Suchergebnisse

Introduction: The abuse of New Psychoactive Substances (NPS) has been a major issue since the 1990s, due to the significant impact this phenomenon has on the society and public health. The Italian legislation, in line with the European legislation, is aimed to limit the NPS use in the Country through different intervention strategies. Methods: The Department for Anti-Drug Policies (DPA) of the Presidency of the Council of Ministers, has activated in Italy the National Early Warning System (SNAP) on NSP reporting, aimed at the early detection of potentially health-threatening phenomena related to the appearance of NPSs in the market. A second aim of the SNAP is the promotion of the technical-analytical expertise of laboratories for the detection of these substances. Since June 2016, the DPA, where the National Focal Point (NFP) for the verification of the phenomenon of drug dependence is established, has entrusted the coordination of SNAP with the National Centre for Addiction and Doping (NCAD) of the "Istituto Superiore di Sanità". The Centre works with several partners, including the Pavia Poison Control Centre for clinical toxicology, the Unit of Forensic Toxicology of Sapienza University of Rome for bio-toxicological aspects, and the Central Directorate for Anti-Drug Services for national coordination of seizures operated by law enforcement agencies. To implement this network of collaboration, the NCAD has developed and activated a strategic 'web application' that simplifies the collection of information across the Country, optimizing incoming/outgoing reports between all the involved authorities. Results: The analysis of the information collected on NPS during 2020 through the SNAP platform, highlighted the circulation of 47 NSPs in Europe, including two synthetic cannabinoids with potential public health risks and the emergence of NPS belonging to the class of benzodiazepines and the benzimidazole family. Conclusions: The SNAP is an important tool for identifying and notifying the spread of NSPs in our ...

La Somatotropina bovina ricombinante (rbST) è un ormone proteico di sintesi prodotto da alcune grandi multinazionali farmaceutiche e commercializzato in ambito zootecnico allo scopo di incrementare la resa lattea degli animali in produzione. Sebbene in alcune parti del mondo il suo utilizzo sia stato autorizzato, l'Unione Europea ne ha bandito il commercio e l'impiego all' interno dei suoi stati membri (Decisione del Consiglio 1999/879/CE). Il recente ritrovamento di farmaci iniettabili a lento rilascio a base di rbST in alcuni allevamenti di bufale italiani ha ulteriormente confermato la necessità di disporre di strumenti analitici in grado di svelare eventuali trattamenti illeciti. Ad oggi, non esistono test di screening validati con questo proposito. Questo lavoro di tesi si inserisce in un progetto che si propone di analizzare la variazione dell'espressione genica in leucociti di bufale alle quali è stata somministrata rbST, con lo scopo di ricercare biomarcatori come indice del trattamento ormonale. L'RNA estratto dalle cellule bianche di 8 animali trattati e 8 controlli, in 6 momenti di campionamento, all'interno di 3 cicli di somministrazione bisettimanale del farmaco a lento rilascio a base di rbST, è stato analizzato mediante microarray, al fine di selezionare i geni differenzialmente espressi in un contesto di circa 44.000 geni totali. In un secondo momento l'analisi è stata approfondita tramite Real Time PCR. I risultati sono stati sottoposti ad analisi discriminante lineare, allo scopo di individuare le variabili (geni) con maggior potere discriminante, utilizzate, a loro volta, per elaborare un sistema lineare classificante. La percentuale di falsi negativi ottenuta è stata circa 6.5%. Anche se la Decisione 2002/657/CE prevede che un test di screening debba essere caratterizzato da un tasso di falsi negativi inferiore al 5%, il test oggetto di questo studio potrebbe essere ulteriormente perfezionato aumentando la popolazione statistica e selezionando algoritmi che privilegino la sensibilità rispetto alla specificità. The recombinant bovine somatotropin (rbST) is a synthetic peptide hormone produced and commercialized by several large international pharmaceutical companies in order to increase milk yield in livestock. Although its use has been authorized in some countries worldwide, the European Union has banned its trade and use within its member States (Council Decision 1999/879 / EC). However, the use in some Italian buffaloes herds of slow- release injectable drugs containing rbST has been recently discovered, further confirming the need for analytical tools capable of revealing any illegal treatment. To date, no validated screening tests exist. This thesis is part of a project that aims to analyze the variation of gene expression in leukocytes of buffaloes treated with rbST, in order to seek hormonal treatment biomarkers. RNA extracted from white cells of 8 treated animals and 8 controls, in 6 sampling times, within 3 cycles of biweekly administration of the rbST slow- release drug, was analyzed by microarray, in order to select differentially expressed genes out of a total of about 44,000 genes. In a second phase, the analysis was refined by Real Time PCR. The results were submitted to linear discriminant analysis, in order to identify the variables with the greatest discriminating power. The identified variables were then used to develop a linear system for classification. A percentage of false negatives of approximately 6.5% was obtained. The classifying algorithm may be further improved to reach a false negative rate of less than 5% as required by the decision 2002/657/EC. This could be done by increasing the statistical population and by selecting algorithms that favor sensitivity rather than specificity.

