Thèses : « Modelli in vivo »

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FARAVELLI, GIULIA. « Meccanismi molecolari dell'amiloidosi sistemica : modelli in vitro e in vivo ». Doctoral thesis, Università degli studi di Pavia, 2019. http://hdl.handle.net/11571/1301267.

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Il mio progetto di ricerca ha riguardato lo studio dei meccanismi di aggregazione di proteine globulari dalla loro forma solubile a quella fibrillare insolubile. In particolare mi sono occupata dello studio di due proteine: transtiretina (TTR) e β2-microglobulina (β2-m). La prima proteina è correlata ad una forma di amiloidosi senile e sistemica denominata Senile Systemic Amyloidosis. Inoltre esistono più di 100 mutazioni di TTR che danno luogo a patologie ereditarie differenti. Dallaltra parte, lelevata concentrazione di β2-m nei pazienti dializzati per lungo tempo può causare una forma di amiloidosi acquisita chiamata Dialysis Related Amyloidosis. Nel 2012, è stata scoperta la prima variante naturale di β2-m, che presenta una sostituzione amminoacidica in posizione 76 (β2-m D76N) ed è responsabile di una forma ereditabile e molto aggressiva della patologia. Per andare a meglio comprendere quali siano i meccanismi molecolari alla base dellinsorgenza dellamiloidosi correlata a queste due proteine, ho sviluppato sia modelli della patologia in vitro sia modelli a complessità biologica maggiore in vivo. Per quanto riguarda la TTR: il mio lavoro di ricerca è stato promosso dalla recente individuazione di un meccanismo di fibrillogenesi in vitro denominato meccano- enzimatico che, utilizzando la tripsina e in condizioni di agitazione meccanica, porta alla formazione di fibrille amiloidi con caratteristiche morfologiche simili a quelle estratte ex vivo da campioni di pazienti affetti da amiloidosi. In particolare ho potuto riesaminare con questo nuovo metodo di aggregazione, lefficacia di alcuni farmaci studiati per evitare la dissociazione della struttura tetramerica di TTR che rappresenta levento scatenante dellaggregazione ottenuta in condizioni di pH estremo. Tra questi ho potuto osservare che ligandi bivalenti, tra cui mds84, legandosi a entrambi i siti di legame della tiroxina, si sono dimostrati più efficaci rispetto a quelli monovalenti. In secondo luogo ho potuto contribuire allidentificazione della proteasi responsabile del taglio proteolitico della TTR e della sua conseguente aggregazione in vivo. La plasmina è stata identificata a partire da un database denominato MEROPS, e la sua abilità di generare fibre amiloidi in vitro con caratteristiche simili a quelle naturali, è stata dimostrata sperimentalmente. Nellottica di comparare le fibrille prodotte con quelle estratte ex vivo, ho effettuato degli studi di termodinamica e ho evidenziato la maggiore stabilità delle fibrille ottenute con il metodo meccano-enzimatico rispetto a quelle ottenute a pH acido con il metodo proposto da Kelly a pH acido. Il secondo progetto di cui mi sono occupata durante questi anni ha riguardato la creazione e la caratterizzazione di una linea transgenica di C. elegans che esprime la variante naturale della β2-microglobulina umana tramite lutilizzo di un sistema termo-inducibile. Infatti con questo sistema, lespressione della proteina è permessa solo quando i vermi vengono coltivati ad alte temperature (23-25°C). Per poter valutare il difetto fenotipico di questa linea transgenica, ho utilizzato un sistema automatizzato, chiamato INVAPP/Paragon, e tramite il suo utilizzo, ho dimostrato in vivo lefficacia della doxiciclina, un noto inibitore dellaggregazione della β2-m in vitro. Ho avuto anche loccasione di effettuare uno screening su una libreria di molecole disegnata a partire da una database disponibile online che raccoglie informazioni riguardo la struttura dei complessi proteina-proteina e dei loro inibitori. Ho quindi testato questa libreria su una linea di C. elegans esprimente il peptide Aβeta1-42, modello della patologia di Alzheimer, e sono riuscita ad identificare 11 molecole in grado di agire sul difetto di motilità presentato dai nematodi transgenici.
Systemic amyloidosis is a fatal disease caused by misfolding of native globular proteins that then aggregate extracellularly as insoluble fibrils, damaging structure and function of affected organs. My PhD project has been designed to address the crucial question of the mechanism of globular to fibrillary conversion of two amyloidogenic proteins, transthyretin (TTR) and β2-microglobulin (β2-m), in conditions that closely resemble the physiological environment. For those reasons, I have been trying to combine in vitro and in vivo methods in order to get a general and broadest comprehension of the process that leads toward the deposition of amyloid fibrils. Wild-type TTR is intrinsically amyloidogenic and tends to form microscopic and clinically silent amyloid deposits in the heart, in the lungs and in the blood vessels wall of the majority of elderly people, causing Senile systemic amyloidosis (SSA). Over 100 mutations codify for protein variants of the WT TTR, causing familial amyloidosis, although they have a rare incidence. Firstly, I investigated the inhibition of transthyretin isoforms by small ligands. The study was based on the discovery of the previously unrecognised mechano enzymatic mechanism in which shear stress and proteolysis play a key role towards the formation of amyloid fibrils. This pathway of aggregation is efficiently inhibited only by ligands that occupy both binding sites in TTR. Mds84, a bivalent ligand of TTR superstabiliser family, has shown to be more potent than the monovalent ligands probably because of its additional interactions of its linker within the TTR central channel. Secondly, I have been actively involved in the identification of the putative protease responsible for proteolysis of transthyretin in vivo. In a comprehensive bioinformatics search for systemically active proteases with tryptic specificity, plasmin was selected as the leading candidate. Indeed, plasmin selectively cleaves TTR in vitro, releasing full length and truncated protomers that rapidly aggregate via nucleation and elongation into genuine amyloid fibrils. Finally, I carried out a comparative analysis of the thermodynamic stability of natural and in vitro made TTR fibrils. The second protein that I studied was β2-m. WT β2-m is associated with the amyloidosis of patients under chronic haemodialytic treatment; known as dialysis-related amyloidosis, but in the presence of a specific mutation (D76N β2-m) causes a familial form of systemic amyloidosis. Recently C. elegans, a nematode model well suited to the investigation of age-related diseases, was used in order to establish three transgenic lines expressing the wild type and two highly amyloidogenic forms, but the expression of the D76N variant was not possible with this system. Therefore, the smg-1 temperature sensitive strain was engineered, in order to express the protein variant only at higher temperatures. Using the INVertebrate Automated Phenotyping Platform (INVAPP) and an algorithm, Paragon, we were able to rapidly detect growth and motility impairment in D76N β2-m expressing worms that were incubated at 25°C. Moreover, the INVAPP/Paragon system enabled demonstration of the efficacy of doxycycline, a drug able to inhibit β2-m fibrillogenesis both in vitro and in vivo. Thus, a useful C. elegans model for D76N β2-m related amyloidosis has been developed and the INVAPP/Paragon system provides a powerful tool with which to undertake high-throughput screening in the search for candidates able to combat amyloid-induced toxicity. Indeed, using the automated system for C. elegans phenotyping, a library-scale screening was performed and 11 molecules, which are interactors of protein-protein complexes, have been selected for their ability to revert the defective phenotype of transgenic worms expressing Aβeta1-42 peptide.

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CHIORAZZI, ALESSIA. « Studio della neurotossicità periferica indotta da farmaci antitubulinici in modelli sperimentali in vitro e in vivo ». Doctoral thesis, Università degli Studi di Milano-Bicocca, 2010. http://hdl.handle.net/10281/18017.

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Lo scopo di questo lavoro è stato quelli di studiare la neurotossicità periferica indotta da due differenti farmaci antitubulinici (epotilone B e sagopilone) in modelli sperimentali in vitro e in vivo In particolare lo studio è stato articolato in due fasi consecutive: la prima fase ha previsto lo studio della tossicità periferica indotta dalla somministrazione di epotilone B attraverso la messa a punto di un modello in vitro su colture organotipiche di gangli delle radici dorsali e un successivo modello in vivo che ha visto l’utilizzo di ratti Fischer; nella seconda fase è stato studiata la tossicità periferica indotta dalla somministrazione di sagopilone attraverso la messa a punto di un modello in vitro su colture organotipiche di gangli delle radici dorsali e due successivi modelli in vivo che hanno visto l’utilizzo di ratti Wistar e sono stati svolti per ricercare la massima dose tollerata di farmaco e la corretta scheda di somministrazione.

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CAVAGNINI, MIRIAM. « STUDIO IN VIVO DEL RUOLO DEGLI iMSN NELLA CODIFICA DELL’ATTIVITÀ MOTORIA IN MODELLI MURINI MEDIANTE CALCIUM IMAGING ». Doctoral thesis, Università degli studi di Modena e Reggio Emilia, 2022. http://hdl.handle.net/11380/1273445.

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Lo striato, il principale nucleo dei gangli della base, è costituito per il 95% da Medium Spiny Neurons (MSN) GABAergici. Gli MSN si suddividono in neuroni della via diretta e della via indiretta. Gli MSN della via diretta (dMSN), esprimono i recettori per la dopamina D1 (D1R), mentre gli MSNs della via indiretta (iMSN) esprimono i recettori dopaminergici D2 (D2R) e per l’adenosina A2A (A2AR) [Gerfen 1990; Gerfen 1992; Gong 2003]. Secondo l’interpretazione classica, queste due vie esercitano un effetto opposto nella regolazione dell’attività motoria: la via diretta promuove l’attivazione della corteccia coinvolta nella codifica dei piani motori, e di conseguenza promuove l’avvio del movimento, mentre la via indiretta inibisce l’attività corticale e il movimento [Albin 1989; Alexander and Crutcher, 1990; DeLong 1990]. Studi più recenti hanno modificato questa interpretazione e dimostrato che entrambe le vie risultano essere attive durante l’avvio del movimento, sebbene con alcune differenze [Bonnavion 2019; Tecuapetla 2016]. Ad oggi, mentre il ruolo di queste due vie è globalmente compreso, gli specifici meccanismi cellulari e la loro interazione in relazione al controllo motorio rimangono parzialmente sconosciuti. Tecnologie moderne, fra cui il calcium (Ca2+) imaging, possono essere applicate per meglio comprendere il coinvolgimento di queste due vie. Quando vi è un’intensa attività cellulare, il Ca2+ entra nelle diramazioni dendritiche e nei corpi cellulari, incrementando la concentrazione intracellulare. Queste fluttuazioni del Ca2+ possono essere monitorate mediante indicatori del Ca2+, come il GCaMP. Lo scopo di questo lavoro di tesi è stato quello di investigare il ruolo dello striato nella locomozione murina, in particolare il coinvolgimento degli iMSN mediante tecnica di Ca2+ imaging in vivo. L’attività motoria è stata studiata mediante test comportamentali, mentre l’attività GABAergica degli iMSN è stata investigata dopo somministrazione di anfetamina, una sostanza psicostimolante, a diverse dosi. In questo studio, il circuito della via indiretta è stato visualizzato mediante calcium imaging in vivo attraverso un microscopio associato ad un endoscopio impiantato cronicamente. Come modelli murini sono stati utilizzati topi A2AGCaMP6S esprimenti GCaMP negli iMSN. L’analisi comportamentale ha rivelato che l’iniezione acuta intraperitoneale di anfetamina alla dose di 3 mg/kg incrementa significativamente la locomozione, con un picco a 15 min dopo l’iniezione, mentre la dose di 1 mg/kg riduce lievemente l’attività motoria, probabilmente a seguito dell’induzione di stereotipie. L’analisi dei transienti del calcio negli iMSN ha evidenziato una decrescita marcata del numero di iMSN attivi e un incremento della durata degli spike a seguito della somministrazione di anfetamina a 3 mg/Kg. La stessa tendenza è stata mantenuta a seguito dell’iniezione ad 1 mg/kg. Inoltre, comparando i dati ricavati dal Ca2+imaging e i dati comportamentali, è emerso come gli iMSN codifichino il movimento, confermando precedenti studi [Parker 2018]. L’analisi dimostra che l’attività degli iMSN aumenta circa 0,5-0,7 secondi prima dell’avvio del movimento e decresce circa 1-0,5 secondi prima dell’arresto del movimento. Tuttavia, non è ancora chiaro se il movimento o l’assenza di movimento siano codificati da cambiamenti di attività distribuiti casualmente nella popolazione di iMSN o se determinate sottopopolazioni di iMSN modifichino con regolarità la loro attività specificatamente durante uno dei due stati di attività motoria. Questo studio ha portato nuovi elementi interpretativi relativi alla complessità dell’attività degli iMSN, portando supporto all’ipotesi che sostiene la necessità di un’attivazione della via indiretta all’avvio del movimento.
Striatum is the main input nucleus of the basal ganglia and 95% of its neurons are GABAergic Medium Spiny Neurons (MSNs). MSNs are subdivided into neurons of the direct and indirect pathway. The direct pathway consists of MSNs (direct MSNs, dMSNs) mainly expressing dopamine D1 receptors (D1R), whereas MSNs of the indirect pathway (iMSNs) express dopamine D2 receptor (D2R) and adenosine A2A receptor (A2AR) [Gerfen et al., 1990; Gerfen et al., 1992; Gong et al. 2003]. According to the classical model, these two pathways exert an opposite effect on movement regulation: the direct pathway promotes activity of the cortex that codes a motor plan, and therefore movement initiation, while the indirect pathway inhibits cortical activity and movement [Albin et al., 1989; Alexander and Crutcher, 1990; DeLong 1990]. Recent studies have challenged this interpretation and demonstrated that both pathways are co-activated during movement initiation but with differential activities [Bonnavion et al., 2019; Tecuapetla et al., 2016]. Yet, while the role of these two pathways is globally understood, cell-specific mechanisms and their interaction in relation to movement control are only partially known. Modern technological advances such as calcium (Ca2+) imaging techniques can be applied to directly observe the neuronal activity in these two pathways. During periods of heightened neural activity, Ca2+ flows into the dendritic branches and cell bodies of neurons, increasing its intracellular concentrations. These activity-dependent fluctuations in intracellular Ca2+ can be monitored by expressing a Ca2+ indicator, such as GCaMP, into the neuronal population of interest. The aim of this study was to investigate the role of murine striatum in locomotion, and in particular the involvement of iMSNs, using recent in vivo Ca2+ imaging technologies. Locomotor activity was tested by behavioural experiments and the GABAergic iMSN activity was stimulated by a psychostimulant drug, d-Amphetamine, at different doses. In this study, in vivo calcium imaging was used to visualize striatal neural circuit dynamics of the indirect pathway during mouse behaviour with head-mounted microscopes and chronically implanted endoscopes. A2AGCaMPs mice expressing GCaMP in iMSNs were used as an animal model. Behavioural analysis showed that acute intraperitoneal injection of d-Amphetamine at 3 mg/kg dose markedly increases locomotor activity with a peak 15 minutes after injection, while a dose of d-Amphetamine at 1 mg/kg modestly decreases locomotor activity, possibly due to the induction of stereotyped behaviour. Traces of iMSN calcium activity were extracted from the imaging data and analysed with custom developed software. The results showed a significant decrease in the average number of active iMSNs and a significant increase in the average spike duration after d-Amphetamine 3 mg/kg injection. At 1mg/kg dose, there was a trend for a significant decrease in the average number of active cells and a significant increase in the average spike duration. Analysis of the acquired datasets showed how iMSNs encode movement, confirming previous results reported by Parker and colleagues [Parker et al, 2018]. In detail, the analysis revealed that iMSN activity rises around 0.5-0.7 seconds before motion onset and falls around 1-0.5 seconds before motion offset, suggesting that increased firing of iMSNs encodes locomotion. However, it is not clear whether movement or rest are encoded by changes in the activity of randomly distributed iMSNs or specific sub-sets of iMSNs change their activity during rest or movement. This study allows us to better understand the complexity of MSN activity, challenging the classical view where the direct and indirect pathway show an opposite pattern of activity during movement.

