Towards a more sustainable aquaculture: diets effects and environmental impact of aquaculture from a microbiome perspective

There is a lot of interest to optimize aquaculture production due to its overexploitation of marine resources, ocean pollution and habitat destruction. Since feed production is one of the greatest issues in aquaculture, feeding strategy optimization is important. The study of several different feed additives or supplementation is important to secure optimal growth, gut health, and function in farmed fish. Feed additives are typically supplied to ensure good health and to help the animal ward off pathogens during both normal and challenging conditions, which could stress animals and promote insurgence of pathologies or pathogens invasions. In this context has an increasing interest the study of host associated microbiome to understand the influence of novel functional feed on the health and physiology of animals. To achieve a more sustainable aquaculture sector, show a great importance the understanding of the environmental impact of this human activity in terms of habitat destruction, ocean pollution and reduction marine environments biodiversity. Marine microbiomes, either free-living or associated with multicellular hosts, is acquiring an increasing interest because their role in supporting the functioning and biodiversity of marine ecosystems, providing essential ecological services. Becoming extremely important to understand how these activities can affect marine microbiomes by altering their function and diversity. In this thesis work, we were able to present a comprehensive evaluation of different functional feeds assessing their effects in terms of growth and gut health of three fish species, Rainbow Trout (Oncorhynchus mykiss), Gilthead seabream (Sparus aurata) and Zebrafish (Danio rerio). We also explored the impact of Aquaculture on the surrounding marine microbiomes, using Patella caerulea as a model holobionts. Finally, we provided a synoptical study on the microbiomes of the water column and surface sediments in North-Western Adriatic Sea (Italy), providing the finest-scale mapping of marine microbiomes in the Mediterranean Sea.

Comportamento elastodinamico di meccanismi in catena chiusa bilanciati: analisi numerica e sperimentale di un quadrilatero articolato

The study presented in this work deals with the investigation of the effects produced by two common techniques of static balancing on the dynamic performances of closed-chain linkages, taking into account the compliance of the mechanism components. The long-term goal of the research consists in determining an optimal balancing strategy for parallel spatial manipulators. The present contribution is a starting point and it focuses on the planar four-bar linkage, intended as the simplest example of closed-chain mechanism. The elastodynamic behaviour of an unbalanced four-bar linkage and two balanced ones, respectively obtained by mass and elastic balancing, is investigated by means of both numerical simulations and experimental tests. The purpose of this work is to obtain preliminary results, to be refined and broadened in future developments

Novel techniques for the remote sensing of photosynthetic processes

Remote sensing (RS) techniques have evolved into an important instrument to investigate forest function. New methods based on the remote detection of leaf biochemistry and photosynthesis are being developed and applied in pilot studies from airborne and satellite platforms (PRI, solar-induced fluorescence; N and chlorophyll content). Non-destructive monitoring methods, a direct application of RS studies, are also proving increasingly attractive for the determination of stress conditions or nutrient deficiencies not only in research but also in agronomy, horticulture and urban forestry (proximal RS). In this work I will focus on some novel techniques recently developed for the estimation of photochemistry and photosynthetic rates based (i) on the proximal measurement of steady-state chlorophyll fluorescence yield, or (ii) the remote sensing of changes in hyperspectral leaf reflectance, associated to xanthophyll de-epoxydation and energy partitioning, which is closely coupled to leaf photochemistry and photosynthesis. I will also present and describe a mathematical model of leaf steady-state fluorescence and photosynthesis recently developed in our group. Two different species were used in the experiments: Arbutus unedo, a schlerophyllous Mediterranean species, and Populus euroamericana, a broad leaf deciduous tree widely used in plantation forestry. Results show that ambient fluorescence could provide a useful tool for testing photosynthetic processes from a distance. These results confirm also the photosynthetic reflectance index (PRI) as an efficient remote sensing reflectance index estimating short-term changes in photochemical efficiency as well as long-term changes in leaf biochemistry. The study also demonstrated that RS techniques could provide a fast and reliable method to estimate photosynthetic pigment content and total nitrogen, beside assessing the state of photochemical process in our plants’ leaves in the field. This could have important practical applications for the management of plant cultivation systems, for the estimation of the nutrient requirements of our plants for optimal growth.

