The pathogenesis of septic shock is due mainly to bacterial toxin stimulation of the immune system, resulting in an excessive production of proinflammatory cytokines. TNF-alpha has been implicated as a major mediator in septic shock. Coinjection of D-galactosamine and LPS or staphylococcal enterotoxin B induced a rapid-onset, low-dose form of septic shock syndrome and ultimately led to death. We found that both the septic shock syndrome and death could be prevented by administration of anti-CD28 Ab. The protection induced by anti-CD28 Ab was associated with a decrease in TNF-alpha levels in the circulation. In addition, serum from anti-CD28 Ab-treated mice was capable of inhibiting the production of TNF-alpha by bone marrow-derived macrophages following treatment with LPS, indicating that anti-CD28 Ab induced production of soluble factors that subsequently inhibited the production of TNF-alpha. We confirmed that one of the factors present in serum was IL-10, because anti-CD28 Ab treatment stimulated the expression of IL-10, both in splenocytes and in T cell lines. Furthermore, injection of anti-IL-10 Abs could abolish the protective effect of anti-CD28 Ab on septic shock. Anti-IL-10 Ab could also suppress the anti-CD28 Ab-induced inhibition of TNF-alpha production, either in vivo or in vitro. Thus, we conclude that ligation of CD28 induces expression of IL-10, which in turn suppresses TNF-alpha production and prevents septic shock.