Sensory neuropeptides and epithelial cell restitution: the relevance of SP- and CGRP-stimulated mast cells.
BACKGROUND
Calcitonin-gene-related peptide (CGRP) and substance P (SP) are neurotransmitters of extrinsic primary afferent neurons located within the dorsal root ganglia. In experimental models of colitis in rats and rabbits, a protective role of SP and CGRP on intestinal mucosa was presumed. The mucosal protection partly depends on a CGRP-mediated modulation of mucosal blood flow. Limited data are available regarding CGRP- or SP-mediated effects on epithelial cell restitution. Having shown earlier that SP-stimulated fibroblasts but not CGRP-stimulated fibroblasts induce epithelial cell migration in vitro, the aim of this study was to explore whether mast cells mediate effects of SP and CGRP on epithelial cell restitution in vitro.


METHODS
Mast cells (C57) were exposed to SP [10(-12)-10(-6 M)] and CGRP [10(-12)-10(-7 M)]. After a 24-h incubation period, the cell supernatants (conditioned media, CDM) were taken from mast cell cultures and directly applied to rat intestinal epithelial cell lines-18 or Caco-2 monolayers, which had been wounded with a razor blade 24 h prior to the experiments. Epithelial cell migration was assessed by counting cells across the wound edge and epithelial cell proliferation was measured using 3-[4, 5-dimethylthiazol-2-yl]-2, 5-diphenyl-tetrazolium bromide test.


RESULTS
CGRP significantly induced epithelial cell migration and proliferation via mast cells when supernatants were directly applied to epithelial cells in vitro. The effects on epithelial cell migration were abolished after neutralizing anti-transforming growth factor-beta (TGF-beta) had been added to the cell cultures. SP had no effects on epithelial cells following stimulation of mast cells.


CONCLUSIONS
CGRP modulates epithelial cell restitution in vitro mediated by mast cells. The CGRP- and mast-cell-induced epithelial cell migration is TGF-beta dependent. This observation underlines an important role for extrinsic primary afferent neurons in mucosal defence and repair and in keeping the mucosal homeostasis. This knowledge leads to a better understanding of the interaction of the enteric nervous system and wound healing and may, in the future, lead to new therapeutic approaches to inflammatory diseases of the intestine.
Bulut
Kerem
K
Department of Internal Medicine I, St. Josef Hospital, Ruhr-University of Bochum, Gudrun Str. 54, 44791, Bochum, Germany.
Felderbauer
Peter
P
Deters
Susanne
S
Hoeck
Karoline
K
Schmidt-Choudhury
Anjona
A
Schmidt
Wolfgang E
WE
Hoffmann
Peter
P
D000818:Animals; D000906:Antibodies; D018938:Caco-2 Cells; D015740:Calcitonin Gene-Related Peptide / Q000378:metabolism*; D002460:Cell Line; D002465:Cell Movement; D049109:Cell Proliferation; D017077:Culture Media, Conditioned / Q000378:metabolism; D004847:Epithelial Cells / Q000378:metabolism*; D006801:Humans; D008407:Mast Cells / Q000378:metabolism*; D051379:Mice; D051381:Rats; D013373:Substance P / Q000378:metabolism*; D013997:Time Factors; D053773:Transforming Growth Factor beta1 / Q000276:immunology / Q000378:metabolism; D014945:Wound Healing
D000906:Antibodies; D017077:Culture Media, Conditioned; D053773:Transforming Growth Factor beta1; D013373:Substance P; D015740:Calcitonin Gene-Related Peptide