The Law and Economics of Hedge Fund Regulation: A Comparison between the U.S. and the EU

This doctoral dissertation seeks to assess and address the potential contribution of the hedge fund industry to financial instability. In so doing, the dissertation investigates three main questions. What are the contributions of hedge funds to financial instability? What is the optimal regulatory strategy to address the potential contribution of hedge funds to financial instability? And do new regulations in the U.S. and the EU address the contribution of hedge funds to financial instability? With respect to financial stability concerns, it is argued that despite their benefits, hedge funds can contribute to financial instability. Hedge funds’ size and leverage, their interconnectedness with Large Complex Financial Institutions (LCFIs), and the likelihood of herding behavior in the industry can potentially undermine financial stability. Nonetheless, the data on hedge funds’ size and leverage suggest that these features are far from being systemically important. In contrast, the empirical evidence on the interconnectedness of hedge funds with LCFIs and their herding behavior is mixed. Based on these findings, the thesis focuses on one particular aspect of hedge fund regulation: direct vs. indirect regulation. In this respect, a major contribution of the thesis to the literature consists in the explicit discussion of the relationships between hedge funds and other market participants. Specifically, the thesis locates the domain of the indirect regulation in the inter-linkages between hedge funds and prime brokers. Accordingly, the thesis argues that the indirect regulation is likely to address the contribution of hedge funds to systemic risk without compromising their benefits to financial markets. The thesis further conducts a comparative study of the regulatory responses to the potential contribution of hedge funds to financial instability through studying the EU Directive on Alternative Investment Fund Managers (AIFMD) and the hedge fund-related provisions of the Dodd-Frank Wall Street Reform and Consumer Protection Act of 2010.

Patogenesi molecolare delle osteomieliti associate all'impianto

L’osteomielite associata all’impianto è un processo infettivo a carico del tessuto osseo spesso accompagnato dalla distruzione dell’osso stesso. La patogenesi delle osteomieliti associate all’impianto si basa su due concetti fondamentali: l’internalizzazione del patogeno all’interno degli osteoblasti e la capacità dei batteri di formare il biofilm. Entrambi i meccanismi consentono infatti di prevenire l’eliminazione del batterio da parte delle difese immunitarie dell’ospite e di ostacolare l’azione della maggior parte degli antibiotici (che non penetrano e non agiscono pertanto su microrganismi intracellulari), così sostenendo ed alimentando l’infezione. Il saggio di invasione messo a punto su micropiastra ha consentito di investigare in modo approfondito e dettagliato il ruolo ed il peso dell’internalizzazione nella patogenesi delle infezioni ortopediche peri-protesiche causate da S. aureus, S. epidermidis, S. lugdunensis ed E. faecalis. Lo studio ha evidenziato che l’invasione delle cellule MG-63 non rappresenta un meccanismo patogenetico delle infezioni ortopediche associate all’impianto causate da S. epidermidis, S. lugdunensis ed E. faecalis; al contrario, in S. aureus la spiccata capacità invasiva rappresenta un’abile strategia patogenetica che consente al patogeno di sfuggire alla terapia sistemica e alla risposta immunitaria dell’ospite. È stato studiato inoltre il ruolo dell’immunità innata nella difesa contro il biofilm batterico. In seguito all’incubazione del biofilm opsonizzato di S. epidermidis con i PMN è stato possibile osservare la formazione delle NETs. Le NETs rappresentano ottime armi nella difesa contro il biofilm batterico, infatti le trappole sono in grado di limitare la diffusione batterica e quindi di confinare l’infezione. La comprensione del ruolo dell’internalizzazione nella patogenesi delle osteomieliti associate all’impianto e lo studio della risposta immunitaria innata a questo tipo di infezioni, spesso caratterizzate dalla presenza di biofilm, sono presupposti per identificare e affinare le migliori strategie terapeutiche necessarie ad eradicare l'infezione.