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DA, SACCO LETIZIA. « Analisi dei profili di espressione di microRNA applicata a modelli sperimentali in vitro e in vivo ». Doctoral thesis, Università degli Studi di Roma "Tor Vergata", 2010. http://hdl.handle.net/2108/1381.

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Nell’ultimo decennio la scoperta dei microRNA ha messo in luce un nuovo e fine meccanismo di regolazione post-trascrizionale, che interviene in molti processi quali lo sviluppo, il differenziamento la proliferazione e la morte cellulare. Inoltre, numerose evidenze hanno dimostrato il coinvolgimento dei microRNA in diverse patologie. Nella maggior parte degli studi effettuati, e' stata utilizzata la tecnologia microarray per identificare i microRNA coinvolti nei meccanismi patogenetici, ma anche per ottenere dei profili di espressione caratteristici di patologia con valore diagnostico o prognostico. Questi studi suggeriscono che l'analisi dei profili di espressione dei microRNA può essere considerata uno strumento utile per comprendere quale ruolo essi svolgono nella regolazione dei processi fisiopatologici. In questo lavoro abbiamo studiato il profilo di espressione di microRNA, mediante tecnologia microarray in due diversi modelli sperimentali: 1) un modello in vitro, utile per la comprensione dei meccanismi molecolari alla base della risposta immunitaria; 2) un modello in vivo, idoneo per lo studio della patogenesi di una patologia epatica molto diffusa nota come NAFLD ("Non-alcoholic fatty liver disease"). E' di recente scoperta la relazione tra infiammazione, immunità innata e microRNA, che sono descritti essere coinvolti nella regolazione della risposta cellulare all’infezione microbica. Noi abbiamo, così, analizzato lo specifico profilo di espressione di microRNA in cellule dendritiche umane, utilizzando un modello in vitro di stimolazione e attivazione mediante agonisti di differenti recettori Toll-like (TLRs): R848/Resiquimod, ligando del TLR7/8; LPS, ligando del TLR4; e poly(I:C), ligando del TLR3. Questa analisi ha permesso di identificare gruppi di microRNA espressi specificamente in risposta a determinati stimoli e puo' risultare utile per chiarire il possibile ruolo dei microRNA nei meccanismi attraverso i quali le cellule dendritiche discriminano i diversi patogeni. La steatosi epatica di origine non-alcolica, o NAFLD, è una patologia emergente caratterizzata da un ampio spettro di condizioni epatiche: dalla semplice steatosi, alla steatoepatite con fibrosi più o meno avanzata (NASH, non-alcoholic steatohepatitis). La patologia nelle sue forme più gravi può evolvere fino alla cirrosi e all’epatocarcinoma. La NAFLD ha una complessa eziopatogenesi ancora poco chiara. Per individuare i possibili meccanismi molecolari coinvolti nello sviluppo della NAFLD abbiamo utilizzato un modello animale, capace di riprodurre i vari aspetti della patologia umana. In prticolare, in questo studio abbiamo effettuato l’analisi dei profili di espressione dei microRNA nel tessuto epatico di ratti sottoposti a diversi regimi dietetici. I nostri risultati hanno dimostrato che il trattamento con i diversi regimi ipercalorici causa un aumento significativo del peso corporeo e del fegato, e di alcuni parametri metabolici rispetto agli animali controllo, come anche differenti danni epatici. L’analisi dei microRNA ha dimostrato la significativa deregolazione di 3 microRNA down-regolati (miR-122, miR-451 e miR-27a) e 3 up-regolati (miR-200a, miR-200b e miR-429) negli animali sottoposti alle diete ipercaloriche rispetto alla dieta standard. Fra i potenziali bersagli di tali microRNA emergono alcune molecole coinvolte nel controllo dell’apoptosi e dell'infiammazione, ma soprattutto proteine del segnale intracellulare, e del metabolismo lipidico e glucidico.
Over the last decade, the discovery of microRNAs revealed a new mechanism of post-transcriptional regulation. MicroRNAs are involved in many biological processes such as development, differentiation, proliferation and cell death. Moreover, several evidences showed the pathogenic role of microRNAs in various diseases. A lot of studies used microarray technology to identify miRNAs involved in the pathogenesis, but also to obtain the expression pattern characteristic of pathology with diagnostic or prognostic assessment. These studies suggest that profiling of microRNAs may be used to understand the role they play in regulating pathophysiological processes. In this work we employed microarray technology to investigate the expression profile of microRNAs in two different experimental models: 1) an in vitro model, useful for understanding the molecular mechanisms underlying the immune response, 2) a in vivo model, suitable for studying the pathogenesis of non-alcoholic fatty liver disease, also known as NAFLD. Recently, has been explored the relationship between inflammation, innate immunity and microRNAs, which are described to be involved in regulating cellular response to microbial infection. Thus, we identified the specific expression profile of microRNAs in human dendritic cells, using an in vitro model of stimulation and activation by agonists of different Toll-like Receptors (TLRs): R848/Resiquimod, ligand of TLR7/8; LPS, ligand of TLR4; and poly(I: C), ligand of TLR3. Analysis of expression profiles identified groups of miRNAs expressed specifically in response to treatments with LPS, R848, or their combination with respect to control dendritic cells. This analysis will help to clarify their possible role in mechanisms of dendritic cells to discriminate pathogens. The non-alcoholic fatty liver disease or NAFLD is an emerging disease characterized by a wide spectrum of liver conditions from simple steatosis, steatohepatitis with or without fibrosis (NASH, non-alcoholic steatohepatitis). The etiopathogenesis of NAFLD is complex and still unclear. To identify possible molecular mechanisms involved in the development of NAFLD, we used an animal model, able to reproduce various aspects of human pathology. In this study we performed the analysis of microRNAs expression profiles in liver tissue of rats subjected to different diets. Our results showed that treatment with different ipercaloric regimens caused a significant increase in body weight and liver, and some metabolic parameters, compared to control animals, as well as different liver damage. The analysis of microRNAs showed the significant downregulation of three microRNAs (miR-122, miR-451 and miR-27a) and the up-regulation of other three microRNAs (miR-200A, miR-429 and miR-200B) in animals treated with ipercaloric diets respect to those with a standard diet. Among the potential targets of these microRNAs we found some molecules involved in the regulation of apoptosis and inflammation, but also intracellular signaling proteins, and lipid and glucose metabolism.

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METO, AIDA. « Approcci innovativi per studi sui patogeni del cavo orale : modelli di studio in vitro ed ex vivo ». Doctoral thesis, Università degli studi di Modena e Reggio Emilia, 2021. http://hdl.handle.net/11380/1246163.

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Negli ultimi anni, sono stati proposti nuovi composti/strumenti per mantenere la salute orale e/o per trattare diversi problemi dentali/parodontali. Come è noto, la carie dentale si pone tra le infezioni più diffuse ed una sua gestione impropria comporta lo sviluppo di malattie rilevanti ed eventualmente all'estrazione dell’elemento dentale. Una vasta letteratura documenta il ruolo patogenetico di diversi microorganismi che sono in grado di persistere nel cavo orale, in quanto capaci di organizzandosi come comunità microbica eterogenea (comprendente batteri, virus e funghi), adesa alle diverse superfici, strettamente racchiusa in una matrice polimerica di origine polisaccaridica. Tale comunità sessile, che se adesa ai denti è detta placca dentale, è notoriamente refrattaria non solo alle comuni procedure di pulizia con collutori e dentifrici/spazzolini, ma anche ai farmaci antimicrobici e alle difese immunitarie dell'ospite. Questo scenario si complica ulteriormente considerando che l’ampio uso di attacchi fissi o rimovibili nei trattamenti ortodontici espande la problematica e la conseguente sfida clinica, essendo tali dispositivi un ulteriore habitat utile per l'adesione microbica, la crescita e la formazione di biofilm. In misura simile, i pazienti con impianti dentali possono sviluppare localmente malattie legate alla produzione di biofilm impianto-associato, consentendo la progressione clinica verso quadri di perimucosite o perimplantite infettiva. Da qui, sorge la necessità di strumenti/composti innovativi per facilitare la rimozione di microrganismi potenzialmente patogeni e il mantenimento dell'omeostasi del cavo orale. Oltre ai patogeni orali più noti, tra cui il gruppo Streptococcus mutans e il "complesso rosso" dei bacilli anaerobi Gram-negativi, anche Candida albicans (C. albicans), Staphylococcus aureus (S. aureus) e Pseudomonas aeruginosa (P. aeruginosa) possono essere agenti eziologici di malattie orali. Il primo germe, spesso ospitato come commensale delle mucose sane, è il principale patogeno fungino coinvolto nella mucosite orale. Gli altri due sono patogeni molto subdoli, responsabili di malattie ad ampio spettro; considerati i loro numerosi fattori di virulenza e l’ampia farmaco-resistenza, S. aureus e P. aeruginosa sono ampiamente utilizzati per studi in vitro come preziosi prototipi di patogeni Gram-positivi e Gram-negativi. Lo scopo della presente tesi era di valutare in vitro ed ex vivo l'efficacia antimicrobica e antibiofilm di approcci innovativi contro i patogeni orali. Questa tesi ha fornito prove in vitro ed ex vivo sull'efficacia antimicrobica di composti nuovi e tradizionali per la cura e l’igiene del cavo orale, da cui possono derivare in prospettiva scelte più razionali e consapevoli. Ad esempio, un nuovo utilizzo del prodotto endodontico Cupral potrebbe essere proposto nelle pratiche di igiene quotidiana, così come potrebbe essere privilegiato il trattamento delle peri-implantiti con il sistema Bic-40, vista la sua particolare efficacia nella pulizia e nella decontaminazione di superfici lisce e ruvide in titanio, senza influire sulla vitalità delle cellule staminali dell’ospite. Inoltre, il nostro lavoro ha aggiunto nuove conoscenze sulle proprietà antimicrobiche di un composto naturale come la propoli e sui suoi possibili meccanismi d'azione, offrendo nuove opportunità nella ricerca di molecole antimicrobiche alternative. Infine, abbiamo dimostrato che il dentifricio e la gomma Biorepair Peribioma possono influenzare profondamente il comportamento dei microorganismi del cavo orale, a favore di condizioni utili al mantenimento dello stato di salute di questo distretto anatomico. Questo lavoro ha fornito nuove evidenze su come contrastare i patogeni, particolarmente se produttori di biofilm; facilitando il disegno di strategie mirate per la prevenzione e/o il trattamento delle infezioni dentali e orali associate al biofilm.
During recent years, novel compounds/tools are being proposed to maintain oral health and/or to treat dental/periodontal problems. As well known, dental caries are among the most diffused infections and their improper management turns towards relevant disease(s) and eventually tooth extraction. Extensive literature documents the pathogenic role of certain microorganisms and their ability to persist in the oral cavity, as a complex microbial community, including bacteria, viruses and fungi, tightly enclosed in a polymeric matrix of polysaccharide origin. Such sessile community, and particularly dental plaque, the first deeply studied human-associated biofilm, is notoriously refractory not only to common cleaning procedures by mouthwashes and tooth-pastes/brushes, but also to antimicrobial drugs and host immune defenses. This scenario becomes further complicated considering that the widely diffused orthodontic treatments, with fixed or removal brackets, extend the clinical challenge, being such devices an additional good habitat for microbial adhesion, growth and biofilm formation. To a similar extent, patients with dental implants may locally develop biofilm-related diseases, allowing clinical progression toward pathogen-related peri-mucositis or peri-implantitis. From here, the need arises for innovative tools/compounds to facilitate microbial removal and maintenance of oral cavity homeostasis. Besides the most investigated oral pathogens, including Streptococcus mutans-group and the “red complex” Gram-negative anaerobe bacilli, also Candida albicans (C. albicans), Staphylococcus aureus (S. aureus) and Pseudomonas aeruginosa (P. aeruginosa) may occur as causative agent of oral diseases. The first, often harbored as commensal of healthy mucosae, is the main fungal pathogen involved in oral mucositis. The latter two are subtle pathogens, responsible of wide-spectrum diseases; they are being extensively used for in vitro studies, because of their numerous virulence factors and wide-spectrum antimicrobial resistance. The aim of the present thesis was to evaluate in vitro and ex vivo, the antimicrobial and antibiofilm efficacy of innovative approaches against oral pathogens. Our data provided in vitro and ex vivo evidence on the antimicrobial efficacy of several dental-care compounds. A novel use of the endodontic product Cupral could be proposed in daily hygiene practices. The Bic-40 treatment was shown as the best approach in cleaning smooth and rough titanium surfaces (without altering their properties); importantly, its device-decontamination efficacy did not affect the biological properties of reparative stem cells. Furthermore, our work added new insights on the anti-microbial properties of a natural compound, such as propolis, and on its possible mechanisms of action. At last, we showed that the Biorepair Peribioma toothpaste and gum deeply affected oral microorganisms’ behavior, drastically impairing their ability to contaminate and produce plaque onto orthodontic devices; interestingly, replacement by beneficial microorganisms was observed. The overall take-home message from this research is that basic science may greatly increase our knowledge on how to counteract biofilm-producing pathogens; in turn, this will facilitate prevention and/or treatment of dental and oral biofilm-associated infections, making a huge difference in terms of health promotion.

