Azione antiossidante del sulforafane: analisi in cellule cardiache in coltura ed in un modello animale di esercizio fisico

Oxidative stress is considered to be of major relevance for a variety of pathological processes. Thus, it is valuable to identify compounds, which might act as antioxidants, i.e. compounds that antagonize the deleterious action of reactive oxygen species (ROS) on biomolecules. The mode of action of these compounds could be either to scavenge ROS directly or to trigger protective mechanisms inside the cell, thereby resulting in improved defense against ROS. Sulforaphane (SF) (1-isothiocyanato-(4R)-(methylsulfinyl)butane) is a naturally occurring cancer chemopreventive agent found as a precursor glucosinolate in Cruciferous vegetables like broccoli. Although SF is not a direct-acting antioxidant, there is substantial evidence that SF acts indirectly to increase the antioxidant capacity of animal cells and their abilities to cope with oxidative stress. Induction of phase 2 enzymes is one means by which SF enhances the cellular antioxidant capacity. Enzymes induced by SF include Glutathione S-transferases (GST) and NAD[P]H:quinone oxidoreductase (NQO1) which can function as protectors against oxidative stress. To protect themselves from oxidative stress, cells are equipped with reducing buffer systems including the GSH and thioredoxin (Trx) reductase. GSH is an important tripeptide thiol which in addition to being the substrate for GSTs maintains the cellular oxidation– reduction balance and protects cells against free radical species. Aim of the first part of this thesis was to investigate the ability of SF to induce the expression and the activity of different phase 2 and antioxidant enzymes (such as GST, GR, GPx, NQO1, TR, SOD, CAT) in an in vitro model of rat cardiomyocytes, and also to define if SF treatment supprts cells in counteracting oxidative stress induced by H2O2 It is well known that acute exhaustive exercise causes significant reactive oxygen species generation that results in oxidative stress, which can induce negative effects on health and well being. In fact, increased oxidative stress and biomarkers (e.g., protein carbonyls, MDA, and 8- hydroxyguanosine) as well as muscle damage biomarkers (e.g. plasmatic Creatine cinase and Lactate dehydrogenase) have been observed after supramaximal sprint exercises, exhaustive longdistance cycling or running as well as resistance-type exercises, both in trained and untrained humans. Markers of oxidative stress also increase in rodents following exhaustive exercise. Moreover, antioxidant enzyme activities and expressions of antioxidant enzymes are known to increase in response to exhaustive exercise in both animal and human tissues. Aim of this project was to evaluate the effect of SF supplementation in counteracting oxidative stress induced by physical activity through its ability to induce phase 2, and antioxidant enzymes in rat muscle. The results show that SF is a nutraceutical compound able to induce the activity of different phase 2 and antioxidant enzymes in both cardiac muscle and skeletal muscle. Thanks to its actions SF is becoming a promising molecule able to prevent cardiovascular damages induced by oxidative stress and muscle damages induced by acute exhaustive exercise.

Analisi degli aspetti di qualità nella gestione dei sistemi di drenaggio urbano attraverso modellazione numerica e indagini di campo