Modelling of ORC components and systems for low-medium temperature heat recovery applications with reduced environmental impact

Although its great potential as low to medium temperature waste heat recovery (WHR) solution, the ORC technology presents open challenges that still prevent its diffusion in the market, which are different depending on the application and the size at stake. Focusing on the micro range power size and low temperature heat sources, the ORC technology is still not mature due to the lack of appropriate machines and working fluids. Considering instead the medium to large size, the technology is already available but the investment is still risky. The intention of this thesis is to address some of the topical themes in the ORC field, paying special attention in the development of reliable models based on realistic data and accounting for the off-design performance of the ORC system and of each of its components. Concerning the “Micro-generation” application, this work: i) explores the modelling methodology, the performance and the optimal parameters of reciprocating piston expanders; ii) investigates the performance of such expander and of the whole micro-ORC system when using Hydrofluorocarbons as working fluid or their new low GWP alternatives and mixtures; iii) analyzes the innovative ORC reversible architecture (conceived for the energy storage), its optimal regulation strategy and its potential when inserted in typical small industrial frameworks. Regarding the “Industrial WHR” sector, this thesis examines the WHR opportunity of ORCs, with a focus on the natural gas compressor stations application. This work provides information about all the possible parameters that can influence the optimal sizing, the performance and thus the feasibility of installing an ORC system. New WHR configurations are explored: i) a first one, relying on the replacement of a compressor prime mover with an ORC; ii) a second one, which consists in the use of a supercritical CO2 cycle as heat recovery system.

Cell-free cryopreserved arterial allografts from multiorgan donors: a new strategy to fabricate artificial blood vessels suited for peripheral vascular surgery

Critical lower limb ischemia is a severe disease. A common approach is infrainguinal bypass. Synthetic vascular prosthesis, are good conduits in high-flow low-resistance conditions but have difficulty in their performance as small diameter vessel grafts. A new approach is the use of native decellularized vascular tissues. Cell-free vessels are expected to have improved biocompatibility when compared to synthetic and are optimal natural 3D matrix templates for driving stem cell growth and tissue assembly in vivo. Decellularization of tissues represent a promising field for regenerative medicine, with the aim to develop a methodology to obtain small-diameter allografts to be used as a natural scaffold suited for in vivo cell growth and pseudo-tissue assembly, eliminating failure caused from immune response activation. Material and methods. Umbilical cord-derived mesenchymal cells isolated from human umbilical cord tissue were expanded in advanced DMEM. Immunofluorescence and molecular characterization revealed a stem cell profile. A non-enzymatic protocol, that associate hypotonic shock and low-concentration ionic detergent, was used to decellularize vessel segments. Cells were seeded cell-free scaffolds using a compound of fibrin and thrombin and incubated in DMEM, after 4 days of static culture they were placed for 2 weeks in a flow-bioreactor, mimicking the cardiovascular pulsatile flow. After dynamic culture, samples were processed for histological, biochemical and ultrastructural analysis. Discussion. Histology showed that the dynamic culture cells initiate to penetrate the extracellular matrix scaffold and to produce components of the ECM, as collagen fibres. Sirius Red staining showed layers of immature collagen type III and ultrastructural analysis revealed 30 nm thick collagen fibres, presumably corresponding to the immature collagen. These data confirm the ability of cord-derived cells to adhere and penetrate a natural decellularized tissue and to start to assembly into new tissue. This achievement makes natural 3D matrix templates prospectively valuable candidates for clinical bypass procedures

Development of 3D Cancer Cell Models to Test Metabolic Interventions as an Anticancer Strategy

In recent years, it has become evident that the role of mitochondria in the metabolic rewiring is essential for cancer development and progression. The metabolic profile during tumorigenesis has been performed mainly in traditional 2D cell models, including cell lines of various lineages and phenotypes. Although useful in many ways, their relevance can be often debatable, as they lack the interactions between different cells of the tumour microenvironment and/or interaction with the extracellular matrix 1,2. Improved models are now being developed using 3D cell culture technology, contributing with increased physiological relevance 3,4. In this work, we improved a method for the generation of 3D models from healthy and tumour colon tissue, based on organoid technology, and performed their molecular and biochemical characterization and validation. Further, in-plate cryopreservation was applied to these models, and optimal results were obtained in terms of cell viability and functionality of the cryopreserved models. We also cryopreserved colon fibroblasts with the aim to introduce them in a co-culture cryopreserved model with organoids. This technology allows the conversion of cell models into “plug and play” formats. Therefore, cryopreservation in-plate facilitates the accessibility of specialized cell models to cell-based research and application, in cases where otherwise such specialized models would be out of reach. Finally, we briefly explored the field of bioprinting, by testing a new matrix to support the growth of colon tumour organoids, which revealed promising preliminary results. To facilitate the reader, we organized this thesis into chapters, divided by the main points of work which include development, characterization and validation of the model, commercial output, and associated applications. Each chapter has a brief introduction, followed by results and discussion and a final conclusion. The thesis has also a general discussion and conclusion section in the end, which covers the main results obtained during this work.

