PubMed Health⌕ Search

Biomedical subjects

J M Zingg

Publications and source records attributed to J M Zingg.

At least 19 recordsLinked to original sources
PubMed2005

Effects of aldehydes on CD36 expression.

INTRODUCTION: During the oil frying process lipid peroxidation compounds are formed. These products can modulate gene expression and alter cellular behaviour. The cellular uptake of oxidized LDL, a key step in the development of atherosclerosis, is mediated by the CD36 scavenger receptor, whose expression is down-regulated by alpha-tocopherol. OBJECTIVE: To determine the effects of water-soluble aldehydes, obtained from thermally oxidized sunflower oil on the expression of CD36 scavenger receptor in human monocytes (THP-1 cells). We also wanted to study the effects of alpha-tocopherol on CD36 expression in the presence of water-soluble aldehydes. MATERIALS AND METHODS: Sunflower oil was heated in a frying pan, at 180--200 degrees C for 40 min, water-soluble aldehydes were isolated, and the content of thiobarbituric acid reacting substances (TBARS) was determined. THP-1 monocytes were cultured in RPMI medium during 24 h and incubated with increasing concentrations of the water-soluble aldehydes (ranging from 0.05 to 1 microM) and with or without 50 microM of alpha-tocopherol. In parallel, THP-1 cells were cultured with the same volume of an extract obtained from non-oxidized oil or distilled water. The CD36 expression at the cell surface was studied with fluorescence-activated cell sorting (FACS). RESULTS: Monocytes incubated in a medium containing water-soluble aldehydes, showed a dose dependent increase in the expression of the CD36 protein on the cell surface, compared to with the control groups. When the cells were treated simultaneously with 50 microM of alpha-tocopherol a significant reduction in the expression of the CD36 protein was observed. CONCLUSION: Water-soluble aldehydes, extracted from thermally oxidized culinary oil, increase the expression of CD36. This effect is partially decreased by the presence of alpha-tocopherol.

Aldehydes↗
PubMed2001

Vitamin E 80th anniversary: a double life, not only fighting radicals.

Recent research on alpha-tocopherol has revealed specific cellular functions of this compound belonging to the vitamin E family. Alpha-tocopherol can act as a radical scavenger, as a pro-oxidant, as an anti-alkylation agent and, most important, by mechanisms that are independent of the above properties. To the last group belong protein kinase C and 5-lipoxygenase inhibition at post-translational level, as well as alpha-tocopherol activation of protein phosphatase 2A and diacylglycerol kinase. Furthermore, at transcriptional level, several genes (CD36, alpha-TTP, alpha-tropomyosin, and collagenase) are modulated by alpha-tocopherol. These effects result in inhibition of smooth muscle cell proliferation, platelet aggregation, and monocyte adhesion and may be related to the alleged protection of atherosclerosis by vitamin E. On the other side, epidemiological and intervention studies have shown some inconsistent results. Rather than disregarding vitamin E as a means to protect against atherosclerosis progression, it would be wiser to better design clinical trials based on current knowledge of the biological properties of the molecule.

Animals↗
PubMed2001

Vitamin E: protective role of a Janus molecule.

Since the discovery of vitamin E in 1922, its deficiency has been associated with various disorders, particularly atherosclerosis, ischemic heart disease, and the development of different types of cancer. A neurological syndrome associated with vitamin E deficiency resembling Friedreich ataxia has also been described. Whereas epidemiological studies have indicated the role of vitamin E in preventing the progression of atherosclerosis and cancer, intervention trials have produced contradictory results, indicating strong protection in some cases and no significant effect in others. Although it is commonly believed that phenolic compounds like vitamin E exert only a protective role against free radical damage, antioxidant molecules can exert other biological functions. For instance, the antioxidant activity of 17-beta-estradiol is not related to its role in determining secondary sexual characters, and the antioxidant capacity of all-trans-retinal is distinguished from its role in rhodopsin and vision. Thus, it is not unusual that alpha-tocopherol (the most active form of vitamin E) has properties independent of its antioxidant/radical scavenging ability. The Roman god Janus, shown in ancient coins as having two faces in one body, inspired the designation of 'Janus molecules' for these substances. The new biochemical face of vitamin E was first described in 1991, with an inhibitory effect on cell proliferation and protein kinase C activity. After a decade, this nonantioxidant role of vitamin E is well established, as confirmed by authoritative studies of signal transduction and gene regulation. More recently, a tocopherol binding protein with possible receptor function has been discovered. Despite such important developments in understanding the molecular mechanism and the targets of vitamin E, its new Janus face is not fully elucidated. Greater knowledge of the molecular events related to vitamin E will help in selecting the parameters for clinical intervention studies such as population type, dose response effects, and possible synergism with other compounds.

