Biomedical subjects

V Garaj-Vrhovac

Publications and source records attributed to V Garaj-Vrhovac.

At least 19 recordsLinked to original sources

PubMed2006

Alkaline comet assay study with breast cancer patients: evaluation of baseline and chemotherapy-induced DNA damage in non-target cells.

The sensitivity of the alkaline comet assay for the evaluation of baseline and treatment-induced DNA damage in white blood cells of breast cancer patients receiving adjuvant chemotherapy according to three conventional anthracycline- and cyclophosphamide-containing protocols was investigated. Additionally, baseline DNA damage in cancer patients was compared with the levels of DNA damage recorded in healthy women. Altogether 30 patients with diagnosed breast cancer and 30 female blood donors with no known familial history of breast cancer participated in the study. Alkaline comet assay was performed according to standard protocol and DNA migration in peripheral blood leukocytes was measured by a computer-based image analysis system. For each subject the frequency of "damaged" cells, i.e., long-tailed nuclei (LTN) with tail length exceeding 95th percentile for the considered parameter among controls, is also reported. Breast cancer patients had significantly increased background levels of DNA damage in their peripheral blood leukocytes as compared to healthy women. Prior to the chemotherapy, a majority of patients showed a statistically significant increase in the number of LTN compared to healthy blood donors. Marked interindividual variations in baseline DNA damage among patients were recorded, some of them related to the disease stage and status. The present study confirmed the alkaline comet assay as a sensitive technique able to detect significantly elevated DNA migration in blood cells of patients already one hour after completion of the first cycle of chemotherapy. Administration of antineoplastic drugs in three chemotherapy protocols studied induced a similar increase of primary DNA damage in nontarget cells. The evaluation of the LTN frequencies indicates the best response to the protocol containing cyclophosphamide, methotrexate and 5-fluorouracil (CMF). Our results point to the significance of simultaneous evaluation of DNA migration and frequency of LTN in the same subject and approved the use of alkaline comet assay as a suitable method for the routine detection of critical DNA lesions produced after administration of antineoplastic drugs in the clinical settings.

Adult↗

PubMed2004

Genotoxicity evaluation of pesticide formulations containing alachlor and atrazine in multiple mouse tissues (blood, kidney, liver, bone marrow, spleen) by comet assay.

Every year, in the European countries more than 2 million tons of pesticides are released into the environment. More than 60% of those substances appear to be herbicides. Due to extensive production and application of this chemical their putative detrimental effect on life should be known and minimized. In this study we applied the comet assay on blood and 4 mouse organs (kidney, liver, bone marrow, and spleen) to evaluate possible genome damage caused by two pesticide formulations (Bravo and Gesaprim) containing alachlor and atrazine as active ingredients. Five male CBA mice were assigned to each of 4 treatment groups and control group. Bravo and Gesaprim were injected intraperitoneally once. Two different doses of Bravo were used: 0.031 ml/kg and 0.021 microl/kg, so that doses of alachlor mice received within the pesticide formulation given were 15 mg/kg and 0.01 mg/kg. Also Gesaprim was given at two different doses: 1.08 ml/kg and 0.07 microl/kg so that the doses of atrazine contained within the pesticide formulation given were 540 mg/kg and 3.5 x 10(-2) mg/kg. Mice were sacrificed 24 hours after treatment. Alkaline comet assay on the blood samples, kidney, liver, bone marrow and spleen was performed. Statistically significant (p<0.01) increase of tail length for all 5 tissues examined in mice treated with both Bravo and Gesaprim compared to the control was found. For both pesticides DNA of kidney and liver showed largest increase in migration. Also, distribution of tail length values for Bravo and Gesaprim for all mouse tissues examined showed a shift to the right when compared to the controls.

Acetamides↗

PubMed2003

Examination of cytotoxicity and mutagenicity of AH26 and AH Plus sealers.