RNA Interference and cyclooxygenase-2 (COX-2) regulation in colon cancer cells

Despite new methods and combined strategies, conventional cancer chemotherapy still lacks specificity and induces drug resistance. Gene therapy can offer the potential to obtain the success in the clinical treatment of cancer and this can be achieved by replacing mutated tumour suppressor genes, inhibiting gene transcription, introducing new genes encoding for therapeutic products, or specifically silencing any given target gene. Concerning gene silencing, attention has recently shifted onto the RNA interference (RNAi) phenomenon. Gene silencing mediated by RNAi machinery is based on short RNA molecules, small interfering RNAs (siRNAs) and microRNAs (miRNAs), that are fully o partially homologous to the mRNA of the genes being silenced, respectively. On one hand, synthetic siRNAs appear as an important research tool to understand the function of a gene and the prospect of using siRNAs as potent and specific inhibitors of any target gene provides a new therapeutical approach for many untreatable diseases, particularly cancer. On the other hand, the discovery of the gene regulatory pathways mediated by miRNAs, offered to the research community new important perspectives for the comprehension of the physiological and, above all, the pathological mechanisms underlying the gene regulation. Indeed, changes in miRNAs expression have been identified in several types of neoplasia and it has also been proposed that the overexpression of genes in cancer cells may be due to the disruption of a control network in which relevant miRNA are implicated. For these reasons, I focused my research on a possible link between RNAi and the enzyme cyclooxygenase-2 (COX-2) in the field of colorectal cancer (CRC), since it has been established that the transition adenoma-adenocarcinoma and the progression of CRC depend on aberrant constitutive expression of COX-2 gene. In fact, overexpressed COX-2 is involved in the block of apoptosis, the stimulation of tumor-angiogenesis and promotes cell invasion, tumour growth and metastatization. On the basis of data reported in the literature, the first aim of my research was to develop an innovative and effective tool, based on the RNAi mechanism, able to silence strongly and specifically COX-2 expression in human colorectal cancer cell lines. In this study, I firstly show that an siRNA sequence directed against COX-2 mRNA (siCOX-2), potently downregulated COX-2 gene expression in human umbilical vein endothelial cells (HUVEC) and inhibited PMA-induced angiogenesis in vitro in a specific, non-toxic manner. Moreover, I found that the insertion of a specific cassette carrying anti-COX-2 shRNA sequence (shCOX-2, the precursor of siCOX-2 previously tested) into a viral vector (pSUPER.retro) greatly increased silencing potency in a colon cancer cell line (HT-29) without activating any interferon response. Phenotypically, COX-2 deficient HT-29 cells showed a significant impairment of their in vitro malignant behaviour. Thus, results reported here indicate an easy-to-use, powerful and high selective virus-based method to knockdown COX-2 gene in a stable and long-lasting manner, in colon cancer cells. Furthermore, they open up the possibility of an in vivo application of this anti-COX-2 retroviral vector, as therapeutic agent for human cancers overexpressing COX-2. In order to improve the tumour selectivity, pSUPER.retro vector was modified for the shCOX-2 expression cassette. The aim was to obtain a strong, specific transcription of shCOX-2 followed by COX-2 silencing mediated by siCOX-2 only in cancer cells. For this reason, H1 promoter in basic pSUPER.retro vector [pS(H1)] was substituted with the human Cox-2 promoter [pS(COX2)] and with a promoter containing repeated copies of the TCF binding element (TBE) [pS(TBE)]. These promoters were choosen because they are partculary activated in colon cancer cells. COX-2 was effectively silenced in HT-29 and HCA-7 colon cancer cells by using enhanced pS(COX2) and pS(TBE) vectors. In particular, an higher siCOX-2 production followed by a stronger inhibition of Cox-2 gene were achieved by using pS(TBE) vector, that represents not only the most effective, but also the most specific system to downregulate COX-2 in colon cancer cells. Because of the many limits that a retroviral therapy could have in a possible in vivo treatment of CRC, the next goal was to render the enhanced RNAi-mediate COX-2 silencing more suitable for this kind of application. Xiang and et al. (2006) demonstrated that it is possible to induce RNAi in mammalian cells after infection with engineered E. Coli strains expressing Inv and HlyA genes, which encode for two bacterial factors needed for successful transfer of shRNA in mammalian cells. This system, called “trans-kingdom” RNAi (tkRNAi) could represent an optimal approach for the treatment of colorectal cancer, since E. Coli in normally resident in human intestinal flora and could easily vehicled to the tumor tissue. For this reason, I tested the improved COX-2 silencing mediated by pS(COX2) and pS(TBE) vectors by using tkRNAi system. Results obtained in HT-29 and HCA-7 cell lines were in high agreement with data previously collected after the transfection of pS(COX2) and pS(TBE) vectors in the same cell lines. These findings suggest that tkRNAi system for COX-2 silencing, in particular mediated by pS(TBE) vector, could represent a promising tool for the treatment of colorectal cancer. Flanking the studies addressed to the setting-up of a RNAi-mediated therapeutical strategy, I proposed to get ahead with the comprehension of new molecular basis of human colorectal cancer. In particular, it is known that components of the miRNA/RNAi pathway may be altered during the progressive development of colorectal cancer (CRC), and it has been already demonstrated that some miRNAs work as tumor suppressors or oncomiRs in colon cancer. Thus, my hypothesis was that overexpressed COX-2 protein in colon cancer could be the result of decreased levels of one or more tumor suppressor miRNAs. In this thesis, I clearly show an inverse correlation between COX-2 expression and the human miR- 101(1) levels in colon cancer cell lines, tissues and metastases. I also demonstrate that the in vitro modulating of miR-101(1) expression in colon cancer cell lines leads to significant variations in COX-2 expression, and this phenomenon is based on a direct interaction between miR-101(1) and COX-2 mRNA. Moreover, I started to investigate miR-101(1) regulation in the hypoxic environment since adaptation to hypoxia is critical for tumor cell growth and survival and it is known that COX-2 can be induced directly by hypoxia-inducible factor 1 (HIF-1). Surprisingly, I observed that COX-2 overexpression induced by hypoxia is always coupled to a significant decrease of miR-101(1) levels in colon cancer cell lines, suggesting that miR-101(1) regulation could be involved in the adaption of cancer cells to the hypoxic environment that strongly characterize CRC tissues.

