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D Steinberg

Publications and source records attributed to D Steinberg.

At least 19 recordsLinked to original sources
PubMed1995

The 94- to 97-kDa mouse macrophage membrane protein that recognizes oxidized low density lipoprotein and phosphatidylserine-rich liposomes is identical to macrosialin, the mouse homologue of human CD68.

We have previously reported the partial purification of a 94- to 97-kDa plasma membrane protein from mouse peritoneal macrophages that binds oxidatively modified low density lipoprotein (OxLDL) and phosphatidylserine-rich liposomes. We have now identified that protein as macrosialin, a previously cloned macrophage-restricted membrane protein in the lysosomal-associated membrane protein family (mouse homologue of human CD68). Early in the course of purification of the 94- to 97-kDa protein, a new OxLDL-binding band at 190-200 kDa appeared and copurified with the 94- to 97-kDa protein. The HPLC pattern of tryptic peptides from this higher molecular mass ligand-binding band closely matched that derived from the 94- to 97-kDa band. Specifically, the same three macrosialin-derived tryptic peptides (9, 9, and 15 residues) were present in the purified 94- to 97-kDa band and in the 190- to 200-kDa band and antisera raised against peptide sequences in macrosialin recognized both bands. An antiserum against macrosialin precipitated most of the 94- to 97-kDa OxLDL-binding material. We conclude that the binding of OxLDL to mouse macrophage membranes is in part attributable to macrosialin. Our previous studies show that OxLDL competes with oxidized red blood cells and with apoptotic thymocytes for binding to mouse peritoneal macrophages. Whether macrosialin plays a role in recognition of OxLDL and oxidatively damaged cells by intact macrophages remains uncertain.

Amino Acid Sequence↗
PubMed1995

Clinical trials of antioxidants in atherosclerosis: are we doing the right thing?

The hypothesis that oxidative modification of low density lipoprotein contributes to the progression of atherosclerosis is supported by an impressive body of in-vitro findings and by persuasive results in animal models of atherosclerosis. The hypothesis was originally proposed specifically to account for foam cell formation but oxidation of LDL has now been shown to confer on it a long list of new biological properties any one of which could in principle enhance its atherogenicity. The relative importance of these altered biological properties in vivo remains uncertain. Whatever the precise mechanisms, we know that antioxidants can slow the atherogenic process in several experimental models, including LDL-receptor-deficient rabbits, cholesterol-fed rabbits, and cholesterol-fed non-human primates. Of 18 published studies, 13 have given strongly positive results, the rate of progression of lesions being reduced by 50-80%; 2 have yielded marginally positive results; and 3 have been negative (see ref 3 for references). Furthermore, the fact that four different antioxidant compounds have been used--probucol, butylated hydroxytoluene, N,N'-diphenyl-phenylenediamine, and vitamin E--supports the conclusion that they are working via the property they all share, namely, their antioxidant potential.

Animals↗
PubMed1995

Enhanced levels of lipoperoxides in low density lipoprotein incubated with murine fibroblast expressing high levels of human 15-lipoxygenase.

There is strong experimental evidence that oxidized low density lipoprotein (Ox-LDL) plays an important role in atherosclerosis. However, the mechanisms by which Ox-LDL is formed in vivo are unknown. To test whether 15-lipoxygenase (15-LO) could play a role in oxidation of LDL by cells, we expressed 15-LO activity in murine fibroblasts, which do not normally have 15-LO activity, and tested their ability to modify LDL. Using a retroviral vector, we prepared fibroblasts that expressed 2- to 20-fold more 15-LO activity than control fibroblasts infected with a vector containing beta-galactosidase (lacZ). Compared with LDL incubated with lacZ cells, LDL incubated with 15-LO-containing cells were enriched with lipid hydroperoxides. When these LDL samples were subsequently subjected to oxidative stress, they were more susceptible to further oxidative modification, as judged by increased conjugated diene formation and by increased ability to compete with 125I-Ox-LDL for uptake by macrophages. These findings establish that cellular 15-LO can contribute to oxidative modification of LDL, but the quantitative significance of these findings to the in vivo oxidation of LDL remains to be established.

Animals↗
PubMed1995

A macrophage receptor for oxidized low density lipoprotein distinct from the receptor for acetyl low density lipoprotein: partial purification and role in recognition of oxidatively damaged cells.

