Cytotoxic and genotoxic assessment of Euterpe oleracea fruit oil and Pentaclethra macroloba oil in...
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S126 Abstracts / Toxicology Letters 221S (2013) S59–S256 their use as a follow-up test in the assessment of the genotoxic hazard of cosmetic ingredients in the absence of in vivo data. http://dx.doi.org/10.1016/j.toxlet.2013.05.228 P10-04 The hen’s egg test for micronucleus-induction (HET-MN): Results of the method transfer prior a validation study K. Reisinger 1,∗ , D. Fieblinger 2 , A. Heppenheimer 3 , J. Kreutz 1 , A. Luch 2 , K. Maul 2 , R. Pirow 2 , A. Poth 3 , P. Strauch 1,2,3,4 , T. Wolf 4 1 Henkel AG & Co KgaA, Duesseldorf, Germany, 2 Federal Institute for Risk Assessment, Berlin, Germany, 3 Harlan Cytotest Cell Research, Rossdorf, Germany, 4 University of Osnabrueck, Osnabrueck, Germany The HET-MN assay is distinguished from other in-vitro geno- toxicity assays by toxicologically important properties such as distribution, metabolic activation, and excretion of the test com- pound. As a promising follow-up to supplement existing in-vitro test batteries for genotoxicity, the HET-MN is currently undergoing a formal validation. The hen’s egg test for micronucleus induction was developed during the last years at the University of Osnabrueck. The method was transferred the first time to the test naive laboratories of Henkel in 2007. To assess its applicability and robustness, a study was carried out at the laboratories of the University of Osnabrueck and Henkel. Following the first method transfer, an interlabora- tory study was conducted analyzing 14 test substances in both labs. Meanwhile, 24 compounds have been tested in Osnabrueck or in both labs which were all predicted correctly. In 2010 the method was transferred to the third (Federal Insti- tute for Risk Assessment, BfR) and fourth lab (Harlan Cytotest Cell Research). In the first phase of the transfer Cyclophosphamide (CP) and 7,12-dimethylbenzanthracene (DMBA) were examined. In the second phase, using CP and DMBA as positive controls, three com- pounds (Ampicillin, Carbendazim, Methotrexate) were tested in a blind study in the laboratories of the BfR, Harlan, and Henkel. These data show good intra- and inter-laboratory reproducibility. The testing of coded chemicals (which have not been tested before) in the HET-MN validation is completed for the first 6 out of 20 chemicals. http://dx.doi.org/10.1016/j.toxlet.2013.05.229 P10-05 Cytotoxic and genotoxic assessment of Euterpe oleracea fruit oil and Pentaclethra macroloba oil in human peripheral lymphocytes E.L. Maistro ∗ , E.S. Marques, M.S.F. Tsuboy Universidade Estadual Paulista (UNESP), Faculdade de Filosofia e Ciências, Departamento de Fonoaudiologia, Marília, São Paulo, Brazil Brazilian Amazon forest is especially rich in medicinal plant species and it is an important source of remedies. Euterpe oler- acea (ac ¸ aí) and Pentaclethra macroloba (pracaxi) are Amazonic plants whose biological properties are little studied. The main use attributed to E. oleracea is its high antioxidant activity, and P. macroloba is used as an aid in healing, in skin cell renewing, and also for treating snakes bites. Due the pharmacological potential of this plants and absence of genetic toxicity studies of them, the aim of the present study was to evaluate the cytotoxic and genotoxic potential of E. oleracea and P. macroloba oils. For these evalu- ations the 3-(4,5-dimethylthiazol-2-y1)-2,5-diphenyltetrazolium bromide (MTT) and trypan blue exclusion cytotoxicity assays, as well as comet assay in human lymphocytes were used. Blood sam- ples were obtained by venepuncture from two healthy donors and lymphocytes were isolated using Histopaque 1077. For the exper- iments, 2 × 10 5 cells were seeded in each well of a 24 cell culture microplate. The cells were treated for 4 h with E. oleracea and P. macroloba oils at concentrations of 2.5, 5 and 10 g/mL, as well as MMS 75 M or DMSO 1% as positive and negative control, respec- tively. The DNA damage in comet assay was visually scored and classified as class 0 to class 3. Both plant oils, at concentrations studied, has not induced cytotoxicity or DNA damage when com- pared with the