Development of novel therapeutics and in vitro models for spinal cord injury

Spinal cord injury (SCI) is a devastating condition, which results from trauma to the cord, resulting in a primary injury response which leads to a secondary injury cascade, causing damage to both glial and neuronal cells. Following trauma, the central nervous system (CNS) fails to regenerate due to a plethora of both intrinsic and extrinsic factors. Unfortunately, these events lead to loss of both motor and sensory function and lifelong disability and care for sufferers of SCI. There have been tremendous advancements made in our understanding of the mechanisms behind axonal regeneration and remyelination of the damaged cord. These have provided many promising therapeutic targets. However, very few have made it to clinical application, which could potentially be due to inadequate understanding of compound mechanism of action and reliance on poor SCI models. This thesis describes the use of an established neural cell co-culture model of SCI as a medium throughput screen for compounds with potential therapeutic properties. A number of compounds were screened which resulted in a family of compounds, modified heparins, being taken forward for more intense investigation. Modified heparins (mHeps) are made up of the core heparin disaccharide unit with variable sulphation groups on the iduronic acid and glucosamine residues; 2-O-sulphate (C2), 6-O-sulphate (C6) and N-sulphate (N). 2-O-sulphated (mHep6) and N-sulphated (mHep7) heparin isomers were shown to promote both neurite outgrowth and myelination in the SCI model. It was found that both mHeps decreased oligodendrocyte precursor cell (OPC) proliferation and increased oligodendrocyte (OL) number adjacent to the lesion. However, there is a difference in the direct effects on the OL from each of the mHeps; mHep6 increased myelin internode length and mHep7 increased the overall cell size. It was further elucidated that these isoforms interact with and mediate both Wnt and FGF signalling. In OPC monoculture experiments FGF2 treated OPCs displayed increased proliferation but this effect was removed when co-treated with the mHeps. Therefore, suggesting that the mHeps interact with the ligand and inhibit FGF2 signalling. Additionally, it was shown that both mHeps could be partially mediating their effects through the Wnt pathway. mHep effects on both myelination and neurite outgrowth were removed when co-treated with a Wnt signalling inhibitor, suggesting cell signalling mediation by ligand immobilisation and signalling activation as a mechanistic action for the mHeps. However, the initial methods employed in this thesis were not sufficient to provide a more detailed study into the effects the mHeps have on neurite outgrowth. This led to the design and development of a novel microfluidic device (MFD), which provides a platform to study of axonal injury. This novel device is a three chamber device with two chambers converging onto a central open access chamber. This design allows axons from two points of origin to enter a chamber which can be subjected to injury, thus providing a platform in which targeted axonal injury and the regenerative capacity of a compound study can be performed. In conclusion, this thesis contributes to and advances the study of SCI in two ways; 1) identification and investigation of a novel set of compounds with potential therapeutic potential i.e. desulphated modified heparins. These compounds have multiple therapeutic properties and could revolutionise both the understanding of the basic pathological mechanisms underlying SCI but also be a powered therapeutic option. 2) Development of a novel microfluidic device to study in greater detail axonal biology, specifically, targeted axonal injury and treatment, providing a more representative model of SCI than standard in vitro models. Therefore, the MFD could lead to advancements and the identification of factors and compounds relating to axonal regeneration.

Candidacidal potential of licorice phenolic extract: emphasis on its mode of action

Medicinal plants bave gained a special attention in the last years, due to its renowned health benefits, such as antimicrobial effects [I]. In fact, several natural matrices bave been increasingly studied, namely for its antifungal activity against opportunistic fungi [2,3]. Candida species, although commensa! microorganisms, have caused severe organic dysfunctions to the host, once current antifungal agents have lost their recognized efficiency [2]. So, numerous studies have been carried out focusing the mechanisms of acquired drug-resistance by Candida species [mechanisms of actions, morphological changes and related kinetic parameters are of major importance. Glycyrrhiza glabra L. (licorice) hydromethanolic extract have evidenced promissory candidacidal effects, and therefore, the involved mechanisms of action need to be clarified. Thus, in the present study these modes of action were assessed, by using flow cytometry.Overall, the licorice extract induced significant and irreversible primary damages on Candida cells, being membrane disruption and consequent unviability one of the main targets. In fact, after membrane destabilization, cells lost their proper homeostasis, their metabolic functions were blocked and, consequently cells lost functionality. The relevance and interest of the achieved results open new insights towards the upcorning use of the present phenolic matrix, being important to evaluate its in viva efficacy. Therefore, further studies are necessary to deepen knowledge on this field, aiming not only to establish therapeutic and prophylactic doses, but also to improve the clinical intervention in Candida infections.

