Nitric oxide inhibits enterocyte migration through activation of RhoA-GTPase in a SHP-2-dependent manner

Selma Cetin, Cynthia L. Leaphart, Jun Li, Irene Ischenko, Michael Hayman, Jeffrey Upperman, Ruben Zamora, Simon Watkins, Henri Ford, James Wang, David J. Hackam

Research output: Contribution to journalArticle

49 Citations (Scopus)

Abstract

Diseases of intestinal inflammation like necrotizing enterocolitis (NEC) are associated with impaired epithelial barrier integrity and the sustained release of intestinal nitric oxide (NO). NO modifies the cytoskeletal regulator RhoA-GTPase, suggesting that NO could affect barrier healing by inhibiting intestinal restitution. We now hypothesize that NO inhibits enterocyte migration through RhoA-GTPase and sought to determine the pathways involved. The induction of NEC was associated with increased enterocyte NO release and impaired migration of bromodeoxyuridine-labeled enterocytes from terminal ileal crypts to villus tips. In IEC-6 enterocytes, NO significantly inhibited enterocyte migration and activated RhoA-GTPase while increasing the formation of stress fibers. In parallel, exposure of IEC-6 cells to NO increased the phosphorylation of focal adhesion kinase (pFAK) and caused a striking increase in cell-matrix adhesiveness, suggesting a mechanism by which NO could impair enterocyte migration. NEC was associated with increased expression of pFAK in the terminal ileal mucosa of wild-type mice and a corresponding increase in disease severity compared with inducible NO synthase knockout mice, confirming the dependence of NO for FAK phosphorylation in vivo and its role in the pathogenesis of NEC. Strikingly, inhibition of the protein tyrosine phosphatase SHP-2 in IEC-6 cells prevented the activation of RhoA by NO, restored focal adhesions, and reversed the inhibitory effects of NO on enterocyte migration. These data indicate that NO impairs mucosal healing by inhibiting enterocyte migration through activation of RhoA in a SHP-2-dependent manner and support a possible role for SHP-2 as a therapeutic target in diseases of intestinal inflammation like NEC.

Original languageEnglish (US)
JournalAmerican Journal of Physiology - Gastrointestinal and Liver Physiology
Volume292
Issue number5
DOIs
StatePublished - May 1 2007
Externally publishedYes

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Enterocytes
GTP Phosphohydrolases
Nitric Oxide
Necrotizing Enterocolitis
Intestinal Diseases
Focal Adhesion Protein-Tyrosine Kinases
Phosphorylation
Non-Receptor Type 11 Protein Tyrosine Phosphatase
Inflammation
Adhesiveness
Stress Fibers
Focal Adhesions
Nitric Oxide Synthase Type II
Bromodeoxyuridine
Knockout Mice
Mucous Membrane

Keywords

  • Bacterial translocation
  • Epithelial barrier
  • Intestinal inflammation
  • Necrotizing enterocolitis
  • Restitution

ASJC Scopus subject areas

  • Physiology
  • Hepatology
  • Gastroenterology
  • Physiology (medical)

Cite this

Nitric oxide inhibits enterocyte migration through activation of RhoA-GTPase in a SHP-2-dependent manner. / Cetin, Selma; Leaphart, Cynthia L.; Li, Jun; Ischenko, Irene; Hayman, Michael; Upperman, Jeffrey; Zamora, Ruben; Watkins, Simon; Ford, Henri; Wang, James; Hackam, David J.

In: American Journal of Physiology - Gastrointestinal and Liver Physiology, Vol. 292, No. 5, 01.05.2007.

Research output: Contribution to journalArticle

Cetin, Selma ; Leaphart, Cynthia L. ; Li, Jun ; Ischenko, Irene ; Hayman, Michael ; Upperman, Jeffrey ; Zamora, Ruben ; Watkins, Simon ; Ford, Henri ; Wang, James ; Hackam, David J. / Nitric oxide inhibits enterocyte migration through activation of RhoA-GTPase in a SHP-2-dependent manner. In: American Journal of Physiology - Gastrointestinal and Liver Physiology. 2007 ; Vol. 292, No. 5.
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T1 - Nitric oxide inhibits enterocyte migration through activation of RhoA-GTPase in a SHP-2-dependent manner

AU - Cetin, Selma

AU - Leaphart, Cynthia L.

