Clinical observations on patients with systemic lupus erythematosus and rheumatoid arthritis and studies in murine models of autoimmune diseases suggest an important role for the sex and pituitary hormones in immune function. Estrogen and testosterone have been shown to modulate B cell functions in vitro. Prolactin (PRL) has been shown to have immunomodulatory functions. Follicle stimulating hormone (FSH) and luteinizing hormone (LH) have homology to thioredoxin, a potentiator of T-cell growth. We therefore studied the proliferative response of peripheral blood mononuclear cells (PBMC) from normal adult males to various stimuli in the presence or absence of estradiol (E2), testosterone (Te), PRL, FSH, and LH and the effect of these hormones on T-cell subsets by flow cytometry. Unfractionated PBMC were stimulated with 1% phytohemagglutinin (PHA) (polyclonal activation), anti-CD3 (T-cell receptor stimulation), or recombinant interleukin 2 (IL-2) (late stages of activation). We assessed the effects of E2 (0.3-30 ng/ml), Te (3-300 ng/ml), PRL (2-200 ng/ml), FSH (1-100 mIU/ml), and LH (1-100 mIU/ml). E2 and Te had no consistent effect on PBMC proliferation in response to any of the stimuli. E2 significantly decreased the percentage of CD4+ cells following PHA stimulation (P = 0.022), while significantly enhancing the percentage of CD8+ cells following IL-2 stimulation (P = 0.007). Te significantly increased the percentage of CD4+ cells following IL-2 stimulation (P = 0.03). PRL enhanced proliferation in response to IL-2 and PHA without subset-specific effects. FSH and LH both enhanced IL-2-induced proliferation, particularly in physiological doses (10 mIU/ml). FSH at 100 mIU/ml significantly decreased the percentage of CD4+ cells in unstimulated cultures (P = 0.003), while it enhanced the percentage of CD8+ cells following PHA stimulation (P = 0.004). The enhancement in CD8+ cells appeared the most marked in the CD8+CD28+ subset. LH at 10 mIU/ml significantly enhanced the percentage of CD4+ cells following IL-2 stimulation (P = 0.009) and at higher doses (100 mIU/ml) enhanced the percentage of CD4+ cells following PHA stimulation (P = 0.011). Thus, sex steroids and adenohypophyseal hormones (PRL, FSH, and LH) have subset-specific effects on T-cell activation which may influence sex-related differences in immune response.