EFFETTI BIOLOGICI PRECOCI DELL'INQUINAMENTO ATMOSFERICO NEI BAMBINI: LO STUDIO RESPIRA E IL PROGETTO MAPEC_LIFE

EARLY BIOLOGICAL EFFECTS OF AIR POLLUTION ON CHILDREN: THE RESPIRA STUDY AND THE MAPEC_LIFE PROJECT Elisabetta Ceretti Abstract of the PhD Thesis Background Air pollution is a global problem: airborne or deposited pollutants can be found worldwide, from highly polluted to remote areas. Epidemiological studies attribute the most severe effects from air pollution to particulate matter, which has been associated with cardiovascular diseases, lung cancer and other chronic diseases. In 2013, the International Agency for Research on Cancer (IARC) classified air pollution and particulate matter as carcinogenic to human. Among the whole population, children are at higher risk of suffering the health consequences of airborne chemicals, for various reason. First, children have higher level of physical activity, spend more time outside and have a higher air intake than adults. Second, children are more vulnerable to the adverse effects of air pollution due to their small body size, fast growth rate and relatively immature organs (lungs, in particular), body function, immune system and cell repair mechanisms. Lastly, some data suggest that genetic damage, caused by environmental pollutants, viruses or lifestyle factors, occurring early in life can increase the risk of carcinogenesis in adulthood. Various studies have analyzed the genotoxic effects of urban air pollution exposure in the general population and in highly exposed subjects. In particular, a significant association was found between high levels of urban pollution (PM10 and ozone) and DNA damage detected by the comet assay in human blood lymphocytes and leukocytes and nasal mucosa cells. As regard children, very few data are available on biomarkers of early effect of air pollution. Methods In this research work, some results of two molecular epidemiology cross-sectional projects are presented. Both of them had the objective of evaluating the associations between air pollution and early biological effects in children. The first is the RESPIRA study (Italian acronym for “Rischio ESPosizione Inquinamento aRia Atmosferica”), a small pilot study performed on pre-school children living and attending pre-school in Brescia, a highly polluted town in Northern Italy. The children were recruited in 6 pre-schools located in different areas of the town and their buccal cells were collected to evaluate two biomarkers of early effects: primary DNA damage, detected by comet assay in salivary leukocytes, and micronucleus frequency, investigated in epithelial buccal cells. Child exposure to air pollution was assessed analyzing PM0.5 samples collected near each school in the same days of biological sampling, and retrieving air quality data from the Regional Agency for Environmental Protection. Furthermore, information about some confounding factors was collected by means of a questionnaire filled in by children’s parents. The second study is the MAPEC_LIFE project (Monitoring Air Pollution Effects on Children for supporting public health policy), funded by EU Life+ Programme (LIFE12 ENV/IT/000614) which, in addition to the evaluation of the associations between air pollution and early biological effects in children, aims to propose a model for estimating the global risk of early biological effects due to air pollutants and other factors in children. The MAPEC_LIFE project was carried out on 6-8-year-old children living in five Italian towns in two different seasons. Two biomarkers of early biological effects, primary DNA damage detected with the comet assay and frequency of micronuclei, were investigated in buccal cells of children. Details of children diseases, socio-economic status, exposures to other pollutants and life-style were collected using a questionnaire administered to children’s parents. Child exposure to urban air pollution was assessed by analysing PM0.5 samples collected in the school areas for PAHs and nitro-PAHs concentration, lung toxicity and in vitro genotoxicity on bacterial and human cells. Data on the chemical features of the urban air during the study period were obtained from the Regional Agency for Environmental Protection. The project created also the opportunity to approach the issue of air pollution with the children, trying to raise their awareness on air quality, its health effects and some healthy behaviors by means of an educational intervention in the schools. Results The RESPIRA study involved six pre-schools in Brescia, in two consecutive winter seasons. During the sampling months, PM10, PM2.5 and NO2 were very often over the EU limit values for daily means. Organic extracts of PM0.5 collected near schools induced genotoxic effects in bacterial and human cells in in vitro tests. Regarding DNA damage in children cells, mean micronucleus frequency in epithelial buccal cells of children was 0.29 ± 0.13%, higher than usually found among children living in areas with low or medium-high levels of air pollution, and significantly associated with the concentration of PM10, PM2.5 and NO2. On the other hand, the preliminary results of the comet assay showed some differences between primary DNA damage detected in children attending schools in the different areas of the town. This difference was not detected with the micronucleus test. However, the level of damage resulted from the comet assay on salivary leukocytes of children was not comparable with other literature data, due to the lack of similar studies. The MAPEC_LIFE project involved 26 primary schools in the five Italian towns. Environmental and biological samplings were repeated in the same schools and on the same children for two different seasons, winter and late spring. A total of 1125 children were recruited and sampled for two times. The results of the various in vitro and in vivo tests are still ongoing. Some preliminary results about the samples of the first season in Brescia were presented here. PM10, PM2,5 and NO2 levels remained high for all the winter period, even if they were lower than those registered in the RESPIRA seasons. The organic extracts of PM0.5 collected near each school induced point mutation in Salmonella typhimurium, particularly in the YG1021 strain, but the number of net revertants per cubic meter seem to be slightly lower than other data found in literature. As regard genotoxicity tests on buccal cells of the 283 children recruited in Brescia, no data were available so far for the comet assay, which encountered some reading difficulties. On the other hand, micronucleus frequency detected in MAPEC_LIFE children (0.06 ± 0.08%) was very lower than micronucleus levels found in the RESPIRA study. Statistical analysis of the results is still ongoing, but at the end they will show if the differences in micronucleus frequency between the two studies are due to the level of exposure to air pollution experienced by the children or to other factors. Conclusions The associations between levels of air pollutants, air mutagenicity and biomarkers of early effects will be investigated. A tentative model to calculate the global absolute risk of having early biological effects for air pollution and other variables together will be proposed and may be useful to support policy-making and community interventions to protect children from possible health effects of air pollutants.