Optimal Supervisory Control of Hybrid Vehicles

Hybrid vehicles (HV), comprising a conventional ICE-based powertrain and a secondary energy source, to be converted into mechanical power as well, represent a well-established alternative to substantially reduce both fuel consumption and tailpipe emissions of passenger cars. Several HV architectures are either being studied or already available on market, e.g. Mechanical, Electric, Hydraulic and Pneumatic Hybrid Vehicles. Among the others, Electric (HEV) and Mechanical (HSF-HV) parallel Hybrid configurations are examined throughout this Thesis. To fully exploit the HVs potential, an optimal choice of the hybrid components to be installed must be properly designed, while an effective Supervisory Control must be adopted to coordinate the way the different power sources are managed and how they interact. Real-time controllers can be derived starting from the obtained optimal benchmark results. However, the application of these powerful instruments require a simplified and yet reliable and accurate model of the hybrid vehicle system. This can be a complex task, especially when the complexity of the system grows, i.e. a HSF-HV system assessed in this Thesis. The first task of the following dissertation is to establish the optimal modeling approach for an innovative and promising mechanical hybrid vehicle architecture. It will be shown how the chosen modeling paradigm can affect the goodness and the amount of computational effort of the solution, using an optimization technique based on Dynamic Programming. The second goal concerns the control of pollutant emissions in a parallel Diesel-HEV. The emissions level obtained under real world driving conditions is substantially higher than the usual result obtained in a homologation cycle. For this reason, an on-line control strategy capable of guaranteeing the respect of the desired emissions level, while minimizing fuel consumption and avoiding excessive battery depletion is the target of the corresponding section of the Thesis.

