Endotoxin differentially modulates the basolateral and apical sodium/proton exchangers (NHE) in enterocytes

Selma Cetin, Joshua Dunklebarger, Jun Li, Patricia Boyle, Orkan Ergun, Faisal Qureshi, Henri Ford, Jeffrey Upperman, Simon Watkins, David J. Hackam

Research output: Contribution to journalArticle

31 Citations (Scopus)

Abstract

Background Maintenance of enterocyte activity during extracellular acidosis requires functional sodium/proton exchangers (NHE), which are present at both basolateral and apical surfaces. Necrotizing enterocolitis is characterized by systemic hypoperfusion, metabolic acidosis, and the apical to basolateral translocation of endotoxin (lipopolysaccharide [LPS]). We hypothesized that LPS differentially impairs NHE activity at the basolateral or apical domains of enterocytes, leading to cellular acidification, and explored the mechanisms involved. Methods Experimental necrotizing enterocolitis (NEC) was induced in newborn rats using a combination of gavage feeds and hypoxia. NHE isoforms were assessed in primary and cultured enterocytes by Western blot analysis and by confocal microscopy in the presence or absence of LPS. NHE activity was detected by single-cell fluorescent ratiometric imaging with the use of the pH-sensitive dye 2′, 7′-bis-(2carboxyethyl) -5-(and-6)-carboxyfluorescein. Results In both NEC and control rats, NHE1 was basolateral and NHE3 was apical. A similar distribution was observed in polarized IEC-6 cells. LPS caused a dose-dependent reduction in basolateral NHE1 activity in IEC-6 cells, but had no effect on apical NHE3 activity. This effect could not be accounted for by reduced expression or impaired plasma membrane localization of NHE isoforms. Strikingly, LPS-mediated NHE1 impairment caused marked cytoplasmic acidification under conditions of extracellular acidosis, whereas functional NHE1 maintained cytoplasmic pH homeostasis in control cells. Conclusions LPS selectively impairs basolateral NHE1 but not apical NHE3, leading to cytoplasmic acidification during extracellular acidosis. This effect could mediate impaired enterocyte function after LPS translocation and suggests a mechanism leading to barrier disruption in NEC.

Original languageEnglish (US)
Pages (from-to)375-383
Number of pages9
JournalSurgery
Volume136
Issue number2
DOIs
StatePublished - Aug 1 2004

Fingerprint

Enterocytes
Endotoxins
Lipopolysaccharides
Protons
Sodium
Necrotizing Enterocolitis
Acidosis
Protein Isoforms
Confocal Microscopy
Homeostasis
Western Blotting
Maintenance
Cell Membrane

ASJC Scopus subject areas

  • Surgery

Cite this

Endotoxin differentially modulates the basolateral and apical sodium/proton exchangers (NHE) in enterocytes. / Cetin, Selma; Dunklebarger, Joshua; Li, Jun; Boyle, Patricia; Ergun, Orkan; Qureshi, Faisal; Ford, Henri; Upperman, Jeffrey; Watkins, Simon; Hackam, David J.

In: Surgery, Vol. 136, No. 2, 01.08.2004, p. 375-383.

Research output: Contribution to journalArticle

Cetin, S, Dunklebarger, J, Li, J, Boyle, P, Ergun, O, Qureshi, F, Ford, H, Upperman, J, Watkins, S & Hackam, DJ 2004, 'Endotoxin differentially modulates the basolateral and apical sodium/proton exchangers (NHE) in enterocytes', Surgery, vol. 136, no. 2, pp. 375-383. https://doi.org/10.1016/j.surg.2004.05.013
Cetin, Selma ; Dunklebarger, Joshua ; Li, Jun ; Boyle, Patricia ; Ergun, Orkan ; Qureshi, Faisal ; Ford, Henri ; Upperman, Jeffrey ; Watkins, Simon ; Hackam, David J. / Endotoxin differentially modulates the basolateral and apical sodium/proton exchangers (NHE) in enterocytes. In: Surgery. 2004 ; Vol. 136, No. 2. pp. 375-383.
@article{9eaf3e2f511147d58399155ecbf3510d,
title = "Endotoxin differentially modulates the basolateral and apical sodium/proton exchangers (NHE) in enterocytes",
abstract = "Background Maintenance of enterocyte activity during extracellular acidosis requires functional sodium/proton exchangers (NHE), which are present at both basolateral and apical surfaces. Necrotizing enterocolitis is characterized by systemic hypoperfusion, metabolic acidosis, and the apical to basolateral translocation of endotoxin (lipopolysaccharide [LPS]). We hypothesized that LPS differentially impairs NHE activity at the basolateral or apical domains of enterocytes, leading to cellular acidification, and explored the mechanisms involved. Methods Experimental necrotizing enterocolitis (NEC) was induced in newborn rats using a combination of gavage feeds and hypoxia. NHE isoforms were assessed in primary and cultured enterocytes by Western blot analysis and by confocal microscopy in the presence or absence of LPS. NHE activity was detected by single-cell fluorescent ratiometric imaging with the use of the pH-sensitive dye 2′, 7′-bis-(2carboxyethyl) -5-(and-6)-carboxyfluorescein. Results In both NEC and control rats, NHE1 was basolateral and NHE3 was apical. A similar distribution was observed in polarized IEC-6 cells. LPS caused a dose-dependent reduction in basolateral NHE1 activity in IEC-6 cells, but had no effect on apical NHE3 activity. This effect could not be accounted for by reduced expression or impaired plasma membrane localization of NHE isoforms. Strikingly, LPS-mediated NHE1 impairment caused marked cytoplasmic acidification under conditions of extracellular acidosis, whereas functional NHE1 maintained cytoplasmic pH homeostasis in control cells. Conclusions LPS selectively impairs basolateral NHE1 but not apical NHE3, leading to cytoplasmic acidification during extracellular acidosis. This effect could mediate impaired enterocyte function after LPS translocation and suggests a mechanism leading to barrier disruption in NEC.",
author = "Selma Cetin and Joshua Dunklebarger and Jun Li and Patricia Boyle and Orkan Ergun and Faisal Qureshi and Henri Ford and Jeffrey Upperman and Simon Watkins and Hackam, {David J.}",
year = "2004",
month = "8",
day = "1",
doi = "10.1016/j.surg.2004.05.013",
language = "English (US)",
volume = "136",
pages = "375--383",
journal = "Surgery (United States)",
issn = "0039-6060",
publisher = "Mosby Inc.",
number = "2",