Administrative justice and the control of bureaucratic decision-making: A study investigating how decision-makers in local authority education departments respond to the work of redress mechanisms

This socio-legal thesis has explored the factors responsible for explaining whether and how redress mechanisms control bureaucratic decision-making. The research considered the three principal institutions of administrative justice: courts, tribunals, and ombudsman schemes. The field setting was the local authority education area and the thesis examined bureaucratic decision-making about admissions to school, home-to-school transport, and Special Educational Needs (SEN). The thesis adopted a qualitative approach, using interviews and documentary research, within a multiple embedded case study design. The intellectual foundations of the research were inter-disciplinary, cutting across law, socio-legal studies, public administration, organization studies, and social policy. The thesis drew on these scholarly fields to explore the nature of bureaucratic decision-making, the extent to which it can be controlled and the way that learning occurs in bureaucracies and, finally, the extent to which redress mechanisms might exercise control. The concept of control was studied across all its dimensions – in relation both to ex post control in specific cases and the more challenging notion of ex ante or structuring control. The aim of the thesis was not to measure the prevalence of bureaucratic control by redress mechanisms, but to understand the factors that might explain its presence or absence in a particular area. The findings of the research have allowed for a number of analytical refinements and extensions to be made to existing theoretical and empirical understandings. 14 factors, along with 87 supporting propositions, have been set out with the aim of making empirically derived suggestions which can be followed up in future research. In terms of the thesis’ contribution to existing knowledge, its comparative focus and its emphasis on the broad notion of control offered the potential for new insights to be developed. Overall, the thesis claims to have made three contributions to the conceptual framework for understanding the exercise of control by redress mechanisms: it emphasizes the importance of ‘feedback’ in relation to the nature of the cases referred to redress mechanisms; it calls attention to the structure of bureaucratic decision-making as well as its normative character; and it discusses how the operational modes of redress mechanisms relate to their control functions.

As relações da tríplice hélice no setor da carcinicultura do Rio Grande do Norte: uma análise a partir das abordagens da imersão social e da dependência de recursos

This work consists in a study of the Shrimp Industry in the state of Rio Grande do Norte, whose central issue relates to the understanding of how the Triple Helix (University, Government and the productive sector) interrelationship limits or expands the industry s innovation process. The study aims to understand how the Triple Helix relationship interferes in the innovation process of shrimp in Rio Grande do Norte. As the knowledge becomes the resource key for production methods, the generation of new technologies, new products and processes which demands joint and integrated action of the institutions comprising the Triple Helix: University, Government and productive sector, which possess the essential resources to innovate the process and can be maximized from cooperative relationships between the referred Institutions. Thus, in this work, it was sharply used the pioneering studies of Sabato and Botana (1968) regarding the cooperation relationship between the scientific-technological sphere, the governmental and the productive base, and studies on the Triple Helix approach, proposed by Etzkowitz and Leydesdorff (2000), in which the university has a key role in the process of technological and innovative development of countries and regions, and under which it is assigned to the very University - the character of the entrepreneurial institution, through the concept of entrepreneurial University. Aiming to overcome the criticism of Cooke (2005), regarding the limitations of the Triple Helix approach, in this study it was used - as analytical perspectives - the perspective of social immersion (Granovetter, 1985, 2005) and the theory of resources dependence (PFEFFER; SALANCIK, 1978). The analytical perspectives presented in here, despite of the different assumptions, are essential to eliminate the bias that one only approach can lead (ASTLEY; VAN DE VEM; 2007). The authors arguments focus on the fact that the integration is possible if the researcher acknowledged that different perspectives may have different descriptions of the same phenomenon. As a research strategy, this study is characterized as a study case, along with the proposed objectives - the qualitative method was used as an approach and, depending on the gathering of the sector s historical, a sectional longitudinal view approach was applied (VIEIRA, 2004). The primary and secondary data were used in order to understand the sector s evolutionary process and its inter-institutional relations - regarding the shrimp culture in Rio Grande do Norte - to promote the development, as the content was used for the technical analysis (BARDIN, 1977). The approach of social immersion and resources addiction dependence made it possible to understand that relationships are established within and between each sphere (university, government and productive sector) characterizing a network of low density relationships and strongly internal and external dependence. Based on the speech of Etzkowitz and Mello (2006), a successful Triple Helix strategy of innovation requires not only the involvement and commitment of the parts, within the institutional sphere and among them, but also the development of mechanisms to coordinate the multiple and complex interactions and interfaces, focusing on promoting both environment and context for innovation and learning; it can be acknowledge from study results that the shrimp in the State of the RN, although there are several institutional mechanisms to promote greater integration and technological development, has been presented disjointed - both internally and between the spheres - and under no legitimate practice when facing the innovational promotion and integration institutions. Due to those factors, the central institutions of the network are crucial to the promotion of innovations, spreading through their direct contacts the importance factor of the sustainable competitive activity in the world market and on the national level. However, it may be concluded, from the data, that the Triple Helix relations are interfering in a negative way on what concerns the promotion of innovations in the shrimp industry in RN

