Ruolo del sistema endocannabinoide e delle molecole ad esso correlate nel controllo delle cellule non neuronali in modelli di neurodegenerazione

Le malattie neurodegenerative costituiscono un insieme di condizioni patologiche caratterizzate da una lenta, progressiva ed irreversibile perdita di neuroni e sinapsi in specifiche aree del sistema nervoso centrale. Secondo recenti ricerche, le cellule non neuronali rivestono un ruolo di primo piano nella patogenesi di queste patologie poichè tramite il rilascio di citochine proinfiammatorie, specie reattive dell’ossigeno e fattori pro-angiogenici all’interno del SNC, favorirebbero la neurodegenerazione. Il sistema endocannabinoide esercita un controllo diretto sulle cellule non neuronali modulandone lo stato di attivazione. Ad esempio, le cellule gliali possono esprimere sia CB1 che CB2, la cui attivazione ne modula negativamente l’attività pro-infiammatoria. Il presente studio ha avuto come obiettivo la valutazione dell'effetto di molecole correlate al sistema endocannabinoide sulle cellule non neuronali ritenute, ad oggi, maggiormente coinvolte nell’eziopatogenesi del morbo di Parkinson e del morbo di Alzheimer. Nel modello murino di morbo di Parkinson indotto da MPTP sono stati valutati gli effetti dell’inibizione dell’enzima MAGL, ottenuta mediante somministrazione di JZL-184, e dell’attivazione del recettore CB2 da parte dell’agonista HU-308.In seguito è stato valutato l’effetto della cannabidivarina, fitocannabinoide non psicoattivo, sulla gliosi presente in un modello chimico di morbo di Alzheimer

A Feasibility Study on the Use of a Structured Light Depth-Camera for Three-Dimensional Body Measurements of Dairy Cows in Free-Stall Barns

Frequent checks on livestock\u2019s body growth can help reducing problems related to cow infertility or other welfare implications, and recognizing health\u2019s anomalies. In the last ten years, optical methods have been proposed to extract information on various parameters while avoiding direct contact with animals\u2019 body, generally causes stress. This research aims to evaluate a new monitoring system, which is suitable to frequently check calves and cow\u2019s growth through a three-dimensional analysis of their bodies\u2019 portions. The innovative system is based on multiple acquisitions from a low cost Structured Light Depth-Camera (Microsoft Kinect\u2122 v1). The metrological performance of the instrument is proved through an uncertainty analysis and a proper calibration procedure. The paper reports application of the depth camera for extraction of different body parameters. Expanded uncertainty ranging between 3 and 15 mm is reported in the case of ten repeated measurements. Coef\ufb01cients of determination R2> 0.84 and deviations lower than 6% from manual measurements where in general detected in the case of head size, hips distance, withers to tail length, chest girth, hips, and withers height. Conversely, lower performances where recognized in the case of animal depth (R2 = 0.74) and back slope (R2 = 0.12)

CHARACTERIZATION OF YB-1 AS A NOVEL STRESS BIOMARKER AND PARACRINE SIGNALING POLYPEPTIDE

Cell signalling is the complex network of connections between cells and inside the single cell. Protein trafficking is deeply involved in cell signalling. Different proteins are ordered in clusters of receptors of extracellular signals, transducers, sensors and biological response effectors. Together, they are arranged in molecular pathways that are relevant for maintenance of cell homeostasis. Protein secretion is a relevant component of eukaryotic cell signalling. The prototype cold-shock Y-box binding protein 1 (YB-1) is a multifunctional protein which regulates a variety of fundamental biological processes. Recently YB-1 has been found in human extracellular fluids and shown to be secreted by different cell types. This PhD project is focused on YB-1 protein secretion to investigate on its role as a potential paracrine signal to elicit changes or responses in nearby cells, altering their behaviour. During the first part of the project I focused on stimuli and conditions triggering YB-1 secretion. Low amounts of extracellular YB-1 are released by HEK293T cells in physiological condition. Interestingly, I found that YB-1 secretion was enhanced following oxidative insults. Secreted YB-1 protein was purified and analysed by mass spectrometry to confirm its identity. In parallel, I verified the assembly of YB-1 in stress granules (SGs), cytoplasmic foci where untranslated mRNAs are sorted or processed for reinitiation, degradation, or packaging into mRNPs. To understand the mechanism beneath YB-1 secretion and uptake by receiving cells, I tried several experimental approaches; among them the production and characterization of a stable HEK293T cell line expressing a YB-1-GFP fusion protein. The properties of this cell line as a stress biosensor were evaluated. The second part of the project was devoted to the production of purified recombinant human YB-1 (rYB-1) in bacterial host. In parallel I produced enriched fractions of YB-1 from HEK293T cell culture medium (CCM-YB-1). I found that both CCM-YB-1 and rYB-1 have anti-proliferative activity on receiving cells. Both forms of YB-1 protein were effective on different recipient cell lines, including HaCaT cells. In particular, inhibition of human keratinocytes proliferation by extracellular YB-1 was associated to a G2/M cell cycle arrest, induction of p21WAF and reduction of Np63 protein level. The obtained results suggest that sustained release of full length YB-1 protein or YB-1 derived peptides, by stress stimuli acts as paracrine/autocrine signal stimulating cell cycle arrest. Finally, I spent six months of my PhD at the Institute of Toxicology and Genetics (ITG-KIT), in Karlsruhe, Germany, where I had the opportunity to study the behaviour of the evolutionarily conserved YB-1 in zebrafish (Danio rerio), one of the most versatile genetic models for environmental studies. Unexpectedly, I found that zebrafish cells assemble YB-1 positive aggregates only in response to heat, but not oxidative stress or copper treatment. YB-1-positive aggregates were confirmed as SGs as they contained the G3BP1 protein, a well assessed SG marker in mammalian. I found that zfYB-1 gene silencing compromised cell viability under heat shock, but not in normal conditions highlighting the essential role played by YB-1 in cell survival following heat shock. My findings point to a fundamental role of YB-1 as a valuable biomarker for thermal stress