The binding and uptake of oxidatively modified low density lipoprotein (OxLDL) by mouse peritoneal macrophages occurs, in part, via the well characterized acetyl LDL receptor. However, several lines of evidence indicate that as much as 30-70% of the uptake can occur via a distinct receptor that recognizes OxLDL with a higher affinity than it recognizes acetyl LDL. We describe the partial purification and characterization of a 94- to 97-kDa plasma membrane protein from mouse peritoneal macrophages that specifically binds OxLDL. This receptor is shown to be distinct from the acetyl LDL receptor as well as from two other macrophage proteins that also bind OxLDL--the Fc gamma RII receptor and CD36. We suggest that this OxLDL-binding membrane protein participates in uptake of OxLDL by murine macrophages and also represents a receptor responsible for macrophage binding and phagocytosis of oxidatively damaged cells.

Animals↗
PubMed1995

Recognition of oxidatively damaged and apoptotic cells by an oxidized low density lipoprotein receptor on mouse peritoneal macrophages: role of membrane phosphatidylserine.

We recently reported that oxidized low density lipoprotein (OxLDL), but not acetyl LDL (AcLDL), inhibited the binding and phagocytosis of nonopsonized, oxidatively damaged red blood cells (OxRBCs) by mouse peritoneal macrophages, implying the involvement of a "scavenger receptor" other than the AcLDL receptor. Numerous studies establish that loss of plasma membrane phospholipid asymmetry, which increases phosphatidylserine expression on the outer leaflet of the membrane, can play a key role in macrophage recognition of damaged and apoptotic cells. We report here that this recognition is in part attributable to the same mouse macrophage receptor that recognizes OxLDL. As described in an accompanying paper, this is a plasma membrane protein of 94-97 kDa. Phosphatidylserine liposomes show strong ligand binding to the same 94- to 97-kDa protein and this binding is inhibited by OxLDL but not by AcLDL. Inhibition of the RBC membrane phospholipid translocase by incubation with sodium vanadate caused a progressive increase in the appearance of phosphatidylserine on the cell surface and a parallel increase in the binding of these RBCs to macrophages, binding that was inhibited by OxLDL. Finally, OxLDL also inhibited the binding of sickled RBCs and apoptotic thymocytes to mouse macrophages. However, the latter was incomplete (approximately 50%), suggesting that other receptors are also involved. We suggest that the OxLDL receptor plays a significant role in recognition of damaged and apoptotic cells.

Animals↗
PubMed1995

Effect of the antioxidant N,N'-diphenyl 1,4-phenylenediamine (DPPD) on atherosclerosis in apoE-deficient mice.

Apolipoprotein (apo) E-deficient mice develop atherosclerotic lesions that contain epitopes formed during the oxidative modification of lipoproteins, and they demonstrate high titers of circulating autoantibodies against such epitopes, suggesting that this murine strain may provide a model to investigate the atherogenic mechanisms of oxidized lipoproteins (Palinski et al, Arterioscler Thromb. 1994; 14:605-616). To test the hypothesis that lipoprotein oxidation contributes to lesion formation in apoE-deficient mice, we studied the effect of the antioxidant N,N'-diphenyl 1,4-phenylenediamine (DPPD) in mice fed a high-fat diet containing 0.15% cholesterol. Animals were divided into two subgroups matched for sex and plasma cholesterol levels, and DPPD (0.5% wt/wt) was added to the diet of one subgroup. Throughout the 6 months of intervention, DPPD treatment had no significant effect on plasma cholesterol. Plasma levels of DPPD at the end of the experiment were 33.1 mumol/L. As judged by resistance to loss of polyunsaturated fatty acids, lipoproteins (d < 1.019 g/mL) from DPPD-treated animals showed greater resistance to copper-induced oxidation than lipoproteins from control animals. In addition, there was a greater than twofold prolongation of the lag time in the formation of conjugated dienes in the LDL and IDL fractions of DPPD-treated mice. Atherosclerosis was significantly reduced, by 36% in the DPPD-treated mice (14.0 +/- 4.53% of aortic surface area versus 21.9 +/- 11.6%; n = 32; P < .02). These results are consistent with the hypothesis that lipoprotein oxidation contributes to atherogenesis in apoE-deficient mice. However, further studies with other antioxidants are needed to validate this hypothesis.

Animals↗
PubMed1995

Mercury levels among dental personnel in Israel: a preliminary study.

