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Ronald Barbaras

Publications and source records attributed to Ronald Barbaras.

11 recordsLinked to original sources
PubMed2006

The low-density lipoprotein receptor plays a role in the infection of primary human hepatocytes by hepatitis C virus.

BACKGROUND/AIMS: The direct implication of low-density lipoprotein receptor (LDLR) in hepatitis C virus (HCV) infection of human hepatocyte has not been demonstrated. Normal primary human hepatocytes infected by serum HCV were used to document this point. METHODS: Expression and activity of LDLR were assessed by RT-PCR and LDL entry, in the absence or presence of squalestatin or 25-hydroxycholesterol that up- or down-regulates LDLR expression, respectively. Infection was performed in the absence or presence of LDL, HDL, recombinant soluble LDLR peptides encompassing full-length (r-shLDLR4-292) or truncated (r-shLDLR4-166) LDL-binding domain, monoclonal antibodies against r-shLDLR4-292, squalestatin or 25-hydroxycholesterol. Intracellular amounts of replicative and genomic HCV RNA strands used as end point of infection were assessed by RT-PCR. RESULTS: r-shLDLR4-292, antibodies against r-shLDLR4-292 and LDL inhibited viral RNA accumulation, irrespective of genotype, viral load or liver donor. Inhibition was greatest when r-shLDLR4-292 was present at the time of inoculation and gradually decreased as the delay between inoculation and r-shLDLR4-292 treatment increased. In hepatocytes pre-treated with squalestatin or 25-hydroxycholesterol before infection, viral RNA accumulation increased or decreased in parallel with LDLR mRNA expression and LDL entry. CONCLUSIONS: LDLR is involved at an early stage in infection of normal human hepatocytes by serum-derived HCV virions.

Adolescent↗
PubMed2006

Accelerated lipid absorption in mice overexpressing intestinal SR-BI.

Dietary cholesterol absorption contributes to a large part of the circulating cholesterol. However, the mechanism of sterol intestinal uptake is not clearly elucidated. Scavenger receptor class B type I (SR-BI), major component in the control of cholesterol homeostasis, is expressed in the intestine, but its role in this organ remains unclear. We have generated transgenic mice overexpressing SR-BI primarily in the intestine by using the mouse SR-BI gene under the control of intestinal specific "apoC-III enhancer coupled with apoA-IV promoter." We found SR-BI overexpression with respect to the natural protein along the intestine and at the top of the villosities. After a meal containing [(14)C]cholesterol and [(3)H]triolein, SR-BI transgenic mice presented a rise in intestinal absorption of both lipids that was not due to a defect in chylomicron clearance nor to a change in the bile flow or the bile acid content. Nevertheless, SR-BI transgenic mice showed a decrease of total cholesterol but an increase of triglyceride content in plasma without any change in the high density lipoprotein apoA-I level. Thus, we described for the first time a functional role in vivo for SR-BI in cholesterol but also in triglyceride intestinal absorption.

Absorption↗
PubMed2006

Ecto-F1Fo ATP synthase/F1 ATPase: metabolic and immunological functions.

PURPOSE OF REVIEW: Until recently, F1Fo ATP synthase expression was believed to be strictly confined to mitochondria where it generates most cellular ATP. This paper reviews the recent evidence for an extra-mitochondrial expression of its components by immunofluorescence, biochemistry and proteomics studies. It discusses its possible implications in an ecto-nucleotide metabolism and its pathophysiological role in normal and tumoral cells. RECENT FINDINGS: F1Fo ATP synthase components have been identified as cell-surface receptors for apparently unrelated ligands in the course of studies carried out on angiogenesis, lipoprotein metabolism, innate immunity, hypertension, or regulation of food intake. SUMMARY: F1Fo ATP synthase is expressed on endothelial cells where it binds angiostatin, regulates surface ATP levels, and modulates endothelial cell proliferation and differentiation. Through binding of apolipoprotein A-I, a similar complex, expressed on hepatocytes, regulates lipoprotein internalization. On tumors, it is recognized in association with apolipoprotein A-I by the antigen receptor of circulating cytotoxic lymphocytes of the gammadelta subtype and thus promotes an innate tumor cell recognition and lysis. It binds enterostatin on brain cells. Biochemistry and proteomics studies indicate an enrichment of F1Fo components in lipid rafts selectively with some other mitochondrial proteins, suggesting intracellular traffic connections between mitochondria and other membrane compartments. Finally, depending on cell type and environment, it can generate ATP or ADP which may transfer a downstream signal to purinergic receptors.