The application of a new psychotherapeutic strategy for enhancing eudaimonic well-being in children with mood and anxiety disorders

The aim of the dissertation was to test the feasibility of a new psychotherapeutic protocol for treating children and adolescents with mood and anxiety disorders: Child-Well-Being Therapy (CWBT). It originates from adult Well-Being Therapy protocol (WBT) and represents a conceptual innovation for treating affective disorders. WBT is based on the multidimensional model of well-being postulated by Ryff (eudaimonic perspective), in sequential combination with cognitive-behavioral therapy (CBT). Results showed that eudaimonic well-being was impaired in children with affective disorders in comparison with matched healthy students. A first open investigation aimed at exploring the feasibility of a 8-session CWBT protocol in a group of children with emotional and behavioural disorders has been implemented. Data showed how CWBT resulted associated to symptoms reduction, together with the decrease of externalizing problems, maintained at 1-year follow-up. CWBT triggered also an improvement in psychological well-being as well as an increasing flourishing trajectory over time. Subsequently, a modified and extended version of CWBT (12-sessions) has been developed and then tested in a controlled study with 34 patients (8 to 16 years) affected by mood and anxiety disorders. They were consecutively randomized into 3 different groups: CWBT, CBT, 6-month waiting list (WL). Both treatments resulted effective in decreasing distress and in improving well-being. Moreover, CWBT was associated with higher improvement in anxiety and showed a greater recovery rate (83%) than CBT (54%). Both groups maintained beneficial effects and CWBT group displayed a lower level of distress as well as a higher positive trend in well-being scores over time. Findings need to be interpret with caution, because of study limitations, however important clinical implications emerged. Further investigations should determine whether the sequential integration of well-being and symptom-oriented strategies could play an important role in children and adolescents’ psychotherapeutic options, fostering a successful adaptation to adversities during the growth process.

Design of experiment in drug development: optimal allocations in multi-arm clinical trials and design space for multi-step processes

In this thesis, we deal with the design of experiments in the drug development process, focusing on the design of clinical trials for treatment comparisons (Part I) and the design of preclinical laboratory experiments for proteins development and manufacturing (Part II). In Part I we propose a multi-purpose design methodology for sequential clinical trials. We derived optimal allocations of patients to treatments for testing the efficacy of several experimental groups by also taking into account ethical considerations. We first consider exponential responses for survival trials and we then present a unified framework for heteroscedastic experimental groups that encompasses the general ANOVA set-up. The very good performance of the suggested optimal allocations, in terms of both inferential and ethical characteristics, are illustrated analytically and through several numerical examples, also performing comparisons with other designs proposed in the literature. Part II concerns the planning of experiments for processes composed of multiple steps in the context of preclinical drug development and manufacturing. Following the Quality by Design paradigm, the objective of the multi-step design strategy is the definition of the manufacturing design space of the whole process and, as we consider the interactions among the subsequent steps, our proposal ensures the quality and the safety of the final product, by enabling more flexibility and process robustness in the manufacturing.

Progettazione, modellazione e integrazione di risorse rinnovabili per promuovere lo sfruttamento dell'energia da moto ondoso e la Blue Growth.

L’energia da onda potrebbe assumere un ruolo fondamentale per la transizione energetica durante i prossimi decenni, grazie alla sua continuità nel tempo molto superiore rispetto ad altre risorse rinnovabili e alla sua vasta distribuzione nello spazio. Tuttavia, l’energia da onda è ancora lontana dall’essere economicamente sostenibile, a causa di diverse problematiche tecnologiche e alle difficoltà finanziarie associate. In questa ricerca, si è innanzitutto affrontata una delle maggiori sfide tecniche, nello specifico la progettazione e modellazione di sistemi di ancoraggio per i dispositivi galleggianti, proponendo possibili soluzioni per la modellazione numerica di sistemi di ancoraggio complessi e per l’ottimizzazione dei dispositivi stessi. Successivamente sono state analizzate le possibili sinergie strategiche di installazioni per lo sfruttamento della energia da onda con altre risorse rinnovabili e la loro applicazione nel contesto di aree marine multiuso. In particolare, una metodologia per la valutazione della combinazione ottimale delle risorse rinnovabili è stata sviluppata e verificata in due diversi casi studio: un’isola e una piattaforma offshore. Si è così potuto evidenziare l’importante contributo della risorsa ondosa per la continuità energetica e per la riduzione della necessità di accumulo. Inoltre, è stato concepito un metodo di supporto decisionale multicriteriale per la valutazione delle opzioni di riuso delle piattaforme offshore alla fine della loro vita operativa, come alternativa al decommissionamento, nell’ottica di una gestione sostenibile e della ottimizzazione dell’uso dello spazio marino. Sulla base dei criteri selezionati, l’inclusione di attività innovative come la produzione di energia da onda si è dimostrata essere rilevante per rendere vantaggioso il riuso rispetto al decommissionamento. Numerosi studi recenti hanno infatti sottolineato che, nell’ambito della “crescita blu”, i mercati come l’oil&gas, le attività offshore e le isole stimoleranno lo sviluppo di tecnologie innovative come lo sfruttamento dell’energia da onda, promuovendo la sperimentazione e fornendo un importante contributo all’avanzamento tecnico e alla commercializzazione.

