Effect of LPS on basal and induced apo E secretion by 25-OH chol and 9cRA in differentiated CaCo-2.
The infection and inflammation process is associated with disturbances in lipid and lipoprotein metabolism. The apolipoprotein E (apo E) plays an important role in the lipoprotein metabolism and has been linked to inflammatory disease such as atherosclerosis and Alzheimer disease. An anti-inflammatory effect has also been suggested. The heterodimer nuclear receptor Liver-X-Receptor(alpha)/Retinoid-X-Receptor (LXR(alpha)/RXR) is considered to be a transcription factor for apo E. The aim of this study was to determine whether lipopolysaccharide (LPS) (principal component of the outer membrane Gram-negative bacteria) has an effect on apo E secretion by intestinal mucosa cells, using the Caco-2 cell line. Differentiated Caco-2 cells grown on filter inserts were incubated apically with LPS and/or 25-hydroxycholesterol (25-OH chol) and 9 cis retinoic acid (9cRA), ligands of LXR and RXR, respectively. The apical and basolateral media were separately collected. Apo E was detected by specific antibodies after protein separation by Two-dimensional nondenaturing gradient gel electrophoresis and apo E secreted in the cell culture media was measured by enzyme linked immunosorbent assay (ELISA). Apo E mRNA was analyzed by reverse transcription-polymerase chain reaction (RT-PCR). LXR(alpha) and RXR mass was analyzed by Western Blot. We demonstrate here that CaCo-2 cells secrete apo E, by either apical or basolateral sides, associated with a high-density like lipoprotein, with a stoke's diameter comprised between 7.10 and 8.16 nm. We show that only apical secretion is decreased by LPS in a dose and time dependent manner. This is associated with a decrease in apo E gene expression contrasting with an increase of Il-8, a chemokine factor. Moreover, we demonstrate that only basolateral apo E secretion by CaCo-2 is significantly increased by 25-OH chol and 9cRA while apical secretion remains unchanged. LPS does not decrease the 25-OH chol and 9cRA mediated apo E secretion in basolateral compartment, while apical secretion is diminished under these circumstances. Our results provide evidence for the polarized secretion of apo E by intestinal epithelium. They also demonstrate that apo E secretion by CaCo-2 cell line is decreased by LPS through an LXR(alpha)/RXR independent signaling pathway.
Ripollés Piquer
B
B
Laboratoire de Biochimie Fondamentale et Appliquée, UFR de Pharmacie, Nantes, France.
Nazih
H
H
Neunlist
M
M
Huvelin
J M
JM
Bard
J M
JM
D000077556:Alitretinoin; D001057:Apolipoproteins E / Q000235:genetics / Q000378:metabolism*; D018938:Caco-2 Cells; D002454:Cell Differentiation; D016764:Cell Polarity; D005786:Gene Expression Regulation / Q000187:drug effects; D006801:Humans; D006888:Hydroxycholesterols / Q000037:antagonists & inhibitors* / Q000494:pharmacology; D016209:Interleukin-8 / Q000378:metabolism; D008070:Lipopolysaccharides / Q000494:pharmacology* / Q000633:toxicity; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D014212:Tretinoin / Q000037:antagonists & inhibitors* / Q000494:pharmacology
D001057:Apolipoproteins E; D006888:Hydroxycholesterols; D016209:Interleukin-8; D008070:Lipopolysaccharides; D012333:RNA, Messenger; D000077556:Alitretinoin; D014212:Tretinoin; C007997:25-hydroxycholesterol