Animals↗
PubMed2005

High-density lipoproteins prevent the oxidized low-density lipoprotein-induced epidermal [corrected] growth factor receptor activation and subsequent matrix metalloproteinase-2 upregulation.

OBJECTIVE: The atherogenic oxidized low-density lipoprotein (oxLDL) induces the formation of carbonyl-protein adducts and activates the epidermal [corrected] growth factor receptor (EGFR) signaling pathway, which is now regarded as a central element for signal transduction. We aimed to investigate whether and by which mechanism the anti-atherogenic high-density lipoprotein (HDL) prevents these effects of oxLDL. METHODS AND RESULTS: In vascular cultured cells, HDL and apolipoprotein A-I inhibit oxLDL-induced EGFR activation and subsequent signaling by acting through 2 separate mechanisms. First, HDL, like the aldehyde scavenger dinitrophenyl hydrazine, prevented the formation of oxLDL-induced carbonyl-protein adducts and 4-hydroxynonenal (HNE)-EGFR adducts. Secondly, HDL enhanced the cellular antioxidant defenses by preventing (through a scavenger receptor class B-1 (SR-BI)-dependent mechanism) the increase of intracellular reactive oxygen species (ROS) and subsequent EGFR activation triggered by oxLDL or H2O2. A pharmacological approach suggests that this protective effect of HDL is independent of cellular glutathione level and glutathione peroxidase activity, but it requires catalase activity. Finally, we report that oxLDL upregulates both membrane type 1 (MT1)-matrix metalloproteinase-1 (MT1-MMP) and MMP-2 through an EGFR-dependent mechanism and that HDL inhibits these events. CONCLUSIONS: HDLs block in vitro oxLDL-induced EGFR signaling and subsequent MMP-2 activation by inhibiting carbonyl adducts formation and cellular oxidative stress. These effects of HDL may participate to reduce cell activation, excessive remodeling, and alteration of the vascular wall.

Aldehydes↗
PubMed2005

Tumor recognition following Vgamma9Vdelta2 T cell receptor interactions with a surface F1-ATPase-related structure and apolipoprotein A-I.

Vgamma9Vdelta2 T lymphocytes, a major gammadelta T lymphocyte subset in humans, display cytolytic activity against various tumor cells upon recognition of yet uncharacterized structures. Here, we show that an entity related to the mitochondrial F1-ATPase is expressed on tumor cell surface and promotes tumor recognition by Vgamma9Vdelta2 T cells. When immobilized, purified F1-ATPase induces selective activation of this lymphocyte subset. The Vgamma9Vdelta2 T cell receptors (TCR) and the F1-ATPase also bind a delipidated form of apolipoprotein A-I (apo A-I), as demonstrated by surface plasmon resonance. Moreover, the presence of apo A-I in the culture medium is required for optimal activation of Vgamma9Vdelta2 T cells by tumors expressing F1-ATPase. This study thus describes an unanticipated tumor recognition mechanism by Vgamma9Vdelta2 lymphocytes and a possible link between gammadelta T cell immunity and lipid metabolism.

Apolipoprotein A-I↗
PubMed2005

Role of apolipoproteins in gammadelta and NKT cell-mediated innate immunity.

Recent findings reveal unanticipated connections between the fields of lipid metabolism and immunology. They concern gammadelta and NKT cells, nonconventional T cell populations that do not recognize protein antigens and are involved in immunity against cancer, defense against infections, or in regulation of classical immune responses. In this review, we summarize data linking perturbations of apolipoprotein levels and nonconventional T cells with inflammatory processes such as autoimmune diseases or atherosclerosis. We integrate and discuss recent findings on the implication of apolipoproteins in antigen recognition by gammadelta and NKT cells, with emphasis on apolipoproteins A-I and E. These findings also provide indications that apolipoproteins influence antitumor immunosurveillance.

Animals↗
PubMed2003

Enterophilin-1, a new partner of sorting nexin 1, decreases cell surface epidermal growth factor receptor.

We previously described enterophilin-1 (Ent-1), a new intestinal protein bearing an extended leucine zipper and a B30.2 domain. Ent-1 expression is associated with growth arrest and enterocyte differentiation. To investigate the importance of Ent-1 in the differentiation, we performed a yeast two-hybrid screening. We identified sorting nexin 1 (SNX1) as a novel partner of Ent-1 and confirmed the specificity of interaction by co-immunoprecipitation experiments in mammalian cells. SNX1 is associated with endosomal membranes and triggers the endosome-to-lysosome pathway of epidermal growth factor receptor (EGFR). We observe by immunofluorescence microscopy that Ent-1 and SNX1 are co-localized on vesicular and tubulovesicular structures, which are different from early endosome antigen 1-containing endosomes. By gel filtration chromatography, we show that Ent-1 and SNX1 co-eluted in macromolecular complexes containing part of EGFR. Furthermore, overexpressed Ent-1 decreases cell surface EGFR. Ent-1 and SNX1 co-overexpression strongly extends EGFR diminution, indicating a synergetic effect of both proteins on cell surface EGFR removal. Interestingly, the increase of endogenous Ent-1 expression correlates with the decrease of EGFR during spontaneous differentiation of Caco-2 cells. We thus propose a role of Ent-1 in the trafficking of EGFR to down-regulate intestinal mitogenic signals, highlighting the mechanisms of cell growth arrest associated with enterocytic differentiation.

