Campi elettromagnetici pulsanti nella terapia dell'artrosi del cane

The aim of this study is to evaluate the effects of the pulsed electromagnetic fields (PEMFS) on pain relief and functional capacity of dogs with osteoarthritis by which a single centre study prospective clinical trial. PEMFs are non ionized, athermic and time varying electromagnetic fields that has been successfully used for the treatment of osteoarthritis in human thanks to their chondroprotective, antinflammatory and analgesic property. 20 dogs were treated with PEMFs , 3 times per week for a total of 20 sessions.We found beneficial effects on pain relief and lameness in the absence of adverse effect. The decrease of pain impacted positively on the health-dogs related quality of life and the grade of satisfaction of their owner was very high. The benefits were obvious at half therapy and lasted for a medium long time. This is the first published report concerning PEMFs treatment on canine osteoarthtitis. The result of this study proves that PEMFs is a non –invasive remedy, lacking in adverse effect , easy to employ and useful for controlling pain and inflammation associated with osteoarthritis.

La funzione limitatrice dei principi - valori costituzionali nella giurisprudenza costituzionale. Riflessi ed implicazioni sulla legalità penale

La tesi di ricerca si propone di indagare il riflesso che i principi/valori producono sul parametro nel sindacato di legittimità costituzionale, al fine di verificarne le implicazioni sulla legalità, in termini di prevedibilità e certezza. In particolare, delineata la connessione tra principi e valori costituzionali e, ricostruito, secondo la teoria dell'ordinamento, il rapporto tra valori e normatività,si analizzano i riflessi prodotti, sul piano interpretativo, dall’apertura del parametro costituzionale alla logica dei valori, enfatizzandone le ricadute sul controllo di costituzionalità delle leggi. Identificato il nesso tra principi e valori nella capacità funzionale dei primi di realizzare i diritti fondamentali, si è inteso rimarcare come la più estesa realizzazione dei principi-valori costituzionali potrebbe compiersi a spese della legge e della certezza del diritto, in una relazione inversamente proporzionale. Ciò apparirebbe evidente dall’ottica privilegiata della materia penale, per cui una legalità materiale, letta alla luce di criteri di adeguatezza e di ragionevole proporzione, seppur vicina alle esigenze di giustizia del caso concreto, se spinta in eccessi interpretativi rischia di invadere il campo del legislatore, unico deputato a compiere scelte di valore.

Physical activity and osseointegration: "step forward" to improve the quality of life of a trans-femoral amputee person

Lower limb amputation is an event that inevitably changes the lifestyle of the person with a significant impact on quality of life. The socket-type prosthesis entails that the residual limb is in direct contact with the socket which often implies numerous disadvantages. Osseointegrated prosthesis is a solution that avoids skin problems because not include the presence of the socket. In this type of prosthesis, a stem is surgically inserted inside the medullary canal and connected with the external prosthetic limb. Therefore, this thesis aims to highlight and explore the main strengths and problems of osseointegrated prostheses and to examine the role of physical activity, with attention to functional capacity and bone quality. The objectives of the thesis will be developed through 5 studies: (I) A gait analysis of a 44 years-old male patient who underwent surgery for the implantation of an osseointegrated prosthesis; (II) A systematic review to investigate the state of stump bone quality in patients with limb amputations; (III) A systematic review of the technologies involved in such devices has been carried out to identify the most fruitful ones in improving bone quality; (IV) A systematic review investigating the topic of physical activity and bone turnover biomarkers; (V) A systematic review to investigate the effects of physical activity interventions combined with drug treatments on bone biomarkers in people with osteopenia and osteoporosis. The integrated prosthesis is a good solution for people with lower limb amputation who cannot use their traditional socket-type prosthesis. Although many objectives have already been achieved, there are still many aspects that we can improve. These include the creation of a multidisciplinary path that support patients along their path, with particular attention to the pre-surgery and the post-rehabilitation phase that is still lacking even if of fundamental impact in determining the quality of life.

