PubMed Health⌕ Search

Biomedical subjects

E Orsó

Publications and source records attributed to E Orsó.

At least 19 recordsLinked to original sources
PubMed2001

Characterization of the ATPase cycle of human ABCA1: implications for its function as a regulator rather than an active transporter.

ABCA1 plays a key role in cellular cholesterol and phospholipid traffic. To explore the biochemical properties of this membrane protein we applied a Baculovirus-insect cell expression system. We found that human ABCA1 in isolated membranes showed a specific, Mg(2+)-dependent ATP binding but had no measurable ATPase activity. Nevertheless, conformational changes in ABCA1 could be demonstrated by nucleotide occlusion, even without arresting the catalytic cycle by phosphate-mimicking anions. Addition of potential lipid substrates or lipid acceptors (apolipoprotein A-I) did not modify the ATPase activity or nucleotide occlusion by ABCA1. Our data indicate that ATP hydrolysis by ABCA1 occurs at a very low rate, suggesting that ABCA1 may not function as an effective active transporter as previously assumed. In the light of the observed conformational changes we propose a regulatory function for human ABCA1.

ATP Binding Cassette Transporter 1↗
PubMed2001

Inhibition of effects of endogenously synthesized histamine disturbs in vitro human dendritic cell differentiation.

Histamine, a principal mediator in various physiological and pathological cell functions is synthesized from L-histidine exclusively by histidine decarboxylase, an enzyme, which is expressed in many tissues of mammalian organism. Histamine plays a role in various cellular functions, including cell differentiation. The aim of this study was to determine the presence and to characterize the role of the endogenously produced histamine during in vitro dendritic cell (DC) differentiation induced by interleukin-4 (IL-4) and granulocyte-monocyte colony stimulating factor (GM-CSF). The changes in intracellular histamine content, biosynthesis and gene expression of histidine decarboxylase were investigated during this process. One also studied how histamine receptor antagonists and a histamine synthesis blocker influence the expression of differentiation antigens on the DC during in vitro maturation. During in vitro differentiation parallel culture incubations were performed by adding H1 receptor antagonist triprolidine, H2 receptor antagonist tiotidine, the tamoxifene derivate DPPE which blocks the intracellular binding of histamine, and an irreversible blocker of histidine decarboxylase, alpha-fluoromethyl histamine (alpha-FMH). The results show simultaneous increase in both histidine decarboxylase level and histamine content during differentiation of elutriated monocytes toward DC. Both blockade of de novo histamine production (by alpha-FMH) and inhibition of histamine binding (by H1 and H2 receptor antagonists, triprolidine and tiotidine, respectively) markedly decreased CD40 expression and that of CD45 from the 3rd day of treatment. DPPE by disturbing intracellular interaction of histamine with cytochrome P-450 moieties was able to decrease the expression of CD45, CD86, HLA-DR, CD33, CD40 and CD11c. Based on the data it is suggested that endogenous histamine is actively synthesized during cytokine-induced in vitro DC differentiation. The functional relevance and autocrine and paracrine action of endogenously produced histamine is supported by the data showing that inhibition of histamine synthesis by HDC, blocking of histamine binding by both 'extracellular' histamine receptors (by specific antagonists, triprolidine and tiotidine) and 'intracellular' antagonists (DPPE) disturb the differentiation of DC. This conclusion is supported by the fact, that by the inhibition of histamine acting in an autocrine/paracrine way, the expression pattern of differentiation markers on DC is markedly changed.

Antigens, CD↗
PubMed2001

Lipopolysaccharide and ceramide docking to CD14 provokes ligand-specific receptor clustering in rafts.

The glycosylphosphatidylinositol-anchored receptor CD14 plays a major role in the inflammatory response of monocytes to lipopolysaccharide. Here, we describe that ceramide, a constituent of atherogenic lipoproteins, binds to CD14 and induces clustering of CD14 to co-receptors in rafts. In resting cells, CD14 was associated with CD55, the Fcgamma-receptors CD32 and CD64 and the pentaspan CD47. Ceramide further recruited the complement receptor 3 (CD11b/CD18) and CD36 into proximity of CD14. Lipopolysaccharide, in addition, induced co-clustering with Toll-like receptor 4, Fcgamma-RIIIa (CD16a) and the tetraspanin CD81 while CD47 was dissociated. The different receptor complexes may be linked to ligand-specific cellular responses initiated by CD14.

Antigens, CD↗
PubMed2001

Increased histidine decarboxylase expression during in vitro monocyte maturation; a possible role of endogenously synthesised histamine in monocyte/macrophage differentiation.