Dioxins are commonly known as highly toxic compounds that are persistent organic pollutants. 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), the most potent dioxin congener, and dioxin-like products are formed during the incomplete combustion of organic compounds in the presence of chlorine (waste incineration, burning of various fuels and poorly controlled combustion sources). In 1997, the International Agency for Research on Cancer (IARC) classified TCDD as carcinogen. Prostate cancer (PC) is an extremely serious disease in dogs. PC also represents the most commonly diagnosed cancer in males in the Western world. In general, animal models of human cancer have evolved in attempts to capture the complexity of the human disease. In particular, as humans, the dogs are the only other mammals that develop PC. The canine prostate gland shares many morphological and functional similarities with the human prostate, so this specie represents thus an attractive model for the study of the prostatic disease in dogs. Consequently, to evaluate the biological effects of TCDD on prostate cancer, in this study we used human prostate cancer cell line, PC3. Until now, very few data are present in literature about PC3 and TCDD exposure. It is known that in PC3 cell line, a hormone-independent prostate cancer cell line, TCDD induces cytochrome P450 (CYP) 1A1 and CYP1B1 via the aryl hydrocarbon receptor (AhR). This effect of TCDD could result in higher elimination rates of concomitant drugs metabolized by these particular CYP isoenzymes. Autophagy is a tightly regulated process playing a normal part in cell growth, development, and homeostasis and helping to maintain a balance between the synthesis, degradation, and subsequent recycling of cellular products through the degradation via the lysosome. Defects of autophagy machinery are responsible for pathogenesis of different diseases, including cancer. The role of autophagy in cancer is controversial. There is evidence that autophagy may play a critical role in cancer progression at later stages, such as dissemination and metastasis, which account for most cancer-associated deaths, whereas in other cases it clearly contributes to tumor suppression by inducing tumor cell death. So, the aim of this study is to evaluate the biological effects of TCDD exposure to a highly metastatic prostate cancer cell line, PC3. The prostate cancer cell line PC3 was exposed to different concentrations of TCDD (0.1, 1 and 100 pg/ml) and after 24, 48, and 72 hours of exposure cell proliferation and viability, cell morphology and cell cycle analysis were performed. Also, autophagy was evaluated using the following: (a) detection of acidic vesicular organelles (AVOs) by acridine orange staining; (b) immunofluorescence analysis (IF) of LC3 (microtubule associated light chain protein 3), one of the well-known autophagy markers; (c) LC3 gene expression by real-time PCR; (d) study of autophagic flux by chloroquine (CQ) (10µM), autophagy inhibitor, by western blot; (e) study the expression of multiple genes involved in autophagy machinery by real-time PCR. When compared with their relative controls, TCDD (100 pg/ml) exposure at 24, 48, and 72 hours caused the following: (1) significant increase of cell proliferation (CP) (CP24h =20.3%, CP48h =54.5% and CP72h =52.4%); (2) significant increase of cell population at S phase; (3) significant increase of autophagy demonstrated by (a) detection of AVOs; (b) LC3 IF positivity; (c) LC3 over expression (expression ratio24h =1.4, expression ratio48h =6.2 and expression ratio72h =11.4); (d) CQ increased LC3-II accumulation; (e) moreover, we demonstrated that the induction of autophagy by TCDD in PC3 cells was accompanied by an increase in the mRNA levels of certain genes that are present in different subnetworks composing the autophagy interactive network, such as: PIK3C3, BECN-1, AMBRA1, MAP1LC3B, ATG4A, ATG4C, ATG5, ATG7, ATG10, ATG16L1, GABARAPL1, PRKAA, WIPI-1. Also, we showed that TCDD negatively influenced genes that are autophagy inhibitor, such as: AKT1 and BCL2. These data suggested a multiple effect of TCDD on autophagy machinery. Furthermore, we revealed that TCDD exposure upregulated TNF whose high levels have been related to prostate cancer progression via stimulation of proliferation, survival of malignant cells and increased resistance to chemotherapeutic agents. From