Sensing of Dietary Lipids by Enterocytes: A New Role for SR-BI/CLA-1

BACKGROUND: The intestine is responsible for absorbing dietary lipids and delivering them to the organism as triglyceride-rich lipoproteins (TRL). It is important to determine how this process is regulated in enterocytes, the absorptive cells of the intestine, as prolonged postprandial hypertriglyceridemia is a known risk factor for atherosclerosis. During the postprandial period, dietary lipids, mostly triglycerides (TG) hydrolyzed by pancreatic enzymes, are combined with bile products and reach the apical membrane of enterocytes as postprandial micelles (PPM). Our aim was to determine whether these micelles induce, in enterocytes, specific early cell signaling events that could control the processes leading to TRL secretion. METHODOLOGY/PRINCIPAL FINDINGS: The effects of supplying PPM to the apex of Caco-2/TC7 enterocytes were analyzed. Micelles devoid of TG hydrolysis products, like those present in the intestinal lumen in the interprandial period, were used as controls. The apical delivery of PPM specifically induced a number of cellular events that are not induced by interprandial micelles. These early events included the trafficking of apolipoprotein B, a structural component of TRL, from apical towards secretory domains, and the rapid, dose-dependent activation of ERK and p38MAPK. PPM supply induced the scavenger receptor SR-BI/CLA-1 to cluster at the apical brush border membrane and to move from non-raft to raft domains. Competition, inhibition or knockdown of SR-BI/CLA-1 impaired the PPM-dependent apoB trafficking and ERK activation. CONCLUSIONS/SIGNIFICANCE: These results are the first evidence that enterocytes specifically sense postprandial dietary lipid-containing micelles. SR-BI/CLA-1 is involved in this process and could be a target for further study with a view to modifying intestinal TRL secretion early in the control pathway

Régulation de l'expression du CRLR et des ramps dans le système cardiovasculaire

PARIS7-Bibliothèque centrale (751132105) / SudocSudocFranceF

RAMP et récepteurs couplés aux protéines G

Les RAMP (receptor activity-modifying protein) modifient le caractère fonctionnel des récepteurs couplés aux protéines G (RCPG) des peptides de la famille de la calcitonine. Il semble que l’on puisse étendre l’interaction des RAMP à d’autres RCPG de classe II, puisque ces protéines s’associent aux récepteurs de l’hormone parathyroïdienne, du glucagon et du VIP/PACAP (vasoactive intestinal peptide/pituitary adenylate cyclase activating polypeptide) dénommé également R-VPAC1 (vasoactive-intestinal peptide PACAP receptor 1). Une fonction nouvelle des RAMP émerge de ces observations, car le complexe RAMP2/R-VPAC1 potentialise la voie de signalisation des phospho-inositides. La régulation de l’expression des RAMP à un niveau transcriptionnel ou post-transcriptionnel revêt une certaine importance, puisqu’elle peut influencer la réponse d’une cellule cible à un ligand, en modifiant le caractère fonctionnel d’un RCPG de classe II ou en affectant le niveau d’une voie de signalisation de ces récepteurs.RAMPs (receptor activity-modifying proteins) were discovered in 1998 as accessory proteins needed to the functionnal activity of CGRP (calcitonin gene-related peptide) receptors. Three RAMPs generated by three different genes are known in human, rat and mice. The coding sequences of such genes are described, but as yet, regulation sequences are unknown. RAMPs interact with GPCR (G protein-coupled receptors) of class II. In the case of the calcitonin/CGRP peptide family, RAMPs determine the functionnal specificity of the receptor, glycosylate and translocate the receptor to the cell surface. CGRP receptors are observed in presence of the RAMP1/calcitonin receptor-like receptor (CRLR), but the association of RAMP2 or RAMP3 with CRLR generates an adrenomedullin receptor. The calcitonin receptor (CTR) is translocated alone to the cell surface, but interactions of RAMPs with CTR forms amylin receptors. If RAMPs can interact with glucagon, parathyroid hormone and VIP/PACAP (vasoactive intestinal peptide/pituitary adenylate cyclase activating polypeptide (VPACR1)) receptors, the functionnal specificity of these receptors remains unaltered. However, the complex VPACR1/RAMP2 enhances specifically the phosphoinoside signaling pathway

Détection luminale des micelles lipidiques

Post-prandial hypertriglyceridemia is a risk factor for metabolic diseases. The intestine, through its role in alimentary lipid absorption, participates in the secretion of lipoprotein rich-triglycerides (TRL) and contributes to the increase in plasma triglyceride levels during the postprandial state. Understanding the molecular mechanisms involved in the secretion of intestinal TRL would allow the identification of new drug targets for treatment of metabolic diseases. The sensing of lipids by intestinal cells represents a promising mechanism allowing the modulation of TRL secretion. While many studies show the importance of enteroendocrine cells in the detection of alimentary lipids, several evidence suggest also the implication of enterocytes, the absorptive intestinal cells, in this process. Recent experimental results on the role of the scavenger receptor SR-BI in the detection of dietary lipids, supplied in their physiological form of postprandial lipid micelles, are reviewed

