Biomedical subjects

T Mazzone

Publications and source records attributed to T Mazzone.

At least 37 records · Page 2Linked to original sources

PubMed1997

Lipoprotein lipase reduces secretion of apolipoprotein E from macrophages.

Macrophages are a significant source of lipoprotein lipase (LPL) and apolipoprotein E (apo E) in the developing arterial wall lesion, and each of these proteins can importantly modulate lipid and lipoprotein metabolism by arterial wall cells. LPL and apo E share a number of cell surface binding sites, including proteoglycans, and we have previously shown that proteoglycans are important for modulating net secretion of apoprotein E from macrophages. We therefore evaluated a potential role for LPL in modulating net secretion of macrophage-derived apo E. In pulse-chase experiments, addition of LPL during the chase period produced a decrease in secretion of apoprotein E from human monocyte-derived macrophages, from the human monocytic THP1 cell line, and from J774 cells transfected to constitutively express a human apo E cDNA. LPL similarly reduced apo E secretion when it was prebound to the macrophage cell surface at 4 degrees C. A native LPL particle was required to modulate apo E secretion; addition of monomers and aggregates did not produce the same effect. Depletion of cell surface proteoglycans by a 72-h incubation in 4-methylumbelliferyl-beta-D-xyloside did not attenuate the ability of LPL to reduce apo E secretion. However, addition of receptor-associated protein attenuated the effect of LPL on apo E secretion. Although LPL could mediate removal of exogenously added apo E from the culture medium, detailed pulse-chase analysis suggested that it primarily prevented release of newly synthesized apo E from the cell layer. Cholesterol loading of cells or antibodies to the low density lipoprotein receptor attenuated LPL effects on apo E secretion. We postulate that LPL sequesters endogenously synthesized apo E at the cell surface by a low density lipoprotein receptor-dependent mechanism. Such post-translational regulation of macrophage apo E secretion by LPL could significantly influence apo E accumulation in arterial vessel wall lesions.

Animals↗

PubMed1997

Cisapride use during human pregnancy: a prospective, controlled multicenter study.

The objective of this prospective multicenter study was to determine whether cisapride is associated with increased risk of malformations, spontaneous abortions, or decreased birthweight when used during pregnancy. Cases were paired for age, smoking, and alcohol consumption with controls exposed to nonteratogens, as well as with disease-paired controls. One hundred and twenty-nine pregnant women were exposed to cisapride during pregnancy, including 88 during the period of fetal organogenesis. There were no differences in maternal history, birthweight, gestational age at delivery, and rates of livebirths, spontaneous or therapeutic abortions, fetal distress, and major or minor malformations among groups. It is concluded that exposure to cisapride during pregnancy is not associated with a major increased risk of malformations or spontaneous abortions or with decreased birthweight.

Abnormalities, Drug-Induced↗

PubMed1997

Mechanisms of enhanced macrophage apoE secretion by oxidized LDL.

Previous studies have demonstrated that atherosclerotic lesions contain apoE synthesized primarily by macrophages. As oxidized LDL has been implicated in the development of atherosclerosis, its effect on macrophage apoE synthesis and secretion was examined. Human monocytic leukemia cells, THP-1, and human monocyte-derived macrophages were exposed to various forms of oxidatively modified LDL for determination of their effect on apoE mRNA and protein levels. Extensively copper oxidized (Cu-oxidized) LDL resulted in a time- and concentration-dependent increase in apoE mRNA and protein as compared to other forms of oxidized LDL, i.e., LDL modified by soybean lipoxygenase (SLO), azoamidinopropane HCl (AAPH), and hypochlorite (HOCl). Consistent with these results, experiments using THP-1 cells transfected with the apoE promoter linked to a luciferase reporter gene indicated that Cu-oxidized LDL was the most potent stimulator of apoE transgene expression. Enhanced apoE expression due to Cu-oxidized LDL was shown to be due to cholesterol accumulation as well as additional factors. HPLC analysis of the various forms of modified LDL revealed that 7-ketocholesterol was the major oxysterol present in Cu-oxidized LDL. AAPH-oxidized LDL contained significantly less 7-ketocholesterol than Cu-oxidized LDL and virtually no 7-ketocholesterol was detected in SLO- or HOCl-oxidized LDL. Northern blot analysis indicated an increase in apoE mRNA in response to increasing concentrations of 7-ketocholesterol. These results elucidate a potential role of oxidized LDL, and specifically 7-ketocholesterol, in the stimulation of macrophage apoE secretion in atherosclerotic lesions.

