Interstitial leukocyte infiltration is a major finding in tubulointerstitial damage (TID). Infiltrating lymphocytes interact with proximal tubular epithelial cells (PTEC) by means of secreted soluble factors and/or cell contact mechanisms. CD40 expressed onto PTEC can be engaged by CD40L present on T cells. PTEC are able to locally secrete complement C3, which may most likely promote TID. The aim of the study was to investigate the putative action of CD40 ligation on enhancement of C3 secretion by PTEC. Primary human PTEC and stabilized HK-2 cells were used in culture experiments. Cells were stimulated by soluble factors IL-1beta, IFN-gamma, and/or CD40L-expressing murine fibroblast L cells. Analysis of C3 gene expression was evaluated by reverse-transcription PCR and Northern blot. Secreted C3 was assayed by ELISA and a functional hemolytic test on supernatants. Intracellular events were explored by the NF-kappaB-specific inhibitor caffeic acid phenetyl ester (CAPE). Among soluble factors, IL-1beta and IFN-gamma increased C3 gene expression and secretion (two-fold to three-fold versus basal) on both HK-2 and PTEC. CD40 engagement by CD40L upregulated HK-2 C3 secretion by four-fold. IL-1beta did not further increase CD40-induced C3 secretion, whereas IFN-gamma associated with CD40L was the strongest stimulus (30-fold increase). Inhibition of NF-kappaB offset CD40L-induced C3 secretion by 70%. CD40 ligation is able to enhance C3 secretion by PTEC. This cell contact mechanism is in synergism with a T cell-derived soluble factor (IFN-gamma). C3 secretion induced by CD40L may represent a mechanism of amplification of TID associated with lymphocyte infiltration.