Substance P mediates antiapoptotic responses in human colonocytes by Akt activation

We examined the hypothesis that substance P (SP) and the neurokinin-1 receptor (NK-1R), both in vitro and in vivo, promote mucosal healing during recovery from colitis by stimulating antiapoptotic pathways in human colonic epithelial cells. For the in vitro experiments, human nontransformed NCM460 colonocytes stably transfected with NK-1R (NCM460-NK-lR cells) were exposed to SP, and cell viability assays, TUNEL assays, and Western blot analyses were used to detect apoptotic and antiapoptotic pathways. SP exposure of NCM460-NK-lR colonocytes stimulated phosphorylation of the antiapoptotic molecule At and inhibited tamoxifeninduced cell death and apoptosis evaluated by the cell viability assay and poly(ADP-ribose) polymerase cleavage, respectively. SP-induced phosphorylation of At and cleavage of poly(ADPribose) polymerase were inhibited by blockade of integrin alpha V beta 3, Jak2, and activation of phosphaticlylinositol 3-kinase. For the in vivo experiments, C57BL/6 mice, administered 5% dextran sulfate (DSS) dissolved in tap water for 5 days followed by a 5-day recovery period, were treated with the NK-1R antagonist 0-12,255 or vehicle. Vehicle-treated mice showed increased colonic At phosphorylation and apoptosis compared with mice that received no DSS. In contrast, daily i.p. administration of 0-12,255 for 5 days post-DSS suppressed At activation, exacerbated colitis, and enhanced apoptosis, and pharmacologic inhibition of Akt, either alone or together with 0-112,255, produced a similar effect. Thus, SP, through NK-1R, possesses antiapoptotic effects in the colonic mucosa by activating Akt, which prevents apoptosis and mediates tissue recovery during colitis.