Functional characterization of Streptococcus pyogenes pili

Group A Streptococcus is a Gram-positive human pathogen able to colonize both upper respiratory tract and skin. GAS is responsible for several acute diseases and autoimmune sequelae that account for half a million deaths worldwide every year (Cunningham et al., 2000). As other bacteria, GAS infections requires the capacity of the pathogen to adhere to host tissues and to form cell aggregates. The ability to persist in distinct host niches like the throat and the skin and to trigger infections is associated with the expression of different GAS virulence factors. GAS pili has been described as important virulence factors encoded by different FCT-operon regions. Based on this information, we decided to study the possible effect of environmental conditions that could regulate the pili expression. In this study we reported the influence of pH environment variations in biofilm formation for strains pertaining to a panel of different GAS FCT-types. The biofilm formation was promoted, excepted in the FCT-1 strains, by a changing in pH from physiological to acidic condition of growth in in vitro biofilm assay. By analyzing the possible association between biofilm formation and pH dependence, we have found that in FCT-2 and FCT-3 strains, the biofilm is promoted by pH reduction leading to an increase of pili expression. These data confirmed a direct link between pH dependent pilus expression and biofilm formation in GAS. As pili are a multi component structure we decided to investigate the functional role of one of its subunits, the AP-1 protein. AP-1 is highly conserved through the different FCT-types and suggests a possible essential role for the pili function. We focused our attention on the AP-1 protein encoded by the FCT-1 strains (M6). In particular this AP-1 protein contains the von Willebrand Factor A (VWFA) domain, which share an homology with the human VWFA domain that has been reported to be involved in adhesion process. We have demonstrated that the AP-1 protein binds to human epithelial cells by its VWFA domain, whereas the biofilm formation is mediated by the N-terminal region of AP-1 protein. Moreover, analyzing the importance of AP-1 in in vivo experiments we found a major capacity of tissue dissemination for the wild-type strain compared to the isogenic AP-1 deletion mutant. Pili have been also reported as potential vaccine candidates against Gram positive bacteria. For these reason we decided to investigate the relationship between cross reaction of sera raised against different GAS and GBS pilin subunits and the presence of a conserved Cna_B domain, in different pilin components. Our idea was to investigate if, using pilus conserved domains, a broad coverage vaccine against streptococcal infection could be possible.

Epidemiological and functional characterization of Streptococcus pneumoniae pili

Streptococcus pneumoniae is an important life threatening human pathogen causing agent of invasive diseases such as otitis media, pneumonia, sepsis and meningitis, but is also a common inhabitant of the respiratory tract of children and healthy adults. Likewise most streptococci, S. pneumoniae decorates its surface with adhesive pili, composed of covalently linked subunits and involved in the attachment to epithelial cells and virulence. The pneumococcal pili are encoded by two genomic regions, pilus islet 1 (PI-1), and pilus islet-2 (PI-2), which are present in about 30% and 16% of the pneumococcal strains, respectively. PI-1 exists in three clonally related variants, whereas PI-2 is highly conserved. The presence of the islets does not correlate with the serotype of the strains, but with the genotype (as determined by Multi Locus Sequence Typing). The prevalence of PI-1 and PI-2 positive strains is similar in isolates from invasive disease and carriage. To better dissect a possible association between PIs presence and disease we evaluated the distribution of the two PIs in a panel of 113 acute otitis media (AOM) clinical isolates from Israel. PI-1 was present in 30.1% (N=34) of the isolates tested, and PI-2 in 7% (N=8). We found that 50% of the PI-1 positive isolates belonged to the international clones Spain9V-3 (ST156) and Taiwan19F-14 (ST236), and that PI-2 was not present in the absence of Pl-1. In conclusion, there was no correlation between PIs presence and AOM, and, in general, the observed differences in PIs prevalence are strictly dependent upon regional differences in the distribution of the clones. Finally, in the AOM collection the prevalence of PI-1 was higher among antibiotic resistant isolates, confirming previous indications obtained by the in silico analysis of the MLST database collection. Since the pilus-1 subunits were shown to confer protection in mouse models of infection both in active and passive immunization studies, and were regarded as potential candidates for a new generation of protein-based vaccines, the functional characterization was mainly focused on S. pneumoniae pilus -1 components. The pneumococcal pilus-1 is composed of three subunits, RrgA, RrgB and RrgC, each stabilized by intra-molecular isopeptide bonds and covalently polymerized by means of inter-molecular isopeptide bonds to form an extended fibre. The pilus shaft is a multimeric structure mainly composed by the RrgB backbone subunit. The minor ancillary proteins are located at the tip and at the base of the pilus, where they have been proposed to act as the major adhesin (RrgA) and as the pilus anchor (RrgC), respectively. RrgA is protective in in vivo mouse models, and exists in two variants (clades I and II). Mapping of the sequence variability onto the RrgA structure predicted from X-ray data showed that the diversity was restricted to the “head” of the protein, which contains the putative binding domains, whereas the elongated “stalk” was mostly conserved. To investigate whether this variability could influence the adhesive capacity of RrgA and to map the regions important for binding, two full-length protein variants and three recombinant RrgA portions were tested for adhesion to lung epithelial cells and to purified extracellular matrix (ECM) components. The two RrgA variants displayed similar binding abilities, whereas none of the recombinant fragments adhered at levels comparable to those of the full-length protein, suggesting that proper folding and structural arrangement are crucial to retain protein functionality. Furthermore, the two RrgA variants were shown to be cross-reactive in vitro and cross-protective in vivo in a murine model of passive immunization. Taken together, these data indicate that the region implicated in adhesion and the functional epitopes responsible for the protective ability of RrgA may be conserved and that the considerable level of variation found within the “head” domain of RrgA may have been generated by immunologic pressure without impairing the functional integrity of the pilus.

