Highlights
- •Binge alcohol exposure caused apoptosis of gut enterocytes in rats and WT mice.
- •Binge alcohol exposure reduced the levels of tight junction and adherent junction proteins.
- •Nitration of the junctional proteins led to ubiquitin-dependent degradation.
- •Apoptosis of enterocytes and reduced junctional proteins led to gut leakiness.
- •Binge alcohol-induced gut leakiness was observed in mice, rats and humans.
Background & Aims
Binge alcohol exposure causes gut leakiness, contributing to increased endotoxemia
and inflammatory liver injury, although the molecular mechanisms are still elusive.
This study was aimed at investigating the roles of apoptosis of enterocytes and nitration
followed by degradation of intestinal tight junction (TJ) and adherens junction (AJ)
proteins in binge alcohol-induced gut leakiness.
Methods
The levels of intestinal (ileum) junctional complex proteins, oxidative stress markers
and apoptosis-related proteins in rodents, T84 colonic cells and autopsied human ileums
were determined by immunoblot, immunoprecipitation, immunofluorescence, and mass-spectral
analyses.
Results
Binge alcohol exposure caused apoptosis of gut enterocytes with elevated serum endotoxin
and liver injury. The levels of intestinal CYP2E1, iNOS, nitrated proteins and apoptosis-related
marker proteins were significantly elevated in binge alcohol-exposed rodents. Differential,
quantitative mass-spectral analyses of the TJ-enriched fractions of intestinal epithelial
layers revealed that several TJ, AJ and desmosome proteins were decreased in binge
alcohol-exposed rats compared to controls. Consistently, the levels of TJ proteins
(claudin-1, claudin-4, occludin and zonula occludens-1), AJ proteins (β-catenin and
E-cadherin) and desmosome plakoglobin were very low in binge alcohol-exposed rats,
wild-type mice, and autopsied human ileums but not in Cyp2e1-null mice. Additionally, pretreatment with specific inhibitors of CYP2E1 and iNOS
prevented disorganization and/or degradation of TJ proteins in alcohol-exposed T84
colonic cells. Furthermore, immunoprecipitation followed by immunoblot confirmed that
intestinal TJ and AJ proteins were nitrated and degraded via ubiquitin-dependent proteolysis,
resulting in their decreased levels.
Conclusions
These results demonstrated for the first time the critical roles of CYP2E1, apoptosis
of enterocytes, and nitration followed by ubiquitin-dependent proteolytic degradation
of the junctional complex proteins, in promoting binge alcohol-induced gut leakiness
and endotoxemia, contributing to inflammatory liver disease.
Lay summary
Binge alcohol exposure causes gut leakiness, contributing to increased endotoxemia
and inflammatory liver injury. Our results demonstrated for the first time the critical
roles of apoptosis of enterocytes and nitration followed by ubiquitin-dependent proteolytic
degradation of the junctional complex proteins in promoting this gut leakiness and
endotoxemia. These results provide insight into the molecular mechanisms of alcohol-induced
inflammatory liver disease.
Graphical abstract
Graphical Abstract
Keywords
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Article info
Publication history
Published online: February 17, 2018
Accepted:
February 7,
2018
Received in revised form:
February 7,
2018
Received:
September 5,
2017
Identification
Copyright
Published by Elsevier B.V. on behalf of European Association for the Study of the Liver.
