATMENT WITH LYMPHOCYTE SUBPOPULATIONS AND RENAL ALLOGRAFT SURVIVAL IN THE RAT1

Abstract

The induction of antigen-induced immunosuppression by highly purified lymphocyte populations of different phenotypes was investigated in a rat renal allograft model (LEW-to-DA). Lymphocyte populations were prepared from Lewis spleen cells by rosette depletion and flow cytometric separation. Rosette depletion was performed using appropriate monoclonal reagents. B cells were prepared from spleen lymphocytes by rosette depletion of T cells, using three monoclonal antibodies: MRC OX19 (anti-T-cell), MRC OX8 (anti-T-cytotoxic/suppressor) and MRC W3/25 (anti-T-helper). T cells were purified by rosette depletion of B cells with MRC OX12 (anti-rat-K-chain). After depletion the lymphocyte populations were analyzed by flow cytometry and the purity of each preparation determined. B cells were 93.6% pure and T cells were 92.4% pure. Intravenous injection of 5×106 of these LEW B or T cells one week before transplantation of a LEW kidney into a DA recipient resulted in was indefinite renal allograft survival (median survival time [MST] > 100 days). Purification of the enriched B and T cell preparations by flow sorting resulted in highly purified populations of spleen B cells (99.3%) and T cells (98.2%). Further fractionation of the T cells into cells of T helper (99.6%) (i.e., W3/25-positive) and T cytotoxic/suppressor phenotype (99.4%) (i.e., MRC OX8-positive) was performed. While purified B cells, T cells, and T helper cells, given at a dose of 106 cells intravenously one week before transplantation resulted in long-term renal allograft survival (MST > 100 days), cells of the T cytotoxic/suppressor phenotype did not prevent graft rejection at this dose (MST 10 days).