Hemeoxygenase (HO)-1 induction following adverse circulatory conditions is known to be protective, and precastrated males have less intestinal damage than sham-operated males following trauma-hemorrhage (T-H). Previous studies have also shown that administration of flutamide up-regulated estrogen receptor (ER) expression in males following T-H. We hypothesized that flutamide administration in males following T-H up-regulates HO-1 via an ER-dependent pathway and protects against intestinal injury. Male Sprague-Dawley rats underwent T-H [mean blood pressure (MBP) 40 mmHg for 90 min and then resuscitation]. A single dose of flutamide (25 mg/kg body weight), with or without an ER antagonist (ICI 182,780), a HO enzyme inhibitor [chromium-mesoporphyrin (CrMP)], or vehicle, was administered subcutaneously during resuscitation. At 2 h after T-H or sham operation, intestinal myeloperoxidase (MPO) activity, intercellular adhesion molecule (ICAM)-1, cytokine-induced neutrophil chemoattractant (CINC)-1, and CINC-3 levels were measured. Intestinal ER-α, ER-β, androgen receptor, and HO-1 mRNA/protein levels were also determined. Results showed that T-H increased intestinal MPO activity, ICAM-1, CINC-1, and CINC-3 levels. These parameters were improved significantly in the flutamide-treated rats subjected to T-H. Flutamide treatment increased intestinal HO-1 and ER-β mRNA/protein levels as compared with vehicle-treated T-H rats. Administration of the ER antagonist ICI 182,780 or the HO inhibitor CrMP prevented the flutamide-induced attenuation of shock-induced intestinal damage. Thus, the salutary effects of flutamide administration on attenuation of intestinal injury following T-H are mediated via up-regulation of ER-β-dependent HO-1 expression. © Society for Leukocyte Biology.