AIM: To study in vitro the cytotoxic and mutagenic effects of AH26 and AH Plus. METHODOLOGY: Cytotoxic effects on Chinese hamster V79 cells were determined by counting viable cells following incubation with eluations of AH26 and AH Plus. In one set of experiments, the materials were mixed, set for 1 h and then eluted with dimethyl sulphoxide (DMSO) for 1 h, 24 h and 7 days. In the other set, AH26 and AH Plus were mixed and set for 1 h, 24 h and 7 days in physiological saline then crushed and eluted in DMSO for 24 h. The cytotoxic effects of these eluates were evaluated. Three concentrations were chosen to examine the mutagenic effects of AH26 and AH Plus: 5.57, 16.7 and 55.7 microg mL(-1). The structural chromosomal aberration analysis and micronucleus test were performed on human lymphocytes according to standard procedures. RESULTS: Dose-response curves of cell survival were obtained. Both materials were shown to be cytotoxic in doses larger than 55.7 microg mL(-1), except for AH26, after 7 days setting time. AH Plus was also shown to be toxic in concentrations of 16.7 microg mL(-1), except after 7 days setting time. Neither AH26 nor AH Plus induced a significant increase of chromosomal aberrations or micronuclei induction at any setting time or concentration. CONCLUSION: There was no mutagenicity found for AH26 and AH Plus on human lymphocytes in highly controlled conditions in vitro.

Animals↗

PubMed2002

Application of the alkaline comet assay in biodosimetry: assessment of in vivo DNA damage in human peripheral leukocytes after a gamma radiation incident.

The alkaline comet assay was employed in the assessment of DNA damage in leukocytes of a worker incidently exposed to gamma radiation (221 mSv, 60Co source). The comet tail lengths and tail moments were studied. By using the alkaline comet assay immediately after accidental exposure a high level of DNA damage was recorded. The highest levels of DNA damage were recorded one day and one week after the radiation incident. Later on, a decrease in both comet parameters was observed. Although the level of DNA damage was diminished during a one year period, it was still elevated compared to normal values recorded in leukocytes of a healthy, unexposed person. The results obtained indicate that the alkaline comet assay is a rapid and sensitive microdosimetric technique and is suitable for in vivo human biomonitoring, especially in cases of incidental exposure to ionising radiation.

Cobalt Radioisotopes↗

PubMed2001

Cytogenetic monitoring of croatian population occupationally exposed to a complex mixture of pesticides.

This paper describes a longitudinal study of possible genetic damage in Croatian workers occupationally exposed to a complex mixture of pesticides. The methods of choice were chromosomal aberration analysis, sister chromatid exchange analysis (SCE), micronucleus assay and comet assay. In order to determine primary genotoxic effects in workers, blood samples were taken after the workers spent 8 months in the production of pesticides. During the production all subjects were simultaneously exposed to a complex mixture of pesticides containing atrazine, alachlor, cyanazine, 2,4-dichlorophenoxyacetic acid, and malathion. To detect DNA repair in lymphocytes of the same subjects the second series of blood samples was taken 8 months after the workers were removed from production. Regardless of the time sampling time the exposed workers showed an increased number of chromosomal aberrations, SCE frequency, micronucleus (MN) frequency, and values of comet assay parameters. After 8 months of non-exposure the workers showed a significantly decreased number of chromosomal aberrations, MN frequency, and DNA migration compared to the results of the first sampling, but it was still significantly higher than in controls. Furthermore, the SCE frequency in the exposed subjects did not drop after the 8 months of non-exposure, which indicates long-term exposure to a mixture of pesticides.

Adult↗

PubMed2001

Application of the alkaline comet assay in human biomonitoring for genotoxicity: a study on Croatian medical personnel handling antineoplastic drugs.

The alkaline comet assay was used to evaluate the genotoxicity towards peripheral blood lymphocytes of medical personnel regularly handling various antineoplastic drugs with different safety precautions. The study population consisted of 50 exposed subjects working in the oncology, pulmonology, gynaecology and haematology units of nine Croatian hospitals and 20 unexposed control subjects. Peripheral blood lymphocytes from the subjects were embedded in agarose on a microscope slide and lysed; the DNA was unwound and subjected to electrophoresis at pH 13. Staining with a fluorescent dye was used to identify cells with DNA damage, as judged by increased migration of genetic material from the cell nucleus. DNA damage was quantified by measuring the displacement between the genetic material of the nucleus and the resulting tail using an image analysis system. Three parameters were used as indicators of DNA damage: i.e. tail length, percentage of DNA in the tail and tail moment. Statistically significant differences in all three parameters were observed between the exposed and control groups. Within the exposed group, there were marked differences between individuals in the comet tail parameters. In the majority of exposed subjects an effect on DNA damage of age or duration of occupational exposure could be excluded. In the exposed group, the highest level of DNA damage was recorded in subjects who used only latex gloves in their work with antineoplastic drugs. The observed DNA damage was lower in exposed subjects who used more than one type of protective equipment and who worked in a well-ventilated safety cabinet. No statistically significant differences were found between the mean values of comet tail parameters for smoking and non-smoking subpopulations from the exposed group. In view of the results obtained, the alkaline comet assay, as a simple, rapid and sensitive method, appears to be a promising additional test for biomonitoring purposes in human populations.

