The present investigation examined the effect of interleukin-1beta (IL-1beta) on progesterone production by human luteal cells and the expression and localization of the IL-1 system in the human corpus luteum (CL). Luteal cells were isolated from corpora lutea collected throughout the luteal phase. After dispersion, luteal cells were treated with a panel of monoclonal antibodies directed to leukocyte-specific molecules. The leukocytes were isolated with immunomagnetic beads. Leukocyte-free luteal cells exhibited greater steroidogenic responsiveness to hCG toward the end of the luteal phase. The treatment of mixed luteal cells (total luteal cells) with IL-1beta inhibited by 60% hCG-stimulated progesterone production. Interestingly, the treatment of leukocyte-free luteal cells with IL-1beta did not affect progesterone production. In addition, the treatment of mixed luteal cells with monoclonal antibodies against IL-1 receptor type I (IL-1RtI) resulted in a 2.5-fold increase in the hCG-supported progesterone production. IL-1RtI and IL-1 receptor antagonist were localized by immunohistochemistry in both somatic and immune cells of the CL. Flow cytometric analysis indicated that both nonleukocyte luteal cells and leukocyte-luteal cells exhibited IL-1Rt-I positive cells, representing 56% and 31% of the total luteal cells, respectively. However, 13% of nonleukocyte luteal cells did not express IL-1Rt-I. Northern analysis demonstrated the presence of the 5.1-kb IL-1RtI messenger ribonucleic acid transcript in CL of different ages. RT-PCR indicated that both leukocyte-free luteal cells and luteal leukocytes express IL-1RtI messenger ribonucleic acid. We conclude that 1) luteal leukocytes have an inhibitory effect on hCG-stimulated progesterone production; 2) IL-1beta inhibits hCG-stimulated progesterone production only in mixed luteal cell cultures, indicating that leukocytes mediate the effect; 3) the somatic and immune cells of the CL are sites of action and expression of the IL-1 system; and 4) interaction between the steroidogenic and immune cells of the CL suggests a functional intraovarian role for IL-1beta in CL physiology.