Differential mechanisms of angiotensin II and PDGF-BB on migration and proliferation of coronary artery smooth muscle cells

Angiotensin II (Ang II) and platelet-derived growth factor-BB (PDGF-BB) are associated with excessive cell migration, proliferation and many growth-related diseases. However, whether these agents utilise similar mechanisms to trigger vascular pathologies remains to be explored. The effects of Ang II and PDGF-BB on coronary artery smooth muscle cell (CASMC) migration and proliferation were investigated via Dunn chemotaxis assay and the measurement of [3H]thymidine incorporation rates, respectively. Both atherogens produced similar degrees of cell migration which were dramatically inhibited by mevastatin (10 nM). However, the inhibitory effects of losartan (10 nM) and MnTBAP (a free radical scavenger; 50 μM) were found to be unique to Ang II-mediated chemotaxis. In contrast, MnTBAP, apocynin (an antioxidant and phagocytic NADPH oxidase inhibitor; 500 μM), mevastatin and pravastatin (100 nM) equally suppressed both Ang II and PDGF-BB-induced cellular growth. Although atherogens produced similar changes in NADPH oxidase, NOS and superoxide dismutase activities, they differentially regulated antioxidant glutathione peroxidase activity which was diminished by Ang II and unaffected by PDGF-BB. Studies with signal transduction pathway inhibitors revealed the involvement of multiple pathways i.e. protein kinase C, tyrosine kinase and MAPK in Ang II- and/or PDGF-BB-induced aforementioned enzyme activity changes. In conclusion, Ang II and PDGF-BB may induce coronary atherosclerotic disease formation by stimulating CASMC migration and proliferation through agent-specific regulation of oxidative status and utilisation of different signal transduction pathways.

Maladaptive defense mechanisms are associated with decoupling of testosterone from sexual desire in women of reproductive age

O presente artigo contém uma errata, disponível em: http://www.tandfonline.com/doi/full/10.1080/15294145.2015.1108503

Fish Reproduction Is Disrupted upon Lifelong Exposure to Environmental PAHs Fractions Revealing Different Modes of Action

Polycyclic aromatic hydrocarbons (PAHs) constitute a large family of organic pollutants emitted in the environment as complex mixtures, the compositions of which depend on origin. Among a wide range of physiological defects, PAHs are suspected to be involved in disruption of reproduction. In an aquatic environment, the trophic route is an important source of chronic exposure to PAHs. Here, we performed trophic exposure of zebrafish to three fractions of different origin, one pyrolytic and two petrogenic. Produced diets contained PAHs at environmental concentrations. Reproductive traits were analyzed at individual, tissue and molecular levels. Reproductive success and cumulative eggs number were disrupted after exposure to all three fractions, albeit to various extents depending on the fraction and concentrations. Histological analyses revealed ovary maturation defects after exposure to all three fractions as well as degeneration after exposure to a pyrolytic fraction. In testis, hypoplasia was observed after exposure to petrogenic fractions. Genes expression analysis in gonads has allowed us to establish common pathways such as endocrine disruption or differentiation/maturation defects. Taken altogether, these results indicate that PAHs can indeed disrupt fish reproduction and that different fractions trigger different pathways resulting in different effects.