Adult↗

PubMed2001

Chromosomal aberration and single cell gel electrophoresis (Comet) assay in the longitudinal risk assessment of occupational exposure to pesticides.

In recent years the use of pesticides in agriculture has been increasing steadily. At present there are more than 1000 chemicals classified as pesticides. Therefore, the widespread use of pesticides and their potential genetic hazard suggests that evaluation of their genotoxicity should be extended using the newer assays now available. In the present study chromosomal aberration analysis and the alkaline single cell gel electrophoresis (Comet) assay were used to evaluate the extent of DNA damage and DNA repair in peripheral blood lymphocytes of subjects employed in pesticide production. In order to determine possible primary genotoxic effects in workers blood samples were taken after an 8 month long period of exposure to a complex mixture of pesticides. To detect the possible occurrence of DNA repair in lymphocytes of the same subjects the second blood sample was taken after an 8 month long period of absence from the pesticide exposure zone. Regardless of the period of sampling, in the exposed group statistically significantly increased numbers of aberrant cells, chromatid and chromosome breaks, acentric fragments and dicentric chromosomes compared with the controls were found. After the workers had spent 8 months out of the pesticide exposure zone the number of aberrant cells and all types of chromatid and chromosome aberrations decreased significantly compared with sampling after the high exposure period, but it still remained significantly higher in comparison with the control group. After the period of high exposure to a mixture of pesticides statistically significantly increased levels of DNA damage in the Comet assay in terms of tail length and tail moment were found. After the workers were removed from production for 8 months both Comet assay end-points decreased significantly compared with the first sampling point, but they remained increased compared with the control.

Adult↗

PubMed2000

Evaluation of DNA damage in workers occupationally exposed to pesticides using single-cell gel electrophoresis (SCGE) assay. Pesticide genotoxicity revealed by comet assay.

The comet assay, also called the single-cell gel electrophoresis (SCGE) assay, is a rapid and sensitive method for the detection of DNA damage (strand breaks and alkali-labile sites) in individual cells. The assay is based on the embedding of cells in agarose, their lysis in alkaline buffer and finally subjection to an electric current. In the present study, alkaline SCGE was used to evaluate the extent of primary DNA damage and DNA repair in peripheral blood lymphocytes of workers employed in pesticide production. After the period of high pesticide exposure, lymphocytes of the occupationally exposed workers manifested increased tail length and tail moment compared to the control group. After the workers spent 6 months out of the pesticide exposure zone, both endpoints were still above that of the control but significantly decreased as compared to the results of the first analysis.

Adult↗

PubMed2000

Application of cytogenetic endpoints and Comet assay on human lymphocytes treated with vincristine in vitro.

The genotoxic potential of vincristine is assessed on human peripheral blood lymphocytes following administration of the drug at a dose 0.0875 microg/ml by use of single cell gel electrophoresis - Comet assay (SCGE), analysis of structural chromosome aberrations (CA), micronucleus assay (MN) and sister chromatid exchange (SCE) analysis. In vitro treatment of human lymphocytes with vincristine was performed on cells in G0 phase, as well on lymphocyte cultures 24 hours after stimulation with mitogen phytohemagglutinine. For the Comet assay at 24, 48 and 72 h the treated cells were embedded in agarose on slides, lysed with alkaline lysis solution and exposed to an electric field. DNA migrated within the agarose and formed comets whose length depends on the amount of DNA damage. For the analysis of structural CA cells were grown on F-10 medium for 48 hours, and for MN and SCE analysis for 72 hours. The results on SCGE showed an increase in tail length compared to control both in cells treated in G0 and in cells treated 24 h after mitogen stimulation. The amount of DNA damage was higher in cells treated with vincristine 24 h after mitogen stimulation. Administered concentration of drug caused total inhibition of lymphocytes growth in 72-h cultures for MN and SCE analysis indicating strong microtubule distruptive effects of vincristine. Analysis of structural CA reveals chromatid breaks and acentric fragments as the main aberration types both in cells treated in G0 and in cells treated 24 h after mitogen stimulation. Number of these aberrations was higher in cells treated in G0 phase. Results obtained in this study by use of different cytogenetic endpoints confirmed that vincristine exhibits both aneugenic and clastogenic effects on human lymphocytes.

