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Biomedical subjects

D R Greaves

Publications and source records attributed to D R Greaves.

At least 19 recordsLinked to original sources
PubMed2000

Adenovirus-mediated gene transfer of a secreted form of human macrophage scavenger receptor inhibits modified low-density lipoprotein degradation and foam-cell formation in macrophages.

BACKGROUND: Macrophage scavenger receptors (MSRs) play an important role in the pathogenesis of atherosclerosis. Therefore, local modulation of MSR activity could have a beneficial effect on atherogenesis. METHODS AND RESULTS: We cloned a secreted "decoy" MSR (sMSR) that contains an extracellular portion of the human MSR type AI and constructed an adenoviral vector that directs high-level expression of sMSR in macrophages under the control of the human CD68 promoter. Expression of the sMSR protein inhibited the degradation of (125)I-labeled acetylated LDL and oxidized LDL by murine macrophages up to 90%. sMSRs also reduced acetylated LDL degradation in MSR knockout mouse peritoneal macrophages by 60% to 80%, which suggests that the decoy construct can compete for the uptake mediated via other related scavenger receptors. In addition, sMSRs inhibited foam-cell formation in murine macrophages in the presence of cytochalasin D. The mechanism of inhibition is through ligand binding to the sMSRs, which prevents the ligand binding to MSRs on cell membranes. CONCLUSIONS: The demonstration that recombinant adenovirus-mediated gene transfer of decoy sMSRs can block foam-cell formation suggests a possible new strategy for gene therapy of atherosclerosis and for the treatment of lipid accumulation after arterial manipulations.

Adenoviridae↗
PubMed2000

Chemokines and myeloid cell recruitment.

Chemokine-chemokine receptor interactions mediate constitutive leukocyte trafficking and leukocyte recruitment to sites of infection and inflammation. We suggest that a multiplicity of leukocyte chemoattractants exists to increase the selectivity of leukocyte recruitment in a range of physiological and pathological settings.

Acute Disease↗
PubMed1999

The human eukaryotic initiation factor 4AI gene (EIF4A1) contains multiple regulatory elements that direct high-level reporter gene expression in mammalian cell lines.

The gene encoding human eukaryotic initiation factor 4A (EIF4A1) is located on chromosome 17p13, 667 bp upstream from the gene encoding the macrophage endosomal protein CD68. The EIF4AI gene contains 10 intervening sequences with the 1397-bp first intron containing a CpG-rich methylation-free island. Sequences capable of enhancing gene expression reside between positions -69 and -371 and positions -504 and -1100 of the EIF4AI 5' flanking sequence and within introns 1, 2, 3, 7, and 9. In macrophage cell lines, EIF4A1 expression vectors give sustained high-level reporter gene expression to levels 10 times higher than that obtained using the human cytomegalovirus immediate-early gene promoter/enhancer. Sequences of the human EIF4AI gene may find application in the development of new vectors for gene therapy and genetic vaccination.

5' Untranslated Regions↗
PubMed1999

Analysis of macrophage scavenger receptor (SR-A) expression in human aortic atherosclerotic lesions.

The class A scavenger receptors (SR-As) are trimeric, integral membrane glycoproteins that exhibit unusually broad ligand-binding properties. A number of studies have suggested that these receptors may play an important role in host defense and in many macrophage-associated pathological processes, including atherosclerosis and Alzheimer's disease. The study of the expression and function of these receptors in human disease has been hampered by the lack of suitable antibodies recognizing human SR-A. This has generated questions regarding the nature of receptors responsible for scavenger receptor activity detected in a variety of cell types, including monocytes, macrophages, smooth muscle cells, and endothelial cells. To address these questions, we have produced high-titer antisera recognizing human SR-A by using mice deficient for SR-A (SR-A -/-). We show that SR-A -/- mice produce a significantly higher-titer immune response than do wild-type (SR-A +/+) littermates, with antisera of the former having a broad species reactivity and recognizing SR-A from humans, mice, and rabbits. The antisera recognize both type I and II SR-A in a wide range of immunological techniques. Using these antisera we show that the expression of SR-A protein is induced during monocyte to macrophage differentiation and that SR-A mediates 80% of the uptake of acetylated low density lipoprotein by human monocyte-derived macrophages. We also establish that human SR-A is expressed by tissue macrophages in liver and lung and by macrophage-derived foam cells within aortic atherosclerotic lesions, with little detectable expression by smooth muscle cells or aortic endothelium.

Actins↗
PubMed1998

Functional comparison of the murine macrosialin and human CD68 promoters in macrophage and nonmacrophage cell lines.