Sensing of Dietary Lipids by Enterocytes: A New Role for SR-BI/CLA-1

Background: The intestine is responsible for absorbing dietary lipids and delivering them to the organism as triglyceriderich lipoproteins (TRL). It is important to determine how this process is regulated in enterocytes, the absorptive cells of the intestine, as prolonged postprandial hypertriglyceridemia is a known risk factor for atherosclerosis. During the postprandial period, dietary lipids, mostly triglycerides (TG) hydrolyzed by pancreatic enzymes, are combined with bile products and reach the apical membrane of enterocytes as postprandial micelles (PPM). Our aim was to determine whether these micelles induce, in enterocytes, specific early cell signaling events that could control the processes leading to TRL secretion. Methodology/Principal Findings: The effects of supplying PPM to the apex of Caco-2/TC7 enterocytes were analyzed. Micelles devoid of TG hydrolysis products, like those present in the intestinal lumen in the interprandial period, were used as controls. The apical delivery of PPM specifically induced a number of cellular events that are not induced by interprandial micelles. These early events included the trafficking of apolipoprotein B, a structural component of TRL, from apical towards secretory domains, and the rapid, dose-dependent activation of ERK and p38MAPK. PPM supply induced the scavenger receptor SR-BI/CLA-1 to cluster at the apical brush border membrane and to move from non-raft to raft domains. Competition, inhibition or knockdown of SR-BI/CLA-1 impaired the PPM-dependent apoB trafficking and ERK activation. Conclusions/Significance: These results are the first evidence that enterocytes specifically sense postprandial dietary lipidcontainin

Détection luminale des micelles lipidiques

Post-prandial hypertriglyceridemia is a risk factor for metabolic diseases. The intestine, through its role in alimentary lipid absorption, participates in the secretion of lipoprotein rich-triglycerides (TRL) and contributes to the increase in plasma triglyceride levels during the postprandial state. Understanding the molecular mechanisms involved in the secretion of intestinal TRL would allow the identification of new drug targets for treatment of metabolic diseases. The sensing of lipids by intestinal cells represents a promising mechanism allowing the modulation of TRL secretion. While many studies show the importance of enteroendocrine cells in the detection of alimentary lipids, several evidence suggest also the implication of enterocytes, the absorptive intestinal cells, in this process. Recent experimental results on the role of the scavenger receptor SR-BI in the detection of dietary lipids, supplied in their physiological form of postprandial lipid micelles, are reviewed

PPM supply induces movement of SR-BI/CLA-1 towards raft microdomains.

<p>(A) Caco-2/TC7 cells were harvested in the presence of Triton X-100 and the lysate fractionated on a 5–40% sucrose gradient. Eleven fractions were collected for immunoblots of SR-BI/CLA-1, EEA1 (early endosome antigen 1) and flottilin-1 (raft marker). (B) Caco-2/TC7 cells were cultured in the absence (control) or presence of PPM or IPM for 10 min and then harvested in the presence of Triton X-100. Cell lysates were applied to a 5–40% sucrose gradient and eleven fractions collected. Fractions 3 to 8 were analyzed by immunoblotting with antibodies against SR-BI/CLA-1 (left panel) and flottilin-1 (right panel). (C) Immunolocalization of SR-BI/CLA-1 and alkaline phosphatase (PLAP, used as raft marker) in the brush border of Caco-2/TC7 cells supplied with PPM. SR-BI/CLA-1 is labelled with anti-rabbit immunoglobulin-gold complexes (18-nm particles) and PLAP with anti-sheep immunoglobulin-gold complexes (12-nm particles). MV, microvilli; bar, 100 nm.</p

Signaling pathways specifically induced by postprandial micelles.

<p>(A) Kinetworks™ phosphoprotein immunoblots from lysates of Caco-2/TC7 cells cultured on filters without micelles (control) or with PPM or IPM for 5 min. Duplicate immunoblots using specific antibodies directed against phosphoproteins were analyzed by Kinexus. Bands corresponding to phosphoproteins specifically up-regulated by PPM are indicated by arrows. The antibodies recognized the phosphosites: S144/S141/S154 for PAK1/2/3, T180/Y182 for p38αMAPK, S129/S133 for CREB1, T202/Y204 for ERK1, T185/Y187 for ERK2 and S338 for PKA Cβ. The phosphosites analyzed for MEK1 are indicated in the figure. Histograms show the amounts of phosphoproteins expressed as the ratio of phosphorylation in the PPM versus the IPM sample (from three independent experiments). (B) Caco-2/TC7 cells cultured on semi-permeable filters were incubated in the absence or presence of PPM or IPM in the apical compartment for the indicated times. Cell lysates were analyzed by immunoblot with antibodies against phospho-ERK1/2 (P-ERK), and phospho-p38MAPK (P-p38 MAPK). Total ERK, p38MAPK and E-cadherin (E-cadh) were used as loading controls. (C) Quantification of normalized P-ERK and P-p38MAPK levels (from three independent experiments) in the absence (c) or presence of PPM or IPM for 10 min (for P-ERK and ERK) or for 5 min (for P-p38MAPK and p38MAPK), **p<0.01. (D) Caco-2/TC7 cells were cultured in the presence of various amounts of PPM supplied in the apical compartment for 10 min. Cell lysates were analyzed by immunoblot with the same antibodies as in (B) and a blot representative of three independent analyses is shown.</p