Apolipoproteins E↗

PubMed1996

Cell surface proteoglycans modulate net synthesis and secretion of macrophage apolipoprotein E.

Using a macrophage cell line that constitutively expresses a human apolipoprotein E (apoE) cDNA, we have investigated the post-translational metabolism of endogenously produced apoE. Inhibition of lysosomal or cysteine proteases led to significant inhibition of apoE degradation but did not increase apoE secretion, indicating that cellular degradation is not limiting for apoE secretion in macrophages. Treatment of macrophages with inhibitors of proteoglycan synthesis (4-methylumbelliferyl-beta-D-xyloside) or sulfation (sodium chlorate) enhanced the release of apoE from cells and significantly attenuated the increase in secretion produced by incubation with phosphatidylcholine vesicles (PV). These observations suggested that a significant fraction of the apoE retained by cells (and released by incubation with PV) was associated with proteoglycans. Treatment of cells with exogenous heparinase led to a greater than 4-fold increase in apoE secretion and similarly attenuated the response to PV, suggesting that apoE was trapped in an extracellular proteoglycan matrix. This conclusion was confirmed in studies showing that PV could enhance the release of apoE from cells during an incubation at 4 degrees C, but this enhanced release was abolished in proteoglycan-depleted cells. Incubation with lactoferrin at 4 or 37 degrees C produced a similar decrement in cellular apoE, again indicating the existence of a cell surface pool of apoE. Pulse-chase studies showed that the apoE trapped in the proteoglycan matrix was susceptible to rapid cellular degradation such that net synthesis of apoE (secreted plus cell-associated) was increased significantly in proteoglycan-depleted cells compared with control cells as early as 45 min during a chase period.

Apolipoproteins E↗

PubMed1996

Prospective assessment of pregnancy outcomes after first-trimester exposure to fluconazole.

OBJECTIVE: Our purpose was to study prospectively the pregnancy outcome after first-trimester exposure to fluconazole, an effective antifungal agent teratogenic in animals. STUDY DESIGN: We conducted a prospective cohort study of women who contacted three Italian teratogen information services. We compared the pregnancy outcomes of 226 women exposed to fluconazole with that of 452 women exposed to nonteratogenic agents, with use of logistic regression to control for potential confounders. RESULTS: Among the 226 pregnancies exposed to fluconazole there were 22 miscarriages, 1 stillbirth, and 7 infants with congenital anomalies. The prevalence of these outcomes and of neonatal growth parameters and the rate of neonatal complications were similar to those in the reference group. Women in the fluconazole group had a fivefold increased occurrence of induced abortions. CONCLUSIONS: First-trimester exposure to fluconazole does not appear to increase the prevalence of miscarriages, congenital anomalies, and low birth weight.

Adult↗

PubMed1996

Apolipoprotein E secretion by macrophages: its potential physiological functions.

Results of numerous studies using varied experimental approaches have established a role for macrophage-derived apolipoprotein E in modulating cholesterol balance in the arterial wall, thereby potentially modulating progression or regression of the vessel wall atherosclerotic lesion. Macrophage-derived apolipoprotein E could also act on the vessel wall by modulating the aggregability of platelets and the proliferation of lymphocytes. In addition, macrophage-derived apolipoprotein E interacts with extracellular matrix and such interactions could influence the retention of lipoproteins in the vessel wall, the bioavailability of cytokines and growth factors sequestered in the extracellular matrix and the regulation of arterial smooth muscle cell growth by matrix components.

Apolipoproteins E↗

PubMed1996

Cholesterol homeostasis is modulated by amphiphiles at transcriptional and post-transcriptional loci.

