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Biomedical subjects

A Azzi

Publications and source records attributed to A Azzi.

At least 73 records · Page 4Linked to original sources
PubMed1997

Biogenic 4-hydroxy-2-nonenal activates transcription factor AP-1 but not NF-kappa B in cells of the macrophage lineage.

A large spectrum of pro-oxidant agents, including molecules with lipoperoxidative effect, can modulate gene expression through modification of the DNA binding activity of the transcription factors activator protein 1 (AP-1) and nuclear factor kappa B (NF-kappa B). In this study the effect on these redox-sensitive factors by 4-hydroxy-2-nonenal (HNE), a major aldehydic product of lipid peroxidation, was examined in two cell lines of the macrophage type. Incubation in the presence of microM concentrations of the aldehyde led to a rapid increase of AP-1 binding with a transient maximum 30 min from HNE addition to the culture medium in both cell lines. On the contrary, HNE did not stimulate nuclear translocation of NF-kappa B. The diverging effect of HNE on the two transcription factors is likely related to the demonstrated differential activation pathway of AP-1 and NF-kappa B in macrophages. The HNE-induced activation of AP-1 suggests the aldehyde's involvement in the regulatory mechanisms of cell proliferation and differentiation.

Aldehydes↗
PubMed1997

Alpha-tocopherol as a modulator of smooth muscle cell proliferation.

The effects of alpha-tocopherol and beta-tocopherol have been studied in rat and human aortic smooth muscle cells. Alpha-tocopherol, but not beta-tocopherol, inhibited smooth muscle cell proliferation and protein kinase C in a dose-dependent manner, at concentrations ranging from 10 to 50 microM. Beta-tocopherol added simultaneously with alpha-tocopherol prevented both proliferation and protein kinase C inhibition. Protein kinase C inhibition was cell cycle-dependent and it was prevented by okadaic acid, a protein phosphatase inhibitor. Protein kinase C activity measured from aortas of cholesterol-fed rabbits was also inhibited by alpha-tocopherol. By using protein kinase C (PKC) isoform-specific inhibitors and immunoprecipitation reactions it was found that PKC-alpha was selectively inhibited by alpha-tocopherol. Further, an activation of protein phosphatase 2A by alpha-tocopherol was found, which caused PKC-alpha dephosphorylation and inhibition. Ultimately, this cascade of events at the level of cell signal transduction leads to the inhibition of smooth muscle cell proliferation.

Animals↗
PubMed1997

Transmission of parvovirus B19 by coagulation factor concentrates exposed to 100 degrees C heat after lyophilization.

BACKGROUND: Double inactivation by solvent/detergent treatment plus heating at 100 degrees C for 30 minutes after lyophilization has been adopted to improve viral safety of factor VIII and factor IX concentrates, particularly with respect to non-lipid-enveloped viruses. The aim of this study was to evaluate the safety of concentrates exposed to these virucidal methods. STUDY DESIGN AND METHODS: Twenty-six previously untreated hemophiliacs, 19 with factor VIII deficiency and 7 with factor IX deficiency, were investigated in a prospective multicenter study over a 12-month follow-up period by the use of serologic and virologic markers for lipid- and non-lipid-enveloped viruses (human immunodeficiency virus types 1 and 2; hepatitis A, B, and C viruses; B19 parvovirus antibodies; and B19 DNA). Overall, 270,000 U of factor VIII and 102,000 U of factor IX concentrate were administered during the study period. RESULTS: None of the 26 patients seroconverted for human immunodeficiency virus or hepatitis C virus. Hepatitis B virus markers remained negative in the 10 unvaccinated hemophiliacs. No hepatitis A virus seroconversion occurred among 17 susceptible patients. B19 seroconversion (IgM) and B19 viremia were observed within 2 weeks of the first concentrate infusion in 8 of 15 susceptible patients, 5 of 11 treated with factor VIII and 3 of 4 with factor IX concentrate. CONCLUSION: This prospective study indicates that very high temperatures applied to lyophilized concentrates appear to prevent the transmission of hepatitis A virus to hemophiliacs. However, B19 parvovirus still contaminates concentrates despite the use of this robust virucidal method.

Adolescent↗
PubMed1997

The effect of alpha-tocopherol and beta-tocopherol on proliferation, protein kinase C activity and gene expression in different cell lines.