}

TY - JOUR

T1 - Endotoxin differentially modulates the basolateral and apical sodium/proton exchangers (NHE) in enterocytes

AU - Cetin, Selma

AU - Dunklebarger, Joshua

AU - Li, Jun

AU - Boyle, Patricia

AU - Ergun, Orkan

AU - Qureshi, Faisal

AU - Ford, Henri

AU - Upperman, Jeffrey

AU - Watkins, Simon

AU - Hackam, David J.

PY - 2004/8/1

Y1 - 2004/8/1

N2 - Background Maintenance of enterocyte activity during extracellular acidosis requires functional sodium/proton exchangers (NHE), which are present at both basolateral and apical surfaces. Necrotizing enterocolitis is characterized by systemic hypoperfusion, metabolic acidosis, and the apical to basolateral translocation of endotoxin (lipopolysaccharide [LPS]). We hypothesized that LPS differentially impairs NHE activity at the basolateral or apical domains of enterocytes, leading to cellular acidification, and explored the mechanisms involved. Methods Experimental necrotizing enterocolitis (NEC) was induced in newborn rats using a combination of gavage feeds and hypoxia. NHE isoforms were assessed in primary and cultured enterocytes by Western blot analysis and by confocal microscopy in the presence or absence of LPS. NHE activity was detected by single-cell fluorescent ratiometric imaging with the use of the pH-sensitive dye 2′, 7′-bis-(2carboxyethyl) -5-(and-6)-carboxyfluorescein. Results In both NEC and control rats, NHE1 was basolateral and NHE3 was apical. A similar distribution was observed in polarized IEC-6 cells. LPS caused a dose-dependent reduction in basolateral NHE1 activity in IEC-6 cells, but had no effect on apical NHE3 activity. This effect could not be accounted for by reduced expression or impaired plasma membrane localization of NHE isoforms. Strikingly, LPS-mediated NHE1 impairment caused marked cytoplasmic acidification under conditions of extracellular acidosis, whereas functional NHE1 maintained cytoplasmic pH homeostasis in control cells. Conclusions LPS selectively impairs basolateral NHE1 but not apical NHE3, leading to cytoplasmic acidification during extracellular acidosis. This effect could mediate impaired enterocyte function after LPS translocation and suggests a mechanism leading to barrier disruption in NEC.

AB - Background Maintenance of enterocyte activity during extracellular acidosis requires functional sodium/proton exchangers (NHE), which are present at both basolateral and apical surfaces. Necrotizing enterocolitis is characterized by systemic hypoperfusion, metabolic acidosis, and the apical to basolateral translocation of endotoxin (lipopolysaccharide [LPS]). We hypothesized that LPS differentially impairs NHE activity at the basolateral or apical domains of enterocytes, leading to cellular acidification, and explored the mechanisms involved. Methods Experimental necrotizing enterocolitis (NEC) was induced in newborn rats using a combination of gavage feeds and hypoxia. NHE isoforms were assessed in primary and cultured enterocytes by Western blot analysis and by confocal microscopy in the presence or absence of LPS. NHE activity was detected by single-cell fluorescent ratiometric imaging with the use of the pH-sensitive dye 2′, 7′-bis-(2carboxyethyl) -5-(and-6)-carboxyfluorescein. Results In both NEC and control rats, NHE1 was basolateral and NHE3 was apical. A similar distribution was observed in polarized IEC-6 cells. LPS caused a dose-dependent reduction in basolateral NHE1 activity in IEC-6 cells, but had no effect on apical NHE3 activity. This effect could not be accounted for by reduced expression or impaired plasma membrane localization of NHE isoforms. Strikingly, LPS-mediated NHE1 impairment caused marked cytoplasmic acidification under conditions of extracellular acidosis, whereas functional NHE1 maintained cytoplasmic pH homeostasis in control cells. Conclusions LPS selectively impairs basolateral NHE1 but not apical NHE3, leading to cytoplasmic acidification during extracellular acidosis. This effect could mediate impaired enterocyte function after LPS translocation and suggests a mechanism leading to barrier disruption in NEC.

UR - http://www.scopus.com/inward/record.url?scp=3843060378&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=3843060378&partnerID=8YFLogxK

U2 - 10.1016/j.surg.2004.05.013

DO - 10.1016/j.surg.2004.05.013

M3 - Article

VL - 136

SP - 375

EP - 383

JO - Surgery (United States)

JF - Surgery (United States)

SN - 0039-6060

IS - 2

ER -