Animals↗
PubMed2000

Vitamin E reduces the uptake of oxidized LDL by inhibiting CD36 scavenger receptor expression in cultured aortic smooth muscle cells.

BACKGROUND: Vitamin E is well known as an antioxidant, and numerous studies suggest that it has a preventive role in atherosclerosis, although the mechanism of action still remains unclear. METHODS AND RESULTS: The original aim of this study was to establish whether alpha-tocopherol (the most active form of vitamin E) acts at the earliest events on the cascade of atherosclerosis progression, that of oxidized LDL (oxLDL) uptake and foam-cell formation. We show here that the CD36 scavenger receptor (a specific receptor for oxLDL) is expressed in cultured human aortic smooth muscle cells (SMCs). Treatment of SMCs and HL-60 macrophages with alpha-tocopherol (50 micromol/L, a physiological concentration) downregulates CD36 expression by reducing its promoter activity. Furthermore, we find that alpha-tocopherol treatment of SMCs leads to a reduction of oxLDL uptake. CONCLUSIONS: This study indicates that CD36 is expressed in cultured human SMCs. In these cells, CD36 transports oxLDL into the cytosol. alpha-Tocopherol inhibits oxLDL uptake by a mechanism involving downregulation of CD36 mRNA and protein expression. Therefore, the beneficial effect of alpha-tocopherol against atherosclerosis can be explained, at least in part, by its effect of lowering the uptake of oxidized lipoproteins, with consequent reduction of foam cell formation.

Aorta↗
PubMed2000

Scavenger receptor regulation and atherosclerosis.

Atherosclerosis and its complications, such as coronary heart disease, heart infarction and stroke, are the leading causes of death in the developed world. High blood pressure, diabetes, smoking and a diet high in cholesterol and lipids clearly increase the likelihood of premature atherosclerosis, albeit other factors, such as the individual genetic makeup, may play an additional role. During atherosclerosis, uncontrolled cholesterol and lipid accumulation in macrophages and smooth muscle cells leads to foam cell formation and to the progression of the atherosclerotic plaque. This review will focus on foam cell formation within the atherosclerotic lesion, the involvement of the scavenger receptor genes in this process, and the possibility to interfere with scavenger receptor function to reduce the progression of atherosclerosis. To date, the regulatory mechanisms for the expression of scavenger receptor genes and their role in atherosclerosis are not well characterized. Knowledge on this subject could lead to a better understanding of the process, prevention and therapy of this disease.

Animals↗
PubMed2000

Scavenger receptors and modified lipoproteins: fatal attractions?

Lipoproteins modified by oxidation, glycation, alkylation, and nitration are generated by oxidative stress during inflammation, diabetes, and inadequate supply of dietary antioxidants. A family of genes, the scavenger receptors, recognizes and internalizes modified lipoproteins, making them susceptible to degradation. Clearance of modified lipoproteins by scavenger receptors occurs mainly in macrophages, dendritic cells, and Kupffer cells of the liver. However, scavenger receptor expression also occurs in other cells, such as endothelial cells, aortic smooth muscle cells, neuronal cells, and keratinocytes. Thus, the local clearance of oxidized low-density lipoprotein and the resolution of inflammatory processes may rely in part on the expression of scavenger receptors in "nonprofessional" phagocytes. Uptake of oxidized low-density lipoprotein, without an efficient machinery to degrade them and uncontrolled expression of scavenger receptors, may lead to cellular deregulation, apoptosis, and formation of foam cells. Diseases accompanied by oxidation of lipoproteins, such as atherosclerosis, Alzheimer disease, glomerulosclerosis, ataxia with vitamin E deficiency, and possibly age-dependent lipofuscin deposition, may share a common pathogenetic feature. This review will focus on foam cell formation, mainly within the atherosclerotic lesion, and the possible involvement of aberrant regulation of the scavenger receptor genes. To date, the regulatory mechanisms at the basis of scavenger receptor gene expression and their roles in atherosclerosis and other diseases are not well established. Knowledge on this subject could lead to a better understanding of the pathogenesis, prevention, and therapy of these diseases.

Animals↗
PubMed2000

Specific cellular responses to alpha-tocopherol.

