We investigated whether corticotropin-releasing factor receptor 2 (CRF₂) and its high affinity agonist urocortin 1 (Ucn1) mediate sex-specific signaling and immune responses. Intrarectal trinitrobenzene sulfonic acid (TNBS) was used to induce experimental colitis in wildtype (WT), CRF2 knockout (CRF₂KO), and heterozygous (CRF₂Ht) mice of both sexes. Changes in plasma extravasation, organ weight, survival, immune cell numbers, inflammatory cytokines, and the MAPK signaling pathway were assessed. Stored intestinal biopsies from patients with Crohn's disease (CD) and age- and sex-matched individuals without inflammatory bowel disease (IBD) were examined by immunofluorescence and confocal microscopy to characterize Ucn1 and CRF receptor expression. CRF₂Ht mice of both sexes showed decreased survival during colitis compared to other genotypes. Ucn1 improved survival in male mice alone. Ucn1 restored colon length, spleen and adrenal weight, and decreased colonic TNF-α, IL-6, and IL-1β levels in male CRF₂Ht mice alone. CRF₂Ht mice of both sexes showed decreased phosphorylation of MAPK p38 and heat shock protein 27 (Hsp27) levels. Ucn1 restored p-Hsp27 levels in male CRF₂Ht mice alone. Expression of the chaperone protein Hsp90 decreased during colitis, except in male CRF₂Ht mice. Taken together, our data indicate that sex shows significant interaction with genotype and Ucn1 during colitis. Human duodenal and colonic biopsies revealed that sex-specific differences exist in levels of CRF receptors and Ucn1 expression in patients with CD compared with the matched non-IBD subjects. To conclude, Ucn1 mediates sex-specific immune and cellular signaling responses via CRF₂, emphasizing the need for inclusion of females in preclinical studies.