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P Hrelia

Publications and source records attributed to P Hrelia.

At least 19 recordsLinked to original sources
PubMed2005

Antioxidant effectiveness of organically and non-organically grown red oranges in cell culture systems.

BACKGROUND: Consumers consider plant food products from organic origin healthier than the corresponding conventional plant foods. Clear experimental evidence supporting this assumption is still lacking. AIM OF THE STUDY: To determine if the organic red oranges have a higher phyto-chemical content (i. e., phenolics, anthocyanins and ascorbic acid), total antioxidant activity and in vitro bioactivity, in terms of protective effect against oxidative damage at cellular level, than nonorganic red oranges. METHODS: Total phenolics were measured using the Folin Ciocalteau assay, while total anthocyanins and ascorbic acid levels were determined by spectrophotometric and HPLC analysis, respectively. In addition, the total antioxidant activity of red orange extracts was measured by the ABTS(*+) test. The ability of red orange extracts to counteract conjugated diene containing lipids and free radical production in cultured rat cardiomyocytes and differentiated Caco-2 cells, respectively, was assessed. RESULTS: Organic oranges had significantly higher total phenolics, total anthocyanins and ascorbic acid levels than the corresponding non-organic oranges (all p < 0.05). Moreover, the organic orange extracts had a higher total antioxidant activity than non-organic orange extracts (p < 0.05). In addition, our results indicate that red oranges have a strong capacity of inhibiting the production of conjugated diene containing lipids and free radicals in rat cardiomyocytes and differentiated Caco-2 cells, respectively. Statistically higher levels of antioxidant activity in both cell models were found in organically grown oranges as compared to those produced by integrated agriculture practice. CONCLUSIONS: Our results clearly show that organic red oranges have a higher phytochemical content (i. e., phenolics, anthocyanins and ascorbic acid), total antioxidant activity and bioactivity than integrated red oranges. Further studies are needed to confirm whether the organic agriculture practice is likely to increase the antioxidant activity of other varieties of fruits and vegetables.

Agriculture↗
PubMed2004

A mixture of isothiocyanates induces cyclin B1- and p53-mediated cell-cycle arrest and apoptosis of human T lymphoblastoid cells.

As with other candidate chemopreventive agents, most of our knowledge on the biological effects of isothiocyanates (the many sulfur-containing metabolites found in cruciferous vegetables) comes from studies of single natural or synthetic compounds. To investigate whether the biological/chemopreventive effects of administration of single isothiocyanates can differ from those of a mixture of isothiocyanates, we tested the effects of a mixture of four different isothiocyanates on cell-cycle progression and apoptosis in human T leukemia Jurkat cells, and identified some of the molecular pathways triggered by the mixture. The mixture affected critical points of the cell cycle via modulation of the expression of cyclin B1. Moreover, it induced apoptosis, mediated by an increase in p53 and bax (expression of bcl-2 was unaffected). Comparison of the data with those previously obtained with the single isothiocyanates under identical experimental conditions provides evidence that the quantitative effects of a single, specific isothiocyanate can be significantly different from those of an isothiocyanate mixture at realistic doses.

Apoptosis↗
PubMed2003

Photomutagenic properties of terfenadine as revealed by a stepwise photostability, phototoxicity and photomutagenicity testing approach.

Administration of the second-generation antihistamine, terfenadine, is sometimes associated with photosensitivity and other skin reactions. To obtain information on its photoreactivity, we used a stepwise experimental approach involving tests for photostability, phototoxicity (PT) (mouse fibroblast cell line [3T3] neutral red uptake [NRU] test) and photomutagenicity (with standard Ames salmonella tester strains TA98, TA100 and TA102). Terfenadine was not phototoxic to cultured mammalian cells under the conditions used (i.e. 5000/161 mJ cm(-2) UVA-UVB). Natural sunlight and UV radiations caused considerable drug decomposition and formation of several photoproducts. Addition of the irradiated terfenadine solution (i.e. a mixture of photoproducts) to the tester did not significantly increase background mutation frequency. Irradiation of terfenadine coplated with the TA102 strain induced a clear-cut photomutagenic response, the magnitude of which was dependent upon the precursor compound concentration and the UV dose (212/7 to 339/11 mJ cm(-2) UVA-UVB). These findings demonstrate that in vitro terfenadine is photomutagenic in absence of PT. Further in vitro and in vivo studies are therefore needed to provide an adequate safety assessment of the photochemical genotoxicity--carcinogenicity potential of terfenadine. In the meantime, patients should be advised to avoid excessive exposure to sunlight.

3T3 Cells↗
PubMed2002

Mutagenic and clastogenic activity of gastric juice in human gastric diseases.

