Rat peritoneal macrophages were induced to produce high amounts of nitric oxide (NO) when rats were challenged by MTH68/H, (a live attenuated oncolytic Newcastle disease virus strain). The increase in NO production was observed to be viral particle dose dependent. The higher NO production measured could be due to the enhanced expression of NO synthase II enzyme. In addition, viral administration caused a higher macrophage cell count in the peritoneal cavity of treated rats. Interleukin-1 and granulocyte-monocyte colony stimulating factors were also produced by the induced macrophages. COS 7, a transformed cell line was killed by both NO donors and activated macrophages; the latter effect was markedly decreased in the presence of the inhibitors of NO production. Cytotoxic effect of NO was evidenced by the decrease of cell viability and proliferation of COS 7 cells. Excessive NO production may also be cytotoxic for macrophages themselves as proved by the addition of exogenous NO donors. These results strongly suggested the participation of induced NO synthesis of macrophages in the anti-tumor effect of MTH-68/H vaccine treatment.