Search

Introduction Inhaled particulate pollutants have been shown to produce systemic changes in DNA methylation. Global hypomethylation has been associated to viral sequence reactivation, possibly linked to the activation of pro-inflammatory pathways occurring after exposure. We aimed at evaluating the effects of PM exposure on DNA methylation of the Wp promoter of the Epstein-Barr Virus (EBV-Wp) and the promoter of the human-endogenous-retrovirus w (HERV-w), chosen as a paradigm of an exogenous virus and an endogenous retroviral sequence, in workers in an electric furnace steel plant with well-characterised exposure to metal-rich particulate matter. Methods We measured EBV-Wp and HERV-w DNA methylation through bisulfite PCR Pyrosequencing on peripheral blood leukocytes DNA obtained from 63 male healthy workers, on the first day of a work week (baseline, after 2 days off work) and after 3 days of work (post-exposure). We determined individual exposure to inhalable particles and metals for all subjects. Paired t-test was used to compare baseline and post-exposure samples. Linear mixed models were fitted to evaluate the association between metal-rich particle exposure and DNA methylation. Results Comparing samples obtained at baseline and after 3 days of work, the mean methylation of EBV-Wp was significantly higher at baseline compared to post-exposure (baseline=56.7; postexposure=47.9; p-value=0.009), whereas the mean methylation of HERV-w did not significantly differ. In a regression model adjusted for age, body mass index and smoking, nickel, arsenic and lead had a positive association with EBV-Wp methylation (nickel: β=16.16, p-value Conclusions The difference observed comparing baseline and postexposure samples may be suggestive of a rapid change in EBV methylation induced by air particles, while correlation between EBV methylation and metal exposure may represent an adaptive mechanism that should be further characterised in future investigations.

Introduction Arsenic is an environmental carcinogen that undergoes methylation while metabolised. Subsequent intracellular methyl groups depletion can decrease global DNA methylation, facilitating carcinogenesis. On the other hand, hypermethylation in promoter regions of tumour suppressor genes related to arsenic exposure has been observed in-vivo. The effects of arsenic on enzymes regulating gene-specific methylation (DNA-methyltransferases) have not been fully investigated. We analysed methyltransferase gene expression in workers exposed to high levels of metal-rich particles, and its relationship with the DNA methylation of nine inflammatory and tumour suppressor genes. Methods We recruited 63 healthy male foundry workers. Individual exposure to arsenic and other metals was estimated based on metal components in PM10 collected in each work area (by coupled-plasma mass spectrometer) and on time spent in different areas by each worker. Methyltransferase gene expression was measured by real-time PCR in blood leucocytes DNA. Gene-specific DNA methylation was measured trough bisulfite PCR-pyrosequencing. Multivariable linear regression models adjusted for age, BMI, smoking and drug consumption, were applied to assess the association between exposure and methyltransferase expression and, in turn, between methyltransferase expression and gene-specific methylation. Geometric mean ratios (GMR) were used to express results of log-transformed variables. Results Enhanced methyltransferase (DNMT3B) gene expression was associated with increased exposure to arsenic (GMR=1.52, 95% CI: 1.06 to 2.20) and to other contaminants (Cu, Mo, Sn, Sb). DNMT3B expression was in turn associated with hypermethylation of the RASSF1A tumour suppressor gene (β=0.54, 95% CI: +0.15 to +0.94) and with hypomethylation of the Et-1 and IL-6 genes. Discussion Our preliminary data suggest the possible role of methyltransferase gene overexpression in the pathway linking metal exposure to oncogene regulation. In particular, we found an increased DNMT3B expression in arsenic-exposed workers, that resulted also in hypermethylation (down-regulation) of RASSF1A. Interestingly, RASSF1A is a tumour suppressor gene involved in the development of cancers related to As exposure (bladder, lung).

