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Effect of storage conditions of blood on radiation-induced chromosomal aberrations and apoptosis in human lymphocytes

Thu, 31 Dec 2009 23:00:00 GMT

Title: Effect of storage conditions of blood on radiation-induced chromosomal aberrations and apoptosis in human lymphocytes Authors: Belloni, Paola; Pepe, Gaetano; Palitti, Fabrizio Abstract: To evaluate the effect of storage conditions of blood on the direct relationship between radiation-induced chromosome aberrations and apoptosis in human peripheral blood lymphocytes,whole bloodwas irradi- atedwith 3Gy X-rays. Directly after irradiation, a sample of bloodwas analyzed for chromosome damage and proliferation index, after phytohaemagglutinin stimulation and incubation at 37 ◦ C for 56 h. Blood samples were stored for 48 h at 4 and 20 ◦ C with or without phytohaemagglutinin and analyzed for cell viability and apoptosis at 0, 24 and 48 h storage time. After 48 h of storage, unstimulated cultures were stimulated to proliferate. These samples and cultures stimulated immediately before storage were incubated at 37 ◦ C for 56 h and analyzed for chromosome damage and proliferation index. Metaphases were examined for the presence of dicentrics, excess acentrics, and rings. Storage at 20 ◦ Cwithout phyto- haemagglutinin for 48 h increases signiﬁcantly the yield of apoptosis and decreases signiﬁcantly the yield of dicentrics. During 48 h of storage time the presence of phytohaemagglutinin and the temperature of 4 ◦ C protected the irradiated lymphocytes from apoptosis allowing accurate estimation of the real yield of radiation-induced chromosome damage. Therefore these blood-storage conditions enable analysis in metaphase and may offer some advantages for biodosimetry of absorbed radiation dose. Description: L'articolo è disponibile sul sito dell'editore: http://www.sciencedirect.com

A selective de-O-methylation of guaiacyl lignans to corresponding catechol derivatives by 2-iodoxybenzoic acid (IBX). The role of the catechol moiety on the toxicity of lignans

Wed, 31 Dec 2008 23:00:00 GMT

Title: A selective de-O-methylation of guaiacyl lignans to corresponding catechol derivatives by 2-iodoxybenzoic acid (IBX). The role of the catechol moiety on the toxicity of lignans Authors: Bernini, Roberta; Barontini, Maurizio; Mosesso, Pasquale; Pepe, Gaetano; Willfoer, Stefan M.; Sjoeholm, Rainer E.; Eklund, Patrick C.; Saladino, Raffaele Abstract: We report here the first selective de-O-methylation of a large panel of guaiacyl lignans to the corresponding catechol derivatives by using IBX as primary oxidant under green conditions (dimethyl carbonate–H2O solvent) through an in situ reduction procedure. The influence of the catechol moiety on the cytotoxicity and genotoxicity of new lignan derivatives has been investigated. The results obtained indicated that the presence of the catechol moiety sharply enhances the clastogenic potential (e.g. induction of chromosomal aberrations), the cytotoxicity and the modulation of cell cycle progression with respect to the parent compounds. Thus, despite the in vitro antioxidant activity usually described for catechol derivatives, our results show for the first time the generation of a clastogenic potential, highly indicative of a long-term genetic and cancer risk Description: L'articolo è disponibile sul sito dell'editore: http://www.rsc.org

The novel human gene aprataxin is directly involved in DNA single-strand-break repair.

Fri, 31 Dec 2004 23:00:00 GMT

Title: The novel human gene aprataxin is directly involved in DNA single-strand-break repair. Authors: Mosesso, Pasquale; Piane, Maria; Palitti, Fabrizio; Pepe, Gaetano; Penna, Sabrina; Chessa, Luciana Abstract: The cells of an ataxia-oculomotor apraxia type 1 (AOA1) patient, homozygous for a new aprataxin mutation (T739C), were treated with camptothecin, an inhibitor of DNA topoisomerase I which induces DNA single-strand breaks. DNA damage was evaluated by cytogenetic analysis of chromosomal aberrations. The results obtained showed marked and dose-related increases in induced chromosomal aberrations in the patient and her heterozygous mother compared to the intrafamilial wild-type control. The alkaline comet assay confirmed this pattern. Moreover, the AOA1 cells did not show hypersensitivity to ionizing radiation, i.e. X-rays. These findings clearly indicate the direct involvement of aprataxin in the DNA single-strand-break repair machinery. Description: L'articolo è disponibile sul sito dell'editore: http://www.springerlink.com/

