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Biomedical subjects

I Emerit

Publications and source records attributed to I Emerit.

At least 37 records · Page 2Linked to original sources
PubMed1989

Mutagenic effects of TPA-induced clastogenic factor in Chinese hamster cells.

Previous work in our laboratory has shown that the clastogenic and SCE-inducing effects of 12-O-tetradecanoylphorbol 12-acetate (TPA) are mediated by secondary products formed by the cell in response to the tumor promoter. A low-molecular-weight clastogenic factor (CF) was isolated from the medium of TPA-treated human leukocytes and caused chromosome aberrations and sister-chromatid exchanges (SCE) in fresh cultures not exposed to TPA itself. In the present study, we show that Chinese hamster fibroblasts (V79 cells) also produce CF when exposed to TPA. CF from V79 cells induced SCE not only in hamster cells, but also in human lymphocytes. Vice versa, CF from human leukocyte cultures induced SCE in hamster cells. It also increased the frequency of 6-thioguanine-resistant mutants in this cell system. All cyto- and geno-toxic effects of TPA-induced CF were prevented if the cells were treated with superoxide dismutase before exposure. The lipophilic CF seems to be derived from arachidonic acid of cell membranes released as a consequence of oxidative damage and subsequently degraded to genotoxic aldehydes in an autoxidative process. CF is formed only under culture conditions with low antioxidant content in culture media and sera. This may explain the discordant results obtained by different laboratories with regard to the genotoxic effects of TPA.

Animals↗
PubMed1989

Monocyte-derived clastogenic factor in rheumatoid arthritis.

Blood or lymphocyte cultures from patients with rheumatoid arthritis show increased chromosome breakage. This is due to the presence of a clastogenic factor (CF) inducing also chromosome damage in blood cultures of healthy persons. CF may be isolated not only from patients' plasma or synovial fluid, but also from the supernatant of blood or lymphocyte cultures. No CF was detectable, if the lymphocyte cultures were free of other contaminating blood cells. Addition of neutrophils did not considerably influence the production of CF, and platelets were without any effect. However, addition of increasing numbers of monocytes resulted in increasing clastogenic activity. Also monocytes in adherence, in absence of lymphocytes and without any chemical stimulant, produced CF. This indicates that monocytes are responsible for CF production. The protective effect of superoxide dismutase, as well against CF formation as against CF action on cells of normal subjects, suggests a role of the superoxide radical O2-. Inhibitors of arachidonic acid metabolism were only slightly anticlastogenic.

Arthritis, Rheumatoid↗
PubMed1989

2-(2-hydroxy-4-methylphenyl)aminothiazole hydrochloride as a dual inhibitor of cyclooxygenase/lipoxygenase and a free radical scavenger. 1st communication: in vitro studies.

2-(2-Hydroxy-4-methylphenyl)aminothiazole hydrochloride (CBS-113 A) inhibits cyclooxygenase in platelets and 5-lipoxygenase in leukocytes at micromolar concentrations. On the other hand the drug is a potent scavenger of oxygen-derived free radicals. Moreover and possibly consequently, CBS-113 A inhibits the oxygen burst of stimulated leukocytes and the release of an interleukin 1-like compound from vascular endothelial cells in culture. These properties could lead to another activity of therapeutic interest in comparison with anti-inflammatory drugs already available.

Animals↗
PubMed1988

Clastogenic factor in ischemia-reperfusion injury during open-heart surgery: protective effect of allopurinol.

The hypothesis tested was that free radicals generated following ischemia and reperfusion in cardiac operations can produce clastogenic factor that results in chromosomal aberration. Fourteen randomized patients undergoing coronary artery bypass grafting were divided into two groups. In Group 1 (7 patients), myocardial protection was achieved using a cardioplegic solution without allopurinol. In Group 2 (7 patients), 100 mg of allopurinol (xanthine oxidase inhibitor) was added to the solution. In both groups, blood samples were taken from the coronary sinus before the aorta was clamped and 20 minutes after myocardial reperfusion was achieved. The blood samples were used to study the patients' chromosomes. The results were given as the percentage of chromosomal aberrations observed in 100 mitoses. There were no significant differences between the preischemic values in both groups and the postischemic values in Group 2. On the other hand, there was a significant difference between the postischemic values in Groups 1 and 2 (p less than 0.01). In conclusion, reperfusion following myocardial ischemia in cardiac operations can produce clastogenic aberrations. This clastogenic activity can be reduced by adding allopurinol to the cardioplegic solution.