In the last 10 years precise cellular functions of alpha-tocopherol, some of which are independent of its antioxidant/radical-scavenging ability, have been revealed. Absorption of alpha-tocopherol from the gut is a selective process. Other tocopherols are not absorbed or are absorbed to a lesser extent. At the post-translational level, alpha-tocopherol inhibits protein kinase C and 5-lipoxygenase and activates protein phosphatase 2A and diacylglycerol kinase. Some genes [platelet glycoprotein IV/thrombospondin receptor/class B scavenger receptor (CD36), alpha-tocopherol transfer protein (alpha-TTP), alpha-tropomyosin, connective tissue growth factor and collagenase] are affected by alpha-tocopherol at the transcriptional level. alpha-Tocopherol also inhibits cell proliferation, platelet aggregation, monocyte adhesion and the oxygen burst in neutrophils. Other antioxidants, such as beta-tocopherol and probucol, do not mimic these effects, suggesting a nonantioxidant, alpha-tocopherol-specific molecular mechanism.

Animals↗
PubMed1999

Age-dependent increase of collagenase expression can be reduced by alpha-tocopherol via protein kinase C inhibition.

Total protein kinase C (PKC) activity in human skin fibroblasts increases during in vivo aging as a function of the donor's age. During in vitro aging protein kinase C activity is also increased, as a function of cell passage number. Using PKC isoform specific antibodies, we demonstrate that the increase in total PKC activity is mainly due to the PKC a isoform. PKC alpha protein expression increased up to 8 fold during in vivo aging. Collagenase (MMP-1) gene transcription and protein expression also increased with age, concomitant with the increase in protein kinase C alpha. Furthermore, alpha-tocopherol, which inhibits protein kinase C activity, is able to diminish collagenase gene transcription without altering the level of its natural inhibitor, tissue inhibitor of metalloproteinase, TIMP-1. We propose that an aging program leads to increased protein kinase C alpha expression and activity. This event would induce collagenase overexpression followed by increased collagen degradation. Our in vitro experiments with skin fibroblasts suggest that alpha-tocopherol may protect against skin aging by decreasing the level of collagenase expression, which is induced by environmental insults and by aging.

Adult↗
PubMed1998

Enzyme-mediated cytosine deamination by the bacterial methyltransferase M.MspI.

Most prokaryotic (cytosine-5)-DNA methyltransferases increase the frequency of deamination at the cytosine targeted for methylation in vitro in the absence of the cofactor S-adenosylmethionine (AdoMet) or the reaction product S-adenosylhomocysteine (AdoHcy). We show here that, under the same in vitro conditions, the prokaryotic methyltransferase, M.MspI (from Moraxella sp.), causes very few cytosine deaminations, suggesting a mechanism in which M.MspI may avoid enzyme-mediated cytosine deamination. Two analogues of AdoMet, sinefungin and 5'-amino-5'-deoxyadenosine, greatly increased the frequency of cytosine deamination mediated by M.MspI presumably by introducing a proton-donating amino group into the catalytic centre, thus facilitating the formation of an unstable enzyme-dihydrocytosine intermediate and hydrolytic deamination. Interestingly, two naturally occurring analogues, adenosine and 5'-methylthio-5'-deoxyadenosine, which do not contain a proton-donating amino group, also weakly increased the deamination frequency by M.MspI, even in the presence of AdoMet or AdoHcy. These analogues may trigger a conformational change in the enzyme without completely inhibiting the access of solvent water to the catalytic centre, thus allowing hydrolytic deamination of the enzyme-dihydrocytosine intermediate. Under normal physiological conditions the enzymes M.HpaII (from Haemophilus parainfluenzae), M. HhaI (from Haemophilus hemolytica) and M.MspI all increased the in vivo deamination frequency at the target cytosines with comparable efficiency.

Adenosine↗
PubMed1998

DNA methylation as a target for drug design.

DNA methylation is essential for normal embryonic development. Distinctive genomic methylation patterns must be formed and maintained with high fidelity to ensure the inactivities of specific promoters during development. The mutagenic and epigenetic aspects of DNA methylation are especially interesting because they may lead to the inactivation of genes which are involved in human carcinogenesis. The mutagenicity of 5-Methylcytosine (5mC) and the role of promoter hypermethylation in gene silencing, particularly in cancer, suggest a clinical significance for the design of novel DNA methylation inhibitors which may be utilized to reverse the effects of DNA methylation.

Animals↗
PubMed1997

Identification and characterization of differentially methylated regions of genomic DNA by methylation-sensitive arbitrarily primed PCR.