In order to protect river water quality, highly affected in urban areas by continuos as intermittent immissions, it is necessary to adopt measures to intercept and treat these polluted flows. In particular during rain events, river water quality is affected by CSOs activation. Built in order to protect the sewer system and the WWTP by increased flows due to heavy rains, CSOs divert excess flows to the receiving water body. On the basis of several scientific papers, and of direct evidences as well, that demonstrate the detrimental effect of CSOs discharges, also the legislative framework moved towards a stream standard point of view. The WFD (EU/69/2000) sets new goals for receiving water quality, and groundwater as well, through an integrated immission/emissions phylosophy, in which emission limits are associated with effluent standards, based on the receiving water characteristics and their specific use. For surface waters the objective is that of a “good” ecological and chemical quality status. A surface water is defined as of good ecological quality if there is only slight departure from the biological community that would be expected in conditions of minimal anthropogenic impact. Each Member State authority is responsible for preparing and implementing a River Basin Management Plan to achieve the good ecological quality, and comply with WFD requirements. In order to cope with WFD targets, and thus to improve urban receiving water quality, a CSOs control strategy need to be implemented. Temporarily storing the overflow (or at least part of it) into tanks and treating it in the WWTP, after the end of the storm, showed good results in reducing total pollutant mass spilled into the receiving river. Italian State Authority, in order to comply with WFD statements, sets general framework, and each Region has to adopt a Water Remediation Plan (PTA, Piano Tutela Acque), setting goals, methods, and terms, to improve river water quality. Emilia Romagna PTA sets 25% reduction up to 2008, and 50% reduction up to 2015 fo total pollutants masses delivered by CSOs spills. In order to plan remediation actions, a deep insight into spills dynamics is thus of great importance. The present thesis tries to understand spills dynamics through a numerical and an experimental approach. A four months monitoring and sampling campaign was set on the Bologna sewer network, and on the Navile Channel, that is the WWTP receiving water , and that receives flows from up to 28 CSOs during rain events. On the other hand, the full model of the sewer network, was build with the commercial software InfoWorks CS. The model was either calibrated with the data from the monitoring and sampling campaign. Through further model simulations interdependencies among masses spilled, rain characteristics and basin characteristics are looked for. The thesis can be seen as a basis for further insighs and for planning remediation actions.

Contributo della modellistica idrologica all'analisi di suscettività alle frane superficiali in argilla

L'Appenino Emiliano è tra le zone più franose al mondo; i fenomeni che la interessano sono abbastanza lenti e quindi mai catastrofici, ma, vista la loro diffusione, molto dannosi per le infrastrutture. La ricerca della pericolosità associata alle frane si distingue in previsione spaziale e previsione temporale; la prima, detta anche suscettività, è il tema del presente lavoro. La suscettività è volta alla realizzazione di carte di propensione al dissesto, relative e indipendenti dal tempo. Dall'inizio degli anni '90 sono disponibili in letteratura diversi modelli per rispondere a questa esigenza, i quali sono generalmente costituiti da una componente geo-meccanica (di solito il modello del Pendio Infinito) e una idrologica. Il presente lavoro si concentra su quest'ultima che, nei diversi modelli, presenta la maggiore varietà, e cerca di capire quale sia il contributo che questa componente può dare all'analisi di suscettività in un'area argillosa. Per valutare questo contributo, sono stati applicati ad un'area di studio rappresentativa, diversi modelli fisicamente basati noti in letteratura o creati appositamente. Le informazioni dinamiche dei modelli transitori sono state integrate nel tempo secondo diversi metodi che tengono conto della permanenza delle condizioni critiche nel versante. I risultati dell'analisi suggeriscono che, nell'area di studio, e presumibilmente nelle aree a prevalenza argillosa in genere, per la determinazione della suscettività alle frane, il contributo di un modello fisicamente basato, completo di componente geo-meccanica e componente idrologica accoppiate, è assolutamente trascurabile rispetto ad un semplice modello geo-meccanico basato sulla sola pendenza come quello del Pendio Infinito. Le indicazioni provenienti da un modello completo possono essere ridondanti o addirittura fuorvianti se questo non è adatto alle caratteristiche dell'area in studio e ben calibrato.

Altinópolis, una utopia brasiliana. Analisi di una esperienza di educazione alla pace e non violenza come pratica alternativa dell'educazione popolare

The present thesis has as its object of study a project called “Education towards Peace and No Violence”, developed in the town of Altinópolis, in the inner part of the state of São Paulo, Brazil. Based on the analysis and the studies that were carried out, it was possible to identify the difficulties in the implementation of the project, as well as the positive results evidenced by the reduction of the reports of violence in the town. Through a careful research of the historical facts, it shall be demonstrated the progress of education, particularly in Brazil, throughout the years. In addition, a panoramic view of the studies and programs for peace developed in North America and Occidental Europe will be presented. The education for peace based in the holistic concept of human development appeared as a new educational paradigm that works as a crucial instrument in the construction process towards a peaceful and more human society based in social justice.