A 3D biomimetic in vitro model: a novel strategy to investigate glioma biology

Gliomas are one of the most frequent primary malignant brain tumors. Acquisition of stem-like features likely contributes to the malignant nature of high-grade gliomas and may be responsible for the initiation, growth, and recurrence of these tumors. In this regard, although the traditional 2D cell culture system has been widely used in cancer research, it shows limitations in maintaining the stemness properties of cancer and in mimicking the in vivo microenvironment. In order to overcome these limitations, different three-dimensional (3D) culture systems have been developed to mimic better the tumor microenvironment. Cancer cells cultured in 3D structures may represent a more reliable in vitro model due to increased cell-cell and cell-extracellular matrix (ECM) interaction. Several attempts to recreate brain cancer tissue in vitro are described in literature. However, to date, it is still unclear which main characteristics the ideal model should reproduce. The overall goal of this project was the development of a 3D in vitro model able to reproduce the brain ECM microenvironment and to recapitulate pathological condition for the study of tumor stroma interactions, tumor invasion ability, and molecular phenotype of glioma cells. We performed an in silico bioinformatic analysis using GEPIA2 Software to compare the expression level of seven matrix protein in the LGG tumors with healthy tissues. Then, we carried out a FFPE retrospective study in order to evaluate the percentage of expression of selected proteins. Thus, we developed a 3D scaffold composed by Hyaluronic Acid and Collagen IV in a ratio of 50:50. We used two astrocytoma cell lines, HTB-12 and HTB-13. In conclusion, we developed an in vitro 3D model able to reproduce the composition of brain tumor ECM, demonstrating that it is a feasible platform to investigate the interaction between tumor cells and the matrix.

Growth performance, gut health, and metabolism of broilers under thermoneutral and heat stress conditions: multidisciplinary studies on the effects of nutritional strategies and genotype

This PhD project aimed to (i) investigate the effects of three nutritional strategies (supplementation of a synbiotic, a muramidase, or arginine) on growth performance, gut health, and metabolism of broilers fed without antibiotics under thermoneutral and heat stress conditions and to (ii) explore the impacts of heat stress on hypothalamic regulation of feed intake in three broiler lines from diverse stages of genetic selection and in the red jungle fowl, the ancestor of domestic chickens. Synbiotic improved feed efficiency and footpad health, increased Firmicutes and reduced Bacteroidetes in the ceca of birds kept in thermoneutral conditions, while did not mitigate the impacts of heat stress on growth performance. Under optimal thermal conditions, muramidase increased final body weight and reduced cumulative feed intake and feed conversion ratio in a dose-dependent way. The highest dose reduced the risk of footpad lesions, cecal alpha diversity, the Firmicutes to Bacteroidetes ratio, and butyrate producers, increased Bacteroidaceae and Lactobacillaceae, plasmatic levels of bioenergetic metabolites, and reduced the levels of pro-oxidant metabolites. The same dose, however, failed to reduce the effects of heat stress on growth performance. Arginine supplementation improved growth rate, final body weight, and feed efficiency, increased plasmatic levels of arginine and creatine and hepatic levels of creatine and essential amino acids, reduced alpha diversity, Firmicutes, and Proteobacteria (especially Escherichia coli), and increased Bacteroidetes and Lactobacillus salivarius in the ceca of thermoneutral birds. No arginine-mediated attenuation of heat stress was found. Heat stress altered protein metabolism and caused the accumulation of antioxidant and protective molecules in oxidative stress-sensitive tissues. Arginine supplementation, however, may have partially counterbalanced the effects of heat stress on energy homeostasis. Stable gene expression of (an)orexigenic neuropeptides was found in the four chicken populations studied, but responses to hypoxia and heat stress appeared to be related to feed intake regulation.