Bilaterally nephrectomized Lewis recipients of Fisher 344 (F344) kidney allografts, treated with CyA (1.5 mg/kg/day x 10), develop progressive changes of chronic rejection. Treated F344-to-F344 acted as isograft controls. Proteinuria was determined sequentially. Grafts were harvested 8, 12 and 16 weeks after transplantation (n = 9/group/time period). Infiltrating host cells and their products were assessed in chronically rejecting grafts by histology and immunostaining using mAbs for monocyte/macrophages, T-cells, ICAM-1, LFA-1 and cytokines. For in vitro binding studies, snap-frozen sections of transplanted kidneys were incubated with monocytes/macrophages and lymphocytes isolated from peripheral blood (PBL) of naive animals. For in vivo migration studies, naive cell populations were labeled with Bis-Benzamide and transferred i.v. to grafted animals at weeks 8, 12 and 16 (n = 3/group); grafts were harvested 24 h later and cell localization assessed under immunofluorescence. Increasing numbers of ED1 + monocytes/macrophages in allografted kidneys peaked at 16 weeks, localizing preferentially in glomeruli, where IL-1, IL-6 and TNF-alpha expression had also become intense and correlated with progressive glomerulosclerosis. Binding studies corroborated these results. In vitro, a few monocytes/macrophages bound to glomeruli and vessels at 8 weeks; by 12 weeks, binding to glomeruli was high (72% of cells). In vivo, large numbers of transferred labelled monocytes/macrophages were found in kidney allografts at 12 weeks (23%, isografts; < 7%, P < 0.01). In contrast T cells (primarily CD4+) were a consistent feature in allografts elevated as compared to isografts and correlating with in vitro and in vivo binding patterns; associated cytokines included IL-2, IFN- and TNF-alpha. Functional data followed these results: urine protein excretion by allograft recipients increased from baseline at 8 weeks (12 mg/day) to > 50 mg/day at 16 weeks at which point animals were beginning to die of renal failure; proteinuria in isografted rats did not increase during this time period. These results suggest that monocyte/macrophage and CD4+ T cells and their products are important in chronic kidney allograft rejection, contributing to the progressive sclerosis and fibrosis.