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F Palitti

Publications and source records attributed to F Palitti.

At least 55 records · Page 3Linked to original sources
PubMed1996

DNA damage induced by UV light affects restriction endonuclease recognition sites: correlation between effects at chromosomal level and naked DNA.

Cytogenetic and molecular analyses were performed in G1 Chinese hamster ovary cells treated with 254 nm UV light, to test the hypothesis that UV may affect the recognition site of specific restriction endonucleases (RE). Since short-wavelength UV light induces mainly cyclobutane dimers (CPD) at TT and CT sequences, RE were selected according to the presence or absence of thymine at the recognition site (DraI TTT/AAA, AluI AG/CT and HaeIII GG/CC). A drastic reduction of DraI-and AluI-induced chromosome-type aberrations was found in cells pretreated with UV. Conversely, such a reduction was not observed with HaeIII. To better understand this phenomenon, a molecular analysis was carried out at both the genome level and at the hypoxanthine phosphoribosyl transferase gene level, showing that the cutting pattern of DraI on isolated DNA from UV-irradiated cells was strongly reduced compared with an untreated sample, whereas HaeIII was not able to modify the cutting pattern of irradiated cells. Our data demonstrate a good correlation between the results obtained with cytogenetic and molecular approaches, suggesting that cyclobutane dimers are the main lesions responsible for the observed reduction of the cleaving activities of RE, at both the chromatin and naked DNA levels.

Animals↗
PubMed1996

Simultaneous myogenin expression and overall DNA hypomethylation promote in vitro myoblast differentiation.

Two clones of the L5 myoblast line (M6 and the fusion-defective M12) were examined for the expression of myogenin, one of the regulatory genes involved in the regulation of differentiation to myofibers after treatment with 3-deazaadenosine, a metabolic inhibitor of methyl transfer reactions. Cultures treated with 3-deazaadenosine showed, using Northern blot hybridization, a conspicuous increase in myogenin expression, which in clone M6 correlated to the extent of cell differentiation under fusing conditions but was evident also in growth medium, although the drug was unable to start the myogenic program. We also tested the extent of total DNA methylation to verify whether the activation of the regulatory cascade could be correlated to the decrease of the overall number of 5-methylcytosines present in the genome. The results show that the loss of 5-methylcytosine from newly synthesized DNA, but not from preexisting DNA, is evident in fusing conditions and enhanced by 3-deazaadenosine. It appears that there is a positive correlation between the passive demethylation of newly synthesized DNA, the activation of the myogenin gene by demethylation, and the differentiation of myoblasts. However, in fusing conditions, the defective clone M12, although it is able to express myogenin and its DNA is hypomethylated, fuses only in the presence of 3-deazaadenosine, suggesting some alternative way of induction.

Animals↗
PubMed1994

Clastogenic effects of the dithiocarbamate fungicides thiram and ziram in Chinese hamster cell lines cultured in vitro.

We report here the results obtained using the dithiocarbamate fungicides thiram and ziram to investigate the induction of chromosomal aberrations (CAs) in Chinese hamster ovary (CHO) cells both in the absence and presence of S9 metabolism, and in a Chinese hamster epithelial liver (CHEL) cells which retain metabolic competence to activate different classes of promutagens/procarcinogens. Both thiram and ziram proved to be strong chromosome breaking agents in the CHEL cells and CHO cells in the presence of S9 metabolism. These findings suggest that thiram and ziram require metabolic conversion to become genetically active, and corroborate the evidence that CHEL cells are suitable to activate and detect a broad spectrum of chemical procarcinogens including these two pesticides.

Animals↗
PubMed1994

Lack of effect of inhibitors of DNA synthesis/repair on the ionizing radiation-induced chromosomal damage in G2 stage of ataxia telangiectasia cells.

