Culture supernatants (SUPS) of endotoxin (LPS)-activated human mononuclear cells (MNL) stimulated greater production of cAMP by thymocytes than by spleen cells of C3H/HeJ or nude ( nu nu) mice. Similarly, the addition of prostaglandin E2 (PGE2) stimulated higher levels of cAMP in thymocytes and progressively lower levels in spleen cells from C3H/HeJ mice and nu nu spleen cells, respectively. Partial purification on Bio-Gel P100 of the LPS-induced MNL SUPS yielded peaks of thymocyte proliferative activity characteristic of lymphocyte activation factor (LAF) but these fractions failed to stimulate cAMP levels in thymocytes. Moreover, MNL SUPS induced with LPS in the presence of indomethacin retained their LAF activity but no longer increased thymocyte cAMP levels. Radioimmunoassay of the SUPS for PGE2 revealed significantly higher levels of PGE2 in the media of those MNL cultures stimulated by LPS than when stimulated by phorbol myristic acetate, phytohemagglutin, or extracted cell wall fraction of Actinomyces viscosus. Thus, PGE2 is produced by human MNL and may exert considerable immunoregulatory effects mediated by elevation of lymphocyte cAMP levels. © 1980.