Endothelial TLR4 activation impairs intestinal microcirculatory perfusion in necrotizing enterocolitis via eNOS-NO-nitrite signaling

Ibrahim Yazji, Chhinder Sodhi, Elizabeth K. Lee, Misty Good, Charlotte E. Egan, Amin Afrazi, Matthew D. Neal, Hongpeng Jia, Joyce Lin, Congrong Ma, Maria F. Branca, Thomas Prindle, Ward M. Richardson, John Ozolek, Timothy R. Billiar, David G. Binion, Mark T. Gladwin, David Hackam

Research output: Contribution to journalArticle

Abstract

Necrotizing enterocolitis (NEC) is a devastating disease of premature infants characterized by severe intestinal necrosis and for which breast milk represents the most effective protective strategy. Previous studies have revealed a critical role for the lipopolysaccharide receptor toll-like receptor 4 (TLR4) in NEC development through its induction of mucosal injury, yet the reasons for which intestinal ischemia in NEC occurs in the first place remain unknown. We hypothesize that TLR4 signaling within the endothelium plays an essential role in NEC development by regulating perfusion to the small intestine via the vasodilatory molecule endothelial nitric oxide synthase (eNOS). Using a unique mouse system in which we selectively deleted TLR4 from the endothelium, we now show that endothelial TLR4 activation is required for NEC development and that endothelial TLR4 activation impairs intestinal perfusion without effects on other organs and reduces eNOS expression via activation of myeloid differentiation primary response gene 88. NEC severity was significantly increased in eNOS-/- mice and decreased upon administration of the phosphodiesterase inhibitor sildenafil, which augments eNOS function. Strikingly, compared with formula, human and mouse breast milk were enriched in sodium nitrate - a precursor for enteral generation of nitrite and nitric oxide - and repletion of formula with sodium nitrate/nitrite restored intestinal perfusion, reversed the deleterious effects of endothelial TLR4 signaling, and reduced NEC severity. These data identify that endothelial TLR4 critically regulates intestinal perfusion leading to NEC and reveal that the protective properties of breast milk involve enhanced intestinal microcirculatory integrity via augmentation of nitrate-nitrite-NO signaling.

Original languageEnglish (US)
Pages (from-to)9451-9456
Number of pages6
JournalProceedings of the National Academy of Sciences of the United States of America
Volume110
Issue number23
DOIs
StatePublished - Jun 4 2013
Externally publishedYes

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Necrotizing Enterocolitis
Toll-Like Receptor 4
Nitric Oxide Synthase Type III
Nitrites
Perfusion
Human Milk
Infant, Premature, Diseases
Small Intestine
Endothelium
CD14 Antigens
Sodium Nitrite
Phosphodiesterase Inhibitors
Nitrates
Nitric Oxide
Necrosis
Ischemia
Wounds and Injuries

Keywords

  • Infant formula
  • Neonatal inflammation
  • Neonatal nutrition
  • Prematurity
  • Sepsis

ASJC Scopus subject areas

  • General

Cite this

Endothelial TLR4 activation impairs intestinal microcirculatory perfusion in necrotizing enterocolitis via eNOS-NO-nitrite signaling. / Yazji, Ibrahim; Sodhi, Chhinder; Lee, Elizabeth K.; Good, Misty; Egan, Charlotte E.; Afrazi, Amin; Neal, Matthew D.; Jia, Hongpeng; Lin, Joyce; Ma, Congrong; Branca, Maria F.; Prindle, Thomas; Richardson, Ward M.; Ozolek, John; Billiar, Timothy R.; Binion, David G.; Gladwin, Mark T.; Hackam, David.

In: Proceedings of the National Academy of Sciences of the United States of America, Vol. 110, No. 23, 04.06.2013, p. 9451-9456.

Research output: Contribution to journalArticle

Yazji, I, Sodhi, C, Lee, EK, Good, M, Egan, CE, Afrazi, A, Neal, MD, Jia, H, Lin, J, Ma, C, Branca, MF, Prindle, T, Richardson, WM, Ozolek, J, Billiar, TR, Binion, DG, Gladwin, MT & Hackam, D 2013, 'Endothelial TLR4 activation impairs intestinal microcirculatory perfusion in necrotizing enterocolitis via eNOS-NO-nitrite signaling', Proceedings of the National Academy of Sciences of the United States of America, vol. 110, no. 23, pp. 9451-9456. https://doi.org/10.1073/pnas.1219997110
Yazji, Ibrahim ; Sodhi, Chhinder ; Lee, Elizabeth K. ; Good, Misty ; Egan, Charlotte E. ; Afrazi, Amin ; Neal, Matthew D. ; Jia, Hongpeng ; Lin, Joyce ; Ma, Congrong ; Branca, Maria F. ; Prindle, Thomas ; Richardson, Ward M. ; Ozolek, John ; Billiar, Timothy R. ; Binion, David G. ; Gladwin, Mark T. ; Hackam, David. / Endothelial TLR4 activation impairs intestinal microcirculatory perfusion in necrotizing enterocolitis via eNOS-NO-nitrite signaling. In: Proceedings of the National Academy of Sciences of the United States of America. 2013 ; Vol. 110, No. 23. pp. 9451-9456.
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T1 - Endothelial TLR4 activation impairs intestinal microcirculatory perfusion in necrotizing enterocolitis via eNOS-NO-nitrite signaling

