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Sabrine H.,Regional Research Center on Horticulture and Organic Agriculture | Afif H.,CSIC - Aula Dei Experimental Station | Mohamed B.,Laboratory of Biochemistry and Environmental Toxicology | Hamadi B.,Laboratory of Biochemistry and Environmental Toxicology | Maria H.,CSIC - Aula Dei Experimental Station
Scientia Horticulturae | Year: 2010

In this work, pea (Pisum sativum) plants exposed to increasing cadmium and copper concentrations were tested for heavy metals accumulation in flowers and for '. in vivo' pollen germination. Based on the Cd and Cu accumulation amounts in the flowers, an evaluation of the same metals effects on '. in vitro' pollen germination was achieved. Moreover, the effects of both metals on fruits number and weight and on seed set and yield at individual plant level were examined. While cadmium concentrations did not affect '. in vivo' pollen germination, only higher copper concentrations rendered a significant reduction. This is in contrast with the clear negative effect on pollen germination in vitro and might be explained by the different dynamic and bioavailability of both metals. A clear effect of Cd and Cu was observed on two important yield components ie, fruit weight and seed set. Although results obtained herein cannot give a clear cut relationship between the effect of Ca and Cu on reproductive development and its consequences on yields, they represent emerging results on the potential consequences of metals contamination on reproductive development in plants. © 2010 Elsevier B.V.

Oliveri C.,University of Piemonte Orientale | Peric L.,Ruder Boskovic Institute | Sforzini S.,University of Piemonte Orientale | Banni M.,University of Piemonte Orientale | And 4 more authors.
Comparative Biochemistry and Physiology - Part D: Genomics and Proteomics | Year: 2014

Mollusc haemolymph proteins are known to play several important physiological roles in the immune system, heavy metal transport and the tissue distribution of lipophilic compounds. In this study, we analysed acetone-extracted proteins from mussel haemolymph by one- and two-dimensional gel electrophoresis. The proteins were identified by comparing mass spectrometry data with the invertebrate EST database, allowing us to establish the mussel haemolymph serum proteome. Extrapallial protein (EP) precursor represents the most abundant serum protein; astacin and CuZn superoxide dismutase were also detected. Slight contamination from muscle proteins, due to the sampling method, was also found. No differences were observed in the profiles obtained for male and female serum proteins. One aspect of interest was the previously reported finding that alkali-labile phosphate (ALP) from haemolymph serum may be representative of vitellogenin (vtg)-like protein content in the circulatory fluid of molluscs. In our analysis of mussel haemolymph serum, vitellogenin-like proteins were never found. To confirm these data, a typical methyl-tert-butyl-ether (MTBE) extraction, which is specific for vtg-like proteins, was performed, and the results of the electrophoretic analyses were compared with those obtained by acetonic precipitation. The results showed that the electrophoretic profiles are similar and that vtg-like proteins cannot be identified. Moreover, the main phosphoprotein present in female and male extracts is EP protein precursor. In addition, agarose gel electrophoresis demonstrates that high-molecular-weight forms of vtg-like proteins are not detectable. © 2014 Elsevier Inc.

Banni M.,Laboratory of Biochemistry and Environmental Toxicology | Sforzini S.,University of Piemonte Orientale | Balbi T.,University of Genoa | Corsi I.,University of Siena | And 2 more authors.
Environmental Research | Year: 2016

Despite the growing concern over the potential biological impact of nanoparticles (NPs) in the aquatic environment, little is known about their interactions with other pollutants. In the marine mussel Mytilus galloprovincialis, exposure to nanosized titanium dioxide (n-TiO2), one of the most widespread type of NPs in use, in combination with and 2,3,7,8-tetrachlorodibenzo-p-dioxins (TCDD), chosen as model organic xenobiotic, was shown to induce significant changes in different biomarkers in hemocytes, gills and digestive gland, with distinct effects depending on cell/tissue and type of response measured. In this work, the interactive effects of n-TiO2 and TCDD at the tissue level were further investigated in mussel digestive gland using an integrated approach transcriptomics/immunohistochemistry. Mussels were exposed to n-TiO2 (100μgL-1) and TCDD (0.25μgL-1), alone and in combination, for 96h. Transcriptomic analysis identified 48-, 49- and 62 Differentially Expressed Genes (DEGs) in response to n-TiO2, TCDD and n-TiO2/TCDD, respectively. Gene Ontology (GO) term analysis revealed distinct biological processes affected in different experimental conditions. n-TiO2 mainly up-regulated cytoskeletal genes, while TCDD up-regulated endocrine and signal transduction related processes. Co-exposure induced transcriptional changes common to individual treatments, and identified a newly generated process, response to chemical stimulus. Transcription of selected genes was verified by qPCR. Moreover, expression of tubulin, as an example of target protein of interest identified by gene transcription data, was evaluated in tissue sections by immunolabelling. Tissue TCDD accumulation was evaluated by immunofluorescence with an anti-dioxins antibody.The results demonstrate both distinct and interactive effects of n-TiO2 and TCDD in mussel digestive gland at the molecular and tissue level, identify the main molecular targets involved, and underline how exposure to the n-TiO2/TCDD mixture does not result in increased TCDD accumulation and overall stressful conditions in the tissue. These represent the first data on transcriptional responses of marine invertebrates to exposure not only to n-TiO2 as a model of NP, but also to a legacy contaminant like TCDD. © 2015 Elsevier Inc.