Introduction Brazil is one of the major producers of chrysotile, a well-known carcinogen, but few studies evaluated its impact locally. For these reasons, a joint project, supported by CAPES, Brasilia, was started between the Federal University of Parana (UFPR), Curitiba, Brazil, and the Fondazione IRCCS Ca’ Granda, Milan, Italy. Methods The objects of the joint project include: a cohort mortality study among asbestos–cement workers in the Curitiba Metropolitan Region; the implementation of a registry of malignant mesothelioma (MM) in Curitiba based on the Italian model. MM cases are already collected (1998–2012) in the Curitiba Population-Based Cancer Registry (RCBP); but information on asbestos exposure is unavailable and diagnostic accuracy of MM has never been evaluated. For this reason, we translated the Italian standardised questionnaire on asbestos exposure and are implementing the database for MM cases in Portuguese. Clinical documentation of adult subjects with ICD-10 codes C45 (mesothelioma), C38 (mediastinal and pleural cancers), and C48 (peritoneal cancers) in the RCBP will be examined to evaluate diagnostic accuracy. Results The questionnaire has been preliminarily tested on 40 workers in Curitiba. The software has been translated and adapted. Review of clinical records of 269 subjects is in progress; we identified about 20% of death certificate only cases. For the cohort study, we obtained lists of workers employed since the beginning of production in three asbestos-cement factories. We identified about 4000 workers, 2300 with valid date of birth. Conclusion For the cohort study, we are exploring the existence of additional sources to complete missing information and contacting relevant institutions to perform the mortality follow-up. Within 2018 we foresee to have the questionnaire ready for use, the software implemented, and the clinical documentation review completed. This international collaboration is an important step towards a better assessment of the impact of chrysotile use in Brazil.

Aim of special session To highlight examples and best practice in the area of occupational disease and modelling morbidity.

Introduction Lung cancer (LC) is the leading oncologic cause of death among males. The role of occupational history with exposure to well-established carcinogens is very important, but usually deeply underestimated. To undervalue the occupational origin of cancer can affect the efficacy of preventive measures and preclude patients to receive insurance benefits. We performed a systematic occupational medicine evaluation of a cases-series of lung cancer cases to properly quantify the proportion of LC cases with previous occupational exposure to carcinogens. Methods We systematically evaluated all consecutive LC cases hospitalised in a large university hospital, in Milan. An active systematic search was carried out for 24 months, by trained occupational physicians, using a standardised questionnaire. Results We collected 123 consecutive LC cases (66% males). Former and current smoking habit was found in 61% and 22%, respectively. A clear exposure to occupational carcinogens (in details: asbestos, polycyclic aromatic hydrocarbons, paintings, diesel exhaust) was recognised in 10% of males cases. Discussion Previous occupational exposure to carcinogens was frequent among males cases in Northern Italy. The observed prevalence was even lower than the one reported in a previous systematic search in Lombardy (Porru S , et al. Int Arch Occ Env Health 89:981–9). If we applied our observed proportion of occupational cases to the entire incident LC cases in Lombardy population, we should observe about 460 male occupational LC cases per year (AIRTUM-AIOM report 2016), actually more than the entire number of cancer (all sites, both gender) annually notified to National Institute for Insurance (INAIL Rapporto regionale 2015).

Introduction IARC defined shift work as probably carcinogenic to humans (Group 2A), after investigations highlighted an increased breast cancer risk in night-shift female workers. The biological mechanisms underlying this association are still unclear. Hence, we evaluated the relationship between night-shift work and molecular alterations potentially related to increased cancer risk, in detail: DNA methylation of the oestrogen receptor gene (ER-Beta) and tumour suppressor genes (BRCA1, p53, p16), global DNA methylation estimated in repeated elements (LINE-1, Alu) and telomere length (TL). Methods 46 female nurses (age: 35–45 years) who had been working in night-shifts for at least two years (length of service ≥five years) were recruited at the Policlinico Hospital (Milan, Italy) and matched for age, sex and length of service to 51 colleagues not working in night-shifts. Each subject was administered a structured questionnaire and withdrawn a 12 mL blood sample. Linear regression models adjusted for age, BMI, parity, smoking habit and oral contraceptive use were then applied. Results Working in night-shifts (yes/no) was associated with BRCA1 hypomethylation (β: −0.512, 95% CI: −1.039 to 0.015). When considering also former night-shift workers, the number of years in night-shifts (NYNS) was associated with hypomethylation of BRCA1 (β: −0.084, 95% CI: −0.127 to −0.042), p53 (β: −0.072, 95% CI: −0.133 to −0.011) and LINE-1 (β: −0.043, 95% CI: −0.083 to −0.002). After graphically inspecting the NYNS-TL relationship, we stratified our study population by NYNS Conclusion Our results show epigenetic alterations that might play a role in cellular ageing, genomic instability and carcinogenesis. We are currently extending our study to other molecular targets involved in the cascade of events that might bring from night-shift exposure to cancer.