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CUTULI, Marco Alfio. « Modelli preclinici non convenzionali : vantaggi, limiti e applicabilità della proof of concept agli aspetti regolatori ». Doctoral thesis, Università degli studi del Molise, 2021. http://hdl.handle.net/11695/100701.

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Le nuove regole globali e una maggiore coscienza etica implicano controlli sempre più rigorosi sull’impiego dei vertebrati negli studi in vivo. Di conseguenza, negli ultimi anni, per ovviare ai rilevanti costi e ai tempi necessari ad ottenere le autorizzazioni per condurre studi sui vertebrati, sono stati proposti modelli alternativi che non utilizzano o riducono l’utilizzo dei vertebrati. Nel presente lavoro sono stati proposti e messi a puto tre modelli preclinici non convenzionali per condurre studi sull’attività e sulla tossicità in vivo di sostanze chimiche e biologiche. Risultati promettenti sono stati ottenuti dal lepidottero Galleria mellonella che si è dimostrato un modello preclinico affidabile sia per lo screening di sostanze antimicrobiche naturali, probiotici che per lo studio del rapporto ospite-parassita. Anche per il coleottero Tenebrio molitor sono stati ottenuti dati incoraggianti su un suo utilizzo come potenziale modello di infiammazione intestinale. Infine è stato messo a punto un modello di mucosa impiegando la specie di lumaca Limacus flavus per determinare il potenziale irritante di sostanze chimiche. Questi modelli, nonostante alcune limitazioni, si sono dimostrati strumenti efficaci per condurre screening rapidi e riproducibili che non richiedono strumenti sofisticati ma che sono in linea con le normative vigenti sulla tutela della sperimentazione animale
New global regulations and greater ethical conscience lead to stronger controls on the use of vertebrates in pre-clinical studies. To overcome the significant costs and time required to obtain authorisation for mammalian studies, alternative animal models have been proposed in recent years to reduce the utilisation of vertebrates. Three non-conventional pre-clinical models were proposed and tested in the present work to evaluate in vivo activity and toxicity of chemical and biological substances. Promising results were obtained from the Lepidoptera Galleria mellonella, which proved to be a reliable pre-clinical model for screening natural antimicrobial substances, probiotics, and the host-parasite relationship. Encouraging data were also observed using the Coleoptera Tenebrio molitor as a potential model of intestinal inflammation. In addition, the snail species Limacus flavus was developed as an in vivo mucosal model to assess the irritant potential of chemicals. Although the proposed models have some limitations, they have proven to be effective for rapid and reproducible pre-clinical screening. These assays do not require sophisticated instruments and are aligned with current legislation on the protection of animals used for scientific purposes.

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PANNELLA, Micaela. « Valutazione degli effetti del Dexametasone e del Cisplatino nei modelli sperimentali in vitro (OC-k3)e in vivo(Ratti Wistar) ». Doctoral thesis, Università degli studi di Ferrara, 2011. http://hdl.handle.net/11392/2388742.

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In this thesis I want to evaluate the Dexamethasone and Cisplatin effects in vitro (OC-k3) and in vivo (Rats Wistar) model. On more I want to get the protocol for future gene therapy, using the new non-viral transposable element Sleeping Beauty in vitro model (Human Fetal Auditory Stem Cells (hFASC). The Cisplatin is an antineoplastic agent used to contrast different types of malignant tumors (testicolaris, ovaric, bladder, neck and head). This drug is very toxic and it generates many side effects like ototoxicity. This agent destroys the inner ear tissues, but they are not able to regenerate for this manner it is very important to study the otoprotective drugs mechanisms. There are many agents otoprotective and I want to remember the Dexamethasone. Dexamethasone is a glucocorticoid and usually it is used to contrast the inflammatory processes. The aim of this thesis is evaluating the drugs effects and protective effect of Dexamethasone against the Cisplatin toxicity. I used the cell line OC-k3 for in vitro model, because these cells derive from immortal mouse inner ear. The OC-k3 don’t show the neurological markers, but epidermal markers. For evaluating the drugs toxicity I used the flow citometry and I noted that the Cisplatin toxicity is time dependent. Dexamethasone is not toxic and is able to protect the cells from Cisplatin action. Another method for toxicity and protection evaluation is studying the cytoskeleton morphology. I noted that after 24h Cisplatin and Dexamethasone are not toxic for OC-k3, but after 48h Cisplatin destroys the cytoskeleton, while Dexamethasone is not toxic and protects the cells from Cisplatin action. Finally I studied the apoptosis marker after Cisplatin and Dexamethasone treatment. Cisplatin generates apoptosi intrinsecal via, but Dexamethasone is not toxic and it protects the cells, because they don’t show the apoptosis marker. I can conclude that Cisplatin is toxic for OC-k3 and Dexamethasone is not toxic and it is able to protect the cells from Cisplatin action. In vivo model I used the Wistar rats and I injected the drugs introperitoneal via alone and in co-treatmennt. To evaluate the drugs effects I used the microscope (SEM) and Immunoistochemistry. I noted that Cisplatin destroys the inner ear tissues and Dexamethasone is not toxic and protects the tissues from Cisplatin action. In vitro and in vivo model, Cisplatinand Dexamethasone generate the same effects; Cisplatin is toxic as for the cells as for the tissues. Dexamethasone is not toxic and it is protective as for cells as for tissues. For gene therapy I used hFASC and I trasfected them with new non viral transposable element: Sleeping Beauty. Two plasmids derive from this transposable element (pT2/veus: gene for GFP, SB100: gene for transposase). I used different concentrations of these plasmids for evaluating their toxicity, I noted that high concentrations of plasmids are toxic for the cells and the green cells number decreases during the incubation time. I decided to decrease the plasmids concentrations and I noted that the green cells number increases. I can conclude that high concentrations of plasmids are toxic for the cells, but I can transfect the cells without problem if I use low concentrations. After I evaluate if the cells are able to express the typical marker of undifferentiated cells (SOX 2, PAX 2, OCT4). With immunoistochemistry I noted the cells show these typical markers. Finally I decided to differentiate the transfected cells to bipolar neurons and cells like hair cells. I got that no transfected cells are able to differentiate to neuronal cells and cells like hair cells only. It is not significant result because I had one only single cell transfected in that cells pool.

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Marchi, Enrica <1977&gt. « Pralatrexate sinergizza in vitro ed in vivo con l'inibitore del Proteosoma Bortezomib in modelli pre-clinici di Linfoma Non Hodkin T ». Doctoral thesis, Alma Mater Studiorum - Università di Bologna, 2010. http://amsdottorato.unibo.it/2430/1/Tesi.pdf.

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Marchi, Enrica <1977&gt. « Pralatrexate sinergizza in vitro ed in vivo con l'inibitore del Proteosoma Bortezomib in modelli pre-clinici di Linfoma Non Hodkin T ». Doctoral thesis, Alma Mater Studiorum - Università di Bologna, 2010. http://amsdottorato.unibo.it/2430/.

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BAZZINI, CHIARA. « STUDY OF MOLECULAR MECHANISMS AND NEW STRATEGIES AGAINST A CYTOTOXICITY AND NEUROINFLAMMATION IN EX VIVO CELLULAR MODELS FROM ALZHEIMER’S DISEASE PATIENTS ». Doctoral thesis, Università degli Studi di Milano-Bicocca, 2021. http://hdl.handle.net/10281/306480.

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La malattia di Alzheimer (AD) rappresenta una delle principali problematiche per la salute pubblica ed è stata identificata come una priorità per la ricerca. Le due caratteristiche patologiche fondamentali della malattia sono le placche amiloidi e i grovigli neuro fibrillari che sono alla base della neuroinfiammazione e del deterioramento cognitivo.Le forme solubili degli oligomeri sono la specie più tossica della β-amiloide (Aβ) e interagiscono con diverse chinasi proteiche coinvolte nella trasduzione del segnale intracellulare come Ras/MAPK e PI3K/AKT che regolano molti processi cellulari e funzioni cognitive, e alcuni meccanismi molecolari coinvolti nella degenerazione neuronale, come l'iperfosforilazione di tau e l'eccitotossicità del glutammato. Negli ultimi anni molta attenzione è stata focalizzata sull'utilizzo di composti naturali come agenti neuroprotettivi. Il luppolo (Humulus Lupulus) contiene flavonoidi, molecole aromatiche che hanno proprietà antiossidanti e antinfiammatorie. È stato dimostrato che l'estratto di luppolo ha effetti antiaggreganti sull’Aβ e sembra impedire la sua produzione nelle cellule in coltura. L'accumulo di Aβ induce anche l'attivazione della proteina 3 del recettore Nod-like receptor 3 (NLRP3) dell’inflammosoma e il conseguente rilascio di citochine proinfiammatorie, il quale svolge un ruolo fondamentale nella neuroinfiammazione associata all'AD. NLRP3 attivato induce la produzione e il rilascio di mediatori infiammatori, tra cui i complessi proteici ASC (ASC specks), IL-1β e IL-18, che facilitano la deposizione di Aβ in un ciclo che si auto alimenta. Impedire l’assemblaggio e l'attivazione del complesso dell’inflammosoma potrebbe essere una possibile strategia per la terapia dell'AD. L'obiettivo generale di questo studio è quello di indagare i meccanismi molecolari coinvolti nelle malattie neurodegenerative e nella neuroinfiammazione utilizzando modelli cellulari periferici ex vivo di AD.Al fine di caratterizzare le interazioni Aβ e vie di trasduzione del segnale MAPK e AKT, abbiamo utilizzato fibroblasti di pazienti AD sporadici con diversa gravità della malattia. Per valutare i meccanismi molecolari che potrebbero prevenire o modulare la tossicità indotta da Aβ, sono stati studiati anche i potenziali effetti citoprotettivi dell'estratto di luppolo e il relativo signaling intracellulare. Inoltre, è stato dato particolare interesse alla via di attivazione del NLRP3-infiammasoma. Abbiamo studiato il coinvolgimento dell'attivazione di NLRP3 sulle vie MAPK e AKT e sui loro bersagli a valle, utilizzando una combinazione di studi in vitro e di campioni ottenuti dai pazienti. In particolare, abbiamo utilizzato monociti umani THP-1 di derivazione macrofagica e monociti derivati da cellule mononucleate del sangue periferico (PBMC) di soggetti sani (HC) e pazienti affetti da AD, per analizzare la modulazione autofagica e gli effetti della Stavudina (D4T), un inibitore nucleosidico della trascrittasi inversa, che riduce l'attivazione dell'inflammosoma bloccando il recettore purinergico P2X7R. Inoltre, abbiamo analizzato il pathway di attivazione dell'inflammosoma NLRP3 e il ruolo di CRID3 un inibitore selettivo, per confrontare gli effetti dell’inibizione dell’inflammosoma attraverso due pathway differenti. I monociti derivati da HC e AD sono stati differenziati in cellule microglia-like (MDMIs) e caratterizzati per l'espressione di proteine intracellulari e di superficie tipiche delle cellule mieloidi. Funzioni tipiche della microglia come il rilascio di citochine infiammatorie, la fagocitosi e la degradazione sono state valutate anche in seguito all'esposizione di attivatori dell'inflammosoma con o senza CRID3. MDMIs riflettono molte caratteristiche della microglia e sono un modello cellulare utile per comprendere la patogenesi dell'AD, identificare i target terapeutici e consentire lo screening farmacologico su larga scala dei nuovi composti per uso terapeutico.
Alzheimer's disease (AD) is a major public health concern and has been identified as a priority for research in Life Science. The two core pathological hallmarks of AD are extracellular amyloid plaques and intracellular neurofibrillary tangles which underlie microglial and neuronal damage, neuroinflammation and cognitive impairment. Soluble oligomers are the most toxic species of β-amyloid (Aβ) and interact with several protein kinases such as Ras/MAPK and PI3K/AKT pathways, which regulate many cellular processes and cognitive functions. These pathways mediate Aβ toxicity, regulating some molecular mechanisms involved in neuronal degeneration such as cytoskeletal impairment, glutamate excitotoxicity and neuroinflammation. In the last years much attention has been focused on the potential role of natural compounds as neuroprotective agents. Hop (Humulus Lupulus) contains flavonoids, aromatic molecules which have antioxidant, anti-inflammatory and anti-atherogenic properties. In fact, hop extract has anti-aggregating effects on Aβ, and it seems to prevent its production in cultured cells. Aβ induces also the activation of the pattern recognition receptor Nod-like receptor protein 3 (NLRP3) inflammasome complex in microglia and the consequent release of proinflammatory cytokines, playing a pivotal role in AD-associated neuroinflammation. NLRP3 activation results in the release of inflammatory mediators, including ASC protein complexes (ASC specks), IL-1β and IL-18, that facilitate Aβ deposition and neuroinflammation in a self-feeding pathogenic loop. Since specific therapeutical strategies are still lacking, the dampening of the inflammasome assembly and activation could be a new strategy for AD. The overall focus of this study is to investigate molecular mechanisms involved in neurodegenerative diseases and in neuroinflammation, using peripheral ex vivo cellular models from AD, to check new potential therapeutical targets. In order to characterize the complex interactions among Aβ, MAPK and AKT signaling, we used fibroblasts from sporadic AD patients with different disease severity. To evaluate any molecular mechanisms that could prevent or modulate Aβ-induced toxicity, the potential cytoprotective effects of Hop extract and related intracellular signaling were also investigated. Fibroblasts provide a useful cellular model for studying AD, since they could be differentiated into patient-specific neural cell lines, using iPSC technologies. Moreover, particular interest was given to NLRP3-inflammasome activation pathway. We investigated the involvement of NLRP3 inflammasome activation on intracellular pathways and their downstream targets, using a combination of in vitro studies and patient-derived samples. In particular, we used macrophage-derived THP-1 human monocytes and peripheral blood mononuclear cells (PBMC)-derived monocytes from healthy control (HC) subjects and AD patients, to analyse phagocytosis, autophagy and apoptosis modulation and the effects of the nucleoside reverse transcriptase inhibitor Stavudine (D4T), that reduces NLRP3 inflammasome activation blocking the purinergic receptor P2X7R. Furthermore, we analyzed the NLRP3 inflammasome pathway and the role of the selective NLRP3 inhibitor CRID3, to compare the effects of inflammasome inhibition through two different mechanisms. At this purpose, HC and AD-derived monocytes were differentiated into microglia-like cells (MDMIs) and characterized for myeloid surface and intracellular proteins expression. Key microglia functions such as inflammatory cytokines release, Aβ phagocytosis and degradation were evaluated upon exposure to NLRP3 inflammasome activators with or without CRID3. MDMIs reflected many features of microglia and, as fibroblasts-derived iPSCs, they are attractive cellular models helpful to understand AD pathogenesis, identify therapeutic targets and allow large-scale drug screening of the novel therapeutic candidates.

