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Analysis of microRNAs in lung cells exposed to particulate matter (PM10)
Abstracts / Toxicology Letters 259S (2016) S73–S247
P1: Air Pollution PO1.1 Comparative analysis of toxicity of diesel engine particles generated from the combustion of 1st and 2nd generation biodiesel fuels in vitro A. Lankoff 1,4 , M. Kowalska 1 , A. Wegierek-Ciuk 1 , H. Lisowska 1 , J. Czarnocka 2 , M. Odziemkowska 2 , R. Mruk 3 , J. Gromadzka-Ostrowska 3 , K. Dziendzikowska 3 , M. Oczkowski 3 , S. Meczynska-Wielgosz 4 , M. Wojewodzka 4 , J. Øvrevik 5 , M. Oddvar 5 , P. Magnusson 5 , M. Kruszewski 4,6 1
Jan Kochanowski University, Kielce, Poland 2 Automotive Industry Institute, Warsaw, Poland 3 Warsaw University of Life Sciences, Warsaw, Poland 4 Institute of Nuclear Chemistry and Technology, Warsaw, Poland 5 Norwegian Institute of Public Health, Oslo, Norway 6 Institute of Rural Health, Lublin, Poland Introduction: Currently biodiesel fuels are used at low levels throughout Europe, but vision plans has been proposed to increase the shear of biocomponents in diesel fuels considerably in the near future. Hence, the potential health consequences of these new fuels need to be critically assessed before large-scale introduction. Objective: Taking this into consideration the aim of our study was to determine the toxic effects of diesel engine particles (DEPs) generated from the combustion of 1st and 2nd generation biodiesel fuels in vitro in BEAS-2B cells. Materials and methods: The uptake kinetics of DEPs into BEAS2B cells was determined by flow cytometry and TEM microscopy. Cytotoxicity was determined by the Sulforodamine assay. Genotoxicity was evaluated by the alkaline comet assay and micronucleus assay. Results: The results of our study showed that DEPs generated from the combustion of B7, B20 and SHB biodiesel fuels have been effectively taken up by the BEAS-2B cells and that the concentration of DEPs in cells was a dose- and time-dependent. All three kinds of DEPs significantly decreased a cell protein content and increased a rate of DNA damage in BEAS-2B cells. Conclusions: The induction of cytotoxic and genotoxic response was dependent on the ratio to which biofuel was blended and on the chemical composition of 1st and 2nd generation biofuels. DEPs from the combustion of B7 biofuel were found to be the most toxic, whereas DEPs from the combustion of B20 were found to be the least toxic to BEAS-2B cells. To conclude, our study clearly highlights that a chemical composition of organic species adsorbed on DEPs plays a crucial role in their toxicity. Financial support: This project is funded from Norway Grants in the Polish-Norwegian Research Programme operated by the National Centre for Research and Development: PolNor/201040/72/2013 [FuelHealth]. http://dx.doi.org/10.1016/j.toxlet.2016.07.178
S73
PO1.2 Analysis of microRNAs in lung cells exposed to particulate matter (PM10 ) E.M. Quezada-Maldonado 1 , C.M. García-Cuellar 1 , F. Vaca-Paniagua 1,2 , Y. Sánchez-Pérez 1 , Y.I. Chirino 2 1 Subdirección de Investigación Básica, Instituto Nacional de Cancerología, Ciudad de México, Mexico 2 Unidad de Biomedicina, Facultad de Estudios Superiores Iztacala, UNAM, Estado de México, Mexico
Introduction: Air pollution has been classified as a human carcinogen since 2013. Particulate matter with a mean aerodynamic diameter of 10 m (PM10 ) is one of the main pollutants which contains organic, inorganic and biological components. After PM10 inhalation biological effects include inflammatory cytokines release, genotoxic damage, oxidative stress and cytoskeleton remodeling. At epigenetic level induce changes in gene methylation and modification of histones. Thus far it is known that the chronic exposure to PM represents a risk factor for development lung cancer, although previous studies provide relevant information about the effects of the PM, the carcinogenic molecular mechanisms still need to be further described. In this sense, the effects of particulate matter on microRNAs (miRNA) expression regulation have not been sufficiently explored. miRNA molecules can regulate more than 60% of human genes at post-transcriptional level, including oncogenes and tumor suppressor genes, in addition miRNAs participate in different biological processes involved in tumors development. Objective: Evaluate global changes in miRNA expression levels and their target genes in A549 cells exposed to PM10 from Mexico City. Material and methods: The human epithelial A549 cell line was exposed to 10 g/cm2 of PM10 for 72 h, the total RNA was extracted and RNA samples were hybridized in Affymetrix GeneChip microarrays to evaluate the expression level of 2578 human miRNAs. Data normalization (RMA, quantile, median polish, fold change of 1.5, p ≤ 0.05) and statistical analysis (ANOVA) were performed using Partek Genomics Suite (Partek Inc). DIANA-mirPathv.3 was used to predict miRNA targets. Results: We identified 74 miRNAs that were significantly deregulated (45 up-regulated and 29 down-regulated). The bioinformatics evaluations indicated that the target genes of the deregulated miRNAs participate in molecular processes like regulation of cell cycle, DNA damage response, proliferation and regulatory antiapoptotic process. After to review the literature, we determine that some of this miRNAs are associated with lung, gastric, liver and bladder neoplasms. Conclusion: PM10 exposure induced changes in the expression of various miRNAs that may play important roles in the regulation of different pathways associated with carcinogenic process. Financial support: This work received support from SEPCONACYT 180471 and a Master Scholarship from CONACyT (CVU 628260). http://dx.doi.org/10.1016/j.toxlet.2016.07.179