We have developed a simple and reproducible fingerprinting method for screening the genome for regions of DNA that have altered patterns of DNA methylation associated with oncogenic transformation. Restriction enzymes with different sensitivities to cytosine methylation in their recognition sites were used to digest genomic DNAs from primary tumors, cell lines, and normal tissues prior to arbitrarily primed PCR amplification. Fragments that showed differential methylation were cloned and sequenced after resolving the PCR products on high-resolution polyacrylamide gels. The cloned fragments were then used as probes for Southern analysis to confirm differential methylation of these regions in colon tissues and cell lines. Forty-four DNA fragments associated with a total of five different regions of genomic DNA containing methylation sites were detected in 10 matched sets of normal and tumor colon DNAs and 7 colon cancer cell lines. A novel CpG island was also isolated that was found to be frequently hypermethylated in bladder and colon tumors. We have demonstrated that this technique is a rapid and efficient method that can be used to screen for altered methylation patterns in genomic DNA and to isolate specific sequences associated with these changes.

Blotting, Southern↗
PubMed1996

Methylation inhibitors can increase the rate of cytosine deamination by (cytosine-5)-DNA methyltransferase.

The target cytosines of (cytosine-5)-DNA methyltransferases in prokaryotic and eukaryotic DNA show increased rates of C-->T transition mutations compared to non-target cytosines. These mutations are induced either by the spontaneous deamination of 5-mC-->T generating inefficiently repaired G:T rather than G:U mismatches, or by the enzyme-induced C-->U deamination which occurs under conditions of reduced levels of S-adenosylmethionine (AdoMet) and S-adenosylhomocysteine (AdoHcy). We tested whether various inhibitors of (cytosine-5)-DNA methyltransferases analogous to AdoMet and AdoHcy would affect the rate of enzyme-induced deamination of the target cytosine by M.HpaII and M.SssI. Interestingly, we found two compounds, sinefungin and 5'-amino-5'-deoxyadenosine, that increased the rate of deamination 10(3)-fold in the presence and 10(4)-fold in the absence of AdoMet and AdoHcy. We have therefore identified the first mutagenic compounds specific for the target sites of (cytosine-5)-DNA methyltransferases. A number of analogs of AdoMet and AdoHcy have been considered as possible antiviral, anticancer, antifungal and antiparasitic agents. Our findings show that chemotherapeutic agents with affinities to the cofactor binding pocket of (cytosine-5)-DNA methyltransferase should be tested for their potential mutagenic effects.

Bacteria↗
PubMed1996

The rate of CpG mutation in Alu repetitive elements within the p53 tumor suppressor gene in the primate germline.

Cytosine to thymine transition mutations at the CpG dinucleotide are the most common point mutations in cancer and genetic disease. We calculated the in vivo rate of CpG mutation in the primate germline by deriving a primordial consensus sequence for an Alu repetitive element which inserted into intron 6 of the primate p53 gene 35 to 55 million years ago. Comparison of this primordial sequence to the Alu sequence in intron 6 of present-day primates was used to determine the nature and rate of mutations which occurred during evolution. We estimate the half-life of a CpG nucleotide to be 24 to 60 million years, and the rate constant for mutation at this dinucleotide to be 1.2 x 1O(-8) to 2.9 x 1O(-8) years(-1). These results were confirmed by the analysis of a second Alu sequence in intron 10 of the p53 gene. The in vivo mutation rate is at least 1250-fold slower than the in vitro chemical rate of 5-methylcytosine deamination in double-stranded DNA, showing that current estimates of CpG mutation repair have been significantly underestimated. Furthermore, the mutability of the CpG dinucleotide has led to the depletion of this dinucleotide from the vertebrate genome, and calculations in this study suggest that current levels of the CpG dinucleotide in the primate genome are very close to a steady state equilibrium in which the rate of CpG mutation is equal to the rate of CpG formation by random mutation.

Animals↗
PubMed1995

A mutant HpaII methyltransferase functions as a mutator enzyme.