Allelopathy: driving mechanisms governing its activity in agriculture.

Allelopathy determines the dynamics of plant species in different environments. Understanding this biological phenomenon could help to develop applications in both natural and agricultural systems. This review summarizes the genetic and environmental characteristics that control the production and release of allelochemicals in agroecosystems. This study highlights the current understanding of the environmental changes caused by allelochemicals and summarizes the knowledge about the mechanisms of action of these compounds. Finally, it reviews novel applications of allelopathy in agricultural production systems, including the role of allelochemicals in consortia and their potential use in no-tillage cropping systems through cover crops or mulches.

Anticonvulsant and sedative effect of Fufang Changniu pills and probable mechanism of action in mice

Purpose: To investigate the anticonvulsant and sedative effects of Fufang Changniu Pills (FCP) and its probable mechanism of action in mice. Methods: The water decoction of FCP was prepared and the main constituents were determined by high performance liquid chromatography (HPLC). The anticonvulsant activities of FCP were evaluated by maximal electroshock (MES) and pentylenetetrazole (PTZ)-induced seizures in mice. Pentobarbital sodium-induced sleeping time and locomotor activity measurements were performed to evaluate the sedative effects of FCP in mice. Finally, PTZ-induced chronic seizures were established, and expressions of gamma-aminobutyric acid A receptor (GABA-A) and glutamic acid decarboxylase 65 (GAD65) in the brains of the mice were assayed by western blot in order to explore the probable mechanisms of action of the drug. Results: Gallic acid, liquiritin, cinnamyl alcohol, cinnamic acid and glycyrrhizic acid were detected in FCP decoction. FCP (50, 100 and 200 mg/kg) showed significant anticonvulsant and sedative effects on epileptic mice induced by MES (p < 0.05) and PTZ (p < 0.05). Moreover, pentobarbital sodium-induced sleeping time and locomotor activity tests showed that FCP possesses sedative effect (p < 0.05). Western blot data indicate that FCP significantly up-regulated GABA-A and GAD 65 in the brains of chronic epileptic rats (p < 0.05). Conclusion: FCP has significant anticonvulsant and sedative effects, and the mechanism of its action may be related to the up-regulation of GABA-A and GAD 65 in mice brain.

Rogue waves generation via nonlinear soliton collision in multiple-soliton state of a mode-locked fiber laser

We report for the first time, rogue waves generation in a mode-locked fiber laser that worked in multiple-soliton state in which hundreds of solitons occupied the whole laser cavity. Using real-time spatio-temporal intensity dynamics measurements, it is unveiled that nonlinear soliton collision accounts for the formation of rogue waves in this laser state. The nature of interactions between solitons are also discussed. Our observation may suggest similar formation mechanisms of rogue waves in other systems.

Potential Mechanisms Underlying Response to Effects of the Fungicide Pyrimethanil from Gene Expression Profiling inSaccharomyces cerevisiae

Pyrimethanil is a fungicide mostly applied in vineyards. When misused, residue levels detected in grape must or in the environment may be of concern. The present work aimed to analyze mechanisms underlying response to deleterious effects of pyrimethanil in the eukaryotic model Saccharomyces cerevisiae. Pyrimethanil concentration-dependent effects at phenotypic (inhibition of growth) and transcriptomic levels were examined. For transcriptional profiling, analysis focused on two sublethal exposure conditions that inhibited yeast growth by 20% or 50% compared with control cells not exposed to the fungicide. Gene expression modifications increased with the magnitude of growth inhibition, in numbers and fold-change of differentially expressed genes and in diversity of over-represented functional categories. These included mostly biosynthesis of arginine and sulfur amino acids metabolism, as well as energy conservation, antioxidant response, and multidrug transport. Several pyrimethanil-responsive genes encoded proteins sharing significant homology with proteins from phytopathogenic fungi and ecologically relevant higher eukaryotes.

