GC-MS analysis of soil faecal biomarkers uncovers mammalian species and the economic management of the archeological site "Le Colombare di Negrar"

: The identification of the mammalian species based on faecal sediments in modern and ancient environments is the aim of the research of archaeologists, forensic scientists and ecologists. Here, we set up and validated an optimized gas chromatography-mass spectrometry (GC-MS) method, characterized by a time-saving sample preparation protocol, for the simultaneous analysis of faecal biomarkers (6 sterols/stanols and 5 bile acids) in 14 soil samples from the archaeological site of "Le Colombare di Negrar" in northern Italy. Although the archaeological sediment samples examined are numerically exiguous, a comparative reading of our faecal biomarkers findings with new studies on faunal materials collected in the same stratigraphic detail during recent excavation campaigns will allow to better clarify the economic interest of the animal species farmed in the Colombare site (such as bovines, goats, sheep and pigs) and to shed light on the management of breeding. Together with archaeozoological and archaeobotanical analyses, the investigation of faecal biomarkers can increase our knowledge of how ancient local communities exploited natural resources and may allow us to deduce what their impact on the landscape was

The "Brunella" project: preparation and study of a mysticete from the Early Pliocene of Tuscany

• A project is being carried out at Poggio alle Mura that will allow the preparation and study of a balaenopterid whale from the early Pliocene of Tuscany together with its associated biota. • Up to now, the project resulted in the preparation of 18 vertebrae, the skull and an ulna of the whale. • The associated biota includes 18 mollusk species, 2 sea urchin species, 10 shark teeth and thousands of fish remains from the sediment surrounding the skull. • A wealth of educational activities is being carried out at the laboratory of Poggio alle Mura that are directed to schools and citizens

The arsenic in mice as experimental model for risk modifiers.

Studies on the relevance of host factors in modulating the physiological responses following chronic exposure to xenobiotics were carried out according to a \u201cToxicogenomic Model on Arsenic in Mice\u201d developed at thte JRC. This model is focused on chronic exposure to arsenate given alone or in combination with other xenobiotics, to assess potential \u201ccocktail effects\u201d and related cumulative risks. DNA-macroarrays technology is applied to evaluate physiological responses at transcriptional level and assessing possible biochemical responses. A cluster of 1200 cancer genes was used for screening purposes, while quantitative PCR on selected genes applied for validation. The exposure varied from in-utero and post-lactation up to adult age (4 months), the chemical forms (arsenate and dimethylarsenate) and doses from 0.1 up to 10 mg As/L in drinking water. Comparison between acute single doses and chronic exposure was also performed. Chronic exposure to arsenate and atrazine in drinking water was selected as an example of multiple chronic exposure. The liver, kidney, lung, bone marrow, adrenals, uterus, and testis were the tissues considered. In the tissues of mice chronically exposed to arsenate, the modulation of gene expression was not only depending on the levels, types and length of exposure, while differently regulated also by the sex, age and diet. The main gene functional families modulated were covering a wide range of biochemical and physiological regulations, like cell cycle modulation, cell adhesion, apoptosis, xenobiotic metabolism, DNA repair, protein turnover, and proto-oncogenes. The patterns of gene expression were strongly influenced by co-exposure to other xenobiotics like atrazine and naphthalene, particularly for genes involved in the metabolism and in neuroendocrine regulation. These effects varied according to the tissue considered, supporting the needs for coherent and specifically designed studies to assess relevant biomarkers of long-term exposure to low levels of xenobiotics and their mixtures

Sex as a major determinant of gene expression in tissues of mice exposed to arsenate.