In vitro cytogenetic results supporting a DNA nonreactive mechanism for ochratoxin A, potentially relevant for its carcinogenicity

Mon, 31 Dec 2007 23:00:00 GMT

Title: In vitro cytogenetic results supporting a DNA nonreactive mechanism for ochratoxin A, potentially relevant for its carcinogenicity Authors: Mosesso, Pasquale; Cinelli, Serena; Piñero, Joaquin; Bellacima, Raffaela; Pepe, Gaetano Abstract: Ochratoxin A (OTA) is a widespread mycotoxin of cereals and many agricultural products and causes high incidences of renal tumors in rodents. Although its carcinogenic properties have been known since the eighties, the precise mechanism of action is still relatively undefined. At present, increasing evidence suggests that OTA does not act with a direct genotoxic mechanism, opposed to other previous evidence where the formation of DNA adducts by 32P-postlabeling was observed. The genotoxic activity of OTA assessed in a variety of in vitro and in vivo studies was very low if genotoxic at all. In this study, we clearly show that OTA does not bear any clastogenic or aneugenic activity based on the absence of the induction of chromosome aberrations, sister chromatid exchanges, and micronuclei in human lymphocytes and V79 cells in vitro in both the absence and the presence of S9 metabolism. Alternatively, cytogenetic analyses evidenced significant increases in endoreduplicated cells and highly condensed metaphases with separated chromatids. This implies that OTA or its possible metabolites do not covalently bind DNA through the formation of adducts since structural chromosome aberrations are a very sensitive end points to detect chemical carcinogens with electrophilic substituents. Alternatively, induction of endoreduplication and chromatid separation provides strong evidence for a DNA nonreactive mechanism of OTA carcinogenicity involving the disruption of mitosis by interfering with key regulators of chromosome separation and progression of mitosis. This causes a temporary arrest of mitoses and premature exit from it (mitotic slippage) to generate endoreduplication and polyploidy accompanied by increased risk of aneuploidy and subsequent tumor formation. Description: The present paper is dedicated to Prof. A. T. Natarajan on the occasion of his 80th birthday.; L'articolo è disponibile sul sito dell'editore: http://pubs.acs.org/

Ochratoxin A causes DNA damage and cytogenetic effects but no DNA adducts in rats

Fri, 31 Dec 2004 23:00:00 GMT

Title: Ochratoxin A causes DNA damage and cytogenetic effects but no DNA adducts in rats Authors: Mally, Angela; Pepe, Gaetano; Ravoori, Srivani; Fiore, Mario; Gupta, Ramesh C.; Dekant, Wolfgang; Mosesso, Pasquale Abstract: Ochratoxin A (OTA) is a potent nephrotoxin and renal carcinogen in rats, but the mechanism of OTA tumorigenicity is unknown. Ochratoxin A has been shown to be negative in many genetic toxicology test in vitro. However, the potential of OTA to induce genotoxic effects has not been investigated in male rats, the most sensitive species for OTA-induced tumor formation. In this study, male F344 rats were repeatedly administered OTA (0, 250, 500, 1000, and 2000 íg/kg of body wt) or the non-chlorinated analogue ochratoxin B (OTB; 2000 íg/kg of body wt) for 2 weeks (5 days/week), and DNA breakage was analyzed in target and nontarget tissues using the comet assay both in the absence and presence of formamidopyrimidine-DNA (Fpg) glycosylase. Potential DNA-adduct formation was also analyzed in the target organ kidney by 32P-postlabeling using two different solvent systems. DNA-strand breaks were evident in liver, kidney, and spleen of animals treated with OTA, and a similar degree of DNA damage was observed in rats treated with OTB, despite the lower toxicity of OTB. Moreover, the presence of DNA damage did not correlate with histopathological alterations, which were evident in the kidney but not in the liver. In liver and kidney, the extent of DNA damage was further enhanced in the presence of Fpg glycosylase, which is known to convert oxidative DNA damage into strand breaks, suggesting the presence of oxidative DNA damage. Oxidative DNA damage as a mechanism of OTA-dependent DNA damage is consistent with the absence of lipophilic DNA adducts as assessed by 32P-postlabeling analysis. No spots indicative of OTA-related DNA adducts were observed in kidney DNA extracted from OTA-treated animals by 32P-postlabeling analysis, despite the use of synthetic standard for postulated adducts. A small, but not significant, increase in the incidence of chromosomal aberrations (essentially chromatid and chromosome-type deletions) was observed in splenocytes from rats treated with OTA in vivo and subsequently cultured in vitro to express chromosomal damage. These aberrations are also compatible with oxidative DNA lesions since they are not typically caused by chemical carcinogens which form covalent DNA adducts. Together, with the lack of evidence for formation of lipophilic DNA adducts as assessed by postlabeling, these data suggest that OTA may cause genetic damage in both target and nontarget tissues independent of direct covalent binding to DNA. Description: L'articolo è disponibile sul sito dell'editore: http://www.pubs.acs.org