Use of sub-lethal high pressure homogenization (HPH) treatments to enhance functional properties of lactic acid bacteria probiotic strains

The aim of this PhD thesis was to evaluate the effect of a sub-lethal HPH treatment on some probiotic properties and on cell response mechanisms of already-known functional strains, isolated from Argentinean dairy products. The results achieved showed that HPH treatments, performed at a sub-lethal level of 50 MPa, increased some important functional and technological characteristics of the considered non intestinal probiotic strains. In particular, HPH could modify cell hydrophobicity, autoaggregation and resistance to acid gastric conditions (tested in in vitro model), cell viability and cell production of positive aroma compounds, during a refrigerate storage in a simulated dairy product. In addition, HPH process was able to increase also some probiotic properties exerted in vivo and tested for two of the considered strains. In fact, HPH-treated cells were able to enhance the number of IgA+ cells more than other not treated cells, although this capacity was time dependent. On the other hand, HPH treatment was able to modify some important characteristics that are linked to the cell wall and, consequently, could alter the adhesion capacity in vivo and the interaction with the intestinal cells. These modifications, involving cell outermost structures, were highlighted also by Trasmission Electron Microscopy (TEM) analysis. In fact, the micrographs obtained showed a significant effect of the pressure treatment on the cell morphology and particularly on the cell wall. Moreover, the results achieved showed that composition of plasma membranes and their level of unsaturation are involved in response mechanisms adopted by cells exposed to the sub-lethal HPH treatment. Although the response to the treatment varied according to the characteristics of individual strains, time of storage and suspension media employed, the results of present study, could be exploited to enhance the quality of functional products and to improve their organoleptic properties.

The inhibition of aerobic glycolysis as a therapeutic approach to improve cancer chemotherapy

The aim of the research project discussed in this thesis was to study the inhibition of aerobic glycolysis, that is the metabolic pathway exploited by cancer cells for the ATP generation. This observation has led to the evaluation of glycolytic inhibitors as potential anticancer agents. Lactate dehydrogenase (LDH) is the only enzyme whose inhibition should allow a blocking of aerobic glycolysis of tumor cells without damaging the normal cells which, in conditions of normal functional activity and sufficient oxygen supply, do not need this enzyme. In preliminar experiments we demonstrated that oxamic acid and tartronic acid, two LDH competitive inhibitors, impaired aerobic glycolysis and replication of cells from human hepatocellular carcinoma. Therefore, we proposed that the depletion of ATP levels in neoplastic cells, could improved the chemotherapeutic index of associated anticancer drugs; in particular, it was studied the association of oxamic acid and multi-targeted kinase inhibitors. A synergistic effect in combination with sorafenib was observed, and we demonstrated that this was related to the capacity of sorafenib to hinder the oxidative phosphorylation, so that cells were more dependent to aerobic glycolysis. These results linked to LDH blockage encouraged us to search for LDH inhibitors more powerful than oxamic acid; thus, in collaboration with the Department of Pharmaceutical Sciences of Bologna University we identified a new molecule, galloflavin, able to inhibit both A and B isoforms of LDH enzyme. The effects of galloflavin were studied on different human cancer cell lines (hepatocellular carcinoma, breast cancer, Burkitt’s lymphoma). Although exhibiting different power on the tested cell lines, galloflavin was constantly found to inhibit lactate and ATP production and to induce cell death, mainly in the form of apoptosis. Finally, as LDH-A is able to bind single stranded DNA, thus stimulating cell transcription, galloflavin effects were also studied on this other LDH function.

