Guido Perra

The influence of endocrine disruptors in a selected population of infertile women

Abstract Several studies report that endocrine disrupting chemicals (EDC) able to interfere with endocrine homeostasis may affect women’s reproductive health. We analyzed EDC serum levels and nuclear receptors (NRs) expression in order to have an indication of the internal dose of biologically active compounds and a measurement of indicators of their effects, as a result of the repeated uptake from environmental source. The percentage of patients with detectable bisphenol A (BPA) concentrations was significantly higher in the infertile patients compared with fertile subjects. No significant difference was found between the groups with regard to perfluorooctane sulfonate (PFOS), perfluorooctanoic acid (PFOA), mono-ethylhexyl phthalate (MEHP) and di-(2-ethylhexyl) phthalate (DEHP) concentrations. Among infertile women, the mean expression of estrogen receptor alpha (ERα) and beta (Erβ), androgen receptor (AR) and pregnane X receptor (PXR) was significantly higher than fertile patients. The mean expression of aryl hydrocarbon receptor (AhR) and peroxisome proliferator-activated receptor gamma (PPARγ) did not show significant differences between two groups. Patients with endometriosis had higher levels of PPARγ than all women with other causes of infertility. This study led further support to EDC exposure as a risk factor for women’s fertility.

Levels and spatial distribution of polycyclic aromatic hydrocarbons (PAHs) in sediments from Lenga Estuary, central Chile

The Lenga Estuary is a small brackish wetland located southwest of San Vicente Bay, Region VIII, Chile. Surface sediment from nine sites in the estuary were analysed for PAHs and compared to Sediment Quality Guidelines (SQG). Sediment samples were freeze dried and soxhlet extracted for 16 h using DCM. Identification and quantification was carried out by HPLC. Organic carbon was also determined. Results showed total PAH concentrations ranged from 290 to 6118 (2025 ± 1975)ng g(-1) d.w. (2025 ± 1975). Results for organic carbon percentages ranged from 1% to 7%. Statistical analysis showed a significant positive correlation (Pearson test) between organic carbon percentage PAHs. Comparison of contaminant levels and international Sediment Quality Guidelines (SQG) (ERL and ER) suggested that sediment of the Lenga estuary did not show any ecotoxicological risk for benthic organisms where high levels of PAHs were detected. Monitoring of this and other contaminants is recommended in Chile.

Pilot study on levels of perfluorooctane sulfonic acid (PFOS) and perfluorooctanoic acid (PFOA) in selected foodstuffs and human milk from Italy

Despite the health risks associated with perfluorinated compounds (PFC) exposure and the detection of these compounds in many countries around the world, little is known on their occurrence in Italy. The results of a study on levels of perfluorooctane sulfonate (PFOS) and perfluorooctanoic acid (PFOA), analysed by HPLC-ESI-MS, in human milk and food samples from the city of Siena and its province (central Italy) are here reported. PFOS was found in 13 out of 49 breast milk samples (0.76±1.27 ng/g), while PFOA was detected in one sample (8.04 ng/g). Only PFOS was found in food samples. Fish were the most contaminated samples (7.65±34.2 ng/g); mean concentrations in meat and milk and dairy products were similar (1.43±7.21 ng/g and 1.35±3.45 ng/g, respectively). In all cereal-based food, eggs, vegetables, honey and beverages PFOS concentration was <LOD. These data show that consumption of most breast milk analysed would not result in children exceeding their total daily intakes for perfluorinated compounds.

Hepatic biotransformation genes and enzymes and PAH metabolites in bile of common sole (Solea solea, Linnaeus, 1758) from an oil-contaminated site in the Mediterranean Sea: A field study

The aim of the present field study was to evaluate the response of hepatic biotransformation genes and enzymes of the common sole collected from an area characterized by the presence of an oil-refinery. Based on ∑PAHs levels detected in sediments, three sites were identified: an high-impact site in front of the refinery, a moderate impact site and a reference site at increasing distances from the refinery. Transcription of cyp1a, udpgt and gst genes and related enzyme activities, such as EROD, BROD, MROD, B(a)PMO, UDPGT and GST, were assessed in sole liver. PAHs bile metabolites were measured. The link between phases I and II is discussed with regard to levels of PAHs measured in sediments and fillets. Results provide sequencing data on biotransformation genes essential for further studies on transcriptional responses in common sole and confirm phase I enzyme activities as useful tools for future biomonitoring studies in marine coastal areas.