OBJECTIVE: In this study the expression of histidine decarboxylase (HDC), the pivotal enzyme in histamine formation and the effect of endogenously produced histamine on differentiation antigens was examined during in vitro differentiation of human monocytes. MATERIAL AND TREATMENT: Human elutriated monocytes from healthy volunteers were incubated with macrophage colony stimulating factor (M-CSF) and the expression of HDC was followed at both mRNA and protein levels. To study the possible function of histamine we followed the expression of some cell surface markers (CD14, CD16, CD91, CD49d and CD11c) relevant for phagocytic differentiation upon incubation in the presence of different histamine inhibitors, an HDC inhibitor: S(+)-alpha-fluoromethyl-histidine HCl, (alphaFMH), a compound that disturbs the interaction of histamine with intracellular cyp450 moieties: N,N-diethyl-2-[4-(phenylmethyl) phenoxy]-ethanamine HCI, (DPPE); and H1 and H2 receptor antagonists, Triprolidine and Cimetidine. RESULTS: During in vitro culture of elutriated human monocytes, in the presence of M-CSF, the gene expression and biosynthesis of HDC was considerably increased. The various antihistamine agents decreased the expression of the cell surface markers examined in this study. CONCLUSIONS: These data support the elevation of HDC expression during human monocytic differentiation and the possibility that monocyte-derived histamine is partially involved in regulation of M-CSF induced in vitro human monocyte/macrophage phagocytic differentiation.

Antigens, CD↗
PubMed2001

Apolipoprotein AI and HDL(3) inhibit spreading of primary human monocytes through a mechanism that involves cholesterol depletion and regulation of CDC42.

The objective of the current study was to characterize the influence of high density lipoproteins (HDL) on processes related to the vascular recruitment of human monocytes, which may contribute to the anti-atherogenic properties of these lipoproteins. We show that HDL(3) and apo AI inhibit the following processes in primary human monocytes: (1) M-CSF induced cell spreading; (2) M-CSF stimulated expression of surface molecules involved in adhesion, migration, and scavenging; (3) fMLP induced chemotaxis. These processes are obviously modulated by the regulation of cellular cholesterol pools as indicated by the following findings. In Tangier monocytes with defective apo AI induced cholesterol efflux, apo AI had no influence on the spreading response. In control cells, stimulation of cholesterol efflux by p-cyclodextrin mimicked the effect of apo AI and HDL(3) on spreading and chemotaxis, whereas cholesterol loading with enzymatically modified LDL (E-LDL) showed the opposite effect. Finally, a similar inverse regulation by E-LDL and apo AI/HDL(3) was also observed in regard to the surface expression of beta(1)- and beta(2)-integrins as well as the hemoglobin/haptoglobin scavenger receptor CD163 and the Fcgamma-IIIaR CD16. CDC42 was identified as a potential downstream target linking changes in cellular cholesterol content to monocyte spreading and chemotaxis. Thus, CDC42 antisense markedly reduced spreading and, in parallel with their influence on monocyte spreading, HDL(3), apo AI and p-cyclodextrin down-regulated CDC42 expression while E-LDL had the inverse effect. The apo AI induced decrease of CDC42 protein expression was paralleled by the reduction of active GTP-bound CDC42. In summary, we provide evidence that HDL(3) and apo AI are able to inhibit processes in primary human monocytes, which are related to the recruitment of monocytes into the vessel wall and probably involve regulation of cellular cholesterol pools and CDC42 function.

Apolipoprotein A-I↗
PubMed2001

Intracellular cholesterol and phospholipid trafficking: comparable mechanisms in macrophages and neuronal cells.

During the past ten years considerable evidences have accumulated that in addition to monocytes/macrophages, that are implicated in innate immunity and atherogenesis, neuronal cells also exhibit an extensive cellular metabolism. The present study focuses on the major protein players that establish cellular distribution of cholesterol and phospholipids. Evidences are provided that neuronal cells and monocytes/macrophages are equipped with comparable intracellular lipid trafficking mechanisms. Selected examples are presented that trafficking dysfunctions lead to disease development, such as Tangier disease and Niemann-Pick disease type C, or contribute to the pathogenesis of diseases such as Alzheimer disease and atherosclerosis.

Animals↗
PubMed2000

Identification of a novel human sterol-sensitive ATP-binding cassette transporter (ABCA7).