Salinity Effect on Horticultural Crops: Morphological, Physiological, and Biomolecular Elements of Salinity Stress Response

Among abiotic stresses, high salinity stress is the most severe environmental stress. High salinity exerts its negative impact mainly by disrupting the ionic and osmotic equilibrium of the cell. In saline soils, high levels of sodium ions lead to plant growth inhibition and even death. Salt tolerance in plants is a multifarious phenomenon involving a variety of changes at molecular, organelle, cellular, tissue as well as whole plant level. In addition, salt tolerant plants show a range of adaptations not only in morphological or structural features but also in metabolic and physiological processes that enable them to survive under extreme saline environments. The main objectives of my dissertation were understanding the main physiological and biomolecular features of plant responses to salinity in different genotypes of horticultural crops that are belonging to different families Solanaceae (tomato) and Cucurbitaceae (melon) and Brassicaceae (cabbage and radish). Several aspects of crop responses to salinity have been addressed with the final aim of combining elements of functional stress response in plants by using several ways for the assessment of plant stress perception that ranging from destructive measurements (eg. leaf area, relative growth rate, leaf area index, and total plant fresh and dry weight), to physiological determinations (eg. stomatal conductance, leaf gas exchanges, water use efficiency, and leaf water relation), to the determination of metabolite accumulation in plant tissue (eg. Proline and protein) as well as evaluation the role of enzymatic antioxidant capacity assay in scavenging reactive oxygen species that have been generated under salinized condition, and finally assessing the gene induction and up-down regulation upon salinization (eg. SOS pathway).

Emerging plant-based proteins: comparison from extraction to functional properties and food application

The increasing demand for alternatives to meat food products, which is linked to ethical and environmental reasons, highlights the necessity of using different protein sources. Plant proteins provide a valid option, thanks to the relative low costs, high availability and wide supply sources. The current process used to produce plant concentrates and isolates is the alkaline extraction followed by isoelectric precipitation. However, despite the high purity of the proteins, it presents some drawbacks. Innovative protein extraction processes are emerging, with the aim of reducing the environmental impact and the costs, as well as improving the functional properties. In this study, the traditional wet protein extraction and another simplified wet process were used to obtain protein-rich extracts out of different plants. The sources considered in the project were de-oiled sunflower and canola, chickpea, lentils, and the camelina meal, an emerging oleaginous seed interesting for its high content of omega 3. The extracts obtained from the two processes were then analysed for their capacities to hold water and fat, to form gel and a stable foam. Results highlighted strong differences concerning the protein content, yield and functionalities. The extracts obtained with the alkaline process confirmed the literature data about the four plant sources (sunflower, canola, chickpea and lentils) and allow to obtain a camelina concentrate with a protein content of 63 % and a protein recovery of 41 %. The second easiest process was not effective to obtain a protein enrichment in oleaginous sources, whereas an enrichment of 10 and 15 % was obtained in chickpea and lentils, respectively. The functional properties were also completely different: the easiest process produced protein ingredients completely water-soluble at pH 7, with a discrete foaming capacity compared to the extracts obtained with alkaline process. These characteristics could make these extracts suitable for the plant milk-analogue products.

Alpha oscillations index the functionality and the plastic changes of the visual system

Alpha oscillations are linked to visual awareness and to the periodical sampling of visual information, suggesting that alpha rhythm reflect an index of the functionality of the posterior cortices, and hence of the visual system. Therefore, the present work described a series of studies investigating alpha oscillations as a biomarker of the functionality and the plastic modifications of the visual system in response to lesions to the visual cortices or to external stimulations. The studies presented in chapter 5 and 6 showed that posterior lesions alter alpha oscillations in hemianopic patients, with reduced alpha reactivity at the eyes opening and decreased alpha functional connectivity, especially in right-lesioned hemianopics, with concurrent dysfunctions in the theta range, suggesting a specialization of the right hemisphere in orchestrating alpha oscillations and coordinating complex interplays among different brain rhythms. The study presented in chapter 7 investigated a mechanism of rhythmical attentional sampling of visual information in healthy participants, showing that perceptual performance is influenced by a rhythmical mechanism of attentional allocation, occurring at lower-alpha frequencies (i.e., 7 Hz), when a single spatial location is monitored, and at lower frequencies (i.e., 5 Hz), when attention is allocated to two spatial locations. Moreover, the right hemisphere seemed to have a dominance in this rhythmical attentional sampling, distributing attentional resources to the entire visual field. Finally, the study presented in chapter 8 showed that prolonged visual entrainment induce long-term modulations of resting-state alpha activity in healthy participants, suggesting that persistent modifications in the functionality of the visual system are possible. Altogheter, these findings show that functional processes and plastic changes of the visual system are reflected in alpha oscillatory patterns. Therefore, investigating and promoting alpha oscillations may contribute to the development of rehabilitative protocols to ameliorate the functionality of the visual system, in brain lesioned patients.