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BASTIOLI, GUENDALINA. « Studio dei meccanismi neurotossici coinvolti nella neurodegenerazione indotta da mutazione Lrrk2 o da α-sinucleina in modelli ex-vivo o in vitro di malattia di Parkison ». Doctoral thesis, Università Politecnica delle Marche, 2018. http://hdl.handle.net/11566/253148.

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Studio dei meccanismi neurotossici coinvolti nella neurodegenerazione indotta da mutazione Lrrk2 o da α-sinucleina in modelli ex-vivo o in vitro di malattia di Parkison La malattia di Parkinson (PD) è una malattia neurodegenerativa multifattoriale caratterizzata dalla degenerazione dei neuroni dopaminergici della substantia nigra. Tra le anomalie genetiche identificate nella malattia di Parkinson (PD), le mutazioni del gene della ripetizione della leucina kinase2 (Lrrk2), come la mutazione missenso G2019S legata all'aumento dell'attività della chinasi, sono le più comuni. Mentre il complesso ruolo di Lrrk2 non è stato completamente chiarito, sono state riportate evidenze che l'attività della chinasi mutata influenza la trasmissione sinaptica. L'iperattivazione del dominio della chinasi di Lrrk2 potrebbe rappresentare un fattore predisponente sia per il rilascio glutammatergico striatale potenziato sia per la vulnerabilità mitocondriale ai fattori ambientali osservati nel PD. Per indagare su possibili alterazioni della suscettibilità striatale alla disfunzione mitocondriale, abbiamo eseguito registrazioni elettrofisiologiche dal nucleo striato di un modello di PD Lrrk2 G2019S, e inoltre abbiamo indagato su possibili alterazioni precoci della neurotrasmissione prodotta dalla mutazione Lrrk2 di G2019S nel PD. Un altro fattore genetico è la presenza di inclusioni intracellulari denominate corpi di Lewy costituiti da aggregati α-Synuclein (α-Syn). Diversi studi hanno dimostrato che l'accumulo di α-Syn nei neuroni dopaminergici umani riduce l'attività del complesso I mitocondriale, aumenta la produzione di specie reattive dell'ossigeno e provoca cambiamenti dei livelli di Ca2+. Lo scambiatore Na+/Ca2+ (NCX) è un importante regolatore delle concentrazioni di Ca2+ citoplasmatiche e mitocondriali. Abbiamo quindi studiato il possibile ruolo svolto da NCX nella tossicità mitocondriale in un modello in vitro del PD precoce. Abbiamo trovato che in G2019S-Lrrk2 (KI), mentre la trasmissione glutamatergica spontanea basale, facilitazione sinaptica e rapporti NMDA / AMPA erano invariati, la stimolazione del recettore DA D2 da parte del quinpirolo ha ridotto le correnti postsinaptiche eccitatorie spontanee ed evocate (EPSC). E anche che la stimolazione del recettore D2 ha avuto un effetto neuroprotettivo sulla funzione mitocondriale. Mentre l'inibizione di mNCX esercita un effetto protettivo sul danno neuronale in un modello di PD precoce.
Studio dei meccanismi neurotossici coinvolti nella neurodegenerazione indotta da mutazione Lrrk2 o da α-sinucleina in modelli ex-vivo o in vitro di malattia di Parkison La malattia di Parkinson (PD) è una malattia neurodegenerativa multifattoriale caratterizzata dalla degenerazione dei neuroni dopaminergici della substantia nigra. Tra le anomalie genetiche identificate nella malattia di Parkinson (PD), le mutazioni del gene della ripetizione della leucina kinase2 (Lrrk2), come la mutazione missenso G2019S legata all'aumento dell'attività della chinasi, sono le più comuni. Mentre il complesso ruolo di Lrrk2 non è stato completamente chiarito, sono state riportate evidenze che l'attività della chinasi mutata influenza la trasmissione sinaptica. L'iperattivazione del dominio della chinasi di Lrrk2 potrebbe rappresentare un fattore predisponente sia per il rilascio glutammatergico striatale potenziato sia per la vulnerabilità mitocondriale ai fattori ambientali osservati nel PD. Per indagare su possibili alterazioni della suscettibilità striatale alla disfunzione mitocondriale, abbiamo eseguito registrazioni elettrofisiologiche dal nucleo striato di un modello di PD Lrrk2 G2019S, e inoltre abbiamo indagato su possibili alterazioni precoci della neurotrasmissione prodotta dalla mutazione Lrrk2 di G2019S nel PD. Un altro fattore genetico è la presenza di inclusioni intracellulari denominate corpi di Lewy costituiti da aggregati α-Synuclein (α-Syn). Diversi studi hanno dimostrato che l'accumulo di α-Syn nei neuroni dopaminergici umani riduce l'attività del complesso I mitocondriale, aumenta la produzione di specie reattive dell'ossigeno e provoca cambiamenti dei livelli di Ca2+. Lo scambiatore Na+/Ca2+ (NCX) è un importante regolatore delle concentrazioni di Ca2+ citoplasmatiche e mitocondriali. Abbiamo quindi studiato il possibile ruolo svolto da NCX nella tossicità mitocondriale in un modello in vitro del PD precoce. Abbiamo trovato che in G2019S-Lrrk2 (KI), mentre la trasmissione glutamatergica spontanea basale, facilitazione sinaptica e rapporti NMDA / AMPA erano invariati, la stimolazione del recettore DA D2 da parte del quinpirolo ha ridotto le correnti postsinaptiche eccitatorie spontanee ed evocate (EPSC). E anche che la stimolazione del recettore D2 ha avuto un effetto neuroprotettivo sulla funzione mitocondriale. Mentre l'inibizione di mNCX esercita un effetto protettivo sul danno neuronale in un modello di PD precoce.

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Calo', R. « STUDIO DEI MECCANISMI DI DANNO DA RAGGI UVA E UVB E DEGLI EFFETTI PROTETTIVI DA PARTE DI COMPOSTI POLIFENOLICI IN SISTEMI CELLULARI E MODELLI EX VIVO DI CUTE UMANA ». Doctoral thesis, Università degli Studi di Milano, 2014. http://hdl.handle.net/2434/244829.

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Sun radiation consists of several spectrum regions, depending on the wavelengths, such as ultraviolet, visible and infrared portions. The ultraviolet rays (UV) are associated with both positive and negative effects on humans’ health. Ultraviolet light is used in the process of disinfection, it shows immunoregolatory activity and it stimulates vitamin D and serotonin production. It is well known that artificial sources of UVR are employed in the health field for sterilization procedures and for therapeutical purposes, such as treatment of dermatological pathologies like psoriasis, vitiligo and icterus neonatorum, as well as for aesthetic purposes (sunbed). In the last decade, besides these positive effects, UV side effects on skin have been demonstrated by different studies which correlated the intense and prolonged exposure to UV light with skin cancer development. Based on wavelength, UV spectrum can be divided into three segments: UVA (320-400 nm), UVB (280-320 nm) and UVC (100-280 nm). Radiation in the UVC range, considered to be the most cytotoxic and harmful, is largely absorbed by the stratospheric ozone layer, so UV regions mainly involved in photocancerogenesis development are considered to be ultraviolet radiation A (UVA) and ultraviolet radiation B (UVB). The amount and the UVA/UVB ratio that reach the Earth’s surface is influenced by a lot of environmental factors as latitude, seasons, weather/atmospheric conditions and the time of day. UVA is by far the most abundant solar UV radiation that reaches the Earth (about 90-95% of total UV radiation), penetrates the human epidermis down to the proliferative basal cells and even further to the derma and mediate biological events generating reactive oxygen species (ROS) in cells. ROS oxide other cellular constituents, particularly carbohydrates, lipids, proteins and DNA. Interactions between ROS and DNA induce formation of single strand breaks (SSBs), DNA-proteins crosslink and oxidized bases. The most abundant oxidized base is the guanine that produces the 8-oxoguanine, considered as promoter of carcinogenesis. UVB represents only 5% of total UV radiation that reaches the Earth. Besides it acts mainly at the epidermis basal layer of the skin, it is considered extremely damaging because of its high energy. UVB photons are directly absorbed by DNA and cause the formation of cyclobutane pyrimide dimers (CPDs) and pyrimidine (6-4) pyrimidone photoproducts (6-4PPs), which in turn can lead to apoptotic events or carcinogenesis development. Also tryptophan and tyrosine are modified by UVB inducing the alteration of structure and function of proteins. Moreover, UVB is able to produce reactive oxygen species (ROS) and reactive nitric species (RNS). Cells have developed defense mechanisms and endogenous repair systems in order to reduce genotoxic damage UV-induced: complexes of antioxidant enzymes which take advantage of the presence of a metal to inactivate reactive oxygen species, and DNA repair systems like the base excision repair system (BER), used for oxidized bases, and the nucleotide excision repair (NER), used to repair photolesions like CPD and 6-4 PP. Recently, several molecules have been studied for their ability to give a contribution to these antioxidant defenses. For these reasons, there is a considerable interest in the concept of the use of phytochemicals and micronutrients present in the diet such as carotenoids, vitamin E and C, and polyphenols. This last group of compounds are well known for their several biological properties, such as their antioxidant activity, their ability to enhance the activity of DNA repair systems and also their skills in inflammatory and apoptotic events modulations. Several epidemiological studies have demonstrated their biological effects and their efficiency in the prevention and treatment of several chronic diseases like cardiovascular, respiratory, neurodegenerative and inflammatory diseases and cancer. So, in view of these reasons, the aim of my study was double: first, to confirm literature data regarding the different mechanisms used by UVA and UVB rays to cause damage in various models, then to investigate whether different plant-derivatives compounds, all belonging to the pholiphenols category, were able to prevent UVA- and UVB-induced damage. I reproduced the experiments on in vitro and ex vivo models. In vitro models consist of two human keratinocytes cell lines (HaCaT and NCTC 2544 cells), generally used for routine toxicological tests and photogenotoxic evaluations. Both cell lines are non tumorigenic, but showed a different degree of differentiation, higher in HaCaT cells compared to NCTC 2544. The ex vivo model consist of a three-dimensional model of organotypic human skin cultures useful to reproduce the physiological conditions. I tested the efficiency of three compounds: a water soluble extract of Bilberry only in the in vitro models (Vaccinium Myrtillus extract, rich in polyphenols and considered to be the plant with the highest content of anthocyanins), a Thymus Vulgaris leaf extract (Thyme specie) and the synthetic Thymol (its major component), used in comparable dosages, in both models. All experimental models were pretreated in serum free medium for 1 hour with each compound and then irradiated with UVA or UVB. Control samples were treated in the same way, but not UV exposed. In order to confirm mechanisms of UV damage and to compare results obtained in the experimental models, I performed several tests, each with a specific endpoint. I evaluated the intracellular redox status, quantifying the ROS formed and the lipid peroxidation (malondialdeide levels, MDA), immediately after the end of UVA and UVB exposure and only in the in vitro models; after, I performed some genotoxic tests (the alkaline comet test to detect single and double strand breaks in single cells, the immunostaining of the histone H2AX phosphorilated to detect only double strand breaks and the micronucleous test to detect alterations to mitotic apparatus or to chromosomes); finally I investigated effects of rays on apoptotic events, using the Annexin V test and evaluating the alteration of mitochondrial membrane potential UV-induced. After, using this same assays, I tested the protective effect of Vaccinium Myrtillus extract, thymol and Thymus Vulgaris L. extract. Vaccinium Myrtillus’ effect was evaluated only in the in vitro models, the protection of thymol and Thymus Vulgaris L. extract in both models (in vitro and ex vivo). Results obtained confirmed the different machanisms of action of UVA and UVB in all models. Particularly, confirming the different mechanisms of damage used by UVA and UVB rays. My results showed the more oxidant UVA effect and the more genotoxic and apoptotic UVB effect. Three natural compounds showed their different protective effect against UVA- and UVB- induced damage, in in vitro models. Moreover, thymol and Thymus Vulgaris L. extract reduced damage UV-related both in the NCTC 2544 and HaCaT cell lines but also in the ex vivo model. Vaccinium Myrtillus extract, thymol and Thymus Vulgaris L. extract reduced, to a different extent, UVA-caused damage, on account of their antioxidant properties. Compounds revealed their ability to reduce also UVB-induced damage. The efficiency against UVB damage was lower compared to the one against UVA rays. Probably the protection was related with the ability of pholyphenols, and because of my compounds, to repair DNA damage UVB-induced increasing the expression of genes involved in NER system (specifically involved in UVB-induced damage repair). Vaccinium Myrtillus, thymol and Thymus Vulgaris L. extract showed also their protection in the apoptotic assays, avoiding the increase of extremely damaged cells which could lead to mutagenesis development. Results obtained in my thesis activity should be considered as preliminary data useful to investigate, in detail, mechanisms used by the three natural compounds to prevent genotoxic damage UVA- and UVB-induced. Moreover, it should be very interesting to investigate the bioavailability and the permeability of tested compounds in order to use them as food supplements for a diet enriched in antioxidants (systemic effect) or in sunscreens (topical effect).