The influence of actions on auditory perception: cognitive and neural mechanisms

It is well known that self-generated stimuli are processed differently from externally generated stimuli. For example, many people have noticed since childhood that it is very difficult to make a self-tickling. In the auditory domain, self-generated sounds elicit smaller brain responses as compared to externally generated sounds, known as the sensory attenuation (SA) effect. SA is manifested in reduced amplitudes of evoked responses as measured through MEEG, decreased firing rates of neurons and a lower level of perceived loudness for self-generated sounds. The predominant explanation for SA is based on the idea that self-generated stimuli are predicted (e.g., the forward model account). It is the nature of their predictability that is crucial for SA. On the contrary, the sensory gating account emphasizes a general suppressive effect of actions on sensory processing, regardless of the predictability of the stimuli. Both accounts have received empirical support, which suggests that both mechanisms may exist. In chapter 2, three behavioural studies concerning the influence of motor activation on auditory perception were presented. Study 1 compared the effect of SA and attention in an auditory detection task and showed that SA was present even when substantial attention was paid to unpredictable stimuli. Study 2 compared the loudness perception of tones generated by others between Chinese and British participants. Compared to externally generated tones, a decrease in perceived loudness for others generated tones was found among Chinese but not among the British. In study 3, partial evidence was found that even when reading words that are related to action, auditory detection performance was impaired. In chapter 3, the classic SA effect of M100 suppression was replicated with MEG in study 4. With time-frequency analysis, a potential neural information processing sequence was found in auditory cortex. Prior to the onset of self-generated tones, there was an increase of oscillatory power in the alpha band. After the stimulus onset, reduced gamma power and alpha/beta phase locking were found. The three temporally segregated oscillatory events correlated with each other and with SA effect, which may be the underlying neural implementation of SA. In chapter 4, a TMS-MEG study was presented investigating the role of the cerebellum in adapting to delayed presentation of self-generated tones (study 5). It demonstrated that in sham stimulation condition, the brain can adapt to the delay (about 100 ms) within 300 trials of learning by showing a significant increase of SA effect in the suppression of M100, but not M200 component. Whereas after stimulating the cerebellum with a suppressive TMS protocol, the adaptation in M100 suppression disappeared and the pattern of M200 suppression reversed to M200 enhancement. These data support the idea that the suppressive effect of actions on auditory processing is a consequence of both motor driven sensory predictions and general sensory gating. The results also demonstrate the importance of neural oscillations in implementing SA effect and the critical role of the cerebellum in learning sensory predictions under sensory perturbation.

Integration of Multiple Simultaneous Stimuli within and between Somatosensory sub-Modalities

Psychological characterisation of the somatosensory system often focusses on minimal units of perception, such as detection, localisation, and magnitude estimation of single events. Research on how multiple simultaneous stimuli are aggregated to create integrated, synthetic experiences is rarer. This thesis aims to shed a light on the mechanisms underlying the integration of multiple simultaneous stimuli, within and between different sub-modalities of the somatosensory system. First, we investigated the ability of healthy individuals to perceive the total intensity of composite somatosensory patterns. We found that the overall intensity of tactile, cold, or warm patterns was systematically overestimated when the multiple simultaneous stimuli had different intensities. Perception of somatosensory totals was biased towards the most salient element in the pattern. Furthermore, we demonstrated that peak-biased aggregation is a genuine perceptual phenomenon which does not rely on the discrimination of the parts, but is rather based on the salience of each stimulus. Next, we studied a classical thermal illusion to assess participants’ ability to localise thermal stimuli delivered on the fingers either in isolation, or in uniform and non-uniform patterns. We found that despite a surprisingly high accuracy in reporting the location of a single stimulus, when participants were presented with non-uniform patterns, their ability to identify the thermal state of a specific finger was completely abolished. Lastly, we investigated the perceptual and neural correlates of thermo-nociceptive interaction during the presentation of multiple thermal stimuli. We found that inhibition of pain by warmth was independent from both the position and the number of thermal stimuli administered. Our results suggest that nonlinear integration of multiple stimuli, within and between somatosensory sub-modalities, may be an efficient way by which the somatosensory system synthesises the complexity of reality, providing an extended and coherent perception of the world, in spite of its deep bandwidth limitations.

