Sex-based differences in inflammatory responses to exercise may be mediated by estrogen through increased muscle membrane stability and/or inhibited cytokine production. In this study, in vivo effects of estrogen on systemic inflammatory-related responses to exercise were assessed in healthy men. In a double-blind, placebo-controlled, cross-over design, 11 men cycled for 90 min @ 65% VO_2max following eight days of 17-estradiol supplementation (ES; 2 mg/day) or placebo (PL; glucose polymer). Following a 2-week washout, exercise was repeated after eight days on the alternate treatment. Blood was collected pre- and post-exercise to determine IL-6, sICAM-1, neutrophil counts, and cortisol. Pre-exercise serum was assayed for sex hormones. ES increased estradiol (133 ± 71 to 840 ± 633 pmol/l, P = 0.005) and reduced testosterone (19.9 ± 3.7 to 16.1 ± 3.9 nmol/l, P = 0.007). Exercise increased cortisol (P = 0.02), IL-6 (P < 0.001) and neutrophil counts (P < 0.001) with no influence on sICAM-1 (P = 0.34) and no effect of ES on these changes. Post-exercise IL-6 and neutrophil counts were correlated (r = 0.58, P = 0.005); post-exercise IL-6 and cortisol (r = 0.18, P = 0.43) and post-exercise cortisol and neutrophil counts (r = 0.06, P = 0.78) were not. Post-exercise sICAM-1 was not correlated with the above variables (P 0.79). In conclusion, eight days of ES in healthy men did not influence systemic inflammatory-related responses to acute exercise. Future studies should investigate 17-estradiol effects on IL-6 production and neutrophil infiltration within skeletal muscle during and following exercise.