the 1980s, several illegal and uncontrolled sites of urban, toxic, and industrial waste disposal have been active in Campania region, Southern Italy, with the highest concentration reached in Naples and Caserta provinces. In 1994, Campania region is under a declared State of Emergency, because of the saturation of regional waste treatment facilities. There are growing national and international evidences that the accumulation of waste, illegal and legal, urban and industrial, has contaminated soil, water, and the air with a range of toxic pollutants including dioxins. A high correlation between incidences of cancer, respiratory illnesses, and genetic malformations and the presence of industrial and toxic waste landfills was also found. In Spring 2002, a dioxin emergency emerged in this region, as result of National Control Program of Residues in Foodstuff carried out by Health Minister in 2001, indeed, levels of TCDD exceeding the European Union tolerance were detected in dairy products and milk from cow and water buffalo, raised on in some areas of Campania Region. Cancer mortality descriptive studies demonstrated an increase of incidents rates of the prostate cancer in the same areas where dioxin levels were elevated. Furthermore, studies performed in animal model suggested that TCDD exposure is associated with abnormal prostate development, altered prostate pathology and increase susceptibility to prostate cancer. The administration of TCDD to a variety of cultured cells may alter their ability to proliferate and die. In a previous study we demonstrated that TCDD induced proliferation in Madin-Darby Bovine Kidney (MDBK), an epithelial cell line, in which analysis of MDBK cell morphology revealed some alterations in a large number of exposed cells, where neither signs of apoptosis nor necrosis were detected, but we found that TCDD activated cell death with autophagy. Herein, TCDD exposure in PC3, a cancer cell line, induced no signs of cell death. So, taken together, our results support the idea that TCDD, may induce the progression of prostate cancer enhancing cell proliferation, inducing autophagy, deregulating the expression of genes related to the autophagy machinery, and upregulating TNF resulting in an increased risk for both animal and human health.

Riassunto Nell'Unione Europea, l'uso di promotori di crescita ad azione ormonale, quali ormoni sessuali e corticosteroidi, è vietato nell'allevamento dei bovini da carne, al fine di tutelare la salute dei consumatori dalla presenza di residui potenzialmente pericolosi nei prodotti derivati. Ogni Stato Membro, allo scopo di verificare l'ottemperanza al divieto imposto, predispone annualmente un piano di controllo (PNR), per la ricerca nei tessuti e nelle matrici biologiche, mediante metodi chimici quali-quantitativi, di residui di sostanze che si presume possano essere utilizzate illecitamente. Per quanto riguarda i bovini, è stato recentemente preso in considerazione l'esame istopatologico, quale metodo di screening, poiché in grado di rilevare le gravi lesioni provocate dai trattamenti ormonali a carico degli organi bersaglio, le quali permangono per un tempo abbastanza lungo da poter essere svelate al momento della macellazione. In tal modo, sarebbe possibile evidenziare l'impiego illecito di promotori della crescita, anche se tale approccio analitico non consente un'identificazione a livello di singola molecola. Lo scopo di questa tesi è stato quello di effettuare un'indagine sulla prevalenza di bovini da carne potenzialmente trattati con sostanze ormonali illecite, in uno stabilimento di macellazione operante sul territorio nazionale. A tal fine, è stato realizzato uno studio multilivello su 90 animali (50 maschi e 40 femmine, di età compresa tra gli 11 e i 18 mesi), basato su osservazioni cliniche e comportamentali di ogni soggetto, sull'eventuale riscontro di lesioni anatomo-patologiche macroscopiche a carico di alcuni organi bersaglio (timo, prostata, ghiandola bulbo-uretrale) e sull'esame istologico, quale metodo di screening, di campioni di tali organi. Mentre non sono state riscontrate alterazioni alla visita ante-mortem e post-mortem, all'esame istologico sono stati evidenziati 6 casi (6,7%) di grave atrofia timica, caratterizzata da una notevole deplezione linfocitaria con sostituzione di cellule adipose. Questo quadro istopatologico, in assenza di un'eventuale