Recipient CD4 T regulatory cells inhibit the acute T cell-mediated rejection of renal allografts in wild type mice. chains offered by Kb and Db class I MHC molecules. Allograft-primed CD8 T cells from CCR5-deficient allograft recipients were activated during culture either with pro-inflammatory cytokine stimulated wild type endothelial cells pulsed with the I-Abm12 peptides or with proinflammatory cytokine simulated bm12 endothelial cells, indicating their LP-533401 inhibitor presentation of the I-Abm12 chain peptide/class I MHC complexes. In addition to induction by bm12 renal allografts, the I-Abm12 chain-reactive CD8 T cells were induced in CCR5-deficient, but not wild type C57BL/6, mice by immunization with the peptides. These results reveal novel alloreactive CD8 T cell specificities in CCR5-deficient recipients of single class II MHC renal allografts that mediate rejection of the allografts. 0.05 being considered significant. Results B6.H-2bm12 kidney rejection by B6.CCR5?/? recipients Single class II MHC disparate B6.H-2bm12 (bm12) renal allograft rejection by C57BL/6 (n = 12) vs. B6.CCR5?/? (n = 11) recipients was compared. Consistent with the inability of C57BL/6 mice to reject total MHC-mismatched A/J renal allografts (21), all bm12 renal allografts survived longer than 100 days in wild type recipients. In contrast, CCR5-deficient recipients rejected all bm12 renal allografts by day 42 post-transplant (Physique 1A). Neither C57BL/6 nor B6.CCR5?/? recipients rejected syngeneic renal grafts (data not shown and (21)). Acute injury and dysfunction of bm12 renal allografts in CCR5?/? recipients was indicated by sharp rises in serum creatinine levels beginning on day 25 post-transplant (Physique 1B). C57BL/6 bm12 renal allograft recipients experienced a modest rise in serum creatinine levels, first detected at day 56 post-transplant and managed through day 100 post-transplant. Open in a separate window Physique 1 Single class II MHC-disparate renal allograft rejection by CCR5-deficient, but not wild type C567BL/6, recipients. Groups of wild type C57BL/6 and B6.CCR5?/? mice received single class II MHC disparate renal grafts from B6.H-2bm12 donors. (A) B6.CCR5?/? mice rejected the allografts within 40 days post-transplant (MST=28, n=11) whereas wild type LP-533401 inhibitor recipients did not reject the allografts. (B) Serum creatinine levels in the renal allograft recipients were measured weekly after transplant and the mean serum concentration for each group is shown at each time point SEM. C57BL/6 kidney isografts were managed by both wild type C57BL/6 and B6.CCR5?/? recipients long-term without any rise in serum creatinine levels (data not shown). *p 0.05. Histological evaluation of bm12 renal allografts at day 14 post-transplant indicated intense mononuclear infiltration in grafts from B6.CCR5?/?, but not C57BL/6, recipients LP-533401 inhibitor (Physique 2C vs. B, respectively). Neutrophil, macrophage and T cell infiltration into bm12 renal allografts was detected as early as day GLI1 7 post-transplant (Physique 2A). Numbers of LP-533401 inhibitor CD4 T cells infiltrating bm12 allografts were nearly identical in C57BL/6 and B6.CCR5?/? recipients. Surprisingly, compared to the low CD8 T cell infiltration into bm12 renal allografts in wild type recipients, intense CD8 T cell infiltration into allografts in B6.CCR5?/? recipients was observed as early as day 7 post-transplant and increased markedly thereafter (Physique 2A, B and C). The intense CD8 T cell infiltration into the bm12 renal allografts in B6.CCR5?/? recipients was accompanied by high mRNA levels of all proinflammatory cytokine genes tested, including TNF and IFN- when assessed on day 14 post-transplant where as expression of these proinflammatory mediators was low-absent in allografts from wild type recipients (Physique 3). Open in a separate window Physique 2 Leukocyte infiltration into single class II MHC-disparate renal allografts in CCR5-deficient and wild type C57BL/6 recipients. (A) bm12 renal allografts were harvested from groups of wild type C57BL/6 and B6.CCR5?/? recipients around the indicated days post-transplant. Following digestion of the allografts and bm12 kidneys from non-transplanted B6.H-2bm12 mice, aliquots of prepared single cells suspensions were stained with antibody and analyzed by circulation cytometry to determine the numbers of graft infiltrating leukocyte populations, expressed as figures/mg graft tissue SEM. *p 0.05; **p 0.01. Renal.