A variety of amphiphiles inhibit plasma membrane cholesterol esterification and induce 3-hydroxy-3-methylglutaryl-coenzyme A reductase accumulation in cultured cells; among these are steroids, hydrophobic amines, phenothiazines, ionophores, colchicine, and lysophosphatides. It has been proposed that these amphiphiles signal a sterol deficiency to regulatory sites by blocking the movement of plasma membrane cholesterol into the cell (Lange, Y., and Steck, T. L. 1994. J. Biol. Chem. 269: 29371-29374). If this were the case, these agents also should enhance transcription of sterol responsive genes and stabilize 3-hydroxy-3-methylglutaryl-coenzyme A reductase. As a test of this hypothesis, the effect of the amphiphiles on such transcriptional and post-transcriptional events was assessed. A mouse embryo cell line was transfected with a construct containing the promoter for the human low density lipoprotein receptor upstream of the DNA sequence coding for chloramphenicol acyltransferase (CAT). Incubation of these cells for 7-18 h with the aforementioned agents caused the level of expression of the promoter/CAT construct to increase 2- to 9-fold. We showed further that the amphiphiles stimulated 3-hydroxy-3 methylglutaryl-coenzyme A reductase activity by increasing gene transcription as well as by decreasing degradation of the enzyme. These are the predicted homeostatic responses to cell cholesterol deficiency. These findings support the hypothesis that certain amphiphiles falsely signal a cholesterol deficiency to the intracellular sites regulating cholesterol homeostasis.

3T3 Cells↗

PubMed1995

Isoform-specific induction of the low-density lipoprotein receptor gene by platelet-derived growth factor.

We investigated the effect of recombinant platelet-derived growth factor (PDGF) isomers on low-density lipoprotein (LDL) receptor gene expression and compared this with two indexes of cell growth response: expression of the immediate early gene c-myc and the rate of DNA synthesis. In human skin fibroblasts and NIH 3T3 cells, the PDGF-BB homodimer was more effective in inducing the LDL receptor gene and cell growth response compared with the PDGF-AA homodimer. The second messenger pathways utilized by PDGF receptors for enhancing LDL receptor gene response could, however, be dissociated from those utilized for enhancing c-myc gene response and were insensitive to inhibitors of tyrosine phosphorylation. Inhibition of tyrosine kinase activity inhibited c-myc gene response to PDGF-BB at 10(-8) M but had little effect on LDL receptor gene response. Such inhibition increased expression of the LDL receptor gene in the presence of the PDGF-AA isomer. Our results indicate that the response of the LDL receptor gene to PDGF isoforms reflects cellular growth response. However, different transduction pathways are utilized for PDGF activation of the c-myc and LDL receptor genes in mesenchymal cells.

3T3 Cells↗

PubMed1995

Platelet-derived growth factor enhances Sp1 binding to the LDL receptor gene.

We have previously demonstrated that growth activation of quiescent cells enhances LDL receptor gene transcription and that the proximal 5' flanking region of the LDL receptor gene could transduce a platelet-derived growth factor (PDGF) response. This portion of the LDL receptor gene encompasses a previously characterized sterol response element and an adjacent Sp1 binding site. By use of mobility shift analyses we show that PDGF activation of quiescent cells enhances binding of Sp1 to the LDL receptor gene. Transfection analyses indicated that the Sp1 site, but not the sterol response element binding protein site, could confer PDGF responsiveness to a heterologous promoter in quiescent cells. Furthermore, cotransfection of an LDL receptor reporter gene (containing -141 to +35 bp of the LDL receptor gene promoter) along with an expression construct coding for high-level constitutive expression of an Sp1 cDNA led to marked enhancement in expression of the LDL receptor reporter gene in quiescent cells. Increased Sp1 binding due to PDGF could be due to enhanced production of Sp1; alternatively, posttranslational activation of binding could be involved. Western blot analysis showed no difference in Sp1 abundance in quiescent cells versus PDGF-stimulated cells, suggesting a posttranslational mechanism for activation of Sp1 binding by growth induction. Our data demonstrate that PDGF stimulation of quiescent cells leads to enhanced Sp1 binding to the LDL receptor gene. This enhanced binding could participate in PDGF induction of LDL receptor gene transcription.