denuncia di somministrazione farmacologica a scopo terapeutico, può essere associato ad un trattamento sospetto con corticosteroidi. Pertanto, i risultati di questa indagine suggeriscono che, nonostante una rigida legislazione a riguardo e l'attuazione del PNR, il problema dei trattamenti illeciti con sostanze ormonali sia ancora di preoccupante attualità. Parole chiave: trattamenti ormonali illeciti, bovini da carne, residui, istopatologia Abstract In the European Union, the use of growth promoters with hormonal action, such as sex hormones and corticosteroids, has been banned in beef cattle breeding, in order to protect the consumer from the presence of potentially dangerous residues in food products. Each Member State, to verify the enforcement of the ban imposed, carries out an annual control plan on residues (PNR), which, using quali-quantitative chemical analytical methods, is aimed at uncovering the presence of forbidden substances in tissues and biological matrices. Recently, in cattle, the histopathological examination has been introduced with screening purposes, due to the fact that it allows to detect the typical lesions provoked by the hormonal treatments to their target organs. Such lesions persist for a time long enough to be revealed at the slaughterhouse. In such a way, it would be possible to highlight the illegal use of growth promoters, although such an approach does not permit an identification at the level of single molecule. The aim of this thesis was to perform a survey on the prevalence of beef cattle potentially treated with illegal hormonal substances, in a slaughterhouse operating on the national territory. To this aim, a multilevel study was carried out on 90 animals (50 males and 40 females, 11-18 months old), based on clinical and behavioral observations of each subject, on the possible detection of anatomo-pathological macroscopic lesions of some target organs (thymus, prostate, bulbo-urethral gland) and on the histological examination on samples of these organs, as method of screening. During the ante-mortem and post-mortem inspection, no relevant alterations were found. At the histological analysis, 6 cases (6.7%) of severe thymic atrophy were highlighted, characterized by a remarkable lymphocyte depletion with replacement of fat cells. This histopathological picture, in the absence of any allegation of therapeutic drug administration, represents a strong indication of treatments with corticosteroids. Therefore, the results of this survey suggest that, despite the existence of a strict legislation and the implementation of the PNR, the problem of illegal treatments with hormonal substances is still of topical concern. Keywords: illegal hormonal treatments, beef cattle, residues, histopathological screening

L'utilizzo di tecnologie sanitarie nello scenario sanitario attuale caratterizzato da un progressivo incremento della spesa sanitaria, da un aumento dei bisogni assistenziali a causa dell'invecchiamento della popolazione e dai rapidi progressi nelle conoscenze scientifiche e tecnologiche impone che il processo decisionale che le accompagna sia "informato" e "consapevole", data la difficoltà del sistema sanitario nel reperimento e nell'allocazione delle risorse per le attività di prevenzione, cura e riabilitazione della popolazione assistita. Le valutazioni economiche rappresentano gli strumenti necessari per guidare le scelte circa l'adozione di una tecnologia sanitaria innovativa versus la tecnologia sanitaria tradizionale integrando aspetti clinici e di efficacia con considerazioni economiche. La prospettiva è estesa nel caso dell'Health Technology Assessment ad aspetti di sicurezza, di impatto sociale, legale, etico e politico per effettuare una valutazione globale delle tecnologie sanitarie. La tesi illustra il quadro di riferimento metodologico specifico delle valutazioni economiche in sanità attraverso la descrizione della progettazione di due tipologie di indagine quali gli studi di revisione sistematica della letteratura (studi secondari) e gli studi campionari (studi primari) per valutare i costi sociali e la costo efficacia di tecnologie sanitarie alternative. La metodologia per la progettazione degli studi di valutazione economica è applicata a casi diversi di tecnologie sanitarie per la diagnosi o il trattamento di patologie diverse quali i farmaci biologici per l'artrite