Biomedical subjects

Sylvie Demignot

Publications and source records attributed to Sylvie Demignot.

5 recordsLinked to original sources

PubMed2025

Kinetic Redox Shotgun Proteomics Reveals Specific Lipopolysaccharide Effects on Intestinal Epithelial Cells, Mitigated by a Mn Superoxide Dismutase Mimic.

Overproduction of reactive oxygen species and antioxidant superoxide dismutases (SOD1, SOD2) dysregulation contribute to chronic inflammation such as generated in inflammatory bowel diseases (IBD). A kinetic redox shotgun proteomic strategy (OcSILAC for Oxidized cysteine Stable Isotope Labelling by Amino acids in Cell culture) was used to explore the lipopolysaccharide (LPS) effects including LPS-induced oxidation and inflammation cascades on a dedicated intestinal epithelial cell line (HT29-MD2) together with the potential mitigating role of a Mn-based SOD-mimic Mn1. While LPS induced transient oxidative damages at early times (15 min), cells incubated with Mn1 showed, in this time frame, a significantly reduced cysteine oxidation, highlighting Mn1 antioxidant properties. Over time, cysteine oxidation of LPS-treated cells was counteracted by an overexpression of antioxidant proteins (SOD1, NQO1) and a late (6 h) preponderant increase in SOD2 level. Mn1, when co-incubated with LPS, attenuated the level of most LPS-modified proteins, that is, proteins involved in the inflammatory response. Our results highlight Mn1 as a potentially effective antioxidant and anti-inflammatory agent to consider in the treatment of IBD, as well as a useful tool for exploring the interconnection between oxidative stress and inflammation.

Lipopolysaccharides↗

PubMed2006

Adaptation of enterocytic Caco-2 cells to glucose modulates triacylglycerol-rich lipoprotein secretion through triacylglycerol targeting into the endoplasmic reticulum lumen.

Enterocytes are responsible for the absorption of dietary lipids, which involves TRL [TG (triacylglycerol)-rich lipoprotein] assembly and secretion. In the present study, we analysed the effect on TRL secretion of Caco-2 enterocyte adaptation to a differential glucose supply. We showed that TG secretion in cells adapted to a low glucose supply for 2 weeks after confluence was double that of control cells maintained in high-glucose-containing medium, whereas the level of TG synthesis remained similar in both conditions. This increased secretion resulted mainly from an enlargement of the mean size of the secreted TRL. The increased TG availability for TRL assembly and secretion was not due to an increase in the MTP (microsomal TG transfer protein) activity that is required for lipid droplet biogenesis in the ER (endoplasmic reticulum) lumen, or to the channelling of absorbed fatty acids towards the monoacylglycerol pathway for TG synthesis. Interestingly, by electron microscopy and subcellular fractionation studies, we observed, in the low glucose condition, an increase in the TG content available for lipoprotein assembly in the ER lumen, with the cytosolic/microsomal TG levels being verapamil-sensitive. Overall, we demonstrate that Caco-2 enterocytes modulate TRL secretion through TG partitioning between the cytosol and the ER lumen according to the glucose supply. Our model will help in identifying the proteins involved in the control of the balance between TRL assembly and cytosolic lipid storage. This mechanism may be a way for enterocytes to regulate TRL secretion after a meal, and thus impact on our understanding of post-prandial hypertriglyceridaemia.

Acyltransferases↗

PubMed2005

Lipid micelles stimulate the secretion of triglyceride-enriched apolipoprotein B48-containing lipoproteins by Caco-2 cells.