3T3 Cells↗

PubMed1995

Tumor necrosis factor-alpha modulates monocyte/macrophage apoprotein E gene expression.

apo E has been shown to modulate cholesterol balance in arterial wall cells. Production of apo E by macrophages in atherosclerotic plaques could thereby influence the development of the plaque lesion. Cytokines, including TNF alpha, have been identified in human lesions, therefore, we undertook a series of studies to evaluate the effect of TNF alpha on monocyte/macrophage apo E production. The addition of TNF alpha to freshly isolated human monocytes led to a four- to fivefold increase of apo E mRNA abundance. The addition of TNF alpha to fully differentiated macrophages either had no effect or modestly inhibited apo E mRNA expression. THP1 human monocytic cells also responded to TNF alpha in a phenotype-specific manner. Treatment of these cells with TNF alpha produced a dose- and time-dependent increase in apo E mRNA. This increase was reflected in apo E synthesis and was associated with inhibition of DNA synthesis, and with induction of c-fos and ICAM-1 gene expression. Cell-permanent analogues of ceramide did not reproduce TNF alpha effect on apo E, but antagonists of protein kinase C did inhibit its effect. TNF alpha induction of apo E mRNA abundance was associated with stimulation of apo E promoter-dependent gene transcription. In summary, TNF alpha stimulates apo E gene transcription, mRNA abundance, and protein synthesis in the monocyte/macrophage in a phenotype-specific manner. Such regulation could significantly modify the amount of apo E present in vessel wall lesions.

Apolipoproteins E↗

PubMed1995

Progesterone blocks intracellular translocation of free cholesterol derived from cholesteryl ester in macrophages.

Macrophage foam cells must accommodate continuing fluxes of free cholesterol in spite of a greatly expanded store of cholesteryl ester. Though endogenous free cholesterol synthesis is suppressed, free cholesterol continues to enter the cell via endocytosis of oxidized/modified lipoproteins. It has been shown previously that this free cholesterol is released into the lysosomal compartment and rapidly transported to the plasma membrane prior to its esterification. A substantial amount of free cholesterol is also presented via the continuous hydrolysis of cholesteryl ester during the cholesteryl ester cycle. We addressed the question of whether the intracellular free cholesterol derived from the hydrolysis of cholesteryl ester formed a protected pool for rapid re-esterification. Incubation of macrophage foam cells with cyclic AMP to enhance cholesteryl ester hydrolysis, and with S58035 to inhibit acyl-CoA:cholesterol acyltransferase (ACAT) activity, led to conversion of cellular cholesteryl ester to free cholesterol and transport of this free cholesterol to the plasma membrane. Addition of progesterone, previously demonstrated to be an inhibitor of free cholesterol transport in other cell types, also led to conversion of cholesteryl ester to free cholesterol even though progesterone was only a weak inhibitor of ACAT activity. Free cholesterol in the plasma membrane was an important source of ACAT substrate to balance the constitutive hydrolysis of cholesteryl ester in cholesterol-loaded macrophages. Treatment of cells with progesterone, however, prevented free cholesterol derived from cholesteryl ester hydrolysis from moving to the plasma membrane. The sequestration of free cholesterol by progesterone could be reversed by incubation with human HDL3.(ABSTRACT TRUNCATED AT 250 WORDS)

Amides↗

PubMed1994

Evaluation of the role of Ap1-like proteins in the enhanced apolipoprotein E gene transcription accompanying phorbol ester induced macrophage differentiation.

Differentiation of THP1 monocytes to a macrophage phenotype is accompanied by increased apolipoprotein E gene transcription. Using transfection analysis with 5' deletion mutations of the 5' control region of the apo E gene in THP1 cells, we show that the -651 to +86 chloramphenicol acetyltransferase (CAT) construct is efficiently expressed in the monocyte; as has been reported for other cell types. Further, we found that an 176 bp region between -623 to -447 was required for the induction of apolipoprotein E gene transcription during 12-O-tetradecanoylphorbol-13-acetate-induced differentiation of monocytes to macrophages. Gel-retardation patterns of the apolipoprotein E promoter region using nuclear extracts from differentiated or undifferentiated THP1 cells revealed altered binding of Ap1-like nuclear factor/s to the -620 to -583 bp region after macrophage differentiation. Mutation of an Ap1 element at position -602 abolished specific binding of Ap1-like proteins to the -620 to -583 bp fragment of the apo E gene and significantly reduced expression of a -623 to +86 apo E-CAT construct during differentiation. These data indicate that differentiation-related expression of the apolipoprotein E gene following phorbol ester stimulation is transduced by gene elements between -623 and -447. Furthermore, the data indicate that transcriptional activation of the apo E gene during macrophage differentiation is associated with induction of Ap1-like proteins which bind to the Ap1 response element present at -602 in the apolipoprotein E gene and importantly contribute to enhanced gene expression.

