Proteomic characterization of lipid rafts markers from the rat intestinal brush border

Nguyen, Hang Thi Thu; Amine, Adda Berkane; Lafitte, Daniel; Waheed, Abdül; Nicoletti, Cendrine; Villard, Claude; Létisse, Marion; Deyris, Valérie; Rozière, Muriel; Tchiakpe, Léopold; Danielle, Comeau-Druet; Comeau, Louis-Claude; Hiol, Abel

doi:10.1016/j.bbrc.2006.01.141

Cited by 39 publications

(38 citation statements)

References 35 publications

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“…The immunofluorescence method for GP2 stained the brush border of colonic enterocytes in Crohn's disease (16). The localization in the brush border may be supported by the proteomic characterization of lipid raft proteins from the brush border membrane of the rat jejunum (14). However, since there is no direct evidence showing mRNA expression of enterocytes at cellular levels, we cannot deny that GP2 detected in the brush border of enterocytes is derived from GP2 released from the pancreas.…”

Section: Discussionmentioning

confidence: 94%

The broad distribution of GP2 in mucous glands and secretory products

et al. 2016

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GP2, a GPI-anchored glycoprotein that is a useful marker for M cells of Peyer's patches, is functionally related to the uptake of pathogenic bacteria in the gut lumen. Our immunostaining throughout the whole body of mice detected a broader localization than previously found of GP2 in various mucous glands and secretory cells. In the oral cavity, the palatine gland and lingual gland intensely expressed GP2 with immunolabeling along the basolateral membrane of acini and in luminal secretions of ducts. Secretory portions of the duodenal gland as well as the pancreas were immunoreactive for GP2 in the digestive tract. Luminal contents in the small intestine contained aggregations of GP2-immunoreactive substances which mixed with bacteria. The bulbourethral gland of Cowper displayed the GP2 immunoreactivity among the male reproductive organs. The vaginal epithelium contained many GP2-immunoreactive goblet-like cells, the occurrence of which dramatically changed according to the estrous cycle. These findings show that GP2 is a popular secretory product released from mucous glands and secretory cells and may support defense mechanisms against pathogenic bacteria in the tubular organs open to the external milieu.

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Section: Discussionmentioning

confidence: 94%

The broad distribution of GP2 in mucous glands and secretory products

et al. 2016

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“…In addition to its membrane localization, it has also been reported that in response to fat feeding in vivo, the IAP protein is secreted outside of the cell and into the intestinal lumen (21,22). We therefore assayed both the cellular extract (cell lysate) and the surrounding media for IAP activity.…”

Section: Resultsmentioning

confidence: 99%

Intestinal alkaline phosphatase is a gut mucosal defense factor maintained by enteral nutrition

Goldberg

Austen

Zhang

et al. 2008

Proc. Natl. Acad. Sci. U.S.A.

318

323

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Under conditions of starvation and disease, the gut barrier becomes impaired, and trophic feeding to prevent gut mucosal atrophy has become a standard treatment of critically ill patients. However, the mechanisms responsible for the beneficial effects of enteral nutrition have remained a mystery. Using in vitro and in vivo models, we demonstrate that the brush-border enzyme, intestinal alkaline phosphatase (IAP), has the ability to detoxify lipopolysaccharide and prevent bacterial invasion across the gut mucosal barrier. IAP expression and function are lost with starvation and maintained by enteral feeding. It is likely that the IAP silencing that occurs during starvation is a key component of the gut mucosal barrier dysfunction seen in critically ill patients.A mong the most critical functions of the mammalian gut mucosa is to provide a barrier to luminal microbes and toxins while simultaneously allowing for the necessary digestion and absorption of dietary nutrients. The molecular mechanisms that govern barrier function are incompletely understood, but it is clear that under conditions of starvation and disease, the gut barrier becomes impaired, leading to significant morbidity and mortality (1-8). Trophic enteral feeding to prevent gut mucosal atrophy and resultant barrier dysfunction has become part of the standard treatment of intensive care unit patients (9-14). The mechanism(s) responsible for the beneficial effects of trophic feeding are not understood.Intestinal alkaline phosphatase (IAP), a brush-border protein that hydrolyzes monophosphate esters, is expressed exclusively in villus-associated enterocytes and is considered an excellent marker for crypt-villus differentiation (15-18). Narisawa et al. (19) reported that compared with their wild-type (WT) littermates, mice lacking IAP gained more weight under conditions of a high-fat diet. In addition, several studies have shown that the IAP enzyme is capable of detoxifying LPS, likely through dephosphorylation of the lipid A moiety, the primary source of its endotoxic effects (20). Despite these few reports, the physiological role of IAP within the gut has not been elucidated. Results and DiscussionTo examine the functional role of IAP, we developed in vitro model systems using intestinal cell lines (T84, HT-29, and IEC-6) that express little or no IAP under basal conditions. Stable cell lines were created that overexpress IAP (Fig. 1A), and enzyme assays were used to determine the cellular localization of the ectopically expressed IAP protein. The results in Fig. 1B show that parent cells make little, if any, endogenous IAP. In contrast, large amounts of IAP enzyme are seen in the stably transfected cells. Importantly, the vast majority of the IAP activity is in the membrane fraction as opposed to the cytosol. Fig. 1C shows the results of the control experiment used to validate our separation of membranous and cytosolic fractions, confirming that the majority of the MAPK enzyme activity exists within the cytosol rather than the membrane. This membrane ...

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“…The DRM proteomes of at least a dozen different cells and tissues are now available (11,(14)(15)(16)(23)(24)(25)(26)(27)(28)(29)(30)(31)(32)(33)(34)(35)(36)(37)(38)(39)(40)(41). However, it is extremely difficult, if not impossible, to purify any organelle to homogeneity (42), and the single-step centrifugation used to enrich rafts is no exception, so without rigorous controls a DRM proteome cannot be equated with a raft proteome.…”

Section: Resultsmentioning

confidence: 99%

Mitochondria do not contain lipid rafts, and lipid rafts do not contain mitochondrial proteins

Zheng

Berg

Foster

2009

Journal of Lipid Research

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Lipid rafts are membrane microdomains involved in many cellular functions, including transduction of cellular signals and cell entry by pathogens. Lipid rafts can be enriched biochemically by extraction in a nonionic detergent at low temperature, followed by floatation on a sucrose density gradient. Previous proteomic studies of such detergentresistant membranes (DRMs) are in disagreement about the presence of mitochondrial proteins in raft components. Here, we approach the status of mitochondrial proteins in DRM preparations by employing stable isotope labeling by amino acids in cell culture to evaluate the composition of differentially purified subcellular fractions as well as highresolution linear density gradients. Our data demonstrate that F 1 /F 0 ATPase subunits, voltage-dependent anion selective channels, and other mitochondrial proteins are at best partially copurifying contaminants of raft preparations.

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Proteomic characterization of lipid rafts markers from the rat intestinal brush border

Cited by 39 publications

References 35 publications

<b>The broad distribution of GP2 in mucous glands and secretory pr</b><b>oducts </b>

<b>The broad distribution of GP2 in mucous glands and secretory pr</b><b>oducts </b>

Intestinal alkaline phosphatase is a gut mucosal defense factor maintained by enteral nutrition

Mitochondria do not contain lipid rafts, and lipid rafts do not contain mitochondrial proteins

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