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Yu, Tung Fai. « Musculo-skeletal modelling and parameterisation in vivo ». Thesis, University of Warwick, 2014. http://wrap.warwick.ac.uk/60564/.

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This thesis describes the development of an anatomically meaningful musculo-skeletal model of the human arm, incorporating two modified Hill muscle models representing the elbow flexor and extensor muscles. In vivo experimental methods to determine parameter values are presented. The stimulus for this work was to enable the prediction of movement, to support development of prostheses and orthoses such as Functional Electrical Stimulation (FES). A key problem in model based movement studies is that the passive parameter values in the Hill muscle models and the joint had not been experimentally determined in vivo. The result has been an inability for predictive models to generate realistic predictions of human movement dynamics. In the model, movement dynamics of the forearm was described using the Newton-Euler method, which was validated from analysis of physical pendulum. Structural identifiability analyses of the muscle models ensured that values for the model parameters could be uniquely determined from perfect noise free data. A novel experimental procedure termed the passive movement method is described, which exclusively parameterised the model’s passive components. Simulated model dynamics were fitted to measured movements of the freely swinging forearm under gravity. Model values were obtained on an individual subject basis. The average muscle model spring and damping constants for four healthy subjects were 143N/m and 1.73Ns/m respectively. Separately, the force/length characteristics of the muscles’ active component, the contractile element (CE), were obtained from measurements of isometric maximum voluntary contraction (MVC) at different elbow angles. The results for the five healthy subjects showed good agreement with results reported in the literature. A preliminary experiment was performed to predict elbow flexion movement under FES. An electrical stimulus that generated a specified isometric elbow flexion moment (10% of MVC) was applied to generate elbow flexion movement. Simulated FES arm movement was compared with the measured results. The simulated change in elbow angle did not agree with the measured data. A major cause for this was believed to be skin movement causing a change in the current path across the muscle fibres, thus affecting the force generated. The passive movement method described in this thesis filled an important chapter to fully parameterise musculo-skeletal models in vivo. Although in the FES movement experiment, simulated change in elbow angle generated by FES did not agree with measured data, the shape of the dynamic response in the fitted simulated movement showed good agreement with the measured FES movement.

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Saunders, John. « Ex vivo modelling of oesophago-gastric cancer ». Thesis, University of Nottingham, 2017. http://eprints.nottingham.ac.uk/47574/.

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Introduction The response to neoadjuvant chemotherapy in oesophago-gastric (OG) cancer is only 40%, so over half of the patient’s disease will progress, whilst they also suffer the toxic chemotherapy side-effects. A model to predict chemotherapy response would provide a marked clinical benefit, by enabling personalised treatment of OG cancer. Methods Live chemo-naïve tumour biopsies were obtained following informed consent at staging endoscopy, before patients underwent their routine neoadjuvant chemotherapy. Tumour cells from the endoscopic biopsies were expanded, using an in vitro feeder layer system and supplemented medium. With ethics committee approval and under Home Office guidance, these individual patient cancer cells were engrafted into immuno-compromised mice, where they formed representative tumour xenografts. Primary patient tissue, the corresponding individual patient cancer cells and their matching xenografts were analysed using immunohistochemistry, demonstrating that the in vitro and in vivo cells had retained the characteristics of the original patient’s oesophageal adenocarcinoma. To model the human tumour micro-environment (TME), a three dimensional tumour growth assay (3D-TGA) was developed, whereby the individual patient’s primary tumour cells were grown as 3D cancer cell clusters. This was performed by seeding individual patient’s primary tumour cells within a biological basement membrane extract, rich in extracellular matrix (ECM) components, with and without human mesenchymal stem cells to provide stromal support. The individual patient cancer clusters in the 3D-TGA were subjected to detailed chemotherapeutic assessment, to quantify their chemo-sensitivity to the standard chemotherapy which was administered to the patient in the clinic. This 3D-TGA predicted chemo-sensitivity was then compared with the patient’s actual clinical chemotherapy response, as measured by the histological tumour regression grade, which directly relates to prognosis. In combination with standard platinum-based chemotherapy, the 3D-TGA was assessed as a platform for evaluating new chemotherapeutics: the novel emerging HDAC inhibitor Panobinostat, and the phosphodiesterase type 5 inhibitor Vardenafil, which has recently been shown to be active against cancer stroma, were evaluated. Results Individual patient tumours were grown from primary endoscopic biopsy tissue in over half of samples obtained within a clinically applicable timescale of 2-4 weeks. Incorporating human mesenchymal cells into the 3D-TGA significantly changed the growth and drug resistance profiles (p < 0.005). This 3D-TGA chemo-response in the presence of stroma reflected the clinical chemo-sensitivity, with an accurate correlation between the 3D-TGA predicted chemo-resistance and actual clinical response for the patients evaluated. As well as predicting potential chemo-sensitivity for individual patients, the method allows individual drugs and combinations to be evaluated, trends in chemo-sensitivity between patients to be appraised, and analysis of the effect of the TME on tumour growth and chemotherapy resistance. Combination with Panobinostat enhanced response and proved efficacious in otherwise chemo-resistant tumours. Addition of PDE5i demonstrated an overall significantly enhanced chemotherapeutic response (p=0.003), and consequently provided efficacy in 60% of the otherwise chemo-resistant tumours. Discussion The novel method of growing individual patient OG cancers, using a 3D model with specific components of the tumour micro-environment in particular ECM and mesenchymal cells, provides a clinically-relevant oesophageal cancer model with application for chemo-sensitivity testing. Mesenchymal cells have a significant effect enhancing chemotherapy drug resistance in OG cancer, and this 3D model allowed identification of patients in which stromal targeting using PDE5i provided a significant reduction in chemotherapy drug resistance. In these patients, addition of PDE5i to routine chemotherapy could result in a marked change in the clinical efficacy of their chemotherapy regimen. The 3D model’s chemo-response accurately reflects individual patients’ clinical chemo-sensitivity and so this research has direct clinical application: if this assay proves to be predictive across a wider patient population, then following clinical trials, it could potentially be used to routinely guide individual patient therapy in the clinic, with administration of tailored chemotherapy for individual patient benefit.

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Field, Sarah Louise. « Modelling physiological reproductive inflammatory networks in vivo ». Thesis, University of Leeds, 2014. http://etheses.whiterose.ac.uk/7226/.

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The immune and reproductive systems have long been known to be inextricably linked, with components of immune pathways, particularly cytokines, mediating processes such as ovarian/menstrual cyclicity, endometrial remodelling, mating-induced immunomodulation, implantation, pregnancy, parturition and lactation. The nature of this involvement has often been investigated at the level of single mediators, with little consideration of the fact that cytokines are increasingly understood to function as complex networks. This study aimed to characterise inflammatory networks using both traditional and novel machine-learning Bayesian network-based methods in the context of keystone aspects of reproduction, viz., in the endometrial response to seminal plasma, cytokine:hormone interactions during lactation, and oocyte maturation following controlled ovarian hyperstimulation. ‘Traditional’ pathway analyses used to examine the murine endometrial response to seminal plasma revealed previously unidentified mediators and showed compartmentalised epithelium/stroma-specific responses. However, they proved ineffective in describing novel cytokine interactions. This led to the development a highly effective novel Bayesian network-based approach to explore cytokine:hormone networks during murine lactation. This revealed that prolactin, a putative potent immunomodulator, was far less influential than expected in vivo. The method also identified previously unknown cytokine interactions and described features such as synergy and antagonism. Further refinement of these network analyses as modified variational Bayesian state space models enabled the display of core, conserved subnetworks (communities) of human follicular fluid cytokines whose interactions varied with oocyte maturity. Moreover, these cytokine signatures also allowed the prediction of an oocytes’ fertilisability potential, with potential attendant benefits to assisted conception. This thesis represents the first endeavour to model inflammatory networks in vivo in any setting to date. It has revealed their central role, functional conservation and key features of cytokine interactions across a spectrum of reproductive processes. Further development of this methodology appears set to offer invaluable new insights into the complex immune signalling that underpins reproductive biology.

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Chambin, Odile. « Validation d'un modele d'absorption percutanee ex vivo : approche correlative avec des parametres in vivo ». Dijon, 1995. http://www.theses.fr/1995DIJOPE02.

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Strobel, Steffen Peter. « Die ex-vivo-Perfusion hDAF-transgener Kaninchennieren mit Humanblut : ein Modell für die humane Xenotransplantation ». Ulm : Universität Ulm, Medizinische Fakultät, 2004. http://www.bsz-bw.de/cgi-bin/xvms.cgi?SWB11482174.

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Zabkiewicz, Joanna. « In vivo modelling of tumour suppressor gene function ». Thesis, Cardiff University, 2005. http://orca.cf.ac.uk/55388/.

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LKB1 has been implicated in a wide range of cellular functions and is associated with many potential substrates in in vitro studies, however the in vivo role of LKB1 remains unclear and its precise contribution to the prevention of intestinal tumours in the hereditary Peutz-Jegers syndrome is as yet uncharacterised. Conditional deletion of LKB1 in the murine small intestine resulted in significant disruption of intestinal homeostasis, particularly that of the differentiation process, suggesting LKB1 plays a key role in intestinal differentiation and it is loss of this function that predisposes to tumourigenesis

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Hartung, Niklas. « Modelling of metastatic growth and in vivo imaging ». Thesis, Aix-Marseille, 2014. http://www.theses.fr/2014AIXM4763/document.

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Un problème majeur du cancer est l'apparition de métastases, difficiles à détecter par l'imagerie médicale et qui peuvent progresser rapidement. Par le biais de la modélisation mathématique, nous espérons développer de nouveaux outils capables d'anticiper l'état métastatique d'un patient.Les deux premières parties de cette thèse sont dédiées au développement d'un tel outil, l'objectif étant sonutilisation chez l'animal voire en clinique. Dû aux variabilités intra- et inter-individuelles, nous sommes amenés à utiliser des modèles statistiques coûteux en temps de calcul.Dans la partie 1, nous étendons une approche introduite par Iwata et al. et développée dans l'équipe. Nousproposons une résolution numérique plus efficace basée sur la reformulation du modèle sous formed'équation intégrale de Volterra de type convolution, qui s'avère également utile pour montrer despropriétés théoriques du modèle. En outre, nous étudions une extension stochastique de ce modèle déterministe.Dans la partie 2, nous montrons que notre approche est adaptée à la description de données souris. Utilisant le cadre statistique des modèles nonlinéaires à effets mixtes, nous construisons un modèle métastatique identifiable à partir des données et nous interprétons les résultats biologiquement.La partie 3 regroupe des résultats issus de collaborations avec des biologistes. Nous avons commencé àmodéliser la croissance tumorale à partir d'observations par imagerie SPECT en utilisant un modèle deGyllenberg et Webb. D'autre part, afin d'améliorer la précision des observations SPECT, nous testons des techniques dedétection de contours via des méthodes volumes finis basées sur des schémas DDFV
Metastasis is one of the major problems of cancer because metastases areoften difficult to detect by clinical imaging and may develop rapidly. With the help of mathematical modelling, we hope to developnew tools capable of anticipating the metastatic state of a patient.The first two parts of this thesis are dedicated to developing such a tool, destined for a preclinical oreven clinical use. As tumour growth dynamics vary strongly between individuals and since observations are often sparse andnoisy, we need to consider computationally expensive statistical tools.In the first part, we extend an approach introduced by Iwata et al. and developed by Barbolosi et al. In particular, wepropose a more efficient numerical resolution based on a model reformulation into a Volterra integral equation of convolutiontype. This reformulation also permits to prove theoretical model properties (regularity and identifiability). Moreover, we study a stochastic generalisation of this deterministic model.In the second part, we will show that our approach is suitable for the description of experimental data on tumour-bearing mice.Using the statistical framework of nonlinear mixed-effects modelling, we build a metastatic model that is identifiable fromour data. We then interpret the results biologically.The last part of this thesis contains several results obtained in collaboration with biologists. We have started to model tumourgrowth with data obtained from SPECT imaging, using a model by Gyllenberg and Webb. Also, in order to improve the precision ofSPECT data, we have tested contour detection methods via finite volume methods based on DDFV schemes

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SANTORO, MARIO. « Mathematical modelling of HIV-1 dynamics in vivo ». Doctoral thesis, Università degli Studi di Roma "Tor Vergata", 2009. http://hdl.handle.net/2108/1151.