DNA (cytosine-5)-methyltransferases can cause deamination of cytosine when the cofactor S-adenosylmethionine (AdoMet) is limiting and thus function as sequence-specific C-->U mutator enzymes. Here we explored whether mutations causing inactivation of the cofactor binding activity of the HpaII methyltransferase, thus mimicking conditions of limiting AdoMet concentration, could convert a DNA methyltransferase to a C-->U mutator enzyme. We created two mutator enzymes from the HpaII methyltransferase (F38S and G40D) which both showed enhanced cytosine deamination activities in vitro and in vivo. Interestingly, the G:U mispairs generated by these enzymes were not repaired completely in bacteria equipped with uracil-DNA glycosylase-initiated repair machinery, giving rise to a potent mutator phenotype. This is the first report showing the creation of mutator enzymes from a DNA methyltransferase and the demonstration of their mutagenicity in living cells.

Base Sequence↗
PubMed1995

HhaI and HpaII DNA methyltransferases bind DNA mismatches, methylate uracil and block DNA repair.

The hydrolytic deamination of 5-methylcytosine (5-mC) to thymine (T) is believed to be responsible for the high mutability of the CpG dinucleotide in DNA. We have shown a possible alternate mechanism for mutagenesis at CpG in which HpaII DNA-(cytosine-5) methyltransferase (M.HpaII) can enzymatically deaminate cytosine (C) to uracil (U) in DNA [Shen, J.-C., Rideout, W.M., III and Jones, P.A., Cell, 71, 1073-1080, (1992)]. Both the hydrolytic deamination of 5-mC and enzymatic deamination of C create premutagenic DNA mismatches (G:U and G:T) with the guanine (G) originally paired to the normal C. Surprisingly, we found that DNA-(cytosine-5) methyltransferases have higher affinities for these DNA mismatches than for their normal G:C targets and are capable of transferring a methyl group to the 5-position of U, creating T at low efficiencies. This binding by methyltransferase to mismatches at the recognition site prevented repair of G:U mismatches by uracil DNA glycosylase in vitro.

Base Sequence↗
PubMed1994

MyoD1 promoter autoregulation is mediated by two proximal E-boxes.

We show that in mouse myoblasts the MyoD1 promoter is highly stimulated by MyoD1 expression, suggesting that it is controlled by a positive feedback loop. Using deletion and mutation analyses, we identified the targets for MyoD1 promoter autoregulation as the two proximal E-boxes located close to the MyoD1 core promoter. Gel mobility shift competition assays with MyoD1 antibodies as competitor suggest that the MyoD1 protein is binding directly to these E-boxes. Autoregulation did not occur in fibroblasts cotransfected with the expression vector of MyoD1. It is assumed that autoregulation is controlled by the stoichiometry between the MyoD1 protein and negatively regulatory proteins like Id, which is known to be highly expressed in fibroblasts. When the MyoD1 promoter was methylated, autoregulation only occurred when the density of methylated sites was low. The density of DNA methylation, therefore, can determine the accessibility of the MyoD1 promoter to transcription factors and interfere with the auto- and crossregulatory loop. The MyoD1 promoter in vivo was found to be only partially methylated in all tissues tested except in skeletal muscle where it was demethylated. We propose that high level expression of the MyoD1 gene is a result of release from constraints such as negative regulatory factors and/or DNA methylation interfering with MyoD1 autoregulation.

Animals↗
PubMed1994

AP-1 binds to a putative cAMP response element of the MyoD1 promoter and negatively modulates MyoD1 expression in dividing myoblasts.

We have studied the transcriptional activity of the mouse MyoD1 gene promoter in vivo and in vitro using mouse G8 myoblasts and muscle cell nuclear extracts. 5' deletion analysis of the promoter and transcription-competition analysis using oligonucleotides corresponding to several cis-acting elements revealed that the basal activity of the MyoD1 promoter is conferred by two SP1 boxes, an AP-2 box, and a CAAT box. We have identified a negative regulatory sequence located between nucleotide position -342 to -322 with respect to the cap site. The negative regulatory element shows sequence homology with cAMP-responsive element (CRE) and AP-1 binding site (5'-GAGCACTGAGGTCAGTACAG-3'). As determined by gel mobility shift competition analysis, oligonucleotides containing AP-1 binding sites inhibit protein interactions with the MyoD1 CRE-like element. We also show that binding to this element is down-regulated during myogenic differentiation and can be reinduced by the addition of serum. Furthermore, mutation of the CRE-like element induces MyoD promoter activity in diving myoblasts. By using anti-c-Fos antibodies we show that AP-1 is binding to the MyoD1 CRE-like element. Our results indicate that AP-1 negatively modulates MyoD1 expression in growing myoblasts and strongly suggest that c-Fos and c-Jun inhibit myogenesis and MyoD1 expression by direct binding to a negative cis-acting element in the MyoD1 promoter.

Animals↗