Intestinal triglyceride-rich lipoproteins (TRL) are synthesized from dietary lipids. This study was designed to evaluate the effects of lipid micelles, mimicking post-digestive duodenal micelles, on the fate of apolipoprotein B (apoB)48-containing lipoproteins by Caco-2 cells. Such micelles, consisting of oleic acid (OA), taurocholate, 2-monooleoylglycerol (2-MO), cholesterol (Chol), and L-alpha-lysophospatidylcholine, were the most efficient inducers of OA uptake and esterification. The efficiency of TG and apoB48 secretion increased specifically as a function of cell differentiation. PAGE analysis of secreted lipoproteins separated by sequential ultracentrifugation after [35S] labeling revealed differences in the secretion of apoB100- and apoB48-containing lipoproteins. In absence of micelles, apoB48 was secreted mostly in "HDL-like" particles, as observed in enterocytes in vivo. Micelle application increased 2.7-fold the secretion of apoB, resulting in 53 times more apoB48 being recovered as TG-enriched lipoproteins at d < 1.006 g/ml. Electron microscopy revealed the presence of lipid droplets in the secretory pathway and the accumulation of newly synthesized TG in cytoplasmic lipid droplets, as in enterocytes in vivo. We showed that these droplets could be used for secretion. However, apoB48 preferentially bound to newly synthesized TG in the presence of micelles, accounting in part for the functional advantage of apoB editing in the intestine. While Caco-2 cells express both apoB isoforms, our results show that the apical supply of complex lipid micelles favors the physiological route of apoB48-containing TG-enriched lipoproteins.

Apolipoprotein B-48↗

PubMed2004

Apple procyanidins decrease cholesterol esterification and lipoprotein secretion in Caco-2/TC7 enterocytes.

Decrease of plasma lipid levels by polyphenols was linked to impairment of hepatic lipoprotein secretion. However, the intestine is the first epithelium that faces dietary compounds, and it contributes to lipid homeostasis by secreting triglyceride-rich lipoproteins during the postprandial state. The purpose of this study was to examine the effect of apple and wine polyphenol extracts on lipoprotein synthesis and secretion in human Caco-2/TC7 enterocytes apically supplied with complex lipid micelles. Our results clearly demonstrate that apple, but not wine, polyphenol extract dose-dependently decreases the esterification of cholesterol and the enterocyte secretion of lipoproteins. Apple polyphenols decrease apolipoprotein B (apoB) secretion by inhibiting apoB synthesis without increasing the degradation of the newly synthesized protein. Under our conditions, cholesterol uptake, apoB mRNA, and microsomal triglyceride protein activity were not modified by apple polyphenols. The main monomers present in our mixture did not interfere with the intestinal lipid metabolism. By contrast, apple procyanidins reproduced the inhibition of both cholesteryl ester synthesis and lipoprotein secretion. Overall, our results are compatible with a mechanism of action of polyphenols resulting in impaired lipid availability that could induce the inhibition of intestinal lipoprotein secretion and contribute to the hypolipidemic effect of these compounds in vivo.

Apolipoproteins B↗

PubMed2003

Lipid-dependent bidirectional traffic of apolipoprotein B in polarized enterocytes.

Enterocytes are highly polarized cells that transfer nutrients across the intestinal epithelium from the apical to the basolateral pole. Apolipoprotein B (apoB) is a secretory protein that plays a key role in the transepithelial transport of dietary fatty acids as triacylglycerol. The evaluation of the control of apoB traffic by lipids is therefore of particular interest. To get a dynamic insight into this process, we used the enterocytic Caco-2 cells cultured on microporous filters, a system in which the apical and basal compartments can be delimited. Combining biochemical and morphological approaches, our results showed that, besides their role in protection from degradation, lipids control the intracellular traffic of apoB in enterocytes. A supply of fatty acids and cholesterol is sufficient for the export of apoB from the endoplasmic reticulum and its post-Golgi traffic up to the apical brush-border domain, where it remains until an apical supply of complex lipid micelles signals its chase down to the basolateral secretory domain. This downward traffic of apoB involves a microtubule-dependent process. Our results demonstrate an enterocyte-specific bidirectional process for the lipid-dependent traffic of a secretory protein.

Animals↗