Adaptor Protein Complex 1↗

PubMed1994

Clinical evaluation of a semipermeable polymeric membrane dressing for the treatment of chronic diabetic foot ulcers.

OBJECTIVE: To evaluate the utility of a semipermeable polymeric membrane dressing for the treatment of chronic diabetic foot ulcers. RESEARCH DESIGN AND METHODS: Nineteen subjects with either insulin-dependent diabetes mellitus (IDDM) or non-insulin-dependent diabetes mellitus (NIDDM) and foot ulcers were randomly assigned to the polymeric dressing or conventional wet-to-dry saline dressings. Subjects had foot ulcer site measurements performed every 3 weeks. The subjects using conventional therapy were allowed to cross over to polymeric dressing after 2 months. RESULTS: At the end of 2 months, in the patients using the polymeric dressing, ulcer size was reduced to 35 +/- 16% of baseline. The patients on conventional therapy had an ulcer size of 105 +/- 28% of baseline (P < 0.03, polymeric vs. conventional). Patients initially treated with wet-to-dry saline were crossed over into the polymeric membrane treatment and demonstrated a decrease to 35 +/- 11% of baseline size (p < 0.02) after an additional 2 months. CONCLUSIONS: The semipermeable polymeric membrane dressing is a useful therapeutic option for treating uncomplicated chronic diabetic foot ulcers.

Chronic Disease↗

PubMed1994

[Injuries in pregnancy].

From a review of the literature on this subject it emerges that around 6-7% of women are injured during the course of pregnancy. Road accidents, falls and domestic accidents are the most frequent events: it is therefore important to provide pregnant women with information to prevent such events which may endanger both maternal and fetal health.

Accidental Falls↗

PubMed1994

Expression of heterologous human apolipoprotein E by J774 macrophages enhances cholesterol efflux to HDL3.

Expression of apolipoprotein (apo) E by macrophages is tightly regulated by cellular cholesterol content. We have investigated a potential modulating role for apoE on macrophage cholesterol homeostasis by stably transfecting the J774 macrophage, which does not express its endogenous apoE gene, with a human apoE cDNA expression vector and comparing cholesterol homeostasis in this cell line with that of a control line transfected with the neomycin resistance construct only. Incubation in serum-free medium after cholesterol loading produced no difference in cellular cholesterol content between apoE secreting and non-secreting J774 cells. Similarly, in serum-free medium there was no difference in the amount of radiolabeled cholesterol effluxed. Addition of cAMP or S58035 to cholesterol-loaded J774 cells did enhance efflux of radiolabeled cholesterol from apoE secreting compared to non-secreting macrophages but did not detectably alter cellular free cholesterol or cholesteryl ester mass. Incubation with HDL3 alone, however, significantly decreased macrophage cholesteryl ester mass compared to a 24-h incubation in serum-free medium from 10.5 +/- 3.9 to 3.2 +/- 2.0 (P < 0.01) in apoE-secreting J774 cells. During a 24-h incubation in HDL3, cholesteryl ester fell from 6.4 +/- 2.4 to 0.8 +/- 0.7 (delta = 5.6 micrograms/mg) in apoE-secreting cells and from 9.3 +/- 2.2 to 7.7 +/- micrograms/mg (delta = 1.6 micrograms/mg) in non-secreting cells (P < 0.005 apoE-secreting vs. non-secreting cells).(ABSTRACT TRUNCATED AT 250 WORDS)

Apolipoproteins E↗

PubMed1993

The acute-phase response and associated lipoprotein abnormalities accompanying lymphoma.

