Effects of the stress-related peptides urocortins on colonic epithelial barrier function

Background and Aims Urocortins (Ucn I, Ucn II, and Ucn III) belong to the corticotropin‐releasing factor (CRF)‐like peptide family. They bind to two subtypes of CRF receptors (CRF 1 and CRF 2 ) to exert their biological effects. Ucn I binds to both CRF 1 and CRF 2 receptors with equal affinity, whereas Ucn II and Ucn III bind exclusively to CRF 2 . CRF and urocortins are expressed in the gastrointestinal tract. CRF has been implicated in stress‐induced impairment of intestinal barrier function. However, the effects of urocortins on intestinal epithelial barrier function remain unclear. The aims of the present study were to investigate the role of urocortins on intestinal epithelial barrier function, and determine which CRF receptor subtype(s) mediates their effects. Methods Adult C57BL/6J mice (47 males) were used in the present study. Mucosa/submucosa preparations from the proximal colon were mounted in Ussing flux chambers. Transepithelial resistance (TER) was measured as an indicator of paracellular permeability across the intestinal epithelium. Flux of horseradish peroxidase (HRP) from the luminal side to the basolateral side of the mucosa/submucosa preparation was measured as an indicator of transcellular permeability. Results The colonic mucosa/submucosa preparations were exposed to different concentrations of Ucn I, Ucn II, or Ucn III (30 nM, 100 nM, 300 nM, and 1 μM) on the basolateral side for 60 min. All three urocortins significantly decreased TER and the reduction rate was concentration‐dependent. At a concentration of 1 μM, Ucn III caused the greatest decrease in TER (ΔTER: −14.85±4.73 Ohm·cm 2 ; n=5), followed by Ucn II (ΔTER: −10.91±1.11 Ohm·cm 2 ; n=5) and Ucn I (ΔTER: −6.1±1.16 Ohm·cm 2 ; n=5). The effect of 1 μM Ucn III on TER was similar as the effect of 1 μM Ucn II ( P >0.05), but was significantly greater than that caused by 1 μM Ucn I ( P <0.05). Ucn I‐induced ΔTER was inhibited by both the CRF 1 receptor antagonist NBI 27914 and the CRF 2 receptor antagonist antisauvagine‐30. Ucn II and Ucn III‐induced ΔTER was significantly suppressed by antisauvagine‐30, but was not affected by NBI 27914. Ucn I, Ucn II, or Ucn III also caused a concentration‐dependent increase of HRP flux from the luminal side to the basolateral side of the colonic mucosa/submucosa preparation. At 1 μM, Ucn I, Ucn II, and Ucn III caused similar level of increase in HRP flux (Ucn I: 8.24±2.71 ng/ml/h/cm 2 , n=5; Ucn II: 10.94±0.87 ng/ml/h/cm 2 , n=6; Ucn III: 11.29±1.99 ng/ml/h/cm 2 , n=5) ( P >0.05). The increase of HRP flux caused by Ucn I, Ucn II, and Ucn III was significantly inhibited by antisauvagine‐30, but was unaffected NBI 27914. Conclusions Ucn I, Ucn II, and Ucn III all cause disturbances of colonic epithelial barrier function, which mimic the effects of stress and CRF. Ucn I‐induced increase of paracellular permeability was mediated by both CRF 1 and CRF 2 , whereas Ucn II‐ and Ucn III‐induced increase of paracellular permeability was mediated only by CRF 2 . The effects of Ucn I, Ucn II, and Ucn III on transcellular permeability were mediated by CRF 2 exclusively. The results suggest that the CRF 2 receptors play an important role in the impairment of intestinal epithelial barrier function induced by urocortins. Support or Funding Information NIH R15 DK097460‐01A1 (SL) and UWL graduate research service and educational leadership grant (AK).