Animals↗
PubMed2003

Ectopic beta-chain of ATP synthase is an apolipoprotein A-I receptor in hepatic HDL endocytosis.

The effect of high-density lipoprotein (HDL) in protecting against atherosclerosis is usually attributed to its role in 'reverse cholesterol transport'. In this process, HDL particles mediate the efflux and the transport of cholesterol from peripheral cells to the liver for further metabolism and bile excretion. Thus, cell-surface receptors for HDL on hepatocytes are chief partners in the regulation of cholesterol homeostasis. A high-affinity HDL receptor for apolipoprotein A-I (apoA-I) was previously identified on the surface of hepatocytes. Here we show that this receptor is identical to the beta-chain of ATP synthase, a principal protein complex of the mitochondrial inner membrane. Different experimental approaches confirm this ectopic localization of components of the ATP synthase complex and the presence of ATP hydrolase activity at the hepatocyte cell surface. Receptor stimulation by apoA-I triggers the endocytosis of holo-HDL particles (protein plus lipid) by a mechanism that depends strictly on the generation of ADP. We confirm this effect on endocytosis in perfused rat liver ex vivo by using a specific inhibitor of ATP synthase. Thus, membrane-bound ATP synthase has a previously unsuspected role in modulating the concentrations of extracellular ADP and is regulated by a principal plasma apolipoprotein.

Adenosine Diphosphate↗
PubMed2002

Coupled assay of sphingomyelin and ceramide molecular species by gas liquid chromatography.

This study reports a single-step analysis of the molecular species of endogenous ceramides and of the ceramide moiety of sphingomyelins in biological samples, using gas liquid chromatography (GLC). Silylated sphingomyelins were quantitatively converted to monosilylated ceramide upon injection into GLC, whereas the free ceramides were di-silylated on the primary and secondary alcohol function, as confirmed by mass spectrometry. The reproducible shift of the retention times between the mono- and di-silylated derivatives enables simultaneous quantification of the variety of sphingomyelin and ceramide molecular species. Overlapping diacylglycerols were first removed by a mild alkaline treatment of the lipid extract. The lowest detection limit (5 pmol) did not allow for identification of free ceramides in human plasma, but 17 molecular species of ceramides derived from sphingomyelins were quantified, from NC16:0 up to NC24:1. By contrast, three major free ceramides (NC16:0, NC24:0, and NC24:1) were quantified in HEPG2 and Chinese hamster ovary (CHO) cells. Upon induction of apoptosis in CHO cells by C6-ceramide, we could follow the disappearance of the C6-ceramide, its partial conversion to C6-sphingomyelin, and the prominent increase of NC16:0 ceramide. Thus, our method represents a unique procedure of simultaneous analysis of sphingomyelin and ceramide molecular species able to monitor the variation of the different pools in biological samples.

Acids↗
PubMed2002

Hepatic lipase: structure/function relationship, synthesis, and regulation.

Hepatic lipase (HL) is a lipolytic enzyme, synthesized by hepatocytes and found localized at the surface of liver sinusoid capillaries. In humans, the enzyme is mostly bound onto heparan-sulfate proteoglycans at the surface of hepatocytes and also of sinusoid endothelial cells. HL shares a number of functional domains with lipoprotein lipase and with other members of the lipase gene family. It is a secreted glycoprotein, and remodelling of the N-linked oligosaccharides appears to be crucial for the secretion process, rather than for the acquisition of the catalytic activity. HL is also present in adrenals and ovaries, where it might promote delivery of lipoprotein cholesterol for steroidogenesis. However, evidence of a local synthesis is still controversial. HL activity is fairly regulated according to the cell cholesterol content and to the hormonal status. Coordinate regulations have been reported for both HL and the scavenger-receptor B-I, suggesting complementary roles in cholesterol metabolism. However, genetic variants largely contribute to HL variability and their possible impact in the development of a dyslipidemic phenotype, or in a context of insulin-resistance, is discussed.

Amino Acid Sequence↗