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The enhancement of tumor metastasis by concurrent inflammatory processes is mainly due to the cytokines TNF and IL-1. In the case of TNF this effect is not restricted to metastasis models as measured by in vivo colony formation but also found in experimental model systems of spontaneous metastasis. Direct effects on the tumor cells or interference with the host NK cell system did not seem to account for the observed TNF effect. Experimental evidence from different test systems rather points to TNF- or IL-1-induced enhanced adhesion of tumor cells to the endothelial cell layer as the underlying mechanism. Blocking of integrin-matrix interactions with monoclonal antibodies or competing peptides inhibited tumor cell adhesion to endothelioma cells in vitro and lung colony formation of tumor cells in vivo.

Tumor-associated tumor necrosis factor (TNF) production in patients as well as a TNF-inducing membrane constituent of tumor cells have been reported. In a murine fibrosarcoma model we analyzed TNF production during growth of a tumor transplant. In situ hybridization showed that a gradually increasing number of cells within the tumor tissue became positive for TNFmRNA. Also, in spleen cells of tumor-bearing mice TNFmRNA became more abundant in later stages of tumor growth compared to early stages. In plasma of these animals, however, TNF activity was not detected at any time even after stimulation with bacterial endotoxin. Neutralization with monoclonal antibodies of endogenous TNF during tumor growth did not affect the growth rate of the tumor, indicating that either the antibodies did not reach the relevant TNF production and action sites or that endogenously produced TNF did not play a significant role in this tumor model.

By intrasplenic immunization we raised a rat mAb (mAb V1q; IgG2a, kappa) with a potent neutralizing activity against natural mouse TNF (1 microgram/ml mAb V1q/100 U/ml TNF). mAb V1q was used to study the role of endogenous TNF in experimental peritonitis induced by sublethal cecal ligation and puncture. mAb V1q persisted for over 5 days in the serum of mice injected with 100 micrograms of the antibody and, therefore, proved useful for in vivo experiments. As little as 20 micrograms mAb V1q/mouse prevented lethal shock of the animals by 400 micrograms LPS/mouse. In sublethal cecal ligation and puncture i.p. injection of mAb V1q directly and up to 8 h after induction of experimental peritonitis lead to death of the animals within 1 to 3 days. The lethal effect of mAb V1q was compensated by injection of recombinant mouse TNF. Similar mAb V1q effects as in immunocompetent mice were shown in severe combined immune deficiency mice deficient of mature functional B and T cells. Taken together, these data suggest that during the early phase of peritonitis endogenous TNF may stimulate nonlymphoid cells such as granulocytes, macrophages, platelets, and fibroblasts to ingest bacteria and to localize inflammation, respectively. These beneficial effects of TNF may determine survival. Thus, our data may have implications for the therapeutic management of a beginning peritonitis.

Recombinant tumor necrosis factor (TNF) had hypothermic activity in vivo. Intravenous injection of TNF resulted in a hypothermic reaction of mice within 3 to 6 hours. This reaction was not the result of interleukin 1 (IL1) release from macrophages/monocytes. Peritoneal exudate cell cultures from endotoxin low responder mice or human peripheral mononuclear leukocyte cultures did not generate IL1 activity in the supernatant after exposure to TNF. The addition of interferon-gamma (IFN-gamma) or preexposure to IFN-gamma and then stimulation with TNF did also not result in IL1 secretion. No IL1 inhibitor was generated and TNF did not interfere with the IL1 test systems. Therefore, we conclude that the hypothermic activity of TNF is not mediated via the induction of IL1 production by mononuclear phagocytes.