Allopurinol↗
PubMed1987

Phorbol ester-induced formation of clastogenic factor from human monocytes.

Phorbol-12-myristate-13-acetate (PMA) acts as a tumor promoter on mouse skin. It induces inflammation and leukocyte-mediated clastogenicity which appears to be related to rapid changes in lipid metabolism. To identify lipids possessing clastogenic and/or tumor-promoting properties, we have characterized the metabolism and release of arachidonic acid (AA) and related lipids during the formation of lipophilic clastogenic factors by PMA-treated human monocytes. In 1 h, [3H]AA-labeled monocytes spontaneously released significant amounts of their total radioactivity (4%) which increased nearly 4-fold (15%) with PMA (30 ng/ml) treatment. Eighty-five per cent of extracellular 3H-label from both control and PMA-treated monocytes was composed of free AA (plus AA-metabolites), while the remaining radioactivity was incorporated in phospholipids and mono- and diacylglycerols. Treated and non-treated cells released essentially the same kind of metabolites but PMA induced a 3- to 4-fold increase in total amounts. The major products consisted of prostaglandins F2 alpha and E2, thromboxane B2, 12-hydroxy-5,8,10-heptadecatrienoic acid and 5-, 11- and 15-hydroxyeicosatetraenoic acids. PMA also induced increases in the levels of three unidentified products. Neither leukotrienes nor 4-hydroxynonenal, a major alkenal degradation product of AA, were found in medium from PMA-treated monocytes. PMA, in contrast to the first-stage tumor promoter calcium ionophore A23187, failed to stimulate the release of platelet activating factor. The increased formation of phorbol ester-induced AA metabolites was proportional to the increase in free extracellular AA. The source of AA from treated and untreated monocytes consisted of cellular phospholipids with phosphatidylcholine and phosphatidylethanolamine accounting for 85%.

Arachidonic Acid↗
PubMed1985

Treatment of lymphocyte cultures with a hypoxanthine-xanthine oxidase system induces the formation of transferable clastogenic material.

Culture medium of lymphocyte cultures that have been exposed to the superoxide generating system hypoxanthine plus xanthine oxidase (X-XO) contains substances with chromosome damaging properties. This is demonstrated by the ability of ultrafiltrates of such culture media to induce chromosomal aberrations and sister chromatid exchanges in the lymphocytes of blood test cultures. Culture medium becomes active about 15 hours after the addition of X-XO and stimulation by phytohemagglutinin. Concomitant with the accumulation of clastogenic material, assays for conjugated dienes and thiobarbituric acid-reactive material which measure lipid-peroxidation become positive in the culture media. When cells are pretreated with superoxide dismutase or glutathione peroxidase before the addition of X-XO neither clastogenic substances nor lipid peroxidation products are detected. Catalase is a less efficient protector.

Arachidonic Acid↗
PubMed1985

Lipid peroxidation products and clastogenic material in culture media of human leukocytes exposed to the tumor promoter phorbol-myristate-acetate.

The chromosome-damaging effect of PMA in blood cultures is mediated by secondary products which are formed by the cells in response to the interaction with this tumor promoter. Since this effect could be influenced by antioxidant enzymes and by inhibitors of arachidonic acid metabolism, the present study was undertaken in order to determine whether the formation of these clastogenic substances is concomitant with the formation of AA metabolites and other lipid peroxidation products. Besides the clastogenic effect of ethyl-acetate extracts, the similarities of cytogenetic and biochemical results (conjugated dienes and TBA-reactive material) obtained for the influence of other blood cells than lymphocytes in the culture system, the importance of PHA stimulation and the protective effect of antioxidant enzymes were arguments in favour of a causal relationship between chromosome damage and lipid peroxidation (enzymatic or nonenzymatic). If AA release from membrane phospholipids was prevented by inhibition of phospholipase A2, neither conjugated dienes nor TBA-reactive material were found, and chromosome damage was reduced considerably. However, the results obtained with inhibitors of the cyclo- and lipoxygenase pathway were not conclusive, and discrepancies were also observed in the time course of appearance of clastogenic material and lipid peroxidation products.