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Pastore, C. V. « VENTILAZIONE MECCANICA E VOLUTRAUMA : STUDIO IN VIVO IN UN MODELLO SUINO ». Doctoral thesis, Università degli Studi di Milano, 2010. http://hdl.handle.net/2434/150177.

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Mechanical ventilation is an essential support for patients with acute lung pathologies, such as acute respiratory distress syndrome and acute lung injury, and it is generally applied in chirurgical practice. Specialists are however aware that, despite of its “life saving” role, this practice presents several negative side effects. Recently one of the most serious negative effects of mechanical ventilation, called Ventilatory Induced Lung Injury (VILI), has been detected and better analysed. This syndrome, initially associated with barotrauma, has been recently defined as volutrauma, meaning a damage of lung parenchyma caused by mechanical stress deriving from overdistension induced by high tidal volumes (VT). The aim of the present study was to evaluate the onset of ventilatory induced lung injury in a clinically relevant, validated and well-studied model, which closely mimics the human physiology and the ventilator setting currently used in the clinical arena. The study was performed using 18 pigs were involved, divided into three groups (n=6): two groups were mechanically ventilated (VT 20 ml/kg and 8 ml/kg), and one group was spontaneously breathing (SB). The duration of the experiments was 240 minutes. Hemogasanalysis and all main respiratory and circulatory parameters were detected every 30 minutes. Metalloproteinases 2 and 9 expression and activation and ET-1 levels were observed in the bronchoalveolar lavage fluid. At the end of the experiment, the animals were sacrificed and autoptic samples of lung, kidney and liver for histological and zymographic analysis were obtained. The results showed serious alterations of lung mechanics and structure induced by high VT, although the protective strategies as low VT were not immune from negative side effects. Respiratory function worsening was observed in spontaneously breathing subjects, too. Therefore, our study demonstrates that, both animals undergoing mechanical ventilation with high volumes and non-assisted breathing animals develop a massive lung edema, as revealed by extra-vascular lung water values. As expected, the alveolar over-distension induced ultrastructural cellular abnormalities only in animals subjected to high VT and not in those where lung distension was limited, as in our VT8 group, or absent, as in SB group. Our data show irrefutably that the severe edema formation noticed in spontaneously breathing animals was clearly related to the increase in pulmonary arterial pressure, which induced the extravasation of fluid into lung parenchyma. Moreover, we have evaluated the changes in lung mechanics and metalloproteinases production and activation in three different types of lung damages evoked by mechanical, hypoxic and septic stress. Under that, 24 pigs were studied, randomly divided into four groups (n=6): control group (pigs spontaneously breathing), mechanical stress group (pigs ventilated with high VT), hypoxic group (pigs inhaled with an hypoxic gas mixture), septic group (pigs i.v. infused with E.coli LPS). All the animals were studied for 240 minutes. Hemogasanalysis and main respiratory and circulatory parameters were detected every 20 minutes. At the end of the experiment, subjects were sacrificed and autoptic samples of lung for histological and zymographic analysis were obtained. The changes in physiological parameters were in line with morphological lung alterations. Zymographic analysis showed a strong activation of MMP-2 but no activation of MMP-9 in control, mechanical and hypoxic stress groups. The septic stress group has reflected a specular situation with activation of MMP-9 and low levels of MMP-2, which was present only in the inactivated form. The present study has underlined an acute modulation of MMPs in lung tissues and MMPs different behaviour facing different stimulations. In conclusion, it is clear that mechanical ventilation strategies profoundly affects lung parenchyma integrity and functionality, and the choice of a ventilation strategy that avoids these damages, ensuring at the same time an appropriate exchange of gases, is strongly encouraged.

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Rez, Mohammed Fayez al. « Modelling and measurement of the O2-concentration for the ex vivo cultivation of cells and tissues ». Dresden TUDpress, 2007. http://deposit.d-nb.de/cgi-bin/dokserv?id=2960243&prov=M&dok_var=1&dok_ext=htm.

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Wang, Vicky Yang. « Modelling in vivo cardiac mechanics using MRI and FEM ». Thesis, University of Auckland, 2012. http://hdl.handle.net/2292/13043.

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The role of myocardial mechanical properties in the progression of cardiovascular diseases such as heart failure (HF) remains poorly understood. In order to elucidate the mechanisms underlying cardiac dysfunction, mathematical modelling of ventricular mechanics is a useful tool. This thesis presents finite element modelling methods to estimate in vivo mechanical properties of the left ventricle (LV) by incorporating geometric and kinematic information derived from in vivo magnetic resonance image (MRI) tagging, ex vivo microstructural information extracted from diffusion tensor MRI (DTMRI), and LV cavity pressure data recorded during the MRI tagging experiments. This integrative LV modelling framework enabled in vivo characterisation of LV muscle properties and performance on a subject specific basis, and investigation of mechanisms of HF. In order to investigate passive ventricular mechanics, LV FE models were customised to geometric data segmented from in vivo tagged MRI, and myofibre orientation data derived from ex vivo DTMRI of five canine hearts using nonlinear FE fitting techniques. Simulation of diastolic LV mechanics allowed estimation of the passive myocardial mechanical properties, which were tuned to provide the best match between the end diastolic model predictions and the MRI tagging data. Contractile function and regional myocardial work of the LV were then examined by simulating systolic mechanics and estimating the associated contractile properties during the ventricular cycle. Based on these data, the LV efficiency throughout the cardiac cycle was quantified. The FE modelling framework was applied to study HF by investigating the effects of ventricular dilation, loss of anisotropy, and reduced myocardial contractility on LV mechanical performance. By comparing the mechanical function of normal and abnormal LVs, it was found that cardiac performance was most sensitive to the combination of ventricular dilation and compromised muscle contractility. Integrated physiological modelling of this kind enables mechanistic investigations into the pumping function of the LV on an individualised basis. As demonstrated in this thesis, this type of quantitative modelling can be used to analyse the functional implications of changes to the geometry, structure or mechanical properties of normal hearts, and thus can provide insight into mechanisms underlying mechanical dysfunction in the diseased or failing heart.
Whole document restricted until Feb. 2014, but available by request, use the feedback form to request access.

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Tsaopoulos, Dimitrios E. « In vivo knee joint mechanics : implications for musculotendon modelling ». Thesis, Manchester Metropolitan University, 2007. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.485989.

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The general aim of this thesis was to study knee joint mechanics in-vivo in healthy participants with particular attention to the quantification of the patellar tendon moment arm length and the accurate calculation of the actual knee joint moment when dynamometry-based musculotendon loading calculations are perfonned. Following a review of the relevant literature, the next chapter examined the pote!lttal relation between patellar tendon moment ann length and several relevant anthropometric characteristics. In the third chapter the variation in the patellar tendon moment ann length due to the use of different knee joint rotation centre origins was examined using in-vivo lateral X-ray imaging during passive knee extension. In the following study the knee joint moment errors induced during dynamometry testing due to the misalignment ofthe knee joint and dynamometer axes of rotation were examined accurately using X-ray imaging. The fifth chapter presents an experiment which 1) explores the changes in the patellar tendon moment ann length with different modes and intensities of muscle contraction using in vivo lateral X-ray imaging and 2) detennines the amount oferror in the estimation of the patellar tendon force when using resting-state moment arm values instead of the actual patellar tendon moment ann length values during muscle contraction. The results of the work in this thesis suggest that 1) the patellar tendon moment arm length variance cannot be explained by knee joint size differences and hence, existing imaging-based methodologies remain necessary for accurate quantification of the patellar tendon moment arm, 2) when patellar tendon moment ann length data are reported the clear definition of the used reference landmark is required and the limitations of each method should always be taken into account, 3) during dynamometry testing the moment recorded from the dynamometer in most instances overestimates the actual knee joint moment and therefore, these differences cannot be neglected if the musculotendon forces need to be detennined using dynamometry-based musculoskeletal loads calculations and 4) the patellar tendon moment ann length increased significantly with increased muscle contraction intensity.

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Engelschalt, Vivienne. « Mechanismen der antikörpervermittelten T-Zell-Depletion in vivo im Maus-Modell ». Doctoral thesis, Humboldt-Universität zu Berlin, Mathematisch-Naturwissenschaftliche Fakultät I, 2010. http://dx.doi.org/10.18452/16234.

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Monoklonale Antikörper (mAk) werden bereits erfolgreich zur therapeutischen Depletion verschiedener Zellpopulationen in vivo verwendet, die Mechanismen der Depletion sind jedoch unklar geblieben. In dieser Arbeit wurden im Mausmodell die molekularen Grundlagen der CD4+ T-Zelldepletion (CD4 TZD) nach einmaliger Gabe (i.p.) von 100 µg des anti-CD4-mAk YTS191.1 untersucht. Dabei konnte eine starke Korrelation zwischen Depletion und der Modulation des CD4-Moleküls von der Oberfläche beobachtet werden. Gleichzeitig zeigten sich organabhängige Unterschiede, sowohl im zeitlichen Verlauf, als auch in der Effizienz der Depletion. Im Thymus konnten weder Depletion noch Modulation detektiert werden, in Milz und Lymphknoten (Lk) war die CD4 TZD nach starker CD4-Modulation bereits nach 48 h mit 80-90 % maximal, in den Peyer-Plaques jedoch niedriger und verzögert (50-60 % nach 72 h). Anhand C3-defizienter Mäuse konnte ferner kein wesentlicher Beitrag von Komplement an der CD4 TZD beobachtet werden. Im Gegensatz dazu konnte durch die Verwendung verschiedener FcGamma-Rezeptor (FcGammaR)-defizienter Mäuse (FcGammaRI, FcGammaRII, FcGammaRIII, FcGammaRI/III und FcRGamma) wie auch durch die Blockade des FcGammaRIV eine starke, zudem organabhängige Beteiligung von FcGammaR an der CD4 TZD gezeigt werden. Während in der Milz die CD4 TZD von FcGammaRIV vermittelt wurde, waren in den Lk und Peyer-Plaques FcGammaRI/III involviert. Diese Befunde korrelierten mit der starken Expression von FcGammaRIV in Milz, Lunge, Darm, Niere und Leber, während in den Lk nur eine schwache und in Thymus und Peyer-Plaques keine Expression detektiert werden konnte. Innerhalb der Milz konnten erstmalig F4/80hoch Makrophagen als FcGammaRIV+ identifiziert und somit als potenzielle Effektorzellen der CD4 TZD bestimmt werden. Der direkte Vergleich der Depletion von CD4+ T-Zellen mit der Depletion von ICOS+ T-Zellen verdeutlichte darüber hinaus, dass die Effizienz der Zelldepletion nicht nur von den Eigenschaften des verwendeten mAk, sondern auch von denen des Zielmoleküls abhängig ist.
Monoclonal antibodies (mAb) are efficiently used for the therapeutic depletion of various cells in vivo yet the mechanisms of depletion are still unclear. In this work, the molecular principles of CD4+ T cell depletion (CD4 Tcd) by a single application of 100 µg of the anti-CD4 mAb YTS191.1.1 were investigated in the mouse. A strong correlation between the depletion and the surface modulation of the CD4 molecule could be observed. At the same time, organ-dependent differences in the kinetics as well as in the efficiency of depletion could be detected. In the thymus, neither modulation nor depletion were detectable. In the spleen and the lymph nodes (Ln), the modulation was strong and the depletion was maximal (80-90%) 48 h after mAb treatment. Interestingly, both modulation and depletion were decreased and delayed (50-60% after 72 h) in the Peyer`s patches. By using C3-deficient mice, no major contribution of complement to the CD4 Tcd was seen. On the contrary, with the help of different FcGamma-receptor (FcGammaR)-deficient mice (FcGammaRI, FcGammaRII, FcGammaRIII, FcGammaRI/III, and FcRGamma) and through the blockade of FcGammaRIV, a strong organ dependent involvement of FcGammaR could be shown. While the depletion in the spleen was clearly dependent on FcGammaRIV, in the Ln and the Peyer`s patches, FcGammaRI/III were involved. These findings correlated with the strong expression of FcGammaRIV in the spleen, the lung, the colon, the kidney, and the liver, while in the Ln the expression was weak and undetectable in the thymus and the Peyer`s patches. For the first time, F4/80high macrophages in the spleen could be identified as also being FcGammaRIV+, and are therfore considered as the potential effector cells of the CD4 Tcd. The direct comparison of the depletion of T cells via CD4 or ICOS pointed out that the target cell depletion is not only dependent on the properties of the mAb used, but also on those of the target molecule.