Analisi morfologica e molecolare di cellule da colture primarie umane esposte a resine biocompatibili

The cytotoxicity of dental composites has been attributed to the release of residual monomers from polymerized adhesive systems due to degradation processes or the incomplete polymerization of materials. 2-Hydroxyethyl methacrylate (HEMA) is one of the major components released from dental adhesives. Cytotoxic effects due to high concentrations of HEMA have already been investigated, but the influence of minor toxic concentrations for long-term exposition on specific proteins such as type I collagen and tenascin has not been studied in depth. The objective of this project was to study the effect of minor toxic concentrations of HEMA on human gingival fibroblasts (HGFs) and human pulp fibroblasts (HPFs), investigating modification in cell morphology, cell viability, and the influence on type I collagen and tenascin proteins. Different concentrations of the resin monomer and different times of exposition were tested on both cell lines. The cell vitality was determined by MTT assay, and high-resolution scanning electron microscopy analysis was performed to evaluate differences in cell morphology before and after treatment. To evaluate the variability in the expression and synthesis of procollagen α1 type I and tenascin proteins on HGFs and HPFs treated with HEMA at different concentrations immunofluorescence, RT-PCR and western blot analysis, were carried out. The treatments on HGFs with 3mmol/L HEMA, showed a strong reduction of procollagen α1 type I protein at 72h and 96h, demonstrating that HEMA interferes both with the synthesis of the procollagen α1 type I protein and its mRNA expression. The results obtained on HPFs treated with different concentrations of HEMA ranging from 0,5mmol/L to 3mmol/L and for different exposition times showed a strong reduction in cell viability in specimens treated for 96h and 168h, while immunofluorescence and western blotting analysis demonstrated a reduction of procollagen α1 type I and an overexpression of tenascin protein. In conclusion, our results showed that the concentrations of HEMA we tested, effect the normal cell production and activity, such as the synthesis of some dental extracellular matrix proteins.

Analisi tridimensionale priva di contatto delle ulcere cutanee laser assistita

La valutazione progressiva e il monitoraggio di una ferita difficile impongono misurazioni periodiche per valutare in modo qualitativo e quantitativo l'esistenza e l'entita della riepitelizzazione in rapporto alle pratiche terapeutiche effettuate. L'entità della guarigione nei primi 1530 giorni di trattamento, intesa come riduzione di area percentuale, nelle lesioni ulcerative di origine venosa permette secondo alcuni autori della letteratura scientifica, di avere delle informazioni prognostiche di guarigione completa a sei mesi. Altri autori invece mostrano casistiche analoghe indicando come nelle ulcere venose non si possa predire con facilità la guarigione a sei mesi.Le metodiche di misurazione delle ferite possono essere suddivise in bidimensionali e tridimensionali: nel primo gruppo troviamo la misurazione semplice, le metodiche di tracciamento, planimetria, analisi fotografica analogica e digitale. Queste metodiche misurano i diametri, il perimetro e l'area delle ferite. Nel secondo gruppo si collocano l'uso di righello Kundin, le metodiche di riempimento con alginato o con soluzione fisiologica, le metodiche stereofotogrammetriche e l'uso di strumenti laser. Queste metodiche permettono di calcolare con varie approssimazioni anche il volume delle ferite studiate. La tesi ha preso in esame un gruppo di 17 pazienti affetti da ulcere venose croniche effettuando misurazioni con lo strumento più accurato e preciso disponibile (minolta Vivid 900 laser + sensore) e acquisendo i dati con il software Derma. I pazienti sono stati misurati al primo accesso in ambulatorio, dopo 15 giorni e dopo sei mesi. Sono stati acquisiti i dati di area, perimetro, volume, profondità e guarigione a 6 mesi. L'analisi statistica condotta con modalità non parametriche di analisi dei ranghi non ha associato nessuno di questi valori ne' valori derivati ( delta V /delta A; Delta V/ Delta P) alla guarigione a 6 mesi. Secondo le analisi da noi effettuate, sebbene con l'incertezza derivata da un gruppo ridotto di pazienti, la guarigione delle lesioni ulcerative venose non è predicibile a 6 mesi utilizzando sistemi di misurazione di dimensioni, area e volume.

