Adenylyl cyclase 6 is involved in the hyposecretory status of experimental colitis
- 219 Downloads
One of the cardinal symptoms of intestinal inflammation is diarrhea. Acute intestinal inflammation is associated with inhibition of ion absorption and increased secretion, along with fluid leakage due to epithelial injury and changes in permeability. However, in the chronic situation, a downregulation of both absorptive and secretory transport has been reported. We investigated how experimental colitis reduces cAMP levels in intestinal epithelial cells through modulation of adenylyl cyclases (AC). Primary colonic epithelial cells obtained from rats with trinitrobenzenesulfonic acid colitis and non-colitic controls were analyzed for AC expression by RT-qPCR and Western blot, following a preliminary microarray analysis. AC6 and AC5 were found to be expressed in colonocytes, and downregulated by inflammation, with the former exhibiting considerably higher mRNA levels in both cases. To test the hypothesis that inflammatory cytokines may account for this effect, Caco 2 cells were treated with IL-1β, TNF-α, or IFN-γ. All three cytokines inhibited forskolin evoked short-circuit currents in Ussing chambers and lowered intracellular cAMP, but failed to alter AC6 mRNA levels. AC5/AC6 expression was however inhibited in mouse jejunal organoids treated with IFN-γ and TNF-α, but not IL-1β. Gene knockdown of AC6 resulted in a significant decrease of ion secretion in T84 cells. We conclude that the disturbances in ion secretion observed in rat TNBS colitis are associated with low intracellular levels of cAMP in the epithelium, which may be explained in part by the downregulation of AC5/AC6 expression by proinflammatory cytokines.
KeywordsAdenylyl cyclase cAMP Organoids Colitis
The authors want to express their gratitude to Samuel Cantarero and the Centro de Instrumentación Científica of the University of Granada. This work was supported by the Ministry of Economy and Competitivity, partly with Fondo Europeo de Desarrollo Regional FEDER funds [grant numbers SAF2008-01432, AGL2008-04332, SAF2011-22922, SAF2011-22812, BFU2014-57736-P, AGL2014-58883-R]; and by Junta de Andalucía [grants number CTS164, CTS235]. IRC, BO, RGB and CHC were funded by Ministry of Education. CIBERehd is funded by the Instituto de Salud Carlos III. We also appreciate the collaboration of the Plataforma Andaluza de Bioinformática.
- 1.Asfaha S, Bell CJ, Wallace JL, MacNaughton WK (1999) Prolonged colonic epithelial hyporesponsiveness after colitis: role of inducible nitric oxide synthase. Am J Phys 276:G703–G710Google Scholar
- 3.Bell CJ, Gall DG, Wallace JL (1995) Disruption of colonic electrolyte transport in experimental colitis. Am J Phys 268:G622–G630Google Scholar
- 4.Brazma A, Hingamp P, Quackenbush J, Sherlock G, Spellman P, Stoeckert C, Aach J, Ansorge W, Ball CA, Causton HC, Gaasterland T, Glenisson P, Holstege FC, Kim IF, Markowitz V, Matese JC, Parkinson H, Robinson A, Sarkans U, Schulze-Kremer S, Stewart J, Taylor R, Vilo J, Vingron M (2001) Minimum information about a microarray experiment (MIAME)-toward standards for microarray data. Nat Genet 29:365–371CrossRefPubMedCentralGoogle Scholar
- 6.Dekkers JF, Wiegerinck CL, de Jonge HR, Bronsveld I, Janssens HM, de Winter-de Groot KM, Brandsma AM, de Jong NW, Bijvelds MJ, Scholte BJ, Nieuwenhuis EE, van den Brink S, Clevers H, van der Ent CK, Middendorp S, Beekman JM (2013) A functional CFTR assay using primary cystic fibrosis intestinal organoids. Nat Med 19:939–945CrossRefPubMedCentralGoogle Scholar
- 18.Martinez-Moya P, Ortega-Gonzalez M, Gonzalez R, Anzola A, Ocon B, Hernandez-Chirlaque C, Lopez-Posadas R, Suarez MD, Zarzuelo A, Martinez-Augustin O, Sanchez de Medina F (2012) Exogenous alkaline phosphatase treatment complements endogenous enzyme protection in colonic inflammation and reduces bacterial translocation in rats. Pharmacol Res 66:144–153CrossRefPubMedCentralGoogle Scholar