Urocortin II mediates pro-inflammatory effects in human colonocytes via corticotropin-releasing hormone receptor 2

Alan C Moss ¹, Pauline Anton ¹, Tor Savidge ¹, Paul Newman ¹, Adam S Cheifetz ¹, Jerome Gay ¹, Sophia Paraschos ¹, Michael Weinstein Winter ¹, Mary P Moyer ², Katia Karalis ¹, Efi Kokkotou ¹ and Charalabos Pothoulakis ^1*

¹ Harvard Medical School, United States
² Incell Corporation, United States

^* To whom correspondence should be addressed. E-mail: cpothoul{at}bidmc.harvard.edu.

Accepted 20 March 2007

Abstract

Background & Aims: Urocortin II (UcnII) is a neuropeptide that binds with high affinity to the corticotropin-releasing hormone receptor 2 (CRHR2) in peripheral tissues. UcnII is synthesized in the intestine, but its role in human intestinal inflammation is largely unknown.

Methods: We measured responses of human colonic epithelial cells expressing CRHR2 to stimulation by UcnII using ELISA, Western blot analysis, real time RT-CPR, and IL-8 promoter activity. Expression levels of CRHR2 and UcnII in human colitis were determined by immunofluorescence and real time RT-PCR in mucosal biopsies from patients with Crohns and ulcerative colitis, and in human intestinal xenografts after exposure to Clostridium difficile toxin A.

Results: We report here that expression of CRHR2 mRNA and protein in human colonic epithelial cells (HT-29) are increased by exposure to C. difficile toxin A or TNF. Stimulation of non-transformed NCM460 colonocytes overexpressing CRHR2 receptor with UcnII resulted in a time- and concentration-dependent increase in IL-8 production. UcnII stimulation also led to activation of the transcription factor NF-B and MAP kinase in these cells, as evidenced by degradation of IBand phosphorylation of the p65 subunit of NF-B and ERK 1/2. Furthermore, expression of UcnII and CRHR2 mRNA was increased in mucosal samples of patients with inflammatory bowel disease, and after exposure of human intestinal xenografts to C. difficile toxin A.

Conclusions: These results suggest that UcnII has pro- inflammatory effects in human intestinal cells via the CRHR2receptor and may play an important role in the pathophysiology of colitis in humans.

Keywords: colitis, corticotropin-releasing hormone receptor 2, neuropeptides, urocortin II

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