P1: Air Pollution PO1.1 Comparative analysis of toxicity of diesel engine particles generated from the combustion of 1st and 2nd generation biodiesel fuels in vitro A. Lankoff 1,4 , M. Kowalska 1 , A. Wegierek-Ciuk 1 , H. Lisowska 1 , J. Czarnocka 2 , M. Odziemkowska 2 , R. Mruk 3 , J. Gromadzka-Ostrowska 3 , K. Dziendzikowska 3 , M. Oczkowski 3 , S. Meczynska-Wielgosz 4 , M. Wojewodzka 4 , J. Øvrevik 5 , M. Oddvar 5 , P. Magnusson 5 , M. Kruszewski 4,6 1
Jan Kochanowski University, Kielce, Poland 2 Automotive Industry Institute, Warsaw, Poland 3 Warsaw University of Life Sciences, Warsaw, Poland 4 Institute of Nuclear Chemistry and Technology, Warsaw, Poland 5 Norwegian Institute of Public Health, Oslo, Norway 6 Institute of Rural Health, Lublin, Poland Introduction: Currently biodiesel fuels are used at low levels throughout Europe, but vision plans has been proposed to increase the shear of biocomponents in diesel fuels considerably in the near future. Hence, the potential health consequences of these new fuels need to be critically assessed before large-scale introduction. Objective: Taking this into consideration the aim of our study was to determine the toxic effects of diesel engine particles (DEPs) generated from the combustion of 1st and 2nd generation biodiesel fuels in vitro in BEAS-2B cells. Materials and methods: The uptake kinetics of DEPs into BEAS2B cells was determined by flow cytometry and TEM microscopy. Cytotoxicity was determined by the Sulforodamine assay. Genotoxicity was evaluated by the alkaline comet assay and micronucleus assay. Results: The results of our study showed that DEPs generated from the combustion of B7, B20 and SHB biodiesel fuels have been effectively taken up by the BEAS-2B cells and that the concentration of DEPs in cells was a dose- and time-dependent. All three kinds of DEPs significantly decreased a cell protein content and increased a rate of DNA damage in BEAS-2B cells. Conclusions: The induction of cytotoxic and genotoxic response was dependent on the ratio to which biofuel was blended and on the chemical composition of 1st and 2nd generation biofuels. DEPs from the combustion of B7 biofuel were found to be the most toxic, whereas DEPs from the combustion of B20 were found to be the least toxic to BEAS-2B cells. To conclude, our study clearly highlights that a chemical composition of organic species adsorbed on DEPs plays a crucial role in their toxicity. Financial support: This project is funded from Norway Grants in the Polish-Norwegian Research Programme operated by the National Centre for Research and Development: PolNor/201040/72/2013 [FuelHealth]. http://dx.doi.org/10.1016/j.toxlet.2016.07.178
S73
PO1.2 Analysis of microRNAs in lung cells exposed to particulate matter (PM10 ) E.M. Quezada-Maldonado 1 , C.M. García-Cuellar 1 , F. Vaca-Paniagua 1,2 , Y. Sánchez-Pérez 1 , Y.I. Chirino 2 1 Subdirección de Investigación Básica, Instituto Nacional de Cancerología, Ciudad de México, Mexico 2 Unidad de Biomedicina, Facultad de Estudios Superiores Iztacala, UNAM, Estado de México, Mexico
Introduction: Air pollution has been classified as a human carcinogen since 2013. Particulate matter with a mean aerodynamic diameter of 10 m (PM10 ) is one of the main pollutants which contains organic, inorganic and biological components. After PM10 inhalation biological effects include inflammatory cytokines release, genotoxic damage, oxidative stress and cytoskeleton remodeling. At epigenetic level induce changes in gene methylation and modification of histones. Thus far it is known that the chronic exposure to PM represents a risk factor for development lung cancer, although previous studies provide relevant information about the effects of the PM, the carcinogenic molecular mechanisms still need to be further described. In this sense, the effects of particulate matter on microRNAs (miRNA) expression regulation have not been sufficiently explored. miRNA molecules can regulate more than 60% of human genes at post-transcriptional level, including oncogenes and tumor suppressor genes, in addition miRNAs participate in different biological processes involved in tumors development. Objective: Evaluate global changes in miRNA expression levels and their target genes in A549 cells exposed to PM10 from Mexico City. Material and methods: The human epithelial A549 cell line was exposed to 10 g/cm2 of PM10 for 72 h, the total RNA was extracted and RNA samples were hybridized in Affymetrix GeneChip microarrays to evaluate the expression level of 2578 human miRNAs. Data normalization (RMA, quantile, median polish, fold change of 1.5, p ≤ 0.05) and statistical analysis (ANOVA) were performed using Partek Genomics Suite (Partek Inc). DIANA-mirPathv.3 was used to predict miRNA targets. Results: We identified 74 miRNAs that were significantly deregulated (45 up-regulated and 29 down-regulated). The bioinformatics evaluations indicated that the target genes of the deregulated miRNAs participate in molecular processes like regulation of cell cycle, DNA damage response, proliferation and regulatory antiapoptotic process. After to review the literature, we determine that some of this miRNAs are associated with lung, gastric, liver and bladder neoplasms. Conclusion: PM10 exposure induced changes in the expression of various miRNAs that may play important roles in the regulation of different pathways associated with carcinogenic process. Financial support: This work received support from SEPCONACYT 180471 and a Master Scholarship from CONACyT (CVU 628260). http://dx.doi.org/10.1016/j.toxlet.2016.07.179