Introduction In Italy about 45% of people aged ≥18 years are overweight/obese and might thus be more susceptible to the adverse health effects of air pollution exposure. Particulate matter ≤10 µm (PM10) represents a common pollutant of living and working environments and has been associated with increased risk of cardiovascular diseases (CVD) and hypercoagulability. Extracellular vesicles (EV) might play an important role in PM-related CVD, as they can travel in body fluids and transfer miRNAs between cells. We investigated whether PM10 exposure is associated with changes in fibrinogen levels, EV release, and EV-miRNA content in a large sample of overweight/obese adults. Methods EV concentrations were quantified by nanoparticle tracking analysis and flow cytometry. To identify altered levels of EV-miRNAs, we profiled miRNAs of 883 subjects by the QuantStudio 12K Flex Real Time PCR System. The top 40 EV-miRNAs were validated through custom miRNA plates. Statistical analyses included multiple linear regressions, mediation analysis and bioinformatics analysis. Results In a sample of 1630 overweight/obese subjects from the SPHERE (Susceptibility to Particle Health Effects, miRNAs and Exosomes) study, short-term exposure to PM10 was associated with increased release of EVs, especially those from monocyte/macrophage components (CD14+) and platelets (CD61+). Nine EV-miRNAs (let-7c-5p; miR-106a-5p; miR-143–3 p; miR-185–5 p; miR-218–5 p; miR-331–3 p; miR-642–5 p; miR-652–3 p; miR-99b-5p) were downregulated in response to PM10 exposure and exhibited putative roles in CVD. Five of these nine EV-miRNAs were mediators in the positive association between PM10 exposure and fibrinogen levels. Conclusions Our study sheds some light on the potential mechanisms underlying the adverse cardiovascular health effects of air pollution exposure. Our results were obtained in a hypersusceptible population and thus strengthen the relevance of health promotion interventions for both the general public and the working population, as they might be particularly feasible in the workplace.

Introduction Malignant Pleural Mesothelioma (MPM) is an aggressive cancer mainly caused by asbestos exposure. Due to its long latency and insidious onset, MPM is often diagnosed in advanced stages with poor prognosis. In addition, asbestos is still used in many non-European countries and the incidence of MPM is expected to increase. In this context, the need of reliable diagnostic markers for early MPM diagnosis is of paramount importance. Along with the more frequently studied biological markers (mesothelin, fibulin-3), new emerging biomarkers include miRNA expression in peripheral blood. Methods We previously investigated 23 MPM patients and 19 subjects with past asbestos exposure (PAE) to examine if a specific miRNA signature in plasmatic extracellular vesicles (EV) might help to discriminate between MPM and PAE ( PLoS One, 2017 ). Criteria for enrollment, blood collection, miRNA extraction, screening and validation have been previously described. We found a two miRNA (miR-103a-3p and mir-30-3ep) diagnostic signature that discriminates the two groups with high accuracy (AUC 0.942), high sensitivity (95.5%) and good specificity (80.0%). We are currently expanding our study population to additionally include 25 MPM cases, 50 subjects with PAE, and 20 subjects with other respiratory diseases. Alongside miRNA expression, plasma mesothelin and fibulin-3 will be also measured. Results The diagnostic performance (AUC, Sensitivity and Specificity) of the best five miRNAs previously detected in our study will be examined in combination with plasma mesothelin and fibulin-3, taking into account major confounders (e.g. age, gender, BMI and smoking habit). Conclusions The combination of biological markers belonging to different molecular pathways might help in identifying a panel of biomarkers able to improve the overall diagnostic performance as suggested by Weber et al. ( PLoS One, 2014 ), who recently showed an improved AUC of 0.93 when combining mesothelin and miR-103a-3p.