Role of Th1-stimulating cytokines in bacillus Calmette–Guérin (BCG)-induced macrophage cytotoxicity against mouse bladder cancer MBT-2 cells


Yi Luo, University of Iowa, Department of Urology, 3202 MERF, 375 Newton Road, Iowa City, IA 52242, USA.


Previously, we have demonstrated that macrophages exhibited cytotoxicity toward mouse bladder cancer MBT-2 cells upon bacille Calmette–Guérin (BCG) stimulation. In this study, we have investigated the role of Th1-stimulating cytokines in BCG-induced macrophage cytotoxicity. Thioglycollate-elicited peritoneal exudate cells (PECs) were used as a conventional source for macrophages and the induction of PEC effector functions (cytolytic activity and cytokine production) by BCG was evaluated in vitro. The BCG-activated PECs showed potent cytotoxicity and killed MBT-2 cells in a dose-dependent manner. Depletion of T cells, natural killer (NK) cells, or both, in PEC preparations exhibited a marginal or small reduction of MBT-2 cell killing, suggesting that macrophages played a primary role in PEC cytotoxicity. Transwell assays indicated that the maximal PEC cytotoxicity required both direct cell–cell contact and soluble factors such as interferon (IFN)-γ and tumour necrosis factor (TNF)-α. Neutralizing endogenous cytokines interleukin (IL)-12, IL-18, IFN-γ or TNF-α reduced PEC cytotoxicity by 38%, 22%, 15% and 94%, respectively. Supplementation of BCG with recombinant (r)IL-2, rIL-12 or rIL-18 increased PEC cytotoxicity by approximately twofold. Compared with control BCG for PEC stimulation, rBCGs expressing IL-2 or IL-18 showed enhanced MBT-2 cell killing by PECs. Increased cytokine production (IFN-γ, TNF-α and IL-6) was also observed in rBCG-stimulated PEC cultures. Taken together, these results suggest that Th1-stimulating cytokines play an important role in BCG-induced macrophage cytotoxicity and that combination of BCG with selected Th1-stimulating cytokines, either supplemented or expressed by BCG, may enhance the effect of BCG in the treatment of bladder cancer patients.