To investigate whether subjects with low-acid states are exposed to increased genetic risk with respect to controls, we evaluated mutagenicity and presence of clastogenic factors (CF) in the gastric juice of chronic atrophic gastritis and omeprazole-treated patients. Mutagenic gastric juice was found in 8/15 (53%) chronic atrophic gastritis patients, 8/11 (73%) omeprazole-treated patients, and 2/13 (15%) healthy control subjects. The mean mutagenicity ratio of omeprazole-treated patients (1.52+/-0.48/0.1 ml gastric juice) was significantly higher than those of either controls (1.07+/-0.15; P<0.01) or chronic atrophic gastritis patients (1.16+/-0.21; P<0.05). Only chronic atrophic gastritis patients showed an increased clastogenic index with respect to healthy controls (2.67+/-2.13 versus 0.38+/-0.51; P<0.001). These findings expand our knowledge of gastric disease risk factors, and indicate that there may well be a risk of mucosal DNA damage arising from the presence of mutagenic and CF in the gastric juice.

Adult↗
PubMed2002

Cyclin D3 and p53 mediate sulforaphane-induced cell cycle delay and apoptosis in non-transformed human T lymphocytes.

Despite experimental evidence that sulforaphane can exert chemopreventive effects, whether these effects are specific for neoplastic cells is not known. Following our previous demonstration that sulforaphane induces cell cycle arrest and apoptosis in human T lymphoblastoid Jurkat leukemia cells and increases p53 and bax protein expression, we tested sulforaphane on non-transformed phytohemagglutinin-stimulated human lymphocytes. Here, we demonstrate that sulforaphane arrested cell cycle progression in G, phase, through a decrease in the protein expression of cyclin D3. Moreover, sulforaphane induced apoptosis (and also necrosis), mediated by an increase in the expression of p53. These findings suggest that sulforaphane is a growth modulator for T cells. Our in vitro evidence that sulforaphane is active and even cytotoxic in normal as well as transformed lymphocytes raises important questions regarding its suitability for cancer chemoprevention.

Anticarcinogenic Agents↗
PubMed2001

Micronuclei induction, cell cycle delay and apoptosis as markers of cellular stress caused by ursodeoxycholic acid in human lymphocytes.

Ursodeoxycholic acid (UDCA) is a bile acid (BA) used for cholesterol gallstone dissolution. Since epidemiological evidence indicates that BAs can be involved in the etiology of colorectal cancer, we investigated the effects of UDCA and its physiologically produced taurine conjugate tauroursodeoxycholic acid (TUDCA) on human lymphocyte cultures in terms of genetic damage in the form of micronuclei (MN) production, cell cycle modifications and induction of apoptosis. With respect to controls, treatment with UDCA (from 10 microg/ml) caused a dose-related increase in MN, whereas TUDCA caused no significant increase (up to 1000 microg/ml). Fluorescence in situ hybridization (FISH) analysis using pancentromeric probes suggested that UDCA exerts aneugenic activity. Bromodeoxyuridine/Hoechst flow cytometry showed that both BA significantly inhibit cell cycle progression (UDCA at 100 microg/ml, and TUDCA, more markedly at 300-1000 microg/ml). Neither UDCA nor TUDCA affected induction of apoptosis, as evaluated by the Annexin-V-Fluos assay. We conclude that UDCA is potentially genotoxic. However, taking into account the characteristics of other physiological BA, our findings are in line with the concept that long-term UDCA treatment may be safely administered. The multi-assay approach reported here could be useful in the toxicological evaluation of newly developed BA analogs as candidates for pharmacological use.

Apoptosis↗
PubMed2001

Photostability and phototoxicity studies on diltiazem.

The photostability of diltiazem was studied in aqueous solutions at different pH values. Firstly, the hydrolysis of the drug to desacetyldiltiazem in alkaline medium was evaluated and then the drug photodegradation under exposure to UVA-UVB radiation (solar simulator) was monitored by HPLC methods. The main photoproduct was isolated and characterized as diltiazem-S-oxide on the basis of the NMR and mass spectra. The HPLC method was also applied to the selective analysis of diltiazem in commercial formulations. Tests on mutagenicity and photomutagenicity of the drug were also carried out using Salmonella typhimurium TA 102 strain. In this testing the drug neither was mutagenic nor toxic.

Calcium Channel Blockers↗
PubMed2001

Studies on the photostability and in vitro phototoxicity of Labetalol.