Potassium Bromate but not X-ray cause unexpectedly elevated levels of DNA breakage similar to those induced by ultraviolet light in Cockayne syndrome (CS-B) fibroblast

Wed, 31 Dec 2003 23:00:00 GMT

Title: Potassium Bromate but not X-ray cause unexpectedly elevated levels of DNA breakage similar to those induced by ultraviolet light in Cockayne syndrome (CS-B) fibroblast Authors: Mosesso, Pasquale; Penna, Sabrina; Pepe, Gaetano; Lorenti Garcia, Claudia; Palitti, Fabrizio Abstract: It has been previously reported that the elevated accumulation of repair incision intermediates in cells from patients with combined characteristics of xeroderma pigmentosum complementation group D (XP-D) and Cockayne syndrome (CS) XP-D/CS fibroblasts following UV irradiation is caused by an "uncontrolled" incision of undamaged genomic DNA induced by UV-DNA-lesions which apparently are not removed. This could be an explanation for the extreme sensitivity of these cells to UV light. In the present study, we confirm the immediate DNA breakage following UV irradiation also for CS group B (CS-B) fibroblasts by DNA migration in the "comet assay" and extend these findings to other lesions such as 8-oxodeoxyguanosine (8-oxodG), selectively induced by KBrO3 treatment. In contrast, X-ray exposure does not induce differential DNA breakage. This indicates that additional lesions other than the UV-induced photoproducts (cyclobutane pyrimidine dimers, CPD, and 6-pyrimidine-4-pyrimidone products, 6-4 PP), such as 8-oxodG, specifically induced by KBrO3, are likely to trigger "uncontrolled" DNA breakage in the undamaged genomic DNA in the CS-B fibroblasts, thus accounting for some of the clinical features of these patients Description: L'articolo é disponibile sul sito dell'editore: http://www.karger.com

Relationship between chromatin structure, DNA damage and repair following X-irradiation of human lymphocytes

Thu, 31 Dec 2009 23:00:00 GMT

Title: Relationship between chromatin structure, DNA damage and repair following X-irradiation of human lymphocytes Authors: Mosesso, Pasquale; Palitti, Fabrizio; Pepe, Gaetano; Piñero, Joaquin; Bellacima, Raffaela; Ahnström, Gunnar; Natarajan, Adayapalam T. Abstract: Earlier studies using the technique of premature chromosome condensation (PCC) have shown that in human lymphocytes, exchange type of aberrations are formed immediately following low doses (<2 Gy) of X-rays, whereas at higher doses these aberrations increase with the duration of recovery. This reflects the relative roles of slow and fast repair in the formation of exchange aberrations. The underlying basis for slow and fast repairing components of the DNA repair may be related to differential localization of the initial damage in the genome, i.e., between relaxed and condensed chromatin. We have tried to gain some insight into this problem by (a) X-irradiating lymphocytes in the presence of dimethyl sulfoxide (DMSO) a potent scavenger of radiation-induced .OH radicals followed by PCC and (b) probing the damage and repair in two specific chromosomes, 18 and 19, which are relatively poor and rich in transcribing genes by COMET-FISH, a combination of Comet assay and fluorescence in situ hybridization (FISH) techniques. Results obtained show (a) that both fast appearing and slowly formed exchange aberrations seem to take place in relaxed chromatin, since they are affected to a similar extent by DMSO, (b) significant differential DNA breakage of chromosome 18 compared to chromosome 19 in both G0 and G1 phases of the cell cycle as detected by Comet assay, indicating that relaxed chromatin containing high densities of transcriptionally active genes shows less fragmentation due to fast repair (chromosome 19) compared to chromosome 18, and (c) that relaxed chromatin is repaired or mis-repaired faster than more compact chromatin Description: L'articolo é disponibile sul sito dell'editore: http://www.sciencedirect.com