Opinion Dynamics in an Open Community

We here discuss the process of opinion formation in an open community where agents are made to interact and consequently update their beliefs. New actors (birth) are assumed to replace individuals that abandon the community (deaths). This dynamics is simulated in the framework of a simplified model that accounts for mutual affinity between agents. A rich phenomenology is presented and discussed with reference to the original (closed group) setting. Numerical findings are supported by analytical calculations

Dynamical affinity in opinion dynamics modelling

We here propose a model to simulate the process of opinion formation, which accounts for the mutual affinity between interacting agents. Opinion and affinity evolve self-consistently, manifesting a highly non trivial interplay. A continuous transition is found between single and multiple opinion states. Fractal dimension and signature of critical behaviour are also reported. A rich phenomenology is presented and discussed with reference to corresponding psychological implications

Chronic diarrhoea in children.

Chronic diarrhoea in children shows an age related spectrum. In infants and young children a major role is related to persistent intestinal infections, intolerance to specific nutrients such as cow's milk protein, and toddler's diarrhoea. In older children and adolescents, inflammatory bowel diseases are strongly increasing and nonspecific diarrhoea is also frequent. Coeliac disease is a major cause of diarrhoea throughout childhood. In neonates, congenital diarrhoea is a rare but severe syndrome that includes several highly complex diseases. In children, diagnosis should be based on noninvasive techniques. Endoscopy should be decided based on clinical criteria, but also driven by noninvasive tests to assess the digestive absorptive functions and intestinal inflammation. A stepwise approach may reduce the need of endoscopy, also in the light of its relatively limited diagnostic yield compared to adult patients. Treatment of chronic diarrhoea in children is also substantially different from what is generally done in adults and includes a major role for nutritional interventions. Therefore chronic diarrhoea in children is a complex age-specific disorder that requires an age-specific management that is in many aspects distinct from that in adults

Problem Solving: When Groups Perform Better Than Teammates

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Dolutegravir-based anti-retroviral therapy is effective and safe in HIV-infected paediatric patients

BACKGROUND: Treatment of HIV infection in adolescents is challenging due to long duration of therapy and poor adherence. Recently, the integrase strand transfer inhibitor dolutegravir (DTG) has been approved for the use in adolescents with HIV, but evidence in clinical practice is very limited. METHODS: We describe six cases of HIV-infected children/adolescents successfully treated with DTG-based regimen. Data relative to children/adolescents managed at the Referral Center for Pediatric HIV/AIDS of the University of Naples were reviewed. Patients were tested before introduction of DTG, after 1 month and every 3 months in the first 2 years to assess virologic and immunological response, tolerance and development of side effects. Families were asked to report any suspected adverse events. RESULTS: Six patients (2 male, median age 17 years, range 12-18) were started on DTG-based anti-retroviral regimen due to low adherence to anti-retroviral treatment (ART), multiple drug resistance mutations, or development of ART-related side effects. Within 4-8 weeks after DTG treatment onset, a complete viral suppression and a concomitant increase of CD4+ cell count was observed. Four patients showed a persistent suppression after 2 years of follow-up, and 2 patients at about 1 year. One month after the introduction of DTG, the patient enrolled because of severe dyslipidaemia and hyper-transaminasemia showed a complete normalization of laboratory values. During follow-up (median 24 months, range 9-24) no adverse events were reported and most patients demonstrated a good adherence to treatment. CONCLUSIONS: DTG-based treatments demonstrated efficacy and good safety profile in adolescents. All patients demonstrated a rapid virologic and immunological response within 4-8 weeks, with good adherence and absence of side effects