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Exposure to pesticides during pregnancy by the evaluation of their concentration in amniotic fluid and maternal urine
Abstracts / Toxicology Letters 205S (2011) S60–S179
P1004 Potential intracellular tracker capacity of novel synthetic metalloporphyrins C. Constantin 1 , M. Neagu 1,∗ , R. Boscencu 2 , M.E. Hinescu 1 , A. Oliveira 3 , L.F. Vieira Ferreira 3 1
Immunology, Victor Babes National Institute, Bucharest, Romania, Pharmacy, Carol Davila University of Medicine and Pharmacy, Bucharest, Romania, 3 Centro de Química-Física Molecular (CQFM) and Instituto de Nanociencias e Nanotecnologias (IN), Instituto Superior Técnico, Universidade Técnica de Lisboa, Lisbon, Portugal 2
It is well known that photosensitizing agents based on porphyrins structures tend to accumulate preferentially in tumor cells and less in their normal counterparts. Based on this capacity they were utilized in therapy but their fluorescent properties may be also valuable for imagistic purposes. Our aim was to establish the capacity of some porphyrin families to serve as organelles/cell/tissue markers, thus setting up the cytotoxic outline of these compounds was the first objective of the project. In this view, we have synthesized some novel copper (II) and zinc (II) porphyrinic complexes intended for further development in cellular imagistic procedures. We have evaluated the dark cytotoxicity of the compounds on cell lines (U937) and isolated peripheral blood mononuclear cells (PBMC) from healthy and cancer patients. Several experimental protocols were used for cellular viability and proliferation capacity. Cellular uptake yield and efflux capacity of porphyrins were evaluated by flow-cytometry. Obtained data have proven the non-toxic nature of the novel compounds even at high doses. Furthermore, the best cellular uptake was registered after prolonged incubation. Human PBMC data revealed a different pattern of cellular response to the new complexes, suggesting an anti-tumoral action upon PBMC isolated from tumor bearing patients. Data gathered by confocal microscopy suggested that these compounds can be good candidates for cellular markers, especially in the case of normal PBMC cells. Further data will pin-point the exact intracellular compartment and the accurate tracker capacity of these fluorescent probes. The study was financed by MNT EraNet Project (Bio-Mark) no. 7030-5/2010. doi:10.1016/j.toxlet.2011.05.238 Biomarkers P1005 Exposure to pesticides during pregnancy by the evaluation of their concentration in amniotic fluid and maternal urine D. Koutroulakis 1 , A. Tsatsakis 2 , M. Tzatzarakis 3 , A. Alegakis 3,∗ , P. Fragkiadaki 3 , S. Sifakis 1 1
Department of Obstetrics and Gynaecology, University Hospital of Heraklion, Heraklio, Greece, 2 Lab of Toxicology, Medical School, University of Crete, Heraklion, Crete, Greece, 3 Laboratory of Toxicology, Medical School, University of Crete, Heraklion, Crete, Greece Purpose of this study is to associate the pesticides levels measured in amniotic fluid and maternal urine with parameters of pregnancy outcome and fetus development. Amniotic fluid (437) and urine (163) samples were collected from pregnant women who undergone an amniocentesis at the gestational age of 17–24 weeks. Samples were stored until analysis for dialkyl metabolites
S61
of organophosphates (DAPs) by GC–MS. Additional data selected at the time of birth (neonates somatometric measurements, outcome of pregnancy, etc.) Analysis was applied to data from 415 singleton births. Pregnant women were aged 32.7 ± 5.7 years old, and 144 of them located in rural areas. No life delivery was observed in 20 cases (4.8%) including intrauterine fetal death, pregnancy termination (due to chromosomal abnormalities, congenital fetal anomalies). 136 delivered neonates (31%) were preterms. Mean weight of neonates was 3123 ± 591 g, height 50.1 ± 2.3 cm, and head circumference 34.2 ± 2.2 cm. Exposure from pesticides as expressed as sums of DMPs, DAPs, DEPs in amniotic fluid samples were ranged from 0.07 to 222.9, 0.05 to 252.6, 0.19 to 254.3 measured in ng/ml, respectively. Results from urine samples were 10–99, 0.25–60.1, and 0.25–309.4 ng/ml. Increased levels of DEDTP were observed in pregnancies of pre-terms infants (p = 0.015). The existence of positive DMPs amniotic samples shows to negatively correlated with neonate height (r = −0.13) and neonate weight (r = −0.11). The correlation between neonate somatometric parameters and organophospate metabolites resulted may indicate an effect of pesticides exposure during pregnancy to the fetus growth. However, as fetal and neonate development is a multiplex process, further investigation is required to establish an aetiopathogenetic relationship. doi:10.1016/j.toxlet.2011.05.239