Arachidonic Acid↗
PubMed1984

Evidence for membrane-mediated chromosomal damage by aflatoxin B1 in human lymphocytes.

The hepatocarcinogen aflatoxin B1 (AFB1) was found to be a potent clastogen for phytohemagglutinin stimulated human lymphocytes. It also induced sister chromatid exchanges. These types of chromosomal damage were induced at very low levels of covalent AFB1 - DNA adducts suggesting that AFB1 operates in part by indirect action because of its membrane-active character. The membrane-active character of AFB1 is documented by the following results: (i) AFB1 stimulated the excretion of hydroxy- and/or hydroperoxy-arachidonic acid (AA) and free AA into the culture medium; (ii) the phospholipase A2 inhibitor p-bromophenacylbromide was anticlastogenic ; (iii) the inhibitors of the oxidative metabolism of AA indomethacin, flufenamic acid, 5,8,11,14-eicosatetraynoic acid, nordihydroguaiaretic acid and BN 1015 were anticlastogenic . These results are compatible with the induction of DNA damage by indirect action or the formation of covalent adducts via metabolic activation by cooxygenation . The observation that CuZn superoxide dismutase was anticlastogenic indicates the intermediacy of superoxide in DNA damage formation and supports the former mechanism.

Aflatoxin B1↗
PubMed1983

Suppression of tumor promoter phorbolmyristate acetate-induced chromosome breakage by antioxidants and inhibitors of arachidonic acid metabolism.

To gain insight into the mechanism of formation of chromosomal aberrations by the tumor promoter phorbolmyristate acetate (PMA) in human lymphocytes, we investigated the effect of antioxidants and inhibitors of arachidonic acid metabolism. Among the antioxidants bovine erythrocyte CuZn superoxide dismutase, glutathione peroxidase, mannitol (a scavenger of hydroxyl radicals), butylated hydroxytoluene and butylated hydroxyanisole were anticlastogenic while catalase and dimethylfuran (a scavenger of singlet oxygen) were inactive. These results show that the induction of aberrations by PMA occurs via indirect action, i.e. the intermediacy of superoxide and hydroxyl radicals. The following inhibitors of arachidonic acid metabolism were strongly anticlastogenic: the cyclo-oxygenase inhibitors indomethacin and flufenamic acid and the lipoxygenase inhibitor BN1015. Imidazole, nordihydroguaiaretic acid BN1048 and 5,8,11,14-eicosatetraynoic acid were moderately active. The inhibitor of phospholipase A2, fluocinolone acetonide, was also anticlastogenic. We conclude that the oxidative metabolism of arachidonic acid is involved in the induction of chromosomal aberrations by PMA in human lymphocytes. However, because of the limited selectivity of these drugs, it is not yet possible to identify unambiguously the step(s) in the arachidonic acid cascade responsible for PMA clastogenicity.

Anti-Inflammatory Agents↗
PubMed1983

Variation of superoxide dismutase levels in fetal calf serum.

The results of cytogenetic studies and of other experiments based on tissue-culture systems may be influenced by various components of tissue-culture medium and by variations among batches of fetal calf serum used for supplementation of the media. Negative results may be obtained in breakage studies as a consequence of medium components with a protective effect [6]. Attention has been drawn to differences in growth pattern [8] and mitotic indices [11] in lymphocyte cultures set up with different culture media. Variations in the incidence of sister-chromatid exchanges according to differences in media [7] and sera [5] have also been observed. It has been suggested [9] that the failure of some laboratories to detect increases in sister-chromatid exchanges after treatment with the tumor promoter phorbol-myristate-acetate (PMA) may be due to high concentrations of the free-radical-scavenging enzyme superoxide dismutase (SOD) in the sera used and that heat inactivation of the sera may be responsible for these differences. In the following, we report that considerable variation in the SOD content exists between batches of fetal calf serum, up to levels with anticlastogenic effect.