Bacterial lipopolysaccharides (LPS) enhance both production of colony-stimulating factors (CSF) and proliferation of mononuclear phagocytes in vivo. The present study was undertaken to determine whether the effects of LPS on CSF-dependent monopoiesis are due solely to enhanced production of CSF or also to direct effects of LPS on the responding progenitor cell. Addition of LPS to CSF-stimulated macrophage populations had different effects, depending upon the concentration of CSF in the cultures. In the presence of optimal to supraoptimal concentrations of CSF, LPS at doses >/=0.01 mug/ml inhibited macrophage colony formation. This inhibitory activity was not due to cytotoxicity of the LPS and was not mediated through prostaglandin synthesis. In the presence of suboptimal concentrations of CSF, minute concentrations of LPS (10(-7) mug/ml) significantly enhanced macrophage colony formation. Both effects of LPS (inhibition and enhancement) appeared to be properties of lipid A since neither effect was noted with cells from LPS-resistant C3H/HeJ mice, whereas both effects could be neutralized by the addition of the antibiotic polymyxin B, which binds to the lipid A portion of LPS. These results suggest that the effects of LPS on monopoiesis in vivo may not be due solely to its capacity to stimulate production of CSF. Rather, LPS may be involved in stimulating monopoiesis both indirectly through stimulation of CSF production and by its effects on the CSF-responsive progenitor cell.

Macrophage-enriched peritoneal exudate cells from mice infected with Mycobacterium bovis BCG, macrophage-like tumor cells (PU 5-1.8), and peritoneal macrophages propagated in vitro with macrophage growth factor released tumoricidal activity into the culture medium within 2 to 3 h after stimulation with nanogram quantities of bacterial lipopolysaccharide. The cytotoxic activities from each of the macrophage culture supernatants eluted from diethylaminoethyl-Sephacel columns at a sodium chloride concentration of 200 mM exhibited a molecular weight of 50,000 to 60,000 as estimated by gel filtration, were stable at 56 degrees C for 30 min, and were active at a pH range of 6 to 10. A rabbit antiserum directed against serum-derived cytotoxic activity (tumor-necrotizing factor) from BCG-infected and lipopolysaccharide-challenged mice inhibited all of the cytotoxic activities generated in vitro. This suggests that the macrophage-derived cytotoxins are identical with serum-derived cytotoxic factor, which further implies that the macrophage is the cellular source of tumor-necrotizing factor.

Thymocyte cultures from C3H/HeJ mice were stimulated for proliferative responses with purified preparations of interleukin 1 (IL 1) and interleukin 2 (IL 2). Synergistic responses were obtained in the absence of mitogen. In the presence of excess IL 2, the thymocyte proliferation response was strictly dependent on the amount of IL 1 in the cultures. Antibodies to IL 1 inhibited the response in a dose-dependent manner. The combination of IL 1 plus IL 2 induced the appearance of IL 2 receptors on murine thymocytes as detected with a monoclonal antibody directed against the IL 2 receptor. Neither IL 1 nor IL 2 alone had this effect. The thymic subpopulation found to become IL 2 responsive upon IL 1 stimulus was the peanut agglutinin-negative (PNA-) medullary fraction.

We investigated the role of T cell cytotoxicity inducing factor 1 (TCF1) in the induction of a cytotoxic T cell response. We found that help-deficient thymocyte cultures supplied with saturating amounts of purified IL 2 did not develop CTL in a 5-day culture. The expression of cytotoxicity was dependent on the addition of TCF1 derived from the T cell hybridoma K15. TCF1 also induced proliferation of thymocytes in the presence of IL 2. Only the PNA- thymocyte subpopulation responded to TCF1 with proliferation and cytotoxicity in the presence of IL 2. The monokine IL 1 also induced proliferation in this subpopulation but failed to induce cytotoxicity. IL 1 was further distinguished from TCF1 by inhibition of IL 1-induced but not TCF1-induced proliferation by anti-IL 1 antibodies. In addition, using anti-IL 2 receptor antibodies (AMT 13), we showed that TCF1 in the presence of IL 2 substantially increased IL 2 receptor expression in thymocytes. IL 1 had the same effect on induction of IL 2 receptor expression as TCF1. Because some effects of IL 1 and TCF1 are distinct and some overlap, we discuss whether IL 1 and TCF1 induce different subsets of PNA- thymocytes.