The glycoprotein macrosialin is expressed specifically in murine monocytes and macrophages. In the murine genome the macrosialin gene lies 877 bp 3' of the ubiquitously expressed elongation initiation factor 4A1 (eIF-4AI) gene on chromosome 11. The macrosialin gene promoter directs high-level expression in transiently transfected murine macrophage cell lines, but significant levels of expression are seen in nonmacrophage cell lines. The first intron of the gene encoding the human homologue of macrosialin, CD68, directs macrophage-specific expression when cloned downstream of the human CD68 gene promoter. Despite sharing extensive sequence homology with the human CD68 gene first intron, the murine macrosialin gene first intron does not act as a macrophage-specific enhancer when cloned downstream of the human CD68 or murine macrosialin promoters. We conclude that the genetic elements that are responsible for macrophage-specific gene expression are organized differently in the murine and human CD68 genes.

Animals↗
PubMed1998

Immunohistochemical evidence for a macrophage scavenger receptor in Mato cells and reactive microglia of ischemia and Alzheimer's disease.

Macrophage scavenger receptors (MSR) are implicated in the development of atherosclerosis and amyloid b-protein deposition in Alzheimer's disease. However, histopathological studies of MSR expression in human tissues have been hampered by a lack of specific antibodies. Using MSR-deficient mice, we successfully raised a novel monoclonal antibody against human MSR together with high-titer antisera. These antibodies specifically recognized human tissue macrophages and human MSR protein purified from differentiated THP1 cells. In normal brain, MSR staining was mainly distributed to the perivascular cells, which correspond to Mato's fluorescent granular perithelial cells (Mato cells). In the lesions of ischemia and Alzheimer's disease, a subset of microglia stained positive for MSR. These novel antibodies are useful tools for analysis of MSR expression in human tissues.

Alzheimer Disease↗
PubMed1998

Recent progress in defining the role of scavenger receptors in lipid transport, atherosclerosis and host defence.

Scavenger receptors bind and internalize modified lipoproteins. There are several different classes of scavenger receptors in mammalian cells and their relative contribution to lipid transport in normal physiology and pathological conditions such as atherosclerosis has been the subject of intense investigation. Mice with a disruption in the macrophage scavenger receptor SR-A gene exhibit a reduced size of atherosclerotic lesions and also exhibit an enhanced susceptibility to pathogens and endotoxic shock. In addition to their role in lipid transport, scavenger receptors play important roles in host defence and in the regulation of acquired immunity. Recent progress in delineating the mechanisms by which oxidized LDL effects changes in gene expression will be reviewed.

Animals↗
PubMed1998

A naturally occurring isoform of the human macrophage scavenger receptor (SR-A) gene generated by alternative splicing blocks modified LDL uptake.

The class A macrophage scavenger receptors (SR-A) are macrophage-specific trimeric integral membrane glycoproteins that have been implicated in many macrophage-associated physiological and pathological processes including atherosclerosis, Alzheimer's disease, and host defense. There are two forms of the receptor that have been previously cloned, and both are generated by alternative splicing of a single gene. Here we report the cloning of a third, alternatively spliced isoform of the human SR-A gene (type III hSR-A). The novel isoform is expressed in the human monocytic leukemia cell line THP-1 and also in primary human monocyte derived macrophages. When expressed in CHO-K1 cells, type III hSR-A does not internalize AcLDL despite having the domain shown to mediate this function in type I and II hSR-A. We show that type III protein has altered intracellular processing and is trapped within the endoplasmic reticulum, making it unable to perform endocytosis. Type III protein acts as a dominant negative isoform by reducing modified LDL uptake in CHO cells stably expressing either type I or type II SR-A. The demonstration that a naturally occurring splice variant of SR-A mRNA can act as a dominant negative isoform suggests a novel mechanism for regulation of scavenger receptor activity in macrophages.

Alternative Splicing↗
PubMed1997

CCR6, a CC chemokine receptor that interacts with macrophage inflammatory protein 3alpha and is highly expressed in human dendritic cells.

Dendritic cells initiate immune responses by ferrying antigen from the tissues to the lymphoid organs for presentation to lymphocytes. Little is known about the molecular mechanisms underlying this migratory behavior. We have identified a chemokine receptor which appears to be selectively expressed in human dendritic cells derived from CD34+ cord blood precursors, but not in dendritic cells derived from peripheral blood monocytes. When stably expressed as a recombinant protein in a variety of host cell backgrounds, the receptor shows a strong interaction with only one chemokine among 25 tested: the recently reported CC chemokine macrophage inflammatory protein 3alpha. Thus, we have designated this receptor as the CC chemokine receptor 6. The cloning and characterization of a dendritic cell CC chemokine receptor suggests a role for chemokines in the control of the migration of dendritic cells and the regulation of dendritic cell function in immunity and infection.

Amino Acid Sequence↗