Macrophages play important roles during renal inflammation. Infiltrating macrophages produce proinflammatory mediators and induce apoptosis in a variety of target cells. Because proximal tubules are frequently damaged in inflammatory processes, we investigated murine macrophages (J774) in the induction of apoptosis in murine PKSV-PR proximal tubule cells. PKSV-PR cells were co-cultured with activated or non-activated macrophages. Apoptosis was assessed by Annexin-V-FITC/propidium iodide staining and analyzed by fluorescence-activated cell sorting. Macrophages were separated from tubule cells with transwell membranes to distinguish soluble factor-mediated from direct cell-to-cell contact-mediated apoptosis. Cell supernatants from activated and non-activated macrophages were analyzed for induction of apoptosis. Activated (but not non-activated) macrophages induced tubule cell apoptosis in co-culture. Soluble factors were mainly responsible for induction of apoptosis; membrane separation and transfer of cell supernatant from activated macrophages showed similar levels of apoptosis induction. Although tumor necrosis factor (TNF)-alpha and transforming growth factor (TGF)-beta(1), measured by ELISA, increased significantly in supernatants from activated macrophages, blocking TNF-alpha and TGF-beta did not decrease apoptosis in PKSV-PR cells co-cultured with macrophages. Moreover, exogenous addition of TNF-alpha, TGF-beta, anti-Fas antibody, or TRAIL failed to induce apoptosis in tubule cells. We conclude that inflammatory macrophages mediate proximal tubule cell death, directing apoptosis mainly via release of unidentified soluble factors.