Abbandono del territorio e dissesto idrogeologico in Appennino bolognese: analisi delle dinamiche e degli interventi di gestione idraulico-forestale

The abandonment of less productive fields and agro-forest activities has occured in the last decades, interesting large mountain areas in all mediterranean basin. Until the fifties, agricultural practices dealt mainly with soil surface and surface runoff control systems. However, the apparent sustainability of soil use results often in contrast with historical documents, witnessing heavy hydrogeological instability, in naturally fragile areas. The research focused on the dynamics and effects of post-coltural land abandonment in a critical mountain area of the Reno River. The Reno River rappresents a typical Tuscan-Emilian Apennines Watershed where soil erosion occurs under very different conditions depending on interactions between land use, climate, geomorphology and lithology. Landslides are largely rappresented, due to the diffusion of clay hill slopes. Recent researches suggest that climatic variability will increase as a consequence of global climate change, resulting in greater frequency and intensity of extreme weather events, which could increase rates of erosion, landslides reactivations and diffusion of calanchive basins. As far as hill slopes are concerned, instability is today basically due to intrinsic factors, as the Apennine range is a rather young formation, in geological terms, and is mainly formed by sedimentary rocks with high occurrence of clays. Therefore landslides and rockfalls are very frequent, while surface soil erosion is generally low and anyway concentrated in the low Apennine, where intensive farming is still economically worth. The study, supported by GIS use, analyses the main fisical characteristics of the area and the historical changes of land use, and focalizes the dynamics of spontaneous reafforestation. Futhermore, the research studies the results of soil bioengineering and surface water control solutions for the restablishment of landslides occured in the last period. Infact soil bioengineering has been recently used in different situations in order to consolidate slopes and hillsides and prevent erosion; when applied, it gave good results, both in terms of engineering efficiency and vegetational development, expecially if combined with a good hydraulic control, thus proving to be an actual alternative to other techniques with heavier environmental impacts. Research into the specific site features and the use of proper plant species is vital to the success of bioengineering works.

Sviluppo di tecniche di analisi ifenate per la quantificazione di biomarcatori di esposizione a tossici ambientali: gli acidi mercapturici

Human biomonitoring (HBM) is an ideal tool for evaluating toxicant exposure in health risk assessment. Chemical substances or their metabolites related to environmental pollutants can be detected as biomarkers of exposure using a wide variety of biological fluids. Individual exposure to aromatic hydrocarbon compounds (benzene, toluene, and o-xylene –“BTX”) were analysed with a liquid chromatography coupled to electrospray ionisation-mass spectrometry (μHPLC-ESI-MS/MS) method for the simultaneous quantitative detection of the BTX exposure biomarker SPMA, SBMA and o-MBMA in human urine. Urinary S-phenylmercapturic acid (SPMA) is a biomarker proposed by the American Conference of Governmental Industrial Hygienists (ACGIH) for assessing occupational exposure to benzene (Biological Exposure Index of 25 microg/g creatinine). Urinary S-benzylmercapturic (SBMA) and o-methyl S-benzyl mercapturic acid (o-MBMA) are specific toluene and o-xylene metabolites of glutathione detoxicant pathways, proposed as reliable biomarkers of exposure. To this aim a pre-treatment of the urine with solid phase extraction (SPE) and an evaporation step were necessary to concentrate the mercapturic acids before instrumental analysis. A liquid chromatography separation was carried out with a reversed phase capillary column (Synergi 4u Max-RP) using a binary gradient composed of an acquous solution of formic acid 0.07% v/v and methanol. The mercapturic acids were determinated by negative-ion-mass spectrometry and the data were corrected using isotope-labelled analogs as internal standards. The analytical method follows U.S. Food and Drug Administration guidance and was applied to assess exposure to BTX in a group of 396 traffic wardens. The association between biomarker results and individual factors, such as age, sex and tobacco smoke were also investigated. The present work also included improvements in the methods used by modifying various chromatographic parameters and experimental procedures. A partial validation was conducted to evaluate LOD, precision, accuracy, recovery as well as matrix effects. Higher sensitivity will be possible in future biological monitoring programmes, allowing evaluation of very low level of BTX human exposure. Keywords: Human biomonitoring, aromatic hydrocarbons, biomarker of exposure, HPLC-MS/MS.