The relationship between the repair processes occuring at the G2 phase of the cell cycle and cytogenetic damage in ataxia telangiectasia (AT) cells was studied. Lymphoblastoid cells derived from normal, heterozygote AT (HzAT) and three AT patients were exposed to X-rays or fission neutrons and post-treated with inhibitors of DNA synthesis/repair, such as inhibitors of DNA polymerases alpha, delta and epsilon (cytosine arabinoside, ara-C; aphidicolin, APC; buthylphenylen-guanine, BuPdG) or ribonucleotide reductase (hydroxyurea, HU). A strong increase of radiation-induced chromosomal aberrations was observed in normal and HzAT cells post-treated with ara-C, APC and HU, but not in the presence of BuPdG. No enhancing effect was observed in cells derived from AT patients, except for HU post-irradiation treatment. These results suggest that the enzymes that can be inhibited by these agents are not directly involved in the repair of radiation damage induced in G2 cells from AT patients, indicating that probably the AT cells that we used lack the capability to transform the primary DNA lesions into reparable products, or that AT cells might contain a mutated form of DNA polymerase resistant to the inhibitors.

Ataxia Telangiectasia↗
PubMed1993

DNA hypomethylation and differentiation in Friend leukemia cell variants.

The occurrence, upon differentiation, of a transient DNA hypomethylation has been observed in Friend erythroleukemia cells. Treatment with hexamethylenebisacetamide (HMBA) induces within 24 h a 20% hypomethylation of newly synthesized DNA, that is followed by re-methylation before completion of the differentiative process, as measured by the appearance of benzidine-positive cells. We examined a series of mutant clones which continue to grow in the presence of an inducer. Methylcytosine content of DNA was measured by HPLC, after cell labeling with [3H]uridine. We found that one of these continuously growing clones, which was still capable of hemoglobin synthesis, showed the same degree of hypomethylation as the parental one. The re-methylation process did not occur, however, unless erythroid differentiation was reverted by the removal of the inducer. In another clone which had lost the capacity to synthesize hemoglobin, no DNA hypomethylation was detectable. These experiments show that DNA hypomethylation is an early event strictly related to cell differentiation but not to cell growth arrest.

Acetamides↗
PubMed1993

Mechanism of induction of chromosomal aberrations by inhibitors of DNA topoisomerases.

Evidence is provided that antitopoisomerase drugs induce chromosomal aberrations via the formation of DNA double strand breaks. Camptothecin, an inhibitor of DNA topoisomerase I, produced only chromatid-type aberrations, and in order to be effective it has to be present during the DNA replication. The fact that it induces chromosomal aberrations also in the G2 phase of the cell cycle suggests the presence of a residual DNA synthesis in this phase. A pretreatment with aphidicolin, an inhibitor of DNA polymerases, reduces the camptothecin induced aberrations in the G2 phase, confirming the need of DNA synthesis for the induction of aberrations by the inhibitor of DNA topoisomerase I.

Animals↗
PubMed1993

Cytogenetic effects of inhibition of topoisomerase I or II activities in the CHO mutant EM9 and its parental line AA8.

The possible relationship between topoisomerase activities and the occurrence of SCE and chromosomal aberrations was investigated in the CHO ethyl methanesulfonate-sensitive mutant EM9 and its parental line, AA8. The Topo II inhibitor m-AMSA induced fewer SCE in EM9 than in AA8, mainly when given during the first S period. When the Topo I inhibitor camptothecin was used, it showed a higher efficiency to induce both chromosomal aberrations and SCE in EM9 than in AA8. The impact of BrdU incorporated into parental DNA on topoisomerase activity was tested using nuclear extracts from both EM9 and AA8 assayed for their ability to decatenate kinetoplast DNA by Topo II and to relax supercoiled plasmid DNA by Topo I. Taken as a whole, the results seem to indicate that there are differences between the two cell lines, consistent with the hypothesis put forward by other investigators that topoisomerases are involved in the molecular mechanism of SCE.

Amsacrine↗
PubMed1993

Higher G2 sensitivity to the induction of chromosomal damage in the CHO mutant EM9 than in its parental line AA8 by camptothecin, an inhibitor of DNA topoisomerase I.

The induction of chromosomal alterations by camptothecin (CPT), an inhibitor of DNA topoisomerase I, in the G2 stage was studied in a CHO mutant cell line EM9, which has an elevated baseline frequency of SCEs, and in its parental cell line AA8. EM9 cells were found to be more sensitive to CPT than the parental cell line. It is proposed that the effect of camptothecin in this phase of the cell cycle is caused by a 'residual' DNA synthesis which is higher in EM9 than in AA8 cells.