Canesi L.,University of Genoa | Negri A.,University of Piemonte Orientale | Barmo C.,University of Genoa | Banni M.,University of Piemonte Orientale | And 4 more authors.
PLoS ONE | Year: 2011

Background: Many pesticides have been shown to act as endocrine disrupters. Although the potencies of currently used pesticides as hormone agonists/antagonists are low compared with those of natural ligands, their ability to act via multiple mechanisms might enhance the biological effect. The organophosphate Chlorpyrifos (CHP) has been shown to be weakly estrogenic and cause adverse neurodevelopmental effects in mammals. However, no information is available on the endocrine effects of CHP in aquatic organisms. In the digestive gland of the bivalve Mytilus galloprovincialis, a target tissue of both estrogens and pesticides, the possible effects of CHP on the responses to the natural estrogen 17β-estradiol (E2) were investigated. Methodology/Principal Findings: Mussels were exposed to CHP (4.5 mg/l, 72 hrs) and subsequently injected with E2 (6.75 ng/g dw). Responses were evaluated in CHP, E2 and CHP/E2 treatment groups at 24 h p.i. by a biomarker/transcriptomic approach. CHP and E2 induced additive, synergistic, and antagonistic effects on lysosomal biomarkers (lysosomal membrane stability, lysosome/cytoplasm volume ratio, lipofuscin and neutral lipid accumulation). Additive and synergistic effects were also observed on the expression of estrogen-responsive genes (GSTπ, catalase, 5-HTR) evaluated by RT-Q-PCR. The use of a 1.7K cDNA Mytilus microarray showed that CHP, E2 and CHP/E2, induced 81, 44, and 65 Differentially Expressed Genes (DEGs), respectively. 24 genes were exclusively shared between CHP and CHP/E2, only 2 genes between E2 and CHP/E2. Moreover, 36 genes were uniquely modulated by CHP/E2. Gene ontology annotation was used to elucidate the putative mechanisms involved in the responses elicited by different treatments. Conclusions: The results show complex interactions between CHP and E2 in the digestive gland, indicating that the combination of certain pesticides and hormones may give rise to unexpected effects at the molecular/cellular level. Overall, these data demonstrate that CHP can interfere with the mussel responses to natural estrogens. © 2011 Canesi et al.

Banni M.,Laboratory of Biochemistry and Environmental Toxicology | Negri A.,University of Piemonte Orientale | Dagnino A.,University of Piemonte Orientale | Jebali J.,Laboratory of Biochemistry and Environmental Toxicology | And 2 more authors.
Ecotoxicology and Environmental Safety | Year: 2010

In the present study, mussel (Mytilus galloprovincialis) digestive gland biotransformation and detoxification responses to acute exposure to the polycyclic aromatic hydrocarbon benzo[a]pyrene (B[a]P) were investigated. Mussels were exposed to a sublethal dose of B[a]P (75 nM; 19 γg/l per animal) for 24, 48 and 72 h. The following biological responses were measured in the digestive gland tissues: (1) B[a]P hydroxylase (BPH) activity, as phase I biotransformation parameter; (2) glutathione S-transferase (GST) activity as a phase II conjugation enzyme, (3) catalase (CAT) activity as potential biomarker of oxidative stress, (4) acetylcholinesterase (AChE) activity as an indication of possible neurotoxicity response. DNA damage was assessed over time using the single cell gel electrophoresis comet assay and the micronuclei test. BPH and GST activities showed an increasing trend over exposure period. CAT activity showed a symmetrical bell shape response with a maximum at 48 h. AChE activity was significantly depressed after 48 and 72 h exposure to B[a]P. Comet assay and micronuclei test in digestive gland cells suggest that B[a]P exposure induced significant DNA damage with a maximum response after 72 h exposure. © 2009 Elsevier Inc.

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