AU - Yazji, Ibrahim

AU - Sodhi, Chhinder

AU - Lee, Elizabeth K.

AU - Good, Misty

AU - Egan, Charlotte E.

AU - Afrazi, Amin

AU - Neal, Matthew D.

AU - Jia, Hongpeng

AU - Lin, Joyce

AU - Ma, Congrong

AU - Branca, Maria F.

AU - Prindle, Thomas

AU - Richardson, Ward M.

AU - Ozolek, John

AU - Billiar, Timothy R.

AU - Binion, David G.

AU - Gladwin, Mark T.

AU - Hackam, David

PY - 2013/6/4

Y1 - 2013/6/4

N2 - Necrotizing enterocolitis (NEC) is a devastating disease of premature infants characterized by severe intestinal necrosis and for which breast milk represents the most effective protective strategy. Previous studies have revealed a critical role for the lipopolysaccharide receptor toll-like receptor 4 (TLR4) in NEC development through its induction of mucosal injury, yet the reasons for which intestinal ischemia in NEC occurs in the first place remain unknown. We hypothesize that TLR4 signaling within the endothelium plays an essential role in NEC development by regulating perfusion to the small intestine via the vasodilatory molecule endothelial nitric oxide synthase (eNOS). Using a unique mouse system in which we selectively deleted TLR4 from the endothelium, we now show that endothelial TLR4 activation is required for NEC development and that endothelial TLR4 activation impairs intestinal perfusion without effects on other organs and reduces eNOS expression via activation of myeloid differentiation primary response gene 88. NEC severity was significantly increased in eNOS-/- mice and decreased upon administration of the phosphodiesterase inhibitor sildenafil, which augments eNOS function. Strikingly, compared with formula, human and mouse breast milk were enriched in sodium nitrate - a precursor for enteral generation of nitrite and nitric oxide - and repletion of formula with sodium nitrate/nitrite restored intestinal perfusion, reversed the deleterious effects of endothelial TLR4 signaling, and reduced NEC severity. These data identify that endothelial TLR4 critically regulates intestinal perfusion leading to NEC and reveal that the protective properties of breast milk involve enhanced intestinal microcirculatory integrity via augmentation of nitrate-nitrite-NO signaling.

AB - Necrotizing enterocolitis (NEC) is a devastating disease of premature infants characterized by severe intestinal necrosis and for which breast milk represents the most effective protective strategy. Previous studies have revealed a critical role for the lipopolysaccharide receptor toll-like receptor 4 (TLR4) in NEC development through its induction of mucosal injury, yet the reasons for which intestinal ischemia in NEC occurs in the first place remain unknown. We hypothesize that TLR4 signaling within the endothelium plays an essential role in NEC development by regulating perfusion to the small intestine via the vasodilatory molecule endothelial nitric oxide synthase (eNOS). Using a unique mouse system in which we selectively deleted TLR4 from the endothelium, we now show that endothelial TLR4 activation is required for NEC development and that endothelial TLR4 activation impairs intestinal perfusion without effects on other organs and reduces eNOS expression via activation of myeloid differentiation primary response gene 88. NEC severity was significantly increased in eNOS-/- mice and decreased upon administration of the phosphodiesterase inhibitor sildenafil, which augments eNOS function. Strikingly, compared with formula, human and mouse breast milk were enriched in sodium nitrate - a precursor for enteral generation of nitrite and nitric oxide - and repletion of formula with sodium nitrate/nitrite restored intestinal perfusion, reversed the deleterious effects of endothelial TLR4 signaling, and reduced NEC severity. These data identify that endothelial TLR4 critically regulates intestinal perfusion leading to NEC and reveal that the protective properties of breast milk involve enhanced intestinal microcirculatory integrity via augmentation of nitrate-nitrite-NO signaling.

KW - Infant formula

KW - Neonatal inflammation

KW - Neonatal nutrition

KW - Prematurity

KW - Sepsis

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