Levels and spatial distribution of polycyclic aromatic hydrocarbons (PAHs) in superficial sediment from 15 Italian marine protected areas (MPA)

Surface sediment from 15 Italian marine protected areas (MPA) were analysed for polycyclic aromatic hydrocarbons (PAHs). The organic carbon percentage was also determined. Total PAH concentrations (ng g⁻¹ d.w.) ranged from 0.71 (Penisola del Sinis) to 1550 (Miramare). Individual PAH analysis showed that three and four rings PAHs were the most frequently detected isomers and accounted for 60-70% of the PAH total concentrations. PAH ratio analysis showed a prevalence of pyrolytic PAH origin at most of the MPAs with exception of Porto Cesareo and Ustica where a petrogenic origin was detected. Results for organic carbon percentages ranged from 0.3% (Capo Rizzuto) to 2% (Punta Campanella). These results are comparable to other Mediterranean marine environments. However, our results shows that some MPAs, such as Miramare, Porto Cesareo, Isola Capo Rizzuto and Punta Campanella, are subject to strong pressure from urban and industrial activities where high PAH levels were detected.

Transcriptional and post-transcriptional response of drug-metabolizing enzymes to PAHs contamination in red mullet (Mullus barbatus, Linnaeus, 1758): a field study.

Aim of this study was to evaluate the responsiveness of red mullet (Mullus barbatus) liver detoxification enzymes to PAHs at transcriptional and post-transcriptional levels in the field. Fish were captured in the north-eastern Adriatic Sea, close to an oil refinery. Sixteen PAHs (EPA) were determined in sediments and fish fillets; transcription levels of cyp1a, cyp3a and abcc2 genes and EROD, BROD, B(a)PMO, BFCOD, GST and UDPGT enzymatic activities were measured. Levels of PAHs in sediments reflect the oil pollution gradient of the area, with weak correspondence in fish fillets. cyp1a gene transcription and EROD, B(a)PMO and BFCOD activities were significantly induced in the oil refinery site, and a slight up-regulation of cyp3a and abcc2 was also observed. GST and UDPGT remained unchanged. The present study provides the first data on detoxification responses at transcriptional levels in the liver of red mullet and confirms phase I enzymes as suitable biomarkers of exposure to PAHs in field studies.

Levels and spatial distribution of polychlorinated biphenyls (PCBs) in superficial sediment from 15 Italian Marine Protected Areas (MPA)

Human activities such as industry, sewage, coastal sprawl, intensive agriculture and oil spills have a high impact on the Mediterranean marine environment. In Italy, growing awareness of how man is threatening marine ecosystems has prompted conservation measures, including the designation of Marine Protected Areas (MPAs). MPAs are essential for conservation, providing protection for critical areas and refuges for overexploited species. They can also act as buffers against management miscelation (Allison et al., 1998). Italy has instituted a number of MPAs that are proving effective in protecting wildlife. Most of them contain one or more no-entry zones which coincide with the areas of greatest natural value (Boero et al., 1999). However, MPAs cannot protect populations or assemblages from transboundary movements of chemicals (I.U.C.N, 1981), a phenomenon observed mostly in MPAs located near human activities such as harbours. MPAs can be useful for monitoring release of contaminants. For instance, Terlizzi et al. (2004) reported the effects of organotin compounds such as tributyltin (TBT) and its transboundary movements in species of marine snails living in Italian MPAs. Persistent organic pollutants (POPs) are semivolatile organic compounds of special concern because of their toxicity, persistence, long-range transport and bioaccumulation potential (UNEP, 2001). POPs such as polychlorinated biphenyls (PCBs) have been widely used in industry and are characterized by low vapour pressure, low water solubility and environmental persistence (Mackay et al., 1992). Since they are hydrophobic they bind readily to particulate in water and are transported/deposited with sediment into rivers, lakes and oceans (Bergamaschi et al., 2001). The Stockholm Convention on POPs, coordinated through the United Nations Environment Programme (UNEP), aims to reduce or eliminate the use, discharge and emission of these compounds to the benefit of the environment, reducing risks to humans and wildlife (UNEP, 2001). Initially, 12 chemicals were identified as priority POPs: nine organochlorine pesticides (OCPs), one industrial chemical class, polychlorinated biphenyls (PCBs) and polychlorinated dibenzo-pdioxins/dibenzofurans (PCDD/Fs) associated with various industrial/combustion emissions (UNEP, 2001). Several countries in Europe are parties to the Convention; Italy signed the Stockholm Convention in May 2001, but has not yet ratified it. PCBs were widely used as insulating fluids in electrical equipment, such as transformers and capacitors, and for many other uses. Italian production of PCBs began in 1958. PCB mixtures were

Exposure and effective dose biomarkers for perfluorooctane sulfonic acid (PFOS) and perfluorooctanoic acid (PFOA) in infertile subjects: Preliminary results of the PREVIENI project