Animals↗
PubMed1993

Hypersensitivity of lymphoblastoid lines derived from ataxia telangiectasia patients to the induction of chromosomal aberrations by etoposide (VP-16).

Mammalian DNA topoisomerase II represents the cellular target of many antitumor drugs, such as epipodophyllotoxin VP-16 (etoposide). The mechanism by which VP-16 exerts its cytotoxic and antineoplastic actions has not yet been firmly established, although the unique correlation between sensitivity to ionizing radiation and to topoisomerase II inhibitors suggest the involvement of DNA double-strand breaks. In the present study we analyzed the chromosomal sensitivity of lymphoblastoid cell lines derived from ataxia telangiectasia (AT) patients to low concentrations of the drug. Our results indicate that AT derived cells are hypersensitive to the clastogenic activity of VP-16 either when the drug is present for the whole duration of the cell cycle or specifically in the G2 phase, confirming that the induction of DNA double strand breaks, to which AT cells seem typically sensitive, could have an important role in the biological activity of VP-16.

Ataxia Telangiectasia↗
PubMed1993

Sensitivity of lymphocytes from vulcanizers to the in vitro induction of sister chromatid exchanges.

Spontaneous frequencies of sister chromatid exchanges (SCEs) and SCEs induced in vitro by chemicals with different mechanisms of action such as mitomycin C, 4-nitroquinoline oxide, and 3-aminobenzamide were examined in phytohemagglutinin-stimulated peripheral blood lymphocytes from a group of workers in a rubber plant and a control group, both of which had been analyzed for levels of spontaneous SCEs 2 years earlier. An interindividual variability in the induction of SCEs was found after in vitro treatments with the different mutagens, which did not correlate with occupational exposure. This variability in the sensitivity to the induction of SCEs might be correlated to genetic differences among individuals, which have to be taken into account in environmental monitoring programs.

Adult↗
PubMed1992

Application of an epithelial liver cell line, metabolically competent, for mutation studies of promutagens.

Recently numerous attempts have been made to reduce the use of vertebrate animals in laboratory experiments to evaluate general and acute toxicity, mutagenesis and teratogenesis of new drugs or chemicals. One common approach is to use established, proliferating cell lines that preserve differentiated functions such as the competence to metabolize xenobiotics. To this end a continuous Chinese hamster epithelial liver cell line (CHEL cells) was established, cultured as used for mutagenesis studies. Structurally different promutagens, such as 7,12-dimethylbenz[a]anthracene (7,12-DMBA), benzo[a]pyrene (B(a)P), aflatoxin B1 (AB1) and cyclophosphamide (CP), were used in order to check and validate the test system. anti-Chrysene-1,2-diol 3,4-epoxide (CDE) and mitomycin C (MMC) were taken as representatives of direct mutagens. The genetic change induced by the mutagens was quantified by measuring mutation frequencies at the HGPRT locus. Several parameters, such as mutant expression time for each chemical, cell density for selection of mutants and enzymatic characterization for HGPRT phenotype, were examined to establish the optimal assay conditions. All promutagens analyzed significantly affected either the cloning efficiency and/or the mutant frequency of CHEL cells after 24 h of exposure. In addition, various enzyme activities involved in the metabolism of the promutagens were determined in CHEL cells, under the experimental conditions of chemical exposure used in the mutagenesis assay. The enzyme activities were compared with those found in uninduced Chinese hamster liver.

Animals↗
PubMed1992

Chromosomal aberrations in human lymphocytes induced in vitro by very low doses of X-rays.