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Bein, Matthias. « Molekulare und immunelektronenmikroskopische Untersuchungen zur Expression von sekretorischen Aspartatproteinasen in einem ex vivo Modell der vaginalen Kandidose und in vivo ». Diss., lmu, 2005. http://nbn-resolving.de/urn:nbn:de:bvb:19-40728.

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Porres, Alexandre Torres. « Modelos psicoacústicos de dissonância para eletrônica ao vivo ». Universidade de São Paulo, 2012. http://www.teses.usp.br/teses/disponiveis/27/27157/tde-22022013-141622/.

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O principal problema desta tese é a aplicação da teoria psicoacústica sobre percepção de dissonância em novas ferramentas de computação musical em tempo real. Os objetivos e contribuições incluem: Prover uma revisão critica do Estado da Arte em teoria psicoacústica de modelos de dissonância, projetar trabalhos futuros; Explorar um potencial criativo negligenciado de modelos psicoacústicos de dissonância em Eletrônica ao Vivo; Desenvolver novas ferramentas computacionais baseadas na teoria; Investigar o potencial e limitações da teoria e técnicas. Discutir sua pertinência e impacto; Tornar a teoria e técnicas mais acessíveis a músicos por meio deste texto e as ferramentas desenvolvidas. A percepção de Dissonância é um fenômeno complexo em que a abordagem psicoacústica cobre apenas uma porção. Além disso, ainda há muito debate na área da psicoacústica sobre o desenvolvimento de modelos de dissonância. Portanto, a tese apresenta uma revisão crítica do Estado da Arte da teoria psicoacústica, apontando pontos fracos e fortes do atual conhecimento, e futuros desenvolvimentos no campo. Um teste perceptivo também foi elaborado para gerar ponderações relevantes. Os modelos foram implementados em Pure Data, e uma série de patches foi desenvolvida para testar o potencial criativo em eletrônica ao vivo. Esse processo culminou em um sistema desenvolvido em Pure Data que agrega diversas das técnicas baseadas nos modelos psicoacústicos. Suas possibilidades incluem, por exemplo, encontrar intervalos consonantes de acordo com um espectro, ou alterar componentes espectrais para que estejam de acordo com uma escala musical ou afinação particular. Essas ferramentas foram empregadas em um duo de improvisação livre com o autor e um saxofonista. O autor também compôs uma peça orquestral que utilizou o sistema desenvolvido na parte de eletrônica ao vivo.
The problem of this research is to apply the state of the art in psychoacoustic theories about dissonance perception in the development of novel creative computer music tools for composition and live electronics. Goals and contributions include: To provide a critical review of the State of the Art in Psychoacoustic theory regarding dissonance modeling, project further work; To explore neglected creative potential of psychoacoustic dissonance models in Live Electronics; To develop novel computer music tools based on the theory; To investigate the potential and limitations of the theory and techniques; To discuss its creative musical impact and pertinence; To make the theory and techniques more accessible to musicians through the text and the free developed tools. The perception of Dissonance is a complex phenomenon in which the psychoacoustical approach covers just a portion thereof. Not only that, but psychoacoustic theory is still in debate about the development of dissonance models. Therefore, the thesis provides a critical review of the state of the art in psychoacoustic theory, pointing out weaknesses and strong points of the current knowledge, and future developments in the field. A perceptual test was also designed to generate data for relevant insights. Apart from the theoretical review, the models were implemented in Pure Data, and a series of patches was developed in order to test their creative potential in Live Electronics. This culminated in a system developed as a Pure Data Patch that aggregates several techniques based on the psychoacoustic models. Their possibilities include, for example, finding consonant intervals according to a spectrum, or altering spectral components so they are in accordance to musical intervals and particular tunings. These tools were used in a Free Improvisation duo with the author and a sax player, and the author also composed an orchestral piece that applied the developed system in the Live Electronics section.

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Babiak, Alexander Jurij [Verfasser]. « Die Etablierung eines in vivo Modells zur Untersuchung der Arteriogenese / Alexander Jurij Babiak ». Ulm : Universität Ulm. Medizinische Fakultät, 2004. http://d-nb.info/101543858X/34.

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BUGNON, DENIS. « Simulation de la pharmacocinetique humaine des antibiotiques in vivo ». Nantes, 1997. http://www.theses.fr/1997NANT10VS.

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Nardini, Fabrizio <1985&gt. « Subject Specific Knee Joint Modelling Based on In Vivo Clinical Data ». Doctoral thesis, Alma Mater Studiorum - Università di Bologna, 2016. http://amsdottorato.unibo.it/7588/3/nardini_fabrizio_tesi.pdf.

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The knee is one of the most complex and studied joint of the musculoskeletal system provided its great importance in locomotion. Therefore, a deep understanding of its behaviour and of the role played by each of the structures composing it is fundamental. Knee joint models are an invaluable tool to understand the behaviour of the knee and their usefulness is proved in many fields such as surgical planning and prosthetic design. A huge amount of models has been proposed in the literature focusing on the kinematic, the kinetostatic and the dynamic behavior of the joint. Models can be based on in vivo or in vitro data. While the kinematic and the kinetostatic models are defined properly on in vitro data, the dynamic ones cannot. This discrepancy leads to a gap, a lack of coherence, between the usually in vitro defined kinematic and kinetostatic models and the study of the active structures of the joint. In order to achieve a comprehensive knee joint description in which the kinematic, kinetostatic and dynamic models coherently stem one from the other, the identification of a procedure that allows to obtaining reliable kinematic and kinetostatic models in vivo is needed. In the present dissertation a procedure is defined that allows for the identification of a subject specific knee joint model in vivo starting from standard clinical data obtained by the use of non invasive techniques such as computed tomography (CT), magnetic resonance imaging (MRI) and fluoroscopy. This procedure leads to an accurate identification of the parameters needed to personalize the 5-5 parallel mechanism and its patello-femoral extension on a single patient in order to accurately reply the knee joint original motion. Furthermore, following the sequential approach to the modelling of the joint, a stiffness model of the knee is specialized on the specific subject's anatomy.

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Nardini, Fabrizio <1985&gt. « Subject Specific Knee Joint Modelling Based on In Vivo Clinical Data ». Doctoral thesis, Alma Mater Studiorum - Università di Bologna, 2016. http://amsdottorato.unibo.it/7588/.

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The knee is one of the most complex and studied joint of the musculoskeletal system provided its great importance in locomotion. Therefore, a deep understanding of its behaviour and of the role played by each of the structures composing it is fundamental. Knee joint models are an invaluable tool to understand the behaviour of the knee and their usefulness is proved in many fields such as surgical planning and prosthetic design. A huge amount of models has been proposed in the literature focusing on the kinematic, the kinetostatic and the dynamic behavior of the joint. Models can be based on in vivo or in vitro data. While the kinematic and the kinetostatic models are defined properly on in vitro data, the dynamic ones cannot. This discrepancy leads to a gap, a lack of coherence, between the usually in vitro defined kinematic and kinetostatic models and the study of the active structures of the joint. In order to achieve a comprehensive knee joint description in which the kinematic, kinetostatic and dynamic models coherently stem one from the other, the identification of a procedure that allows to obtaining reliable kinematic and kinetostatic models in vivo is needed. In the present dissertation a procedure is defined that allows for the identification of a subject specific knee joint model in vivo starting from standard clinical data obtained by the use of non invasive techniques such as computed tomography (CT), magnetic resonance imaging (MRI) and fluoroscopy. This procedure leads to an accurate identification of the parameters needed to personalize the 5-5 parallel mechanism and its patello-femoral extension on a single patient in order to accurately reply the knee joint original motion. Furthermore, following the sequential approach to the modelling of the joint, a stiffness model of the knee is specialized on the specific subject's anatomy.

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Steckmeier, Stephanie. « Experimentelle Evaluation der endovenösen Radiofrequenzobliteration und Lasertherapie an einem neuen ex-vivo Modell ». Diss., lmu, 2006. http://nbn-resolving.de/urn:nbn:de:bvb:19-61465.

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Dahler, Anja Christina. « Die Rolle von ICOS auf die B-Zelldifferenzierung in einem in vivo Modell ». Doctoral thesis, Humboldt-Universität zu Berlin, Mathematisch-Naturwissenschaftliche Fakultät I, 2009. http://dx.doi.org/10.18452/15989.2.

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Der induzierbare Kostimulator ICOS ist ein zu CD28 strukturell und funktionell verwandtes Molekül, das eine wichtige regulatorische Rolle bei der T-Zelleffektorfunktion spielt. Eine ICOS-Defizienz beim Mensch manifestiert sich in einer schweren Störung des humoralen Immunsystems. Eine murine ICOS-Defizienz führt ebenfalls zu einer Beeinträchtigung der T-Zell-abhängigen humoralen Immunantwort, bei der kleinere oder komplett fehlende Keimzentren zu beobachten sind. Vielfältige in vitro und in vivo Studien führten diese Phänomene auf die beeinträchtigte Regulation von Kommunikationsmolekülen der Zelloberfläche und der Zytokinexpression durch ICOS-defiziente T-Zellen zurück. Ein Ziel dieser Arbeit war es, mit Hilfe von ICOS KO Mäusen den Einfluss von ICOS auf die B-Zellentwicklung genauer zu untersuchen. Dabei konnte gezeigt werden, dass ICOS erst in der späten Phase der B-Zellentwicklung eine Rolle spielt, da der Interaktionspartner von ICOS erst auf transitionellen B-Zellen der Milz exprimiert wird. Durch die Etablierung eines in vivo adoptiven T-B Transfermodells konnte die Rolle von ICOS erstmalig bei der T-B Kooperation in den frühen Phasen der Immunantwort auf der Ebene Antigen-spezifischer T- und B-Zellen aufgeklärt werden. Dabei konnte beobachtet werden, dass eine ICOS-Defizienz einen dramatischen Einfluss auf die B-Zellexpansion und B-Zellproliferation hat. Zum ersten Mal konnte in vivo gezeigt werden, dass ICOS bei der T-B Kooperation eine entscheidende Rolle bei der Regulation diverser Oberflächenmarker der B-Zellen spielt, wodurch die B-Zellaktivierung, B-Zellproliferation und B-Zelldifferenzierung bei der Keimzentrums- und Plasmazellreaktion beeinflusst werden. Histologische Analysen zeigten, dass bei einer ICOS-Defizienz follikuläre T-Helferzellen nicht in die Keimzentrumsumgebung einwandern und daher keine T-Zellhilfe für die B-Zellen anbieten können. Dadurch kann die Keimzentrumsreaktion nicht weiter aufrechterhalten werden und eine Ausbildung von kleineren Keimzentren ist die Folge. Weiterhin konnte beobachtet werden, dass eine fehlende ICOS-Interaktion zwischen T- und B-Zellen zu einer Störung der Plasmazellgenerierung führt, wodurch auch die Mengen an messbaren Serumimmunglobulinen beeinflusst werden. Eine erhöhte Gabe von ICOS-defizienten T-Zellen kann diese Effekte nicht vollständig ausgleichen. Daher ist erkennbar, dass ICOS eine Vielzahl von zusätzlichen Faktoren beeinflusst, die für die ICOS-abhängigen B-Zelleffekte verantwortlich sind.
The inducible costimulator ICOS, structural and functional similar to CD28, plays an important regulatory role in T cell receptor function. The ICOS deficiency in humans is described as a severe dysfunction of the humoral immune response, resulting in dramatic reduced B cell numbers and impaired antibody response against pathogens. The murine ICOS-deficiency also leads to a disturbed T cell dependent immune response resulting in a reduced germinal center formation. Various in vitro and in vivo studies attributes this phenomenon to impaired upregulation of cell surface communication molecules and cytokine synthesis by ICOS-deficient T cells. In this work the investigations with ICOS KO mice should clarify the impact of ICOS in B cell development. As observed, ICOS can only play a role in the late phase B cell development, because the interaction partner is expressed on transitional B cells in the spleen. The establishment of an in vivo adoptive T-B transfer system could determine for the first time the role of ICOS in T-B cooperation in early immune response stages on antigen specific T and B cell levels. As shown, ICOS deficiency influences in a dramatic extend the B cell expansion and B cell proliferation. For the first time in vivo, we could demonstrate that ICOS plays a significant role by influencing the regulation of various B cell surface markers, which affects the B cell activation, B cell proliferation and B differentiation in germinal center or plasma cell reaction. Histological investigations revealed in the ICOS-deficiency that follicular T helper cells could not migrate into the germinal center microenvironment and therefore could not provide T cell help for B cells. As a result, the germinal center reaction could not maintained and therefore the formation of little germinal centers occurred. The missing interaction between T and B cells leads to a dysfunction in plasma cell generation and also influences the detectable amounts of serum immunglobulines. An administration of higher ICOS KO T cell numbers could not fully compensate these effects. Therefore, ICOS bias multitudes of additional factors, which are responsible for the ICOS dependent B cell effects.

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Dahm, Christian Oliver [Verfasser]. « Beurteilung von kryopräservierten Rattenarterien im In-vivo-Modell nach Transplantation / Christian Oliver Dahm ». Hamburg : Staats- und Universitätsbibliothek Hamburg, 2011. http://d-nb.info/1010760920/34.

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Gassert, Felix [Verfasser]. « Geweberegeneration in einem bovinen ex vivo-Knorpeltrauma-Modell : Analysen therapeutischer Effekte / Felix Gassert ». Ulm : Universität Ulm, 2020. http://d-nb.info/1217715371/34.