AU - Li, Jun

AU - Ischenko, Irene

AU - Hayman, Michael

AU - Upperman, Jeffrey

AU - Zamora, Ruben

AU - Watkins, Simon

AU - Ford, Henri

AU - Wang, James

AU - Hackam, David J.

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N2 - Diseases of intestinal inflammation like necrotizing enterocolitis (NEC) are associated with impaired epithelial barrier integrity and the sustained release of intestinal nitric oxide (NO). NO modifies the cytoskeletal regulator RhoA-GTPase, suggesting that NO could affect barrier healing by inhibiting intestinal restitution. We now hypothesize that NO inhibits enterocyte migration through RhoA-GTPase and sought to determine the pathways involved. The induction of NEC was associated with increased enterocyte NO release and impaired migration of bromodeoxyuridine-labeled enterocytes from terminal ileal crypts to villus tips. In IEC-6 enterocytes, NO significantly inhibited enterocyte migration and activated RhoA-GTPase while increasing the formation of stress fibers. In parallel, exposure of IEC-6 cells to NO increased the phosphorylation of focal adhesion kinase (pFAK) and caused a striking increase in cell-matrix adhesiveness, suggesting a mechanism by which NO could impair enterocyte migration. NEC was associated with increased expression of pFAK in the terminal ileal mucosa of wild-type mice and a corresponding increase in disease severity compared with inducible NO synthase knockout mice, confirming the dependence of NO for FAK phosphorylation in vivo and its role in the pathogenesis of NEC. Strikingly, inhibition of the protein tyrosine phosphatase SHP-2 in IEC-6 cells prevented the activation of RhoA by NO, restored focal adhesions, and reversed the inhibitory effects of NO on enterocyte migration. These data indicate that NO impairs mucosal healing by inhibiting enterocyte migration through activation of RhoA in a SHP-2-dependent manner and support a possible role for SHP-2 as a therapeutic target in diseases of intestinal inflammation like NEC.

AB - Diseases of intestinal inflammation like necrotizing enterocolitis (NEC) are associated with impaired epithelial barrier integrity and the sustained release of intestinal nitric oxide (NO). NO modifies the cytoskeletal regulator RhoA-GTPase, suggesting that NO could affect barrier healing by inhibiting intestinal restitution. We now hypothesize that NO inhibits enterocyte migration through RhoA-GTPase and sought to determine the pathways involved. The induction of NEC was associated with increased enterocyte NO release and impaired migration of bromodeoxyuridine-labeled enterocytes from terminal ileal crypts to villus tips. In IEC-6 enterocytes, NO significantly inhibited enterocyte migration and activated RhoA-GTPase while increasing the formation of stress fibers. In parallel, exposure of IEC-6 cells to NO increased the phosphorylation of focal adhesion kinase (pFAK) and caused a striking increase in cell-matrix adhesiveness, suggesting a mechanism by which NO could impair enterocyte migration. NEC was associated with increased expression of pFAK in the terminal ileal mucosa of wild-type mice and a corresponding increase in disease severity compared with inducible NO synthase knockout mice, confirming the dependence of NO for FAK phosphorylation in vivo and its role in the pathogenesis of NEC. Strikingly, inhibition of the protein tyrosine phosphatase SHP-2 in IEC-6 cells prevented the activation of RhoA by NO, restored focal adhesions, and reversed the inhibitory effects of NO on enterocyte migration. These data indicate that NO impairs mucosal healing by inhibiting enterocyte migration through activation of RhoA in a SHP-2-dependent manner and support a possible role for SHP-2 as a therapeutic target in diseases of intestinal inflammation like NEC.

KW - Bacterial translocation

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KW - Necrotizing enterocolitis

KW - Restitution

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