alpha-Tocopherol, but not beta-tocopherol, negatively regulates proliferation of A7r5 vascular smooth muscle cells at physiological concentration. The HeLa cell line was not affected whereas the Chinese hamster ovary cell line (CHO) was slightly inhibited by both alpha-tocopherol and beta-tocopherol. In A7r5 cells alpha-tocopherol inhibited protein kinase C activity, and this correlated with inhibition of proliferation. beta-Tocopherol did not inhibit either protein kinase C or proliferation. In HeLa cells no inhibition by alpha-tocopherol or beta-tocopherol of protein kinase C activity and cell proliferation was observed. In Chinese hamster ovary cells both tocopherols inhibited protein kinase C activity but not proliferation. Thus in the latter cells proliferation was not protein kinase C-dependent. In A7r5 cells alpha-tocopherol but not beta-tocopherol activated AP-1-mediated gene expression. In HeLa cells no change in gene expression was observed in agreement with the finding that also protein kinase C was not affected. In CHO cells gene-expression was activated by both alpha-tocopherol and beta-tocopherol. In this case also a positive correlation was found with similar inhibition of protein kinase C activity. In these cells, however, the changes at the level of protein kinase C activity and gene expression did not result in proliferation changes. The effect of alpha-tocopherol and beta-tocopherol on protein kinase C activity and gene expression suggest a cause-to-effect relationship. Inhibition of proliferation, however, correlates in the case of A7r5 and HeLa cells but not in the case of CHO suggesting a different proliferation pathway for these cells.

Animals↗
PubMed1997

Aldose reductase induction: a novel response to oxidative stress of smooth muscle cells.

Hydrogen peroxide (H2O2) or 4-hydroxy-2,3-trans-nonenal (HNE) treatment of rat vascular smooth muscle cells (A7r5) caused induction of aldose reductase mRNA. Induction was dose (10-100 microM H2O2, 1-10 microM HNE) and time dependent, reaching a maximum (three- to fourfold) after 7-12 h. Treatment of cells with actinomycin D confirmed de novo synthesis of aldose reductase mRNA. H2O2-induced expression was prevented by catalase but unaffected by Desferal, indicating that metal catalyzed degradation of peroxide was not involved. Induction of enzymatically active aldose reductase by H2O2 and HNE was confirmed using Western blotting and enzyme assays. Aldose reductase can metabolize several aldehyde compounds including HNE, a major toxic product of lipid peroxidation. Inclusion of Sorbinil, an aldose reductase inhibitor, in toxicity assays resulted in a significant (twofold) enhancement of HNE-mediated killing of A7r5 cells, suggesting a protective role of aldose reductase against HNE-induced cell death. These data indicate that the induction of aldose reductase during oxidative stress might represent an important cellular antioxidant defense mechanism.

Aldehyde Reductase↗
PubMed1997

Signalling functions of alpha-tocopherol in smooth muscle cells.

alpha-Tocopherol but not beta-tocopherol, activates protein phosphatase 2A, decreases protein kinase C activity and attenuates smooth muscle cell proliferation at physiological concentrations. beta-Tocopherol prevents the effects of alpha-tocopherol. Inhibition of protein kinase C alpha, but not of the other isoforms, by the inhibitor Gö6976 prevents the effect of alpha-tocopherol. Protein kinase C alpha, immunoprecipitated from alpha-tocopherol treated cells, is less phosphorylated and inactive. It is proposed that the specific activation of protein phosphatase 2A by alpha-tocopherol results in dephosphorylation and inactivation of protein kinase C alpha. Finally, this cascade of events leads to smooth muscle cell proliferation inhibition.

Animals↗
PubMed1996

Dietary cholesterol-induced changes of protein kinase C and the effect of vitamin E in rabbit aortic smooth muscle cells.

The changes occuring in smooth muscle cells during the development of atherosclerosis in rabbits fed 2% cholesterol and the effect of vitamin E treatment were investigated. Ex-vivo smooth muscle cells obtained from the aorta of cholesterol-fed rabbits exhibited a 2-fold increase of protein kinase C expression and activity. The cholesterol induced changes in protein kinase C were equally present in the membrane bound and cytosolic fraction of the enzyme. The amount of a control protein alpha-actin was not affected in smooth muscle cell by the high cholesterol diet treatment, indicating that protein kinase C increase was specific. The increase of protein kinase C expression and activity was not significantly affected by vitamin E treatment although a constant trend was noted. The data are discussed in the light of previous smooth muscle cell in vitro experiments.

Animals↗
PubMed1996

High rates of extracellular superoxide generation by cultured human fibroblasts: involvement of a lipid-metabolizing enzyme.