Amalgam, the most prevalent dental restoration material used in dentistry, is potentially toxic because it contains mercury. Recent international publications confirm that mercury is potentially hazardous to dental personnel, who are exposed to mercury vapors both during their work at the office and from amalgam restorations in their own oral cavities. The purpose of our study was to compare urinary mercury levels of dental personnel with a control group, and to explore possible correlations between environmental factors in the dental office and the urinary level of the personnel. Our results indicate that the urinary mercury levels of the tested dental professionals were significantly higher than those of the control group (2.39 +/- 0.319 vs. 0.899 +/- 0.34 micrograms mercury/g creatinine). Of the dental personnel examined, 72% had detectable levels of urinary mercury, compared to 27% of the control group. Although mercury levels in all participants did not exceed the toxic limit, the above findings clearly point to the need for a continuation of this survey.

Adult↗
PubMed1994

Rabbit and human atherosclerotic lesions contain IgG that recognizes epitopes of oxidized LDL.

Atherosclerotic lesions contain relatively large quantities of IgG. We have previously shown that both human and rabbit sera contain autoantibodies against epitopes of oxidized (Ox) low-density lipoprotein (LDL) and that LDL isolated from atherosclerotic lesions contains small amounts of tightly bound IgG. However, it is not known whether IgG isolated from atherosclerotic lesions recognizes epitopes present in native LDL or Ox-LDL. IgG was isolated from Watanabe heritable hyperlipidemic (WHHL) rabbit atherosclerotic lesions by sequential salt extractions, purified by fast protein liquid chromatography on protein G, and used in a solid-phase radioimmunoassay. IgG and immune complexes were also isolated from the saline extracts of human lesions by adsorption onto latex beads coated with anti-human IgG antibodies or protein A. IgG isolated from rabbit lesions showed significant titers against malondialdehyde (MDA)-modified LDL and LDL oxidized by copper ions for 4 and 18 hours but not against native LDL. On Western blots, lesion IgG stained MDA-LDL and fragments of Ox-LDL. Western blots of immune complexes isolated from human lesions revealed the presence in the isolated complexes of both apoprotein B and apoprotein B fragments, which reacted with antibodies to MDA-lysine. Furthermore, rabbit lesion IgG immunostained epitopes of Ox-LDL present in human atherosclerotic lesions. Immunostains obtained with rabbit lesion IgG were similar to those obtained with a monoclonal antibody specific for MDA-lysine. The results show that human and rabbit atherosclerotic lesions contain IgG that recognizes epitopes characteristic of Ox-LDL. These data suggest that immunologic processes may be an important component of the atherogenic process.

Adult↗
PubMed1994

ApoE-deficient mice are a model of lipoprotein oxidation in atherogenesis. Demonstration of oxidation-specific epitopes in lesions and high titers of autoantibodies to malondialdehyde-lysine in serum.

Apolipoprotein (apo) E-deficient transgenic mice develop marked hyperlipidemia and progressive atherosclerotic lesions. To explore whether oxidative modification of lipoproteins is involved in atherogenesis in this murine model, we performed extensive immunocytochemical studies. Atherosclerotic lesions ranging from early fatty streaks to very advanced plaques were examined from the aortic valve region and the thoracic and abdominal aorta. Using guinea pig antisera against malondialdehyde (MDA)-lysine and 4-hydroxynonenal-lysine, two epitopes generated during the oxidative modification of low-density lipoprotein (LDL), we demonstrated the presence of these "oxidation-specific epitopes" in atherosclerotic lesions. In early lesions, oxidation-specific epitopes were found predominantly in macrophage-rich areas, whereas diffuse extracellular staining predominated in necrotic areas of advanced lesions. We have previously shown that autoantibodies against MDA-lysine are present in the circulation of humans and rabbits and that the immunoglobulin fraction extracted from their lesions contains autoantibodies against several "oxidation-specific" epitopes. Sera from apoE-deficient mice also contained circulating autoantibodies to MDA-lysine, and both early and advanced lesions were rich in murine immunoglobulins. Titers of serum autoantibodies were significantly higher in apoE-deficient mice than in C57BL/6 mice. Autoantibodies in murine plasma recognized MDA-lysine epitopes in atherosclerotic lesions of rabbits, and the immunostaining was competitively inhibited by excess human MDA-LDL. Similar findings were obtained by competitive radioimmunoassay. Finally, a morphometric technique was developed and tested in these mice that allows a quantitative assessment of aortic atherosclerosis. These findings suggest that in apoE-deficient mice, lipoprotein oxidation is involved in atherogenesis and that these transgenic mice constitute an appropriate model with which to study the antiatherogenic effect of antioxidant intervention.