Lipoprotein abnormalities seen in patients with inflammatory diseases are thought to develop secondary to circulating cytokines and the accompanying acute-phase response. Patient's with lymphoma may develop similar lipoprotein abnormalities but the mechanism is unclear. We report a patient with B-cell lymphoma who presented with an HDL cholesterol level of 3 mg dl-1, an ApoA level of 17.4 mg dl-1, elevated triglyceride level (272 mg dl-1) and an elevated ApoB level of 156 mg dl-1. Density gradient analysis of the patient's lipoproteins demonstrated a virtual absence of an identifiable HDL particle. Serum amyloid A and C-reactive protein were also elevated. All of the lipoprotein abnormalities resolved with chemotherapy and resolution of the acute-phase response. The acute-phase response may be associated with striking lipoprotein abnormalities in a subset of patients with lymphoma. Lymphoma should be included in the differential diagnosis of patients with hypertriglyceridaemia and low HDL cholesterol.

Acute-Phase Reaction↗

PubMed1992

Post-translational regulation of macrophage apoprotein E production.

We have transfected the murine macrophage cell line, J774, which does not express its endogenous apoE gene, with a constitutively expressed human apoE cDNA in order to study post-transcriptional and post-translational control of apoE production in macrophages. Loading cells with cholesterol using preincubations in acetylated low density lipoprotein, previously shown to enhance macrophage apoE gene transcription and apoE synthesis, did not increase apoE synthesis or secretion in constitutively expressing transfected cells, suggesting that sterol control of macrophage apoE production occurs predominantly at a transcriptional locus. However, incubation in human high density lipoprotein (HDL3) stimulated apoE secretion and appeared to inhibit degradation of newly synthesized apoE. This effect could be entirely reproduced by incubation with phosphatidylcholine vesicles which increased apoE accumulation in the medium by 2-6-fold. Pulse-chase experiments indicated that the effect of HDL3 or phospholipid vesicles was very rapid (occurring within 15 min) and was independent of changes in apoE synthesis. Furthermore, the increased apoE which accumulated in the medium in the presence of phospholipid vesicles or HDL3 was not due to altered rates of reuptake of labeled apoE since this difference was completely preserved in the absence of extracellular calcium. These results indicate that alteration of sterol content does not regulate macrophage apoE production at a translational or post-translational locus but that incubation with HDL3 or phospholipid vesicles can enhance apoprotein E production independent of changes in apoE gene transcription or apoE synthesis. The nature of the signal generated by the phospholipid vesicles which leads to inhibition of intracellular apoE degradation and enhanced apoE secretion will require further investigation.

Animals↗

PubMed1992

Transcriptional and post-transcriptional control of apolipoprotein E gene expression in differentiating human monocytes.

The present studies examined the mechanisms responsible for the regulation of apolipoprotein (apo) E gene expression during human monocytic differentiation. Levels of apoE mRNA were low in undifferentiated THP1 cells, a human monocytic cell line. Addition of 12-O-tetradecanylphorbol-13-acetate (PMA) induced differentiation of these cells to a macrophage-like phenotype and was associated with increased apoE mRNA abundance in a time-dependent fashion, up to 10-11-fold within 32 h. Results of nuclear run-on transcription assays demonstrated that the apoE gene was transcriptionally active in undifferentiated THP1 cells and that differentiation of monocytes with PMA was associated with a maximal increase of apoE gene transcription rate of only 2-3-fold at 6-12 h. Using actinomycin D as an inhibitor of new transcription, we could demonstrate a more rapid degradation of mature apoE mRNA in undifferentiated compared to differentiated cells, suggesting that the apoE mRNA species was more stable in differentiated THP1 cells. Primer extension assays performed using RNA extracts from undifferentiated and differentiated THP1 cells confirmed the increase of apoE mRNA abundance in the latter but failed to disclose heterogeneity in apoE gene transcription start site between these two phenotypes. These findings indicate that apoE gene expression is controlled at both transcriptional and post-transcriptional loci during human monocyte-macrophage differentiation.

Apolipoproteins E↗