Inorganic arsenic, frequently found as contaminant of ground water used for drinking purposes in many areas of the world, is a well-known potent human toxicant and carcinogen. Chronic exposure to inorganic arsenic has been associated with cancer of skin, lung, bladder and kidney and, probably, liver. The mechanism of arsenic action in vivo is poorly understood, in particular in relation to dose, type of tissue and gender. To elucidate tissue- and gender dependent biological responses in the genome of mice, we have used cDNA macroarrays for investigation on the expression of 1185 cancer-related genes in mice after exposure to arsenate in drinking water. Continuous exposures of mice to arsenate in drinking water modulate the gene expression in tissues. Interestingly, there were remarkable sex differences: male and female mice show completely different changes in the expression of cancer-related genes. The main gene functional families modulated, were covering a wide range of biochemical and physiological regulations, like cell cycle modulation, cell adhesion, apoptosis, xenobiotic metabolism, DNA repair, protein turnover and proto-oncogens. This result demonstrates important gene-environmental interactions: the molecular mechanisms triggered by arsenic levels frequently experienced following exposure via drinking water, are totally different in males and females. The results obtained using cancer-related genes will be compared with the profiles of over 30.000 genes using the Applied Biosystems expression Array System, to clarify the sex-specific gene pathways

Dietary proteins modulates the gene expression in mice chronically exposed to arsenate.

In the frame of a project on the assessment of risk modifying factors modulating the health effects of environmental chemicals we are developing a toxicogenomic approach using an \u201carsenic in mice\u201d experimental model, considering multistressors exposure, genetics, age, levels and length of exposure, etc. In the present study, we used cDNA Macroarrays to investigate the effects of low protein intake on the expression of 1185 cancer-related genes in the liver of male and female mice transplacentary exposed to different levels of arsenate in drinking water during gestation and developmental age. The results of this study support the relevance of dietary factors in modulating the physiological responses in gene expression following chronic exposure to xenobiotics. In mice chronically exposed to arsenate in drinking water, the modulation of gene expression in different tissues was not only depending on the levels of the xenobiotic under investigation, but mainly regulated by the content of proteins in diet

Clonogenicity and gene expression modulation in the bone marrow of mice chronically exposed to arsenic and atrazine.

The clonogenicity of myeloid progenitors (CFU-GM) and the modulation of gene expression of 1185 cancer-related genes by DNA-macroarrays in bone marrow were used to investigate in male and female mice the combined effects of continuous exposure to arsenate and atrazine in drinking water. In male mice, the exposure to arsenate or to atrazine alone and the combined exposure did not change the clonogenicity of the progenitors. In females the percentage of CFU-GM decreased significantly after atrazine exposure, did not change with arsenic treatment, but dramatically increased after the combined exposure to the two chemicals. Results from microarrays indicate that atrazine alone didn\u2019t stimulate the expression of any of the cancer genes analyzed in both male and female. Arsenic induced gene expression modulation only in female and had no effects on male. Major significant changes on the gene expression in bone marrow cells resulted following the co-exposure to arsenic and atrazine in both male and female. These results indicate that co-exposure of mice to atrazine and arsenate induces significant effects at the level of transcriptional activation of genes in bone marrow cells, as well as stimulating the myeloid progenitors to proliferate, particularly when co-administered in drinking water to female mice

Combined in-utero and juvenile exposure of mice to arsenate and atrazine in drinking water modulates the gene expression and clonogenicity of myeloid progenitors in bone marrow.

