Enteric glia protect against Shigella flexneri invasion in intestinal epithelial cells: a role for S-nitrosoglutathione.
BACKGROUND
Enteric glial cells (EGCs) are important regulators of intestinal epithelial barrier (IEB) functions. EGC-derived S-nitrosoglutathione (GSNO) has been shown to regulate IEB permeability. Whether EGCs and GSNO protect the IEB during infectious insult by pathogens such as Shigella flexneri is not known.


METHODS
S flexneri effects were characterised using in vitro coculture models of Caco-2 cells and EGCs (or GSNO), ex vivo human colonic mucosa, and in vivo ligated rabbit intestinal loops. The effect of EGCs on S flexneri-induced changes in the invasion area and the inflammatory response were analysed by combining immunohistochemical, ELISA and PCR methods. Expression of small G-proteins was analysed by western blot. Expression of ZO-1 and localisation of bacteria were analysed by fluorescence microscopy.


RESULTS
EGCs significantly reduced barrier lesions and inflammatory response induced by S flexneri in Caco-2 monolayers. The EGC-mediated effects were reproduced by GSNO, but not by reduced glutathione, and pharmacological inhibition of pathways involved in GSNO synthesis reduced EGC protecting effects. Furthermore, expression of Cdc42 and phospho-PAK in Caco-2 monolayers was significantly reduced in the presence of EGCs or GSNO. In addition, changes in ZO-1 expression and distribution induced by S flexneri were prevented by EGCs and GSNO. Finally, GSNO reduced S flexneri-induced lesions of the IEB in human mucosal colonic explants and in a rabbit model of shigellosis.


CONCLUSIONS
These results highlight a major protective function of EGCs and GSNO in the IEB against S flexneri attack. Consequently, this study lays the scientific basis for using GSNO to reduce barrier susceptibility to infectious or inflammatory challenge.
INSERM U913, Place Alexis Ricordeau, Nantes Cedex, France. michel.neunlist@univ-nantes.fr
Mathurin
M
Flamant
Philippe
P
Aubert
Malvyne
M
Rolli-Derkinderen
Arnaud
A
Bourreille
Margarida Ribeiro
MR
Neunlist
Maxime M
MM
Mahé
Guillaume
G
Meurette
Benoit
B
Marteyn
Tor
T
Savidge
Jean Paul
JP
Galmiche
Philippe J
PJ
Sansonetti
Michel
M
Neunlist
D000818:Animals; D000900:Anti-Bacterial Agents / Q000494:pharmacology; D018988:Bacterial Translocation / Q000502:physiology; D018938:Caco-2 Cells; D018920:Coculture Techniques; D003106:Colon / Q000294:innervation / Q000382:microbiology; D004353:Drug Evaluation, Preclinical / Q000379:methods; D004405:Dysentery, Bacillary / Q000382:microbiology / Q000473:pathology* / Q000503:physiopathology; D017615:Enteric Nervous System / Q000502:physiology; D006801:Humans; D007413:Intestinal Mucosa / Q000187:drug effects / Q000294:innervation* / Q000378:metabolism; D009457:Neuroglia / Q000502:physiology*; D010539:Permeability; D011817:Rabbits; D020133:Reverse Transcriptase Polymerase Chain Reaction; D026422:S-Nitrosoglutathione / Q000378:metabolism* / Q000494:pharmacology; D012763:Shigella flexneri / Q000187:drug effects / Q000502:physiology*; D020764:cdc42 GTP-Binding Protein / Q000378:metabolism
D000900:Anti-Bacterial Agents; D026422:S-Nitrosoglutathione; D020764:cdc42 GTP-Binding Protein