Adult↗

PubMed1999

Possible discrepancies between dicentric chromosome frequencies and recorded ionizing radiation doses: in vivo study.

BACKGROUND: The available data on the effects of radiation on humans are limited to the reports of accidental exposure or studies of patients under diagnostic and therapeutic treatment; few reports refer to a occupationally exposed population groups. METHODS: The research was conducted on 66 subjects employed in the Department of Nuclear Medicine and 41 nonemployed controls. For each of them chromosomal analysis and gamma-spectrometer analysis of 24-hour urine were carried out. Exposure doses were measured using film-badge dosimetry. RESULTS: The comparison of dosimetric data obtained by film-badge measurements and the frequency of dicentric chromosomes in each subject revealed no correlation between the two observed parameters. CONCLUSIONS: In view of the above results, the subjects should be studied within job/task groups as they are far more likely to receive comparable doses.

Adult↗

PubMed1999

Induction of micronuclei in human lymphocytes after occupational exposure to ultrasound.

Micronucleus assay combined with Giemsa and DAPI staining was performed on blood samples of subjects occupationally exposed to ultrasound. Lymphocytes were cultivated in vitro for 72 h. At 44h cytochalasin-B was added in cultures. Frequencies of micronuclei in exposed subjects statistically significant increased compared to control. The frequency of micronucleated cells and micronuclei in exposed subjects shows interindividual variability. Using DAPI staining we observed signal-positive and signal-negative micronuclei. Percentage of signal-positive micronuclei varies between 0 and 66.7% and signal-negative micronuclei between 33.3% and 100%. This study indicate harmful effects of ultrasound on human genome, but further investigations are necessary.

Adult↗

PubMed1999

Micronucleus assay and lymphocyte mitotic activity in risk assessment of occupational exposure to microwave radiation.

The effects of radiofrequency electromagnetic radiation (RFR) on the cell kinetics and genome damages in peripheral blood lymphocytes were determined in lymphocytes of 12 subjects occupationally exposed to microwave radiation. Results showed an increase in frequency of micronuclei (MN) as well as disturbances in the distribution of cells over the first, second and third mitotic division in exposed subjects compared to controls. According to previous reports micronucleus assay can serve as a suitable indicator for the assessment of exposure to genotoxic agents (such as RFR) and the analysis of mitotic activity as an additional parameter for the efficient biomonitoring.

Adult↗

PubMed1999

Incidence of micronuclei in cytokinesis-blocked lymphocytes of medical personnel occupationally exposed to ultrasound.

In order to investigate possible DNA damaging effects of ultrasound, the micronucleus assay on cytokinesis blocked human lymphocytes was performed. Preparations were stained by conventional Giemsa staining technique combined with additional staining techniques using fluorescent dye DAPI and silver nitrate. Blood samples were taken from medical personnel employed on ultrasonic scanning in medical diagnosis and unexposed control subjects from general population. Lymphocytes were cultivated in vitro at 37 degrees C. Cytochalasin-B in final concentration of 6 microg/ml was added 44 h after mitogen stimulation and cultures were harvested 28 h thereafter. Staining with both additional techniques can be used to distinguish micronuclei originating from breakage or mitotic loss of certain human chromosomes bearing DAPI-positively stained or silver-positively stained regions. The results obtained indicate statistically significant increases in total number of micronuclei and changes in their distribution in exposed subjects compared to control. Based on different intensity of DAPI staining signal-positive and signal-negative "type" of micronuclei are distinguished, while silver staining has revealed Ag-NOR+ and Ag-NOR- micronuclei. In exposed subjects a prevalence in number of Ag-NOR+ micronuclei over Ag-NOR-micronuclei compared to control was observed, indicating greater susceptibility of chromosomes from D and G groups to damage caused by continuous occupationally exposure to ultrasound. In spite of their limitations, our results indicate that combination of conventional Giemsa staining of micronuclei with fluorescent dye DAPI and silver nitrate staining techniques can be valuable complement to the standard micronucleus assay.