control. Complementary genotoxic assays are being performed to attest the safe use of this Amazon plant species as phytomedicine by humans. http://dx.doi.org/10.1016/j.toxlet.2013.05.230 P10-06 Determination of lymphocyte DNA damage using the comet assay in sandbalsting workers exposed to crystalline silica dust Esma Söylemez ∗ , Zeliha Kayaaltı, Dilek Kaya Akyüzlü, Tülin Söylemezo˘ glu Ankara University, Institute of Forensic Sciences, Ankara, Turkey Silicosis is the most common disabling occupational lung dis- ease that results from exposure of respirable crystalline silica dust. This disease has been associated with a wide variety of indus- trial activities and occupations such as miners, sandbalsters, stone cutters and ceramic workers. Crystalline silica from occupational sources has been indicated as a carcinogen according to Interna- tional Agency for Research on Cancer (IARC). Studies showed that crystalline silica led to free radical mediated DNA damage. Sin- gle cell gel electrophoresis technique (SCGE) also known as the comet assay has been widely used for environmental genotoxicity and biomonitoring studies. This technique can be used to provide an early indicator for genetic disease or cancer and exposure to a wide variety of genotoxic agents; it is also a sensitive endpoint for detecting DNA damage. In this study, we used the comet assay to evaluate DNA damage in the peripheral lymphocytes of 25 sand- balsting workers exposed to crystalline silica dust and 25 healthy subjects with no history of occupational silica or chemical exposure. The levels of DNA damage were measured by BAB Bs comet Assay system and evaluated by the percentage of DNA in the comet tail (% tail DNA) for each cell. According to our results, increased DNA damage was found in workers exposed to crystalline silica dust compared to controls. % tail DNA in crystalline silica dust-exposed group was significantly higher than the control group (62.36 ± 4.09 vs 47.58 ± 7.93; p < 0.001). http://dx.doi.org/10.1016/j.toxlet.2013.05.231
Transcript of Cytotoxic and genotoxic assessment of Euterpe oleracea fruit oil and Pentaclethra macroloba oil in...
S126 Abstracts / Toxicology Letters 221S (2013) S59–S256
their use as a follow-up test in the assessment of the genotoxichazard of cosmetic ingredients in the absence of in vivo data.
http://dx.doi.org/10.1016/j.toxlet.2013.05.228
P10-04The hen’s egg test for micronucleus-induction(HET-MN): Results of the method transfer priora validation study
K. Reisinger 1,∗, D. Fieblinger 2, A. Heppenheimer 3, J. Kreutz 1, A.Luch 2, K. Maul 2, R. Pirow 2, A. Poth 3, P. Strauch 1,2,3,4, T. Wolf 4
1 Henkel AG & Co KgaA, Duesseldorf, Germany, 2 Federal Institute forRisk Assessment, Berlin, Germany, 3 Harlan Cytotest Cell Research,Rossdorf, Germany, 4 University of Osnabrueck, Osnabrueck, Germany
The HET-MN assay is distinguished from other in-vitro geno-toxicity assays by toxicologically important properties such asdistribution, metabolic activation, and excretion of the test com-pound. As a promising follow-up to supplement existing in-vitrotest batteries for genotoxicity, the HET-MN is currently undergoinga formal validation.
The hen’s egg test for micronucleus induction was developedduring the last years at the University of Osnabrueck. The methodwas transferred the first time to the test naive laboratories ofHenkel in 2007. To assess its applicability and robustness, a studywas carried out at the laboratories of the University of Osnabrueckand Henkel. Following the first method transfer, an interlabora-tory study was conducted analyzing 14 test substances in both labs.Meanwhile, 24 compounds have been tested in Osnabrueck or inboth labs which were all predicted correctly.
In 2010 the method was transferred to the third (Federal Insti-tute for Risk Assessment, BfR) and fourth lab (Harlan Cytotest CellResearch). In the first phase of the transfer Cyclophosphamide (CP)and 7,12-dimethylbenzanthracene (DMBA) were examined. In thesecond phase, using CP and DMBA as positive controls, three com-pounds (Ampicillin, Carbendazim, Methotrexate) were tested in ablind study in the laboratories of the BfR, Harlan, and Henkel. Thesedata show good intra- and inter-laboratory reproducibility.