This paper presents results of a collaborative experiment between six laboratories which examined the yields of unstable chromosomal aberrations in human lymphocytes induced in vitro by X-rays over the dose range 0-300 mGy. The work included data points of nominal doses of 0, 3, 5, 6, 10, 20, 30, 50 and 300 mGy. Cells from 24 donors were examined and a total of about 300,000 metaphases were scored. The work was undertaken to determine the limits of sensitivity of the system taking into account variations in scoring data due to inter-donor sample and inter-laboratory effects. Despite the existence of these effects, aberration yields significantly in excess of control values were seen at doses greater than 20 mGy and these were consistent with a linear extrapolation from higher doses. Below 20 mGy the observed dicentric yields were generally lower than background, but not significantly so. Excess acentric aberrations, on the other hand, and centric rings, were higher than the controls but the increase was usually not significant. It is concluded that the statistical uncertainties are such that below 20 mGy this technique cannot distinguish between a linear or a threshold model.

Chromosome Aberrations↗
PubMed1992

The yield of fission neutron-induced chromatid aberrations in G2-stage human lymphocytes: effect of caffeine, hydroxyurea and cytosine arabinoside post-irradiation.

To evaluate the influence of inhibitors of DNA synthesis/repair on the yield of chromosomal aberrations in the G2 phase of the cell cycle, whole-blood cultures of human lymphocytes were exposed to various doses of fission neutrons or X-rays and treated post-irradiation during the last 2.45 h before harvesting, with 5 mM caffeine, 5 mM hydroxyurea (HU) and 0.05 mM cytosine arabinoside (ara-C). The presence of caffeine and HU strongly potentiated the yield of chromatid-type aberrations induced by both neutrons and X-rays. No potentiating effect, except at the highest dose of neutrons, was observed when irradiated cells were subsequently treated with ara-C. Since ara-C strongly potentiated the frequency of chromatid aberrations induced in G2 lymphocytes by X-rays, the results presented here indicate that fission neutrons produce a smaller proportion of lesions, the repair of which can be inhibited by ara-C, compared with the number produced by X-rays. In addition, neutron-induced mitotic delay was shortened by treatment with caffeine, mainly within the first 2 h after irradiation.

Caffeine↗
PubMed1990

Persistence of azacytidine-induced SCEs and genomic methylation in CHO cells in vitro.

Many carcinogenic agents are able to affect the methylation level in mammalian cells cultivated in vitro. The capacity of azacytidine (AZA) to demethylate DNA can be used to examine the relationship between the genomic methylation level and cytogenetic end-points. Here we compared the sister-chromatid exchange (SCE) level with the genomic % methylcytosine in a Chinese hamster ovary cell line in vitro after giving a single 10-microM pulse of AZA. Both parameters were followed up to 16 cell cycles after the agent was removed. While the SCE level increased starting 2 cycles from the treatment and persisted for the entire 16 cycles, the methylcytosine level, after an initial 50% decrease, approached the control value, completely returning to it after 10 cell cycles. The possibility that the persistence in the SCE increase is an inherited phenomenon is discussed.

5-Methylcytosine↗
PubMed1989

In vitro methylation of CpG-rich islands.

CpG islands are distinguishable from the bulk of vertebrate DNA for being unmethylated and CpG-rich. Since CpG doublets are the specific target of eukaryotic DNA methyltransferases, CpG-rich sequences might be expected to be good methyl-accepting substrates in vitro, despite their unmethylated in vivo condition. This was tested using a partially purified DNA-methyltransferase from human placenta and several cloned CpG-rich or CpG-depleted sequences. The efficiency of methylation was found to be proportional to the CpG content for CpG-depleted regions, which are representative of the bulk genome. However, methylation was much less efficient for CpG frequencies higher than 1 in 12 nucleotides, reaching only 60% of the expected level. That suggests that the close CpG spacing typical of CpG-islands somehow inhibits mammalian DNA methyltransferase. The implications of these findings on the in vivo pattern of DNA methylation are discussed.

Base Sequence↗
PubMed1989

Induction of chromosomal aberrations and SCE by camptothecin, an inhibitor of mammalian topoisomerase I.

The induction of chromosomal aberrations and sister-chromatid exchanges (SCE) was studied in human lymphocyte cultures treated with camptothecin (CM), an inhibitor of mammalian topoisomerase I. While no chromosome-type aberrations were found in G1-treated cells, instead there was a dose-dependent induction of chromatid-type aberrations. These types of chromosomal alteration were not induced during the treatment itself but during the S phase, as CM is not efficiently removed with the normal washing procedure after treatment.

Camptothecin↗