Perfluorooctane sulfonic acid (PFOS) and perfluorooctanoic acid (PFOA) have been used as surfactants in various industry and consumer products. PFOS/PFOA are very persistent in the environment and bioaccumulate in humans. They are potential reproductive and developmental toxicants and are considered to be emerging endocrine disrupters (EDs). The Italian project PREVIENI, funded by the Italian Environment Ministry, aims to link environment and human health through the investigation of selected endocrine disrupters (EDs) exposure and associated biomarkers related to human infertility conditions. In the early PREVIENI phase, PFOS and PFOA were determined in 53 couples affected by an infertility status, enrolled in a metropolitan area, according to established inclusion criteria and informed consensus. Nuclear receptors related to chemical compounds interactions were selected as biomarkers of effect and their gene expression modulations were analyzed in human peripheral blood mononuclear cell (PBMC). Among couples, subjects not presenting infertility factors (IF--) were separated from affected subjects (IF++). Most IF-- serum samples showed PFOS and PFOA concentrations overlapping the limit of detection (LOD) of 0.5 ng/g wet weight (ww). A substantial percentage of IF++ serum samples showed PFOS concentrations >20-fold the LOD, i.e. from 3 to 50 ng/g ww. In male (50%, n=26) and from 3 to 144 ng/g ww in female (37%, n=30) samples. PFOA values were below the LOD levels in 90% of the total samples. Peroxisome proliferator-activated receptor-gamma (PPARγ) and aryl hydrocarbon receptor (AhR) showed a low level of expression in PBMC of both IF++ and IF-- groups. Whereas alpha and beta estrogen receptors (ERα and ERβ), androgen receptor (AR), and pregnane X receptor (PXR) were all upregulated in IF++ of both sexes with respect to IF-- group. Our preliminary results related to the metropolitan area indicate that subjects affected by infertility factors tend to have both higher PFOS levels and higher gene expression of specific nuclear receptors.

Two naphthalene degrading bacteria belonging to the genera Paenibacillus and Pseudomonas isolated from a highly polluted lagoon perform different sensitivities to the organic and heavy metal contaminants

Two bacterial strains were isolated in the presence of naphthalene as the sole carbon and energy source from sediments of the Orbetello Lagoon, Italy, which is highly contaminated with both organic compounds and metals. 16S rRNA gene sequence analysis of the two isolates assigned the strains to the genera Paenibacillus and Pseudomonas. The effect of different contaminants on the growth behaviors of the two strains was investigated. Pseudomonas sp. ORNaP2 showed a higher tolerance to benzene, toluene, and ethylbenzene than Paenibacillus sp. ORNaP1. In addition, the toxicity of heavy metals potentially present as co-pollutants in the investigated site was tested. Here, strain Paenibacillus sp. ORNaP1 showed a higher tolerance towards arsenic, cadmium, and lead, whereas it was far more sensitive towards mercury than strain Pseudomonas sp. ORNaP2. These differences between the Gram-negative Pseudomonas and the Gram-positive Paenibacillus strain can be explained by different general adaptive response systems present in the two bacteria.

Distribution and sources of polycyclic aromatic hydrocarbons (PAHs) in surface sediments of some Italian lagoons exploited for aquaculture and fishing activities.

The concentrations of 16 PAHs in surface sediments collected from four Italian lagoons, exploited for aquaculture and fishing activities, during the period 2004–2007, were analysed. Some molecular ratios and Pearson correlations were used in order to estimate the origin of the pollution and similar transport of PAH sources. Principal Component Analysis (PCA) and Cluster Analysis were used in order to highlight dissimilarities among sampling sites. Concentrations of total 16 PAHs varied significantly among the lagoons as well as within the same basin. Sediments of Orbetello lagoon showed the highest level of contamination (mean of 98.78 ng g−1), followed by that in Santa Giusta (48.15 ng g−1), Lesina (31.06 ng g−1) and Varano (25.19 ng g−1). These results were linked to the greater industrialisation and urbanisation of the catchment area of both Orbetello and Santa Giusta compared with Lesina and Varano. A considerable predominance of 4-rings PAHs was observed for Lesina (52%,), Varano (77%), Orbetello (50%) and Santa Giusta (57%) and Pyrene was the dominant compound. On the other hand, phenanthrene is the dominant compound of low molecular weight contaminants for all the studied lagoons. A meaningful pyrolytic contribution to pollution was found in the sediments of Lesina lagoon, dominated by benzo[a]pyrene (7.27 ng g−1) and benz[a]anthracene (4.14 ng g−1), due to intensive traffic. For Varano, an evident petrogenic contamination was observed along the western area, due probably to accidental oil spillage. All 16 PAHs were found to be correlated in Orbetello and Santa Giusta lagoons and the compounds present in Santa Giusta sediments, near the urban centre, were shown to be mainly of pyrolytic origin. A much more complex situation was observed in Orbetello, where a mixed pattern of pyrolytic and petrogenic inputs was observed.