Consumismo virtuoso? Il rapporto tra politica e consumi in Italia negli anni Settanta

Attraverso l’analisi di riviste femminili, d’attualità e di intrattenimento, fonti raccolte in archivi d’impresa (Eni e Fiat) e fonti legate al mondo pubblicitario, questa tesi vuole indagare il rapporto tra politica e consumo in Italia durante gli anni Settanta. L’analisi è partita dallo studio della crisi petrolifera del 1973, le politiche di austerità e le loro conseguenze sui costumi e i consumi, per poi estendere lo sguardo fino agli anni Ottanta, il decennio definito della “svolta edonistica”. L’analisi delle fonti ha fatto emergere come la sfera del consumo fosse un campo di confronto politico per diverse componenti della società italiana, che appoggiarono o misero in discussione, con modalità e gradi di radicalità differenti, le valutazioni sull’accesso ai consumi proprie della classe politica. I casi studio hanno fatto emergere come sui temi dell’educazione al consumo e l’accesso all’informazione ci sia stato un confronto risultato in una formalizzazione istituzionale, diventata dominante, dell’azione e dei diritti di consumatori e consumatrici. Analizzare questi meccanismi ci permette di complicare la lettura evitando una polarizzazione che veda il consumatore o come totalmente passivo e senza autonomia di pensiero o come investito di un’eccessiva possibilità di azione e di una idealizzata carica progressista. Nel corso degli anni Settanta, infatti, le visioni e prospettive sulla sfera del consumo erano molteplici e complesse e si intrecciavano con l’attualità politica, sia per quel che riguardava gli scandali e le polemiche legate alla élite politica ed economica, sia per quel che concerneva dibattiti come quello sul ruolo della donna nella società e sulla questione ambientale. L’analisi di alcune di queste narrazioni sia problematizza il rapporto tra democrazia e benessere, sia mostra alcune tappe del percorso che ha portato alle definizioni odierne di consumo e consumatore, che sono ancora oggi centrali nel rapporto tra sfera politica e sfera del consumo.

Molecular changes induced by a centrally-induced state of synthetic torpor in multiple organs:a new strategy for radioprotection.

Torpor is a successful survival strategy displayed by several mammalian species to cope with harsh environmental conditions. A complex interplay of ambient, genetic and circadian stimuli acts centrally to induce a severe suppression of metabolic rate, usually followed by an apparently undefended reduction of body temperature. Some animals, such as marmots, are able to maintain this physiological state for months (hibernation), during which torpor bouts are periodically interrupted by short interbouts of normothermia (arousals). Interestingly, torpor adaptations have been shown to be associated with a large resistance towards stressors, such as radiation: indeed, if irradiated during torpor, hibernators can tolerate higher doses of radiation, showing an increased survival rate. New insights for radiotherapy and long-term space exploration could arise from the induction of torpor in non-hibernators, like humans. The present research project is centered on synthetic torpor (ST), a hypometabolic/hypothermic condition induced in a non-hibernator, the rat, through the pharmacological inhibition of the Raphe Pallidus, a key brainstem area controlling thermogenic effectors. By exploiting this procedure, this thesis aimed at: i) providing a multiorgan description of the functional cellular adaptations to ST; ii) exploring the possibility, and the underpinning molecular mechanisms, of enhanced radioresistance induced by ST. To achieve these aims, transcriptional and histological analysis have been performed in multiple organs of synthetic torpid rats and normothermic rats, either exposed or not exposed to 3 Gy total body of X-rays. The results showed that: i) similarly to natural torpor, ST induction leads to the activation of survival and stress resistance responses, which allow the organs to successfully adapt to the new homeostasis; ii) ST provides tissue protection against radiation damage, probably mainly through the cellular adaptations constitutively induced by ST, even though the triggering of specific responses when the animal is irradiated during hypothermia might play a role.