We report the identification of the full-length cDNA for a novel ATP-binding cassette (ABC) transporter from human macrophages. The mRNA is of 6.8 kb size and contains an open reading frame encoding a polypeptide of 2146 amino acids with a calculated molecular weight of 220 kDa. The predicted protein product is composed of two transmembrane domains and two nucleotide binding folds indicating that it pertains to the group of full-size ABC transporters. The novel transporter shows highest protein sequence homology with the recently cloned human cholesterol and phospholipid exporter ABCA1 (54%) and the human retinal transporter ABCR (49%), both members of the ABC transporter subfamily A. In accordance with the currently proposed classification, the novel transporter was designated ABCA7. ABCA7 mRNA was detected predominantly in myelo-lymphatic tissues with highest expression in peripheral leukocytes, thymus, spleen, and bone marrow. Expression of ABCA7 is induced during in vitro differentiation of human monocytes into macrophages. In macrophages, both the ABCA7 mRNA and protein expression are upregulated in the presence of modified low density lipoprotein and downregulated by HDL(3). Our results suggest a role for ABCA7 in macrophage transmembrane lipid transport.

ATP-Binding Cassette Transporters↗
PubMed2000

ABCG1 (ABC8), the human homolog of the Drosophila white gene, is a regulator of macrophage cholesterol and phospholipid transport.

Excessive uptake of atherogenic lipoproteins such as modified low-density lipoprotein complexes by vascular macrophages leads to foam cell formation, a critical step in atherogenesis. Cholesterol efflux mediated by high-density lipoproteins (HDL) constitutes a protective mechanism against macrophage lipid overloading. The molecular mechanisms underlying this reverse cholesterol transport process are currently not fully understood. To identify effector proteins that are involved in macrophage lipid uptake and release, we searched for genes that are regulated during lipid influx and efflux in human macrophages using a differential display approach. We report here that the ATP-binding cassette (ABC) transporter ABCG1 (ABC8) is induced in monocyte-derived macrophages during cholesterol influx mediated by acetylated low-density lipoprotein. Conversely, lipid efflux in cholesterol-laden macrophages, mediated by the cholesterol acceptor HDL(3), suppresses the expression of ABCG1. Immunocytochemical and flow cytometric analyses revealed that ABCG1 is expressed on the cell surface and in intracellular compartments of cholesterol-laden macrophages. Inhibition of ABCG1 protein expression using an antisense strategy resulted in reduced HDL(3)-dependent efflux of cholesterol and choline-phospholipids. In a comprehensive analysis of the expression and regulation of all currently known human ABC transporters, we identified an additional set of ABC genes whose expression is regulated by cholesterol uptake or HDL(3)-mediated lipid release, suggesting a potential function for these transporters in macrophage lipid homeostasis. Our results demonstrating a regulator function for ABCG1 in cholesterol and phospholipid transport define a biologic activity for ABC transporters in macrophages.

ATP Binding Cassette Transporter, Subfamily G, Mem↗
PubMed2000

Multi-color analysis of monocyte and dendritic cell precursor heterogeneity in whole blood.

The coexpression analysis of the 55-kDa lipopolysaccharide receptor (CD14) and the Fc gamma-receptor III (CD16) reveals a broad heterogeneity of blood monocytes which in our previous work could be divided into five subpopulations based on correlated differences in expression of the pan-myeloid antigen CD33 and the adhesion antigens CD11a, CD11b and CD56. An even larger complexity of myeloid cells with antigen presenting capacity in peripheral blood is suggested by the description of small populations of immature and mature precursors of dendritic cells which rapidly develop potent costimulatory activity and a dendritic morphology in in vitro culture. The identity of the subsets of cells which have been described based on heterogeneous analytical approaches, however, remains unclear. The goal of this study, therefore, was the correlated analysis of monocyte subpopulations and dendritic cell precursors in a quantitative whole blood assay. This was achieved based on simultaneous expression analysis of the monocyte markers CD14 and CD16 with antigens such as CD33, HLA-DR, the integrin CD11c, and the interleukin-3 receptor alpha-chain (CD123) which in absence of lineage-related antigens have been used for description of dendritic cell precursors. The selected marker panel revealed identity of cells previously described as CD33bright CD14dim dendritic cell precursors with CD11c+lin-HLA-DR+ cells. Dendritic cell precursors considered to be less mature which have been alternatively described as CD33+ CD14dim CD16- cells or CD123hi dendritic cell precursors, however, were shown to differ in phenotype from each other with regard to expression density of CD33 and expression of CD14. In summary, our study revealed a complex heterogeneity of monocytes and dendritic cell precursors in peripheral blood and indicates that a direct comparison of the analytical approaches of different authors is needed to further clarify the ontogeny of human monocytes and dendritic cells.

Antigens, CD↗
PubMed2000

Transport of lipids from golgi to plasma membrane is defective in tangier disease patients and Abc1-deficient mice.