Expression of NADPH oxidase and low superoxide generation (approx. 0.06 nmol/min per 10(6) cells) by cytokine- or ionophore-stimulated human fibroblasts is known. However, we here show that these cells also contain an ectoplasmic enzyme, distinct from NADPH oxidase, which can generate superoxide (2.19 +/- 0.14 nmol/min per 10(6) cells) at levels similar to phorbol ester-stimulated monocytes on exogenous NADH addition. Superoxide generation was temperature-dependent, insensitive to chelation (desferal), and had a K(m) (app)(NADH) of 11.5 microM. Inhibitor studies showed that there was no involvement of NADPH oxidase (diphenylene iodonium, diphenyl iodonium), prostaglandin H synthase (indomethacin), xanthine oxidase (allopurinol), cytochrome P-450 (metyrapone) or mitochondrial respiration (rotenone, antimycin A). NAD+ was a competitive inhibitor, whereas NADPH supported 40% of the rate seen with NADH. No luminescence was observed after the addition of lactate, malate, pyruvate, GSH or L-cysteine. NADH-stimulated superoxide generation was enhanced by the addition of (3-30 microM) arachidonic acid, linoleic acid or (5S)-hydroxyeicosatetraenoic acid [(5S)-HETE] but not palmitic acid, (15S)-hydroperoxyeicosatetraenoic acid [(15S)-HPETE], (15S)-HETE or (12S)-HETE. Several features suggest involvement of an enzyme related to 15-lipoxygenase, and, in support of this, we show superoxide generation and NADH oxidation by recombinant rabbit reticulocyte 15-lipoxygenase. The large amounts of superoxide measured suggest that the fibroblast extracellular enzyme could be a major source of reactive oxygen species after tissue damage.

Animals↗
PubMed1996

4-hydroxy-2,3-trans-nonenal induces transcription and expression of aldose reductase.

Treatment of A7r5 rat vascular smooth muscle cells with 4-hydroxy-2,3-trans-nonenal (HNE) resulted in increased aldose reductase mRNA transcription. Induction was time dependent, reaching a maximum (2,3-fold) after 7 hours. An enzymatically active aldose reductase analysed by Western blotting and enzyme assays was expressed. Sorbinil, an aldose reductase inhibitor, induced a significant enhancement of HNE cytotoxicity, indicating a protective role of aldose reductase against HNE-induced A7r5 cell death. These data indicate that induction of aldose reductase by HNE may represent an important cellular defence mechanism against oxidative injury.

Aldehyde Reductase↗
PubMed1996

Prevention of necrosis and activation of apoptosis in oxidatively injured human myeloid leukemia U937 cells.

A 3 h exposure to 1 mM H2O2 followed by 6 h post-challenge growth in peroxide-free medium induces necrosis in U937 cells. Addition of the poly(ADP-ribose)polymerase inhibitor 3-aminobenzamide during recovery prevents necrosis and triggers apoptosis, as shown by the appearance of apoptotic bodies, extensive blebbing and formation of multimeric DNA fragments as well as 50 kb double stranded DNA fragments. Thus, the same initial damage can be a triggering event for both apoptotic and necrotic cell death. Furthermore, necrosis does not appear to be a passive response to overwhelming damage.

Apoptosis↗
PubMed1996

The phosphorylation state of MAP-kinases modulates the cytotoxic response of smooth muscle cells to hydrogen peroxide.

Micromolar concentrations of hydrogen peroxide induced the phosphorylation of mitogen-activated protein (MAP) kinases and a lethal response in growth-arrested smooth muscle cells (A7r5). The H202-induced phosphorylation of MAP-kinases was markedly lower in the presence of protein tyrosine kinase (PTK) inhibitors or in protein kinase C (PKC) down-regulated cells. Similarly, the toxicity of H202 was diminished by concomitant addition of either PKC or PTK inhibitors and was also lower in PKC down-regulated cells. These results are consistent with the possibility that phosphorylation of MAP-kinases is a critical event in the toxic response of cultured smooth muscle cells to H202.

Animals↗
PubMed1996

Duplex polymerase chain reaction for the simultaneous detection of the human polyomavirus BK and JC DNA.

With the aim of further simplifying the detection of human polyomaviruses BK (BKV) and JC (JCV), by reducing the time and the cost of the reactions, a duplex polymerase chain reaction (PCR) was developed for the simultaneous detection and identification of BKV and JCV DNA. A nested PCR was performed. In a first amplification reaction the same set of primers was used to amplify a sequence of either BKV or JCV genomes. Two sets of primers, each specific for BKV or JCV DNA, were used in a second amplification. The products of BKV and JCV DNA amplification were distinguishable on the basis of their different size. The sensitivity and the specificity of the duplex PCR was shown to be exactly comparable to that of the single PCR, not only with the BKV and the JCV plasmid but also for BKV and JCV DNA detection in clinical specimens.

BK Virus↗
PubMed1996

Human parvovirus B19 infection within a family and risk for pregnant women.

During an outbreak of parvovirus B19 infection among four related families at least 70% of the household contacts, including a woman at the 33rd week of pregnancy, became infected. Twins were born at the 39th week of pregnancy, both with B19 infection. B19 DNA was detected in their sera by a nested PCR, anti-B19 IgM was detectable only by an immunofluorescence assay, and low levels of maternal anti-B19 IgG were demonstrable by an immunoenzymatic test in the serum of both children. All the haematological parameters were normal at birth and 6 months later, when B19 DNA and anti-B19 antibody were no longer detectable in serum samples. This observation emphasizes the high risk of B19 infection among household contacts and the possibility of a favourable outcome of the foetal infection, possibly related to infection late in the pregnancy.

Adult↗