P1006 The mouse as biomodel in genotoxicity assays, two years of experience, Finlay institute, Cuba D.F. Arencibia 1,∗ , L.A. Rosario 2 , A. Novoa 3 1
Animal Models and Preclinical Toxicology, Researches Vicepresidency (Finlay Institute), La Habana, Cuba, 2 Microbiology, Institute of Pharmacy and Food Science, La Habana, Cuba, 3 Toxicology, Faculty of Biology, Havana University (U.H)., La Habana, Cuba The aim of this work was to evaluate and to compare the spontaneous and induced indexes in mice of both sexes of Balb/c, NMRI, OF-1 and C57/BL6/cenp lines keeping in mind the epididymal sperm concentration, spontaneous frequency of sperm heads abnormal, number of erythrocytes in bone marrow with micronuclei, cytotoxicity index (ratio of young erythrocytes/mature erythrocytes), total of cells with structural chromosome aberrations, mitotic index (number of cells at the stage of division metaphase cell) and the percentage of peripheral lymphocytes undergoing DNA damage as level 1, 2, 3, 4 from low to high damage. This study was carried out to determine the most efficient line, on the base of the significant appearance of lower spontaneous indexes and high induced indexes to mutagenic substances like the cyclophosphamide in the in vivo comet alkaline, micronuclei, chromosome aberrations and sperm head morphology assay according to the standardized protocols and adjusted by Arencibia et al. (2009, 2010). We obtained as a result that the Balb/c line in both sexes differs significantly with the other lines where they met the highest lower and induced spontaneous indexes with cyclophosphamide, keeping in mind the measured indicators. Also the line C57/BL6/cenp was less efficient and less sensitive one to the mutagen, being obtained the lowest higher and induced spontaneous results. This work will allow using the best line of mice like biomodel in the genotoxicity and antigenotoxicity assay according to low spontaneous indexes and sensibility to detect mutagenic substances in a narrow range. doi:10.1016/j.toxlet.2011.05.240
P1004 Potential intracellular tracker capacity of novel synthetic metalloporphyrins C. Constantin 1 , M. Neagu 1,∗ , R. Boscencu 2 , M.E. Hinescu 1 , A. Oliveira 3 , L.F. Vieira Ferreira 3 1
Immunology, Victor Babes National Institute, Bucharest, Romania, Pharmacy, Carol Davila University of Medicine and Pharmacy, Bucharest, Romania, 3 Centro de Química-Física Molecular (CQFM) and Instituto de Nanociencias e Nanotecnologias (IN), Instituto Superior Técnico, Universidade Técnica de Lisboa, Lisbon, Portugal 2
It is well known that photosensitizing agents based on porphyrins structures tend to accumulate preferentially in tumor cells and less in their normal counterparts. Based on this capacity they were utilized in therapy but their fluorescent properties may be also valuable for imagistic purposes. Our aim was to establish the capacity of some porphyrin families to serve as organelles/cell/tissue markers, thus setting up the cytotoxic outline of these compounds was the first objective of the project. In this view, we have synthesized some novel copper (II) and zinc (II) porphyrinic complexes intended for further development in cellular imagistic procedures. We have evaluated the dark cytotoxicity of the compounds on cell lines (U937) and isolated peripheral blood mononuclear cells (PBMC) from healthy and cancer patients. Several experimental protocols were used for cellular viability and proliferation capacity. Cellular uptake yield and efflux capacity of porphyrins were evaluated by flow-cytometry. Obtained data have proven the non-toxic nature of the novel compounds even at high doses. Furthermore, the best cellular uptake was registered after prolonged incubation. Human PBMC data revealed a different pattern of cellular response to the new complexes, suggesting an anti-tumoral action upon PBMC isolated from tumor bearing patients. Data gathered by confocal microscopy suggested that these compounds can be good candidates for cellular markers, especially in the case of normal PBMC cells. Further data will pin-point the exact intracellular compartment and the accurate tracker capacity of these fluorescent probes. The study was financed by MNT EraNet Project (Bio-Mark) no. 7030-5/2010. doi:10.1016/j.toxlet.2011.05.238 Biomarkers P1005 Exposure to pesticides during pregnancy by the evaluation of their concentration in amniotic fluid and maternal urine D. Koutroulakis 1 , A. Tsatsakis 2 , M. Tzatzarakis 3 , A. Alegakis 3,∗ , P. Fragkiadaki 3 , S. Sifakis 1 1