Increasing evidence proves that human fetuses are exposed to multiple risk factors and major concerns have been expressed towards exposure to potential endocrine modulating chemicals at early stage of life and during growth. Understanding that exposures occur as mixture of chemicals and that they converge on other inherent and environmental risk-modulating factors, there is a need to develop experimental models to assess the effects of exposure to multiple chemicals during different stage of life. In the present study, the clonogenicity of myeloid progenitors (CFU-GM) and the modulation of gene expression of 1197 cancer-related genes (DNA macroarrays) in bone marrow were used to investigate in male and female young mice the combined effects of continuous exposure to arsenate and atrazine in drinking water. Female adult mice were treated with arsenate in drinking water (1 mg As/L) for 10 days before mating and during the gestation. Offspring were randomly put into separate groups of males and females. One group of arsenic exposed offspring were exposed for 4 months to atrazine (1mg Atr/L) and arsenate (1 mg As/L) in drinking water (As+Atr). One group of each of arsenic unexposed offspring were exposed for 4 months to atrazine (1mg Atr/L) in drinking water (Atr). Additional arsenate (1 mg As/L) was given to one group of arsenic exposed offspring (As). Control mice without any treatment were also analysed (Ctrl). In male mice the exposure to arsenate or to atrazine alone did not result in significant changes on the gene expression in bone marrow cells, whereas, co-exposure to arsenic and atrazine (As+Atr) resulted in a significant up-modulation of gene expression. The percentage of CFU-GM weakly decreased after exposure to individual compounds, while the co-exposure did not change the clonogenicity of the progenitors. In female mice, the co-exposure to both chemicals resulted in a drastic up-modulation of gene expression, while in these cells the single treatments showed a up-modulation of few genes as well. The percentage of CFU-GM decreased significantly after atrazine exposure, did not change with arsenic treatment, but dramatically increased after the combined administration. These results indicate that in-utero and juvenile co-exposure of mice to atrazine and arsenate induce significant effects at the level of transcriptional activation of genes in bone marrow cells, as well as stimulating the myeloid progenitors to proliferate, particularly when co-administered in drinking water to female mice

Excess of NPM-ALK oncogenic signaling promotes cellular apoptosis and drug dependency.

Most of the anaplastic large-cell lymphoma (ALCL) cases carry the t(2;5; p23;q35) that produces the fusion protein NPM-ALK (nucleophosmin-anaplastic lymphoma kinase). NPM-ALK-deregulated kinase activity drives several pathways that support malignant transformation of lymphoma cells. We found that in ALK-rearranged ALCL cell lines, NPM-ALK was distributed in equal amounts between the cytoplasm and the nucleus. Only the cytoplasmic portion was catalytically active in both cell lines and primary ALCL, whereas the nuclear portion was inactive because of heterodimerization with NPM1. Thus, about 50% of the NPM-ALK is not active and sequestered as NPM-ALK/NPM1 heterodimers in the nucleus. Overexpression or relocalization of NPM-ALK to the cytoplasm by NPM genetic knockout or knockdown caused ERK1/2 (extracellular signal-regulated protein kinases 1 and 2) increased phosphorylation and cell death through the engagement of an ATM/Chk2- and γH2AX (phosphorylated H2A histone family member X)-mediated DNA-damage response. Remarkably, human NPM-ALK-amplified cell lines resistant to ALK tyrosine kinase inhibitors (TKIs) underwent apoptosis upon drug withdrawal as a consequence of ERK1/2 hyperactivation. Altogether, these findings indicate that an excess of NPM-ALK activation and signaling induces apoptosis via oncogenic stress responses. A 'drug holiday' where the ALK TKI treatment is suspended could represent a therapeutic option in cells that become resistant by NPM-ALK amplification.We thank Maria Stella Scalzo for technical support, Dr Emanuela Colombo for kindly providing MEFs that lack NPM1 (MEF NPM−/−p53−/−) and control fibroblasts (MEF p53−/−), Dr Guido Serini for the use of his confocal microscopy unit at the Candiolo Cancer Institute—IRCCS, Torino, Italy. We also thank Ariad Pharmaceutical, Pfizer, Astellas and Novartis that kindly provided all drugs used in this study. This work was supported by the Regione Lombardia (ID14546A) and Fondazione Berlucchi Onlus Grant 2014 (to CGP), and by grants FP7 ERC-2009-StG (Proposal No. 242965—‘Lunely’); Associazione Italiana per la Ricerca sul Cancro (AIRC) Grant IG-12023; Koch Institute/DFCC Bridge Project Fund; Ellison Foundation Boston; Worldwide Cancer Research Association (former AICR) grant 12-0216; the Grant for Oncology Innovation by Merck-Serono and R01 CA196703-01 (to RC).This is the author accepted manuscript. The final version is available from NPG via http://dx.doi.org/10.1038/onc.2015.45