Mutations in the gene encoding ATP-binding cassette transporter 1 ( ABC1) have been reported in Tangier disease (TD), an autosomal recessive disorder that is characterized by almost complete absence of plasma high-density lipoprotein (HDL), deposition of cholesteryl esters in the reticulo-endothelial system (RES) and aberrant cellular lipid trafficking. We demonstrate here that mice with a targeted inactivation of Abc1 display morphologic abnormalities and perturbations in their lipoprotein metabolism concordant with TD. ABC1 is expressed on the plasma membrane and the Golgi complex, mediates apo-AI associated export of cholesterol and phospholipids from the cell, and is regulated by cholesterol flux. Structural and functional abnormalities in caveolar processing and the trans-Golgi secretory pathway of cells lacking functional ABC1 indicate that lipid export processes involving vesicular budding between the Golgi and the plasma membrane are severely disturbed.

ATP Binding Cassette Transporter 1↗
PubMed2000

ABC transporters in cellular lipid trafficking.

ATP-binding cassette (ABC) transporters constitute a group of evolutionary highly conserved cellular transmembrane transport proteins. Recent work has implicated ABC transporters in cellular transmembrane lipid transport and hereditary diseases have been causatively linked to defective ABC transporters translocating lipid compounds. The emerging concept that a defined subset of ABC transporters is intimately involved in cellular lipid trafficking has recently been substantiated convincingly by the finding that ABCA1 plays a central role in the regulation of HDL metabolism and macrophage targeting to the RES or the vascular wall. Differentiation dependent expression of a large number of ABC transporters in monocytes/macrophages and their regulation by sterol flux render these transporter molecules potentially critical players in atherogenesis and other chronic inflammatory diseases.

ATP Binding Cassette Transporter 1↗
PubMed2000

Regulation of scavenger receptor CD163 expression in human monocytes and macrophages by pro- and antiinflammatory stimuli.

CD163, also referred to as M130, a member of the scavenger receptor cysteine-rich family (SRCR) is exclusively expressed on cells of the monocyte lineage. In freshly isolated monocytes the CD14bright CD16+ monocyte subset revealed the highest expression of CD163 among all monocyte subsets. CD163 mRNA and protein expression is up-regulated during macrophage colony-stimulating factor (M-CSF)-dependent phagocytic differentiation of human blood monocytes. In contrast, monocytic cells treated with GM-CSF and interleukin-4 (IL-4) for dendritic differentiation down-regulate this antigen. CD163 expression is also suppressed by proinflammatory mediators like lipopolysaccharide (LPS), interferon-gamma (IFN-gamma), and tumor necrosis factor alpha, whereas IL-6 and the antiinflammatory cytokine interleukin-10 (IL-10) strongly up-regulate CD163 mRNA in monocytes and macrophages. The effects of the immunosuppressants dexamethasone, cyclosporin A (CA), and cortisol differ in their capacity to influence CD163 mRNA levels. Our results demonstrate that CD163 expression in monocytes/macrophages is regulated by proinflammatory and antiinflammatory mediators. This expression pattern implies a functional role of CD 163 in the antiinflammatory response of monocytes.

Antigens, CD↗
PubMed1999

The human apM-1, an adipocyte-specific gene linked to the family of TNF's and to genes expressed in activated T cells, is mapped to chromosome 1q21.3-q23, a susceptibility locus identified for familial combined hyperlipidaemia (FCH).

The human adipocyte-specific apM-1 gene encodes a secretory protein of the adipose tissue that has been suggested to play a role in the pathogenesis of obesity. The regulation of apM-1 was studied along adipocyte differentiation. While apM-1-mRNA and apM-1 protein were absent in preadipocytes and in 48 h differentiated adipocytes, they were found upregulated from day 4 to day 9 of adipocyte differentiation as shown by RNase protection assay and Western blot analysis. These data indicate that apM-1 may be a late marker of adipocyte differentiation. In human sera apM-1 protein is also detectable by Western blots using a polyclonal antibody raised against a synthetic peptide sequence of the human apM-1. The genomic structure of the human apM-1 gene together with a total of 2.7 kb of the 5'-flanking region with putative transcription factor binding sites is presented. Interestingly, sequence comparisons link the apM-1 gene to the family of TNF's and to genes expressed in activated T-cells. The chromosomal localization of apM-1 was investigated by FISH and mapped to human chromosome 1q21.3-1q23, a region that was identified as a susceptibility locus for Familial Combined Hyperlipidaemia (FCH) and polygenic NIDDM. These data and the chromosomal localization on chromosome 1q21.3-q23 raises the possibility that apM-1 as an adipocyte-specific secretory protein may play a role in the pathogenesis of FCH and associated insulin resistance. Exon- and intron-specific primer sequences are presented as a basis for mutation screening of patients affected with FCH.

3T3 Cells↗