Animals↗
PubMed1994

P-selectin mediates the interaction of circulating leukocytes with platelets and microvascular endothelium in response to oxidized lipoprotein in vivo.

BACKGROUND: Oxidized low density lipoprotein (oxLDL) has been demonstrated to stimulate leukocyte/endothelium interaction, an early feature of atherogenesis. Using the skinfold chamber model for intravital microscopy in hamsters and mice, we have shown that oxLDL-induced leukocyte adhesion to microvascular endothelium shares many characteristics with leukocyte adhesion during inflammation and ischemia/reperfusion, including the involvement of beta 2 integrin adhesion molecules. In light of the two-step model of leukocyte adhesion, we have examined the contribution of P-selectin to oxLDL-induced leukocyte/endothelium interaction. P-selectin is an inducible adhesion molecule on platelets and endothelium, mediating the initial steps of leukocyte margination and rolling along the endothelial lining, as well as of aggregate formation between platelets and leukocytes. EXPERIMENTAL DESIGN: For our studies, we used the dorsal skinfold chamber model for intravital fluorescence microscopy on awake Syrian golden hamsters. Hamsters were treated 10 minutes before oxLDL-injection (oxidized by Cu2+, 4 mg/kg body weight, intravenously) with blocking antibodies to P-selectin (2 mg/kg body weight intravenously, N = 7). RESULTS: In seven control animals (pretreated with an irrelevant IgG antibody), oxLDL injection elicited leukocyte rolling and adhesion on both venular and arteriolar endothelium, and also the formation of aggregates tumbling down the microvessels and firmly adhering to the microvascular endothelium. The aggregates consisted of leukocytes and activated, dendritic platelets, as assessed by scanning electron microscopy of the buffy coat isolated by density gradient centrifugation of whole blood taken from hamsters 15 minutes after injection of oxLDL. Leukocyte adhesion to venular and arteriolar endothelium, as well as the formation of leukocyte/platelet aggregates were significantly reduced by pretreatment of the animals with anti-P-selectin antibodies. CONCLUSIONS: These data emphasize the similarities between leukocyte adhesion in response to oxLDL and in other pathophysiologic conditions, identifying P-selectin as a crucial player in the interaction between leukocytes and microvascular endothelium as well as in the formation of circulating leukocyte/platelet aggregates.

Animals↗
PubMed1992

Evidence for a concerted reaction between lipid hydroperoxides and polypeptides.

The events accompanying oxidative modification of low density lipoprotein (LDL) are multiple and complex, and the precise mechanisms remain to be determined. In the present studies, we examined a simple system in which we first prepared large amounts of lipid hydroperoxides (from linoleic acid or from phospholipids containing linoleic acid) by using soybean lipoxygenase (linoleate:oxygen oxidoreductase, EC 1.13.11.12). Linoleoyl hydroperoxide was then incubated with polypeptides in the absence of metal ions. We observed the generation of fluorescent products with a spectrum like that of oxidized LDL. The generation of fluorescent products from incubation of polypeptides with linoleoyl hydroperoxide was manyfold greater than that generated on incubation with preformed 4-hydroxynonenal at the same concentration. Superoxide dismutase (superoxide: superoxide oxidoreductase, EC 1.15.1.1) had no effect on the generation of fluorescent products. Incubation of linoleoyl hydroperoxide with cytochrome c (cyt c) under the same conditions led to progressive reduction of cyt c at a rate determined by the initial linoleoyl hydroperoxide concentration. This reduction was not significantly inhibited by probucol but was inhibited, although never completely, by superoxide dismutase. Even at 100 micrograms/ml, superoxide dismutase inhibited by only 65%. From these results, we are led to suggest a concerted reaction between the peroxy radical and free amino groups of polypeptides or phosphatidylethanolamine to generate fluorescent adducts. During oxidation of LDL or of cell membranes, this mechanism may occur side by side with the conventional Schiff base mechanism.

Cytochrome c Group↗
PubMed1992

The effect of delmopinol on glucosyltransferase adsorbed on to saliva-coated hydroxyapatite.