Transcranial direct current stimulation in psychiatric disorders

The interest in non-invasive brain stimulation techniques is increasing in recent years. Among these techniques, transcranial direct current stimulation (tDCS) has been the subject of great interest among researchers because of its easiness to use, low cost, benign profile of side effects and encouraging results of research in the field. This interest has generated several studies and randomized clinical trials, particularly in psychiatry. In this review, we provide a summary of the development of the technique and its mechanism of action as well as a review of the methodological aspects of randomized clinical trials in psychiatry, including studies in affective disorders, schizophrenia, obsessive compulsive disorder, child psychiatry and substance use disorder. Finally, we provide an overview of tDCS use in cognitive enhancement as well as a discussion regarding its clinical use and regulatory and ethical issues. Although many promising results regarding tDCS efficacy were described, the total number of studies is still low, highlighting the need of further studies aiming to replicate these findings in larger samples as to provide a definite picture regarding tDCS efficacy in psychiatry

Comparison and combination of a hemodynamics/biomarkers-based model with simplified PESI score for prognostic stratification of acute pulmonary embolism: findings from a real world study

Background: Prognostic stratification is of utmost importance for management of acute Pulmonary Embolism (PE) in clinical practice. Many prognostic models have been proposed, but which is the best prognosticator in real life remains unclear. The aim of our study was to compare and combine the predictive values of the hemodynamics/biomarkers based prognostic model proposed by European Society of Cardiology (ESC) in 2008 and simplified PESI score (sPESI).Methods: Data records of 452 patients discharged for acute PE from Internal Medicine wards of Tuscany (Italy) were analysed. The ESC model and sPESI were retrospectively calculated and compared by using Areas under Receiver Operating Characteristics (ROC) Curves (AUCs) and finally the combination of the two models was tested in hemodinamically stable patients. All cause and PE-related in-hospital mortality and fatal or major bleedings were the analyzed endpointsResults: All cause in-hospital mortality was 25% (16.6% PE related) in high risk, 8.7% (4.7%) in intermediate risk and 3.8% (1.2%) in low risk patients according to ESC model. All cause in-hospital mortality was 10.95% (5.75% PE related) in patients with sPESI score ≥1 and 0% (0%) in sPESI score 0. Predictive performance of sPESI was not significantly different compared with 2008 ESC model both for all cause (AUC sPESI 0.711, 95% CI: 0.661-0.758 versus ESC 0.619, 95% CI: 0.567-0.670, difference between AUCs 0.0916, p=0.084) and for PE-related mortality (AUC sPESI 0.764, 95% CI: 0.717-0.808 versus ESC 0.650, 95% CI: 0.598-0.700, difference between AUCs 0.114, p=0.11). Fatal or major bleedings occurred in 4.30% of high risk, 1.60% of intermediate risk and 2.50% of low risk patients according to 2008 ESC model, whereas these occurred in 1.80% of high risk and 1.45% of low risk patients according to sPESI, respectively. Predictive performance for fatal or major bleeding between two models was not significantly different (AUC sPESI 0.658, 95% CI: 0.606-0.707 versus ESC 0.512, 95% CI: 0.459-0.565, difference between AUCs 0.145, p=0.34). In hemodynamically stable patients, the combined endpoint in-hospital PE-related mortality and/or fatal or major bleeding (adverse events) occurred in 0% of patients with low risk ESC model and sPESI score 0, whilst it occurred in 5.5% of patients with low-risk ESC model but sPESI ≥1. In intermediate risk patients according to ESC model, adverse events occurred in 3.6% of patients with sPESI score 0 and 6.65% of patients with sPESI score ≥1.Conclusions: In real world, predictive performance of sPESI and the hemodynamic/biomarkers-based ESC model as prognosticator of in-hospital mortality and bleedings is similar. Combination of sPESI 0 with low risk ESC model may identify patients with very low risk of adverse events and candidate for early hospital discharge or home treatment.