Animals↗
PubMed1983

Clastogenic action of tumor promoter phorbol-12-myristate-13 acetate in mixed human leukocyte cultures.

The tumor promoter phorbol-12-myristate-13-acetate (PMA) induces chromosomal aberrations in mitogen stimulated human lymphocyte cultures containing monocytes, polymorphonuclear cells (PMN) and platelets. Such cultures produce a diffusible clastogenic factor (CF) in response to PMA which causes aberrations in fresh blood cultures which have not been exposed to PMA. We have studied the contribution of monocytes, PMN and platelets to CF formation. 'Pure' lymphocyte cultures (containing no platelets and maximally 1% PMN and 2% monocytes) only produced CF when they were in contact with 1 to 1.8 X 10(6) monocytes attached to plastic during PMA treatment (18.5 +/- 5.3% mitosis with aberrations for CF produced in the presence of monocytes relative to 6.0 +/- 4% in their absence). They also produced CF of increasing potency upon addition of 0.25 to 5 X 10(6) PMN (20.5 +/- 5.9% mitosis with aberrations for CF produced in the presence of 5 X 10(6) PMN). Cultures containing 5-10 platelets/lymphocyte also formed CF upon PMA treatment. Cultures of purified monocytes and PMN were capable of producing CF in the absence of lymphocytes. The presence of bovine erythrocyte CuZn superoxide dismutase during PMA treatment decreased the activity of the resulting CF under all conditions. Catalase prevented CF production from PMN. It is concluded that the presence of monocytes, PMN or platelets is a prerequisite for CF formation by PMA. Neoplastic tissue is usually surrounded by inflammatory leukocytes. CF produced by these cells in response to tumor promoters such as PMA may induce chromosomal damage in the neighboring tumor cells.

Blood Platelets↗
PubMed1983

Immunologic and enzymatic studies of two breeding lines of NZB mice that differ in chromosome breakage.

Two sublines of NZB/BI mice were developed by selective matings according to chromosome breakage frequencies. These sublines--HB, a line with high chromosome breakages, and LB, a line with low or normal breakage rates--were studied in regard to the age of the animals as it related to two different aspects. The first aspect was immunologic: A decreased response to T-cell mitogens was found in old NZB mice, but this response was more pronounced in HB mice. The response to the B-cell mitogen (lipopolysaccharide) was increased in both sublines as compared to that in BALB/c mice. The percentages of IgG-positive and theta-positive spleen cells were evaluated in both sublines: Some increase in IgG-positive cells was observed in the spleens of 2- to 8-month-old NZB mice and a slight decrease was seen after age 8 months. The percentage of theta-positive cells diminished according to the age of the mice, and the decrease occurred earlier in HB than in LB mice. The second aspect studied was enzymatic and concerned the levels of DNA alpha- and beta-polymerases and terminal DNA nucleotidyltransferase in the thymuses and spleens of these animals. The major finding noted was an augmentation of 100-200% in the terminal DNA nucleotidyltransferase levels in HB thymuses by comparison with LB thymuses. The levels of both polymerases were increased in spleen cells of HB mice as compared to those of LB mice.

Age Factors↗
PubMed1982

Tumor promoter phorbol 12-myristate 13-acetate induces a clastogenic factor in human lymphocytes.

The mechanism of the clastogenic action--i.e., the ability to induce chromosomal aberrations--of the tumor promoter phorbol 12-myristate 13-acetate (PMA) was investigated. PMA at 10 and 100 ng/ml induced the formation of a low molecular weight (less than 10,000) clastogenic factor (CF) in phytohemagglutinin-stimulated human blood and lymphocyte cultures. Bovine erythrocyte Cu-Zn superoxide dismutase strongly inhibited PMA clastogenicity, both the formation of CF and the action of previously formed CF. The nonsteroidal anti-inflammatory agents indomethacin, imidazol, and 5,8,11,14-icosatetraynoic acid inhibited PMA clastogenicity and the clastogenic activity of previously formed CF. These results suggest that superoxide radicals and stimulation of the arachidonic acid cascade play a role in PMA-induced clastogenicity and the mechanism of action of the CF. The CF may relate the initial interaction of PMA with the cell membrane to the genome.

Arachidonic Acids↗