The aim was to explore the effects of delmopinol, a substituted amino-alcohol compound recently reported as a potential antiplaque agent, on GTF activity in solution and when adsorbed on to sHA. Delmopinol was without a significant effect on GTF activity in solution. In contrast, a reduction in the bound glucans synthesized by the adsorbed GTF was found in the presence of delmopinol. Delmopinol did not displace the adsorbed GTF from the sHA, nor was there significant desorption of glucans from sHA. The total glucan synthesis (bound and unbound) was reduced in the presence of delmopinol. Inhibition of GTF was not reversed by sucrose. Inhibition of GTF activity by delmopinol apparently results from drug-enzyme interaction on the surface of sHA beads. These observations provide further support for the important differences in the properties of adsorbed GTF and GTF in solution, illustrating that GTF-drug interaction differs between enzyme adsorbed to surfaces and enzyme in solution.

Adsorption↗
PubMed1992

Microbiological evaluation of the efficacy of chlorhexidine in a sustained-release device for dentine sterilization.

The aim of this in-vitro study was to evaluate the effect of chlorhexidine in solution and in a sustained-release device as an intracanal medication. Hollow cylindrical dentine specimens prepared from bovine incisors were incubated with Streptococcus faecalis for a period of 3 weeks. The intracanal medicaments tested were 0.2% chlorhexidine gluconate solution (CH), chlorhexidine in a sustained-release device (1.2 mg) (SBD), camphorated paramonochlorophenol (CMCP), and a control (no medication). Each medicament was introduced into the lumen of one dentine specimen and incubated for 5 min, 24 h, 48 h or 7 days at 37 degrees C. The bacteriological samples were taken by shaving the dentine inside the lumen with dental burs ranging in size from ISO 023-033. The dentine powder was collected in test-tubes containing growth medium and incubated for 24 h. The optical density of the medium was recorded by means of a spectrophotometer at a wavelength of 540 nm. There was a statistically significant difference between the control group and all the medicaments tested.

Animals↗
PubMed1992

Efficacy of a sustained-release device containing chlorhexidine and Ca(OH)2 in preventing secondary infection of dentinal tubules.

This study was conducted to evaluate the efficacy of the antibacterial activity of Ca(OH)2 and a sustained-release device containing chlorhexidine (SRD) in both sterilization and prevention of secondary infection of the root canal system. Bovine root dentine specimens previously incubated with Streptococcus faecalis were used in this experiment. Two different formulations of the SRD (differing in their cross-linkage), Ca(OH)2 and normal saline (control) were evaluated. The degree of bacterial infection of the root canal was tested after incubation periods of 24 h, 72 h and 7 days with these medicaments. Their efficacy in preventing secondary infection after recontamination was tested after 72 h and 7 days. The results demonstrated that both formulations of the SRD significantly reduced the bacterial population in the primary infected groups, as well as preventing secondary infection of the dentinal tubules in the recontaminated group. By contrast, Ca(OH)2 did not show any antibacterial activity, and failed to sterilize the dentinal tubules or prevent secondary infection after recontamination at the time periods examined.

Animals↗
PubMed1992

Dental caries and its determinants among recent immigrants from rural Ethiopia.

Adults (35-45 yr old) and children (5 and 12 yr old) recently arrived in Israel from rural areas in Ethiopia were examined. Caries levels were low: e.g. DMFT = 0.31 among 12-yr-olds, and 1.27 amongst adults. The 5-yr-olds were 86.8% caries-free, while 12-yr-olds were 81.8% and adults 54% caries-free respectively. According to interview data, the diet in Ethiopia had been based on local agricultural products and was almost sugar-free. The mean total count of salivary bacteria, as determined on blood agar, was 3.4 x 10(8); mean count of Streptococcus viridans, on mitis salivarius, was 6.7 x 10(7); and mean count of S. mutans, as determined on mitis salivarius with bacitracin, was 1.7 x 10(7). These levels were all high and were not significantly different from a control group of 20 Israelis. The mean number of lactobacilli, on Rogosa agar, was 2.75 x 10(4), which was significantly higher than among the controls (3.6 x 10(3). Salivary pH levels were generally similar between the Ethiopian group and the controls. Salivary flow was significantly higher for the Ethiopians (1.93 ml/min) than for controls (1.16 ml/min). Low levels of caries in this population can be attributed to an almost sugar-free diet and high salivary flow, but not to the composition of oral microflora.

Adult↗