The purpose of this study was to obtain information on the photochemical and phototoxic properties of Labetalol, a beta-blocker drug. Preliminary information on the drug photoreactivity was achieved using a flow system with a photochemical reactor on-line with a diode array detection system. Photophysical and photochemical investigations on the drug were performed in aqueous solutions at different pH values using spectrophotometric and fluorimetric methods; the photodegradation quantum yield was found to be 2.7 x 10(-3) at pH 5.8 and 1.5 x 10(-2) at pH 11.5. Forced photodegradation of labetalol solutions under exposure to UVA--UVB radiations (xenon arc lamp) was monitored by reversed-phase liquid chromatography. The main photodegradation products were isolated and characterized by NMR and mass spectrometry; labetalol was found to give 3-amino-1-phenylbutane and salicylamide-4-carboxaldehyde as the main photoproducts. Preliminary phototoxic testings on human keratinocyte cultures were performed evaluating the viability of the cells by the neutral-red uptake assay; mutagenic and photomutagenicity tests were also carried out based on Salmonella typhimurium strains. As a result, labetalol was found to be photolabile,mainly in alkaline medium, but evidences of significant phototoxic and photomutagenic effects by the drug were not observed.

3T3 Cells↗
PubMed2001

HUman MicroNucleus project: international database comparison for results with the cytokinesis-block micronucleus assay in human lymphocytes: I. Effect of laboratory protocol, scoring criteria, and host factors on the frequency of micronuclei.

Micronucleus (MN) expression in peripheral blood lymphocytes is well established as a standard method for monitoring chromosome damage in human populations. The first results of an analysis of pooled data from laboratories using the cytokinesis-block micronucleus (CBMN) assay and participating in the HUMN (HUman MicroNucleus project) international collaborative study are presented. The effects of laboratory protocol, scoring criteria, and host factors on baseline micronucleated binucleate cell (MNC) frequency are evaluated, and a reference range of "normal" values against which future studies may be compared is provided. Primary data from historical records were submitted by 25 laboratories distributed in 16 countries. This resulted in a database of nearly 7000 subjects. Potentially significant differences were present in the methods used by participating laboratories, such as in the type of culture medium, the concentration of cytochalasin-B, the percentage of fetal calf serum, and in the culture method. Differences in criteria for scoring micronuclei were also evident. The overall median MNC frequency in nonexposed (i.e., normal) subjects was 6.5 per thousand and the interquartile range was between 3 and 12 per thousand. An increase in MNC frequency with age was evident in all but two laboratories. The effect of gender, although not so evident in all databases, was also present, with females having a 19% higher level of MNC frequency (95% confidence interval: 14-24%). Statistical analyses were performed using random-effects models for correlated data. Our best model, which included exposure to genotoxic factors, host factors, methods, and scoring criteria, explained 75% of the total variance, with the largest contribution attributable to laboratory methods.

Adolescent↗
PubMed2000

Cyto- and genotoxic effects of novel aromatic nitroxide radicals in vitro.

Because of the increasing interest in the use of nitroxide radicals as antioxidants and probes for various applications in biological systems, the question of their toxicity is of paramount importance. Cytotoxicity and mutagenicity studies have been extensively performed with the commercially available aliphatic nitroxides, and the general outcome is that these compounds are nonmutagenic and relatively noncytotoxic. In this study, the cytotoxicity and genotoxicity of a new class of aromatic nitroxides that we have synthesized (i.e., indolinonic and quinolinic nitroxides), whose antioxidant activity has been established in both chemical and biological systems, were evaluated and compared with those of two commercial nitroxides and with that of butylated hydroxytoluene (BHT). The mutagenicity assay was performed using Salmonella typhimurium tester strains TA98, TA100, and TA102, chosen on the basis of their ability to detect various types of mutations and their sensitivity to oxidative damage. None of the compounds tested were found to be mutagenic. The colony-forming assay (CFA) using Chinese hamster ovary (CHO) AS52 cells was employed for determining the cytotoxicity of the test compounds. On comparing the effective dose that inhibits the CFA by 50% (IC(50)), most of the compounds tested on an equal molar concentration basis were less toxic than BHT. Therefore, the overall results obtained correlate well with the data reported in the literature on the toxicity of aliphatic nitroxides and lend support to the possible use of these compounds as therapeutic antioxidants.

Animals↗
PubMed2000

Increased cytogenetic damage detected by FISH analysis on micronuclei in peripheral lymphocytes from alcoholics.