reumatoide, gli impianti cocleari per l'ipoacusia, la chirurgia bariatrica per l'obesità e una combinazione di tecniche di imaging non invasive anatomo-funzionali per la diagnosi e la caratterizzazione della cardiopatia ischemica. La prospettiva sanitaria di valutazione dei costi delle tecnologie sanitarie è ampliata nel lavoro svolto in una di tipo sociale, considerando non solo i costi direttamente assorbiti dalle tecnologie e supportati dal sistema sanitario (costi diretti sanitari) ma anche i costi sostenuti dai pazienti, dalle famiglie e dal sistema produttivo e previdenziale (costi diretti non sanitari e indiretti). I risultati che emergono mettono in luce come i costi delle malattie gravino in modo rilevante su pazienti e famiglie, a causa principalmente dell'assistenza informale e delle perdite di produttività. Quindi l'investimento in tecnologie sanitarie se, da un lato, può incrementare la componente di spesa sanitaria, dall'altro, potrebbe portare benefici oltre che sulla salute dei pazienti, anche sul sistema produttivo, sul sistema socio assistenziale e sul sistema previdenziale. La crescente attenzione da parte dei decisori sanitari verso logiche e politiche che perseguono gli obiettivi di controllo e contenimento dei costi, dati i vincoli di efficacia promossi anche dal movimento dell'Evidence Based Medicine, richiede un sempre maggiore utilizzo delle valutazione economiche nei processi decisionali, incentivando una diffusione delle stesse. I risultati delle valutazioni economiche consentono di supportare le scelte dei decisori sanitari, fornendo la base informativa per le indicazioni di policy. The current health care scenario is characterized by progressive increases in health spending and in care needs due to population aging and to rapid advances in scientific and technology knowledge. It requires that the decision-making process about the use of health care technologies should be "well-informed" and "aware", given the difficulty of the health care system in finding and allocating the resources for prevention, treatment and rehabilitation of the population assisted. Economic evaluations represent the necessary tools to guide decisions about the adoption of innovative health care technologies versus traditional ones by integrating clinical efficacy aspects with economic considerations. The Health Technology Assessment extended the economic analyses to safety, social impact, legal, ethical and political aspects to make a global assessment of health care technologies. The doctoral thesis describes the methodological framework of economic evaluations in health care by analyzing two types of research studies such as the systematic literature reviews (secondary studies) and sample studies (primary studies) aimed to assess the social costs and the cost effectiveness of alternative health care technologies. The methodologies are applied to different cases of health care technologies for the diagnosis or treatment of various diseases such as biologic drugs for rheumatoid arthritis, cochlear implants for hearing loss, bariatric surgery for obesity and a combination of innovative non-invasive anatomo-functional imaging techniques for diagnosis and characterization of coronary artery disease. The health care perspective for cost evaluation of health care technologies has expanded to a social perspective by considering besides the costs directly absorbed by technology and supported by the health care system (direct medical costs) the costs borne by patients, families, the production system and social security system (direct non-medical costs and indirect costs). The results reveal how the costs are borne significantly by patients and families due to informal care and loss of productivity. So if the investment in health care technology, on the one hand, could increase the health care expenditure, on the other, could bring major benefits to the patients health of even on the production system, the social security system and welfare system. The health decision-makers attention towards policies of control and containment of cost given the constraints of efficacy (also promoted by the Evidence-Based Medicine) requires an increasing use of economic evaluation in decision-making process. The results of economic evaluations allow to support the choices of health care decision makers, providing the information basis for policy indications.