The testing of coded chemicals (which have not been testedbefore) in the HET-MN validation is completed for the first 6 outof 20 chemicals.
http://dx.doi.org/10.1016/j.toxlet.2013.05.229
P10-05Cytotoxic and genotoxic assessment of Euterpeoleracea fruit oil and Pentaclethra macroloba oilin human peripheral lymphocytes
E.L. Maistro ∗, E.S. Marques, M.S.F. Tsuboy
Universidade Estadual Paulista (UNESP), Faculdade de Filosofia eCiências, Departamento de Fonoaudiologia, Marília, São Paulo, Brazil
Brazilian Amazon forest is especially rich in medicinal plantspecies and it is an important source of remedies. Euterpe oler-acea (acaí) and Pentaclethra macroloba (pracaxi) are Amazonicplants whose biological properties are little studied. The main useattributed to E. oleracea is its high antioxidant activity, and P.macroloba is used as an aid in healing, in skin cell renewing, andalso for treating snakes bites. Due the pharmacological potential ofthis plants and absence of genetic toxicity studies of them, the aimof the present study was to evaluate the cytotoxic and genotoxic
potential of E. oleracea and P. macroloba oils. For these evalu-ations the 3-(4,5-dimethylthiazol-2-y1)-2,5-diphenyltetrazoliumbromide (MTT) and trypan blue exclusion cytotoxicity assays, aswell as comet assay in human lymphocytes were used. Blood sam-ples were obtained by venepuncture from two healthy donors andlymphocytes were isolated using Histopaque 1077. For the exper-iments, 2 × 105 cells were seeded in each well of a 24 cell culturemicroplate. The cells were treated for 4 h with E. oleracea and P.macroloba oils at concentrations of 2.5, 5 and 10 �g/mL, as well asMMS 75 �M or DMSO 1% as positive and negative control, respec-tively. The DNA damage in comet assay was visually scored andclassified as class 0 to class 3. Both plant oils, at concentrationsstudied, has not induced cytotoxicity or DNA damage when com-pared with the control. Complementary genotoxic assays are beingperformed to attest the safe use of this Amazon plant species asphytomedicine by humans.
http://dx.doi.org/10.1016/j.toxlet.2013.05.230
P10-06Determination of lymphocyte DNA damageusing the comet assay in sandbalsting workersexposed to crystalline silica dust
Esma Söylemez ∗, Zeliha Kayaaltı, Dilek Kaya Akyüzlü, TülinSöylemezoglu
Ankara University, Institute of Forensic Sciences, Ankara, Turkey
Silicosis is the most common disabling occupational lung dis-ease that results from exposure of respirable crystalline silica dust.This disease has been associated with a wide variety of indus-trial activities and occupations such as miners, sandbalsters, stonecutters and ceramic workers. Crystalline silica from occupationalsources has been indicated as a carcinogen according to Interna-tional Agency for Research on Cancer (IARC). Studies showed thatcrystalline silica led to free radical mediated DNA damage. Sin-gle cell gel electrophoresis technique (SCGE) also known as thecomet assay has been widely used for environmental genotoxicityand biomonitoring studies. This technique can be used to providean early indicator for genetic disease or cancer and exposure to awide variety of genotoxic agents; it is also a sensitive endpoint fordetecting DNA damage. In this study, we used the comet assay toevaluate DNA damage in the peripheral lymphocytes of 25 sand-balsting workers exposed to crystalline silica dust and 25 healthysubjects with no history of occupational silica or chemical exposure.The levels of DNA damage were measured by BAB Bs comet Assaysystem and evaluated by the percentage of DNA in the comet tail(% tail DNA) for each cell. According to our results, increased DNAdamage was found in workers exposed to crystalline silica dustcompared to controls. % tail DNA in crystalline silica dust-exposedgroup was significantly higher than the control group (62.36 ± 4.09vs 47.58 ± 7.93; p < 0.001).
http://dx.doi.org/10.1016/j.toxlet.2013.05.231