Alcohol abuse greatly increases the risk of various malignancies, including cancer of the liver and digestive tract. Although it is thought that this may be due, at least partially, to the mutagenic properties of ethanol, little is known about the genotoxic effects of ethanol in humans. We investigated the chromosomal damage in lymphocytes from 20 alcoholics and 20 controls using the micronucleus (MN) assay combined with fluorescence in situ hybridization (FISH) with a pancentromeric DNA probe capable of differentiating centromere positive (C+) from centromere negative (C-) MN. The frequency of MN in binucleate lymphocytes was significantly higher in alcoholics than in controls (12.0 +/- 5.4 and 7.6 +/- 1.6, respectively; P: < 0.05). FISH revealed significantly higher frequencies of C+ MN in alcoholics than in controls (8.2 +/- 4.8 and 3.4 +/- 1.4, respectively; P: < 0.05). In the alcoholics, no association was found between years of alcohol abuse and frequency of MN or C+ MN. However, age influenced MN and C+ MN frequency both in alcoholics and controls. These results indicate that alcohol abuse may well induce chromosome loss in humans, suggesting a possible aneugenic mechanism of alcohol. This effect could contribute to the health hazards related to alcoholism such as cancer risk.

Adult↗
PubMed1999

Bromodeoxyuridine labelling as an alternative method to identify 6-thioguanine-resistant mutant lymphocytes in humans.

6-Thioguanine-resistant (TGR) mutant lymphocytes in human blood are usually enumerated by the cloning assay which allows the molecular characterisation of the HPRT mutations to be detected. A "short-term" alternative approach is provided by the anti-bromodeoxyuridine (anti-BrdU) technique in which TGR lymphocytes are identified immunocytochemically by their ability to synthesise DNA in the presence of 6-thioguanine (TG). We have evaluated the influence of various experimental factors that could affect the frequency of TGR lymphocytes. A standard protocol is proposed, based on 24-h cold storage of isolated lymphocytes at 4 degrees C and 40-h culture with and without TG, the last 16 h with BrdU. The harvested cells are treated with hypotonic (0.075 M) KCl, fixed with methanol:acetic acid (3:1) and put on microscopic slides. For the TG cultures, all cells are prepared on the slides, while slides from the control cultures are made by a 1/50 dilution. DNA is denatured by formamide, and the BrdU label is identified by anti-BrdU antibody detected by immunoperoxidase staining using a peroxidase-conjugated secondary antibody with diaminobenzidine as substrate. In 10 donors, the frequency of TGR lymphocytes (variant frequency, Vf) detected by this protocol ranged from 69.65 x 10(-6) to 83.45 x 10(-6), and split measurements showed a relatively small intra-assay variation in Vf values of each donor. BrdU in DNA was also detected by immunofluorescence using a fluorescein-conjugated anti-BrdU monoclonal antibody. This method, facilitating easy identification of positive cells and rapid microscopic scoring, may serve as a basis for an automated analysis of TGR lymphocytes. Vf values detected by the anti-BrdU assay are higher than mutant frequencies obtained by the cloning assay, which has been assigned to the presence of non-mutant phenocopies considered to represent spontaneously cycling lymphocytes. Although the anti-BrdU assay is rapid and easy and has been shown to respond to genotoxic exposures, its true value could be evaluated only when it can be ascertained that phenocopies do not significantly contribute to the Vf values obtained.

Bromodeoxyuridine↗
PubMed1999

Influence of nitroreductase and O-acetyltransferase on the mutagenicity of substituted nitrobenzothiophenamines in Salmonella typhimurium.

The mutagenic activity of 17 substituted (aryl)(2-nitrobenzo[b]thiophen-3yl)amines has been evaluated in the Ames test with different isogenic strains of Salmonella typhimurium, that varied in their expression of nitroreductase and O-acetyltransferase. Active derivatives induced frameshift mutations in TA98 strain, and differences in the chemical structure resulted in up to 15-fold changes in mutagenic activity. The non-mutagenic compounds are the unsubstituted parent compound and derivatives with para-chloro, para-fluoro, para-diethylamino, meta-bromo and para-dimethylamino groups. They do not show any activity even in strains with higher level of nitroreductase or O-acetyltransferase. The addition of S9 fraction decreases the mutagenic potential or gives comparable results to those obtained without metabolic activation. For electron-donating substituents, the meta-isomers display the greatest mutagenic potency, whereas the transfer of the group to the para-position leads to less active or unactive molecules. All active nitrobenzothiophenes are substrates for bacterial nitroreductase and O-acetyltransferase, as shown by the reduced mutagenicity in the deficient strains and increased mutagenicity in the corresponding overproducing bacteria. Previous reports have pointed out interest in nitrothiophene analogues with para-chloro and para-fluoro substituents as promising anti-inflammatory drugs. The present study further enhances the putative interest in these derivatives, based on absence of mutagenicity.

Acetyltransferases↗
PubMed1999

Flow cytometric analysis of genetic damage, effect on cell cycle progression, and apoptosis by thiophanate-methyl in human lymphocytes.