Adult↗

PubMed1996

A nine-year follow up study of a population occupationally exposed to vinyl chloride monomer.

The presence of structural chromosome aberrations and sister chromatid exchange frequencies (SCE) were studied in lymphocytes from 28 male subjects occupationally exposed to vinyl chloride monomer for a period of 9 years. A significant increase in chromosomal damages and elevated SCE frequencies was detected during the third and fourth year of the follow-up study. During the last 2 years, all examined parameters are approaching control values. This is a result of decreasing vinyl chloride monomer concentrations in the working environment without interrupting the working process.

Adult↗

PubMed1996

[Exposure to low electromagnetic fields and the carcinogenesis process].

In the literature as well as in public media increasing attention is paid to possible health effects of exposure to low-frequency electric and magnetic fields, Experimental studies at the cellular level and mammalian studies in vivo have demonstrated various biological effects that are possibly associated with human exposure to low-frequency radiation. Although the exact mechanism of this relationship is not entirely clear, it has been indicated that low-frequency radiation might promote the process of carcinogenesis, although it does not seem likely to initiate it. A possible link between human low-frequency exposure and occurrence of carcinoma at child and adult age has been examined in some fifty epidemiological studies. A few studies have demonstrated a possible connection between carcinoma at child age and distance from electrical installations. Although the results point to an association between exposure to low-frequency electric and magnetic fields and cancer, further research is called for. In the meantime "cautious avoidance" of extended exposure to low-frequency electromagnetic fields is recommended.

Animals↗

PubMed1995

Relationship between locations of chromosome breaks induced by vinyl chloride monomer and lymphocytosis.

The distribution of vinyl chloride monomer (VCM)-induced chromosome breaks was studied in cultured lymphocytes of subjects occupationally exposed to this gas. In the examined subjects, the mean group value of chromosome aberrations is 6.5% and for sister chromatid exchange (SCE) frequencies, the mean value per cell is 7.9. These values are significantly higher than in the control population. Occupational exposure to VCM caused lymphocytosis together with disturbances of mitogenic activity in lymphocytes stimulated by phytohaemagglutinin. The results of G-banding showed that sites of chromosome breakpoints caused by VCM can be related to the lymphatic tissue disorders.

Adult↗

PubMed1995

Chromosome aberrations in peripheral blood lymphocytes from control individuals.

This study emphasizes the problems encountered in obtaining suitable control levels for comparison with occupational studies of exposure to clastogens. Blood samples taken from 135 individuals (69 females and 66 males) were examined for chromosome aberrations. The data include 15,368 cells scored for chromosome aberrations. The frequencies of chromatid and chromosome breaks, acentric fragments and dicentrics were determined. The frequency of the most common chromosome aberration, chromosome breaks, was 1.1 x 10(-2) and for dicentrics 0.26 x 10(-3). The values obtained were investigated in relation to sex, cigarette smoking habits, diagnostic X-ray exposure and use of antibiotics. In all the parameters, no significant differences were found.

Adolescent↗

PubMed1994

The sensitivity of the micronucleus assay for the detection of occupational exposure to vinyl chloride monomer.

The micronucleus assay was performed in the peripheral lymphocytes of 32 subjects occupationally exposed to vinyl chloride monomer (VCM) divided into two groups according to years of employment. Blood samples were taken in the period from 24 h to 90 days following a transitory exposure to elevated VCM concentrations of 300 ppm due to the technological process. In subjects with a longer period of employment micronucleus frequencies decreased in proportion to the length of the interval after the last exposure to VCM. The results confirm that the micronucleus assay can serve as a suitable indicator of the time elapsed after last exposure to elevated concentrations of environmental mutagen. It can be assumed that duration of employment may contribute to the occurrence of the cumulative effect produced by exposure to elevated VCM concentrations.

Evaluation Studies as Topic↗