CD40 stimulation, by either antibody or ligand, has been shown to inhibit the growth of a variety of neoplastic cells, both in vivo and in vitro. In this study, we assessed the effects of CD40 stimulation using a murine agonistic CD40 monoclonal antibody (MoAb) (FGK115) or a soluble recombinant murine CD40 ligand (srmCD40L) in both lethally irradiated and nonirradiated BALB/c mice. Toxicity after CD40 stimulation was not observed in nonirradiated animals receiving up to 100 microg of the agonist anti-CD40 MoAb. However, as little as 10 microg of the agonistic anti-CD40 MoAb induced acute toxicity resulting in 100% morbidity of lethally irradiated animals by 4 days after irradiation. Histological evaluation of animals receiving anti-CD40 MoAb revealed severe intestinal lesions with disruption of the villi, goblet cell depletion, and crypt hyperplasia of the small intestine, colon, and cecum. Delaying the administration of anti-CD40 MoAb or reducing the amount of irradiation given resulted in increased survival and less severe lesions. Analysis of serum cytokine levels in lethally irradiated mice receiving agonistic anti-CD40 showed a marked increase of interferon (IFN)-gamma. Lethally irradiated IFN-gamma knockout mice given the agonistic anti-CD40 MoAb demonstrated significant increases in survival and minimal gut lesions compared with wild-type mice receiving the same regimen, suggesting that IFN-gamma plays a major role in this toxic reaction. These results indicate that CD40 stimulation using agonistic antibodies following lethal irradiation leads to a fatal, cytokine-induced disease affecting the intestine.