Flow cytometric technique was used to study the effects of the fungicide Thiophanate-methyl on cell proliferation, micronucleus induction, and apoptosis in human peripheral blood lymphocytes treated in vitro. In particular, a combined approach of flow cytometry and fluorescence in situ hybridization (FISH) with a pancentromeric alpha-satellite probe was used to evaluate the mechanism of micronucleus induction by Thiophanate-methyl. Flow sorted micronuclei (MN) induced in human lymphocytes by Thiophanate-methyl were analyzed by FISH and the results were compared with results from FISH analysis on MN in binucleated cells. It could be shown that most MN induced by Thiophanate-methyl did not reveal any centromeric signal, thus demonstrating clastogenic action of this fungicide. Moreover, it was found that as a function of the concentration of Thiophanate-methyl, cellular proliferation was delayed and the frequency of apoptotic cells was increased.

Adult↗
PubMed1998

The pitfall of detoxifying enzymes.

One of the major mechanism of chemical protection against mutagenesis, carcinogenesis and other forms of toxicity is the induction of phase-II metabolizing enzymes such as UDP-glucuronosyl transferases, glutathione S-transferases and NAD(P)H quinone reductase, or inhibition of typical phase-I reactions. The use of selective inducers of conjugating enzymes or inhibitors of CYP- and FAD-dependent monooxygenases revealed the possibility of reducing the expression of certain forms of malignancy. However, the use of some anti-initiating entities devised to reduce tumor initiation, seems to receive invalidated justification. Indeed, considering the double edge-sword nature (activating or detoxifying) of drug metabolizing enzymes as well as the myriad of xenobiotics to which human is exposed, any attempt to modulate such catalysts by dietary components (including drugs) could lead to an increased cancer risk. Paradoxically, it has been recently proposed the use of metabolizing liver preparations, isolated from phase-II induced rodents, as a novel bioactivating model in the field of genetic toxicology. Exogenous microsomal (S9) fraction prepared from 2-(3)-tert-butyl-4-hydroxyanisole (BHA) (monofunctional post-oxidative inducer) treated mice are able to increase the DNA binding and genotoxic response of pre-mutagens. On the whole, the use of enzyme modulators in cancer chemoprevention, for their ability to simultaneously reduce or increase pre-carcinogen bioactivation, should be carefully reconsidered.

Animals↗
PubMed1998

Synthesis, metabolism and structure-mutagenicity relationships of novel 4-nitro-(imidazoles and pyrazoles) in Salmonella typhimurium.

A new series of 4-nitro-(imidazoles and pyrazoles) were synthesized as novel antimycotics and tested for their activation to mutagenic forms using Salmonella typhimurium TA98 and TA100, in the presence and in the absence of metabolic activation. TA100NR, TA100/1,8-DNP6, YG1026 and YG1029 strains were employed to identify a specific metabolic reaction which governs the mutagenic potency. Derivatives in the pyrazole group were generally found to be non mutagenic and active imidazoles were weak-direct-acting mutagens. For most of the compounds the mutagenic responses in TA98 were absent or 12- to 22-fold lower compared to TA100. The presence of a methyl or a benzylic group on the imidazole ring and substituents on the N1 and N3 positions were determinant for mutagenicity. Metabolism by bacterial enzyme systems was important to the expression of genotoxicity. Active compounds showed no mutagenicity toward the strain defective in classical nitroreductase and increased mutagenicity, from 2- to 7-fold depending on the test compound, toward the corresponding overproducing bacteria. On the other hand, compounds displayed reduced mutagenicity to the O-acetyltransferase strain without having increased activity in the corresponding overproducing bacteria, YG1029.

Imidazoles↗
PubMed1997

Immunoprevention of cancer poses a challenge to pharmacological research.

The evolving understanding of the molecular mechanisms of carcinogenesis establishes that the long latency period would offer numerous opportunities for intervention before the final step of fully developed malignancy has been reached. Immunoprevention of cancer may be a new approach to cancer control, by eliminating the cellular minimal deviations that are seen in the early phases of carcinogenesis. Available results of immunotherapy of preneoplastic lesions and data on anticarcinogenesis with immunoregulators in experimental models are in good accordance with theoretical expectations. However, clinical research on immunoregulators still focuses on the treatment of advanced cancer. The major problem hampering a wider application of immunoregulators in cancer prevention is the possibility of potential adverse effects, which are largely unknown. The promotion of future research in this area is essential. Moreover, the ethical issues of intervention trials in cancer must be discussed, by considering the potential psychological and social consequences.

Animals↗