Department of Obstetrics and Gynaecology, University Hospital of Heraklion, Heraklio, Greece, 2 Lab of Toxicology, Medical School, University of Crete, Heraklion, Crete, Greece, 3 Laboratory of Toxicology, Medical School, University of Crete, Heraklion, Crete, Greece Purpose of this study is to associate the pesticides levels measured in amniotic fluid and maternal urine with parameters of pregnancy outcome and fetus development. Amniotic fluid (437) and urine (163) samples were collected from pregnant women who undergone an amniocentesis at the gestational age of 17–24 weeks. Samples were stored until analysis for dialkyl metabolites
S61
of organophosphates (DAPs) by GC–MS. Additional data selected at the time of birth (neonates somatometric measurements, outcome of pregnancy, etc.) Analysis was applied to data from 415 singleton births. Pregnant women were aged 32.7 ± 5.7 years old, and 144 of them located in rural areas. No life delivery was observed in 20 cases (4.8%) including intrauterine fetal death, pregnancy termination (due to chromosomal abnormalities, congenital fetal anomalies). 136 delivered neonates (31%) were preterms. Mean weight of neonates was 3123 ± 591 g, height 50.1 ± 2.3 cm, and head circumference 34.2 ± 2.2 cm. Exposure from pesticides as expressed as sums of DMPs, DAPs, DEPs in amniotic fluid samples were ranged from 0.07 to 222.9, 0.05 to 252.6, 0.19 to 254.3 measured in ng/ml, respectively. Results from urine samples were 10–99, 0.25–60.1, and 0.25–309.4 ng/ml. Increased levels of DEDTP were observed in pregnancies of pre-terms infants (p = 0.015). The existence of positive DMPs amniotic samples shows to negatively correlated with neonate height (r = −0.13) and neonate weight (r = −0.11). The correlation between neonate somatometric parameters and organophospate metabolites resulted may indicate an effect of pesticides exposure during pregnancy to the fetus growth. However, as fetal and neonate development is a multiplex process, further investigation is required to establish an aetiopathogenetic relationship. doi:10.1016/j.toxlet.2011.05.239
P1006 The mouse as biomodel in genotoxicity assays, two years of experience, Finlay institute, Cuba D.F. Arencibia 1,∗ , L.A. Rosario 2 , A. Novoa 3 1
Animal Models and Preclinical Toxicology, Researches Vicepresidency (Finlay Institute), La Habana, Cuba, 2 Microbiology, Institute of Pharmacy and Food Science, La Habana, Cuba, 3 Toxicology, Faculty of Biology, Havana University (U.H)., La Habana, Cuba The aim of this work was to evaluate and to compare the spontaneous and induced indexes in mice of both sexes of Balb/c, NMRI, OF-1 and C57/BL6/cenp lines keeping in mind the epididymal sperm concentration, spontaneous frequency of sperm heads abnormal, number of erythrocytes in bone marrow with micronuclei, cytotoxicity index (ratio of young erythrocytes/mature erythrocytes), total of cells with structural chromosome aberrations, mitotic index (number of cells at the stage of division metaphase cell) and the percentage of peripheral lymphocytes undergoing DNA damage as level 1, 2, 3, 4 from low to high damage. This study was carried out to determine the most efficient line, on the base of the significant appearance of lower spontaneous indexes and high induced indexes to mutagenic substances like the cyclophosphamide in the in vivo comet alkaline, micronuclei, chromosome aberrations and sperm head morphology assay according to the standardized protocols and adjusted by Arencibia et al. (2009, 2010). We obtained as a result that the Balb/c line in both sexes differs significantly with the other lines where they met the highest lower and induced spontaneous indexes with cyclophosphamide, keeping in mind the measured indicators. Also the line C57/BL6/cenp was less efficient and less sensitive one to the mutagen, being obtained the lowest higher and induced spontaneous results. This work will allow using the best line of mice like biomodel in the genotoxicity and antigenotoxicity assay according to low spontaneous indexes and sensibility to detect mutagenic substances in a narrow range. doi:10.1016/j.toxlet.2011.05.240