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Dahler, Anja Christina. « Die Rolle von ICOS auf die B-Zelldifferenzierung in einem in vivo Modell ». Doctoral thesis, Humboldt-Universität zu Berlin, Mathematisch-Naturwissenschaftliche Fakultät I, 2010. http://dx.doi.org/10.18452/15989.

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Der induzierbare Kostimulator ICOS ist ein zu CD28 strukturell und funktionell verwandtes Molekül, das eine wichtige regulatorische Rolle bei der T-Zelleffektorfunktion spielt. Eine ICOS-Defizienz beim Mensch manifestiert sich in einer schweren Störung des humoralen Immunsystems. Eine murine ICOS-Defizienz führt ebenfalls zu einer Beeinträchtigung der T-Zell-abhängigen humoralen Immunantwort, bei der kleinere oder komplett fehlende Keimzentren zu beobachten sind. Vielfältige in vitro und in vivo Studien führten diese Phänomene auf die beeinträchtigte Regulation von Kommunikationsmolekülen der Zelloberfläche und der Zytokinexpression durch ICOS-defiziente T-Zellen zurück. Ein Ziel dieser Arbeit war es, mit Hilfe von ICOS KO Mäusen den Einfluss von ICOS auf die B-Zellentwicklung genauer zu untersuchen. Dabei konnte gezeigt werden, dass ICOS erst in der späten Phase der B-Zellentwicklung eine Rolle spielt, da der Interaktionspartner von ICOS erst auf transitionellen B-Zellen der Milz exprimiert wird. Durch die Etablierung eines in vivo adoptiven T-B Transfermodells konnte die Rolle von ICOS erstmalig bei der T-B Kooperation in den frühen Phasen der Immunantwort auf der Ebene Antigen-spezifischer T- und B-Zellen aufgeklärt werden. Dabei konnte beobachtet werden, dass eine ICOS-Defizienz einen dramatischen Einfluss auf die B-Zellexpansion und B-Zellproliferation hat. Zum ersten Mal konnte in vivo gezeigt werden, dass ICOS bei der T-B Kooperation eine entscheidende Rolle bei der Regulation diverser Oberflächenmarker der B-Zellen spielt, wodurch die B-Zellaktivierung, B-Zellproliferation und B-Zelldifferenzierung bei der Keimzentrums- und Plasmazellreaktion beeinflusst werden. Histologische Analysen zeigten, dass bei einer ICOS-Defizienz follikuläre T-Helferzellen nicht in die Keimzentrumsumgebung einwandern und daher keine T-Zellhilfe für die B-Zellen anbieten können. Dadurch kann die Keimzentrumsreaktion nicht weiter aufrechterhalten werden und eine Ausbildung von kleineren Keimzentren ist die Folge. Weiterhin konnte beobachtet werden, dass eine fehlende ICOS-Interaktion zwischen T- und B-Zellen zu einer Störung der Plasmazellgenerierung führt, wodurch auch die Mengen an messbaren Serumimmunglobulinen beeinflusst werden. Eine erhöhte Gabe von ICOS-defizienten T-Zellen kann diese Effekte nicht vollständig ausgleichen. Daher ist erkennbar, dass ICOS eine Vielzahl von zusätzlichen Faktoren beeinflusst, die für die ICOS-abhängigen B-Zelleffekte verantwortlich sind.
The inducible costimulator ICOS, structural and functional similar to CD28, plays an important regulatory role in T cell receptor function. The ICOS deficiency in humans is described as a severe dysfunction of the humoral immune response, resulting in dramatic reduced B cell numbers and impaired antibody response against pathogens. The murine ICOS-deficiency also leads to a disturbed T cell dependent immune response resulting in a reduced germinal center formation. Various in vitro and in vivo studies attributes this phenomenon to impaired upregulation of cell surface communication molecules and cytokine synthesis by ICOS-deficient T cells. In this work the investigations with ICOS KO mice should clarify the impact of ICOS in B cell development. As observed, ICOS can only play a role in the late phase B cell development, because the interaction partner is expressed on transitional B cells in the spleen. The establishment of an in vivo adoptive T-B transfer system could determine for the first time the role of ICOS in T-B cooperation in early immune response stages on antigen specific T and B cell levels. As shown, ICOS deficiency influences in a dramatic extend the B cell expansion and B cell proliferation. For the first time in vivo, we could demonstrate that ICOS plays a significant role by influencing the regulation of various B cell surface markers, which affects the B cell activation, B cell proliferation and B differentiation in germinal center or plasma cell reaction. Histological investigations revealed in the ICOS-deficiency that follicular T helper cells could not migrate into the germinal center microenvironment and therefore could not provide T cell help for B cells. As a result, the germinal center reaction could not maintained and therefore the formation of little germinal centers occurred. The missing interaction between T and B cells leads to a dysfunction in plasma cell generation and also influences the detectable amounts of serum immunglobulines. An administration of higher ICOS KO T cell numbers could not fully compensate these effects. Therefore, ICOS bias multitudes of additional factors, which are responsible for the ICOS dependent B cell effects.

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Martins, Paulo Sérgio [UNESP]. « A imaginação na elaboração de modelos científicos em Vico ». Universidade Estadual Paulista (UNESP), 2017. http://hdl.handle.net/11449/150522.

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Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)
No século XVIII, o filósofo napolitano Giambattista Vico faz uma crítica ao racionalismo cartesiano, refutando a tese de que as ciências naturais pudessem alcançar a verdade. Seu pensamento reivindica, para a imaginação, a linguagem e a história – dimensões negligenciadas pela corrente racionalista – um status prioritário para a obtenção de conhecimento verossímil. A atenção exacerbada para atividades puramente racionais, como a matemática e a lógica, para a obtenção de conhecimento, segundo Vico, conduziria o homem a uma espécie de “barbárie”, levando-o à desumanização. De acordo com Vico, fazer é conhecer e vice-versa. Somente se pode conhecer aquilo que se faz. Ao homem, não é possível o conhecimento da natureza em sua essência, pelo fato de não ser o seu criador. No entanto, o homem cria a história e, por isso, pode conhecê-la. Os vestígios históricos trazidos pela filologia, somados à reflexão filosófica, poderão conduzir o homem ao conhecimento. Com a sua máxima verum et factum convertuntur (“conheço porque faço, faço porque conheço”), Vico coloca o homem como um produtor de modelos representativos do mundo, o qual lança mão da imaginação e do engenho (criação), para conceber a realidade. Partindo dessa máxima viquiana, visamos, neste trabalho, analisar a relevância das faculdades da imaginação e do engenho nos processos de elaboração de modelos científicos. No primeiro capítulo, buscamos compreender os principais conceitos do pensamento viquiano; em seguida, no segundo capítulo, definimos os conceitos agregados à faculdade imaginativa, como a memória, o engenho e a fantasia. Vico propõe ampliar a validação do conhecimento para além do crivo do raciocínio lógico. Desse modo, no terceiro capítulo investigamos o exercício da imaginação como um fator preponderante para a elaboração de hipóteses e para a geração de novos modelos na ciência.
In the 17th century, Neapolitan philosopher Giambattista Vico criticizes Cartesianism, refuting the thesis that the Natural Sciences could reach the truth. His thought reclaims, to imagination, language and history - dimensions neglected by rationalist thinking - a priority status for the acquisition of credible knowledge. The exacerbated attention to purely rational activities such as mathematics and logic for knowledge acquisition, according to Vico, would lead mankind to a sort of "barbarism" taking it to dehumanization. For Vico, to do is to know and vice-versa. We can only know what we do. To men, it is not possible to know nature in its essence, for they are not its creator. However, man creates history and, therefore, may know it. The historical traces brought by philology, added to the philosophical reflection, may lead man to knowledge. With his maxim verum et factum convertuntur, (I know because I do, I do because I know), Vico places man as a producer of depictive models of the world, who makes use of imagination and ingeniousness (creation) to conceive the reality. From this Vichian maxim, we aim, in this study, to analyze the relevance of imagination and ingeniousness faculties in the processes of scientific models development. In the first chapter, we try to understand the main concepts of the Vichian thinking; after that, in the second chapter, we define the concepts aggregated with the imaginative faculty, like the memory, the ingeniousness and the fantasy. Vico suggests expand the validation of knowledge beyond the scrutiny of logic reasoning. Thus, in the third chapter, we investigate the exercise of imagination as a preponderant factor for the hypotheses elaboration and for the generation of new models in science.

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Dupain, Thibaut. « Modélisation pharmacocinétique "in vitro" et activité bactéricide du cefpirome sur des souches à sensibilité réduite ». Paris 5, 1993. http://www.theses.fr/1993PA05P007.

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Ooms, Astrid [Verfasser]. « Generierung und Charakterisierung eines in vivo Modells zur BAG3P209L-Mutation im Herzmuskel / Astrid Ooms ». Bonn : Universitäts- und Landesbibliothek Bonn, 2017. http://d-nb.info/1124540148/34.

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40

Cavallari, Giuseppe <1972&gt. « Capacità immunomodulatoria delle cellule staminali mesenchimali in un modello di trapianto d'organo in vivo ». Doctoral thesis, Alma Mater Studiorum - Università di Bologna, 2008. http://amsdottorato.unibo.it/1130/1/Tesi_Cavallari_Giuseppe.pdf.

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Cavallari, Giuseppe <1972&gt. « Capacità immunomodulatoria delle cellule staminali mesenchimali in un modello di trapianto d'organo in vivo ». Doctoral thesis, Alma Mater Studiorum - Università di Bologna, 2008. http://amsdottorato.unibo.it/1130/.

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Salas, Cifuentes Valentina Lidia. « Evaluación de la actividad in vitro y de la eficacia experimental in vivo de antibióticos antifúngicos frente a hongos oportunistas emergentes ». Doctoral thesis, Universitat Rovira i Virgili, 2012. http://hdl.handle.net/10803/90246.

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The treatment of invasive mycoses is far to be the optimal and despite advances made in the last decades these infections continue to be associated with high mortality rates. The main goal of this thesis was to evaluate the in vitro and in vivo efficacy of new therapeutical strategies against systemic infections such as candidiasis, aspergillosis and mucoramycosis. The best in vivo efficacy of echinocandins in the treatment of infections by Candida parapsilosis was only observed among those mice infected with the strains that showed the lowest MICs. Posaconazole showed good efficacy against infections by Aspergillus terreus. The voriconazole in vitro data could be useful for predicting the outcome of A. terreus infections; while A. fumigatus isolates showed important variability in their in vivo response to this drug. Amphotericin B was the most effective drug for the treatment of infections by Mucor circinelloides. Posaconazole and amphotericin B showed efficacy in the treatment of Apophysomyces variabilis infections. Posaconazole had the greatest efficacy in our model of disseminated infection by Saksenaea vasiformis.
Las terapias utilizadas en la actualidad para el tratamiento de infecciones fúngicas invasoras causadas por hongos oportunistas están lejos de ser las óptimas, y a pesar de los avances realizados en las últimas décadas, estas infecciones siguen asociadas a altas tasas de mortalidad. El objetivo principal de esta tesis ha sido contribuir al desarrollo experimental de nuevos tratamientos o a la mejora de los ya existentes frente a infecciones sistémicas como las candidiasis, aspergilosis y mucoramicosis. Para ello, se han realizado estudios in vitro e in vivo en modelos animales adecuados que han permitido comprobar la eficacia de diferentes fármacos.

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Heinz, Monika. « Vanadium-Komplexe als Modelle molybdopterinhaltiger Enzyme sowie In-vivo-Einbau von Vanadium in die Sulfitoxidase von E. coli ». Göttingen Cuvillier, 2009. http://d-nb.info/995096570/04.

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Sauter, Andreas. « Evaluation Melanocortin-2-Rezeptor abhängiger Effekte auf das Fettgewebe in einem in vivo Modell ». Diss., Ludwig-Maximilians-Universität München, 2015. http://nbn-resolving.de/urn:nbn:de:bvb:19-178904.

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Drews, Tobias [Verfasser]. « Genauigkeit der computerunterstützten Lungenrundherdvolumetrie an künstlichen pulmonalen Läsionen im Ex-vivo-Modell / Tobias Drews ». Kiel : Universitätsbibliothek Kiel, 2009. http://d-nb.info/1019867647/34.

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Wiesmeier, Michael Wolf Peter [Verfasser], Georg [Akademischer Betreuer] Häcker et Robert [Akademischer Betreuer] Zeiser. « Neutrophilenelastase in der Neutropenie : : Analysen in einem zellulären Modell in vitro und in vivo ». Freiburg : Universität, 2017. http://d-nb.info/1147680647/34.

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Carter, Ross. « In vivo analysis of plant mechanics using atomic force microscopy and modelling ». Thesis, University of Sheffield, 2015. http://etheses.whiterose.ac.uk/11611/.

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Morphogenesis in plants occurs as a combined outcome of hormone pattern formation and mechanical changes involving cell growth and division. In this work a method was developed for measuring mechanical properties of plant cells at sub-cellular resolution using the atomic force microscope. Initial work focussed on the measurement of the mechanical properties of a number of wild type and transgenic Arabidopsis hypocotyls. Mutants in expansin expression were measured and were hypothesised to have cell walls of reduced stiffness. Due to the variation in results, or the wrong hypothesis, trends that fitted with the hypotheses were not seen. A series of experiments were performed to investigate this variance and standardise future measurements. An investigation into the effect of angle of indentation and depth of indentation were carried out. Following on from this work focussed on the stomata. A series of experiments were carried out to quantify the mechanical properties of these cells, comparing them to the properties of the cells surrounding them. Multivariate correlation analysis was performed to test hypotheses that these measurements correlated with other geometrical parameters, such as how open the stomate is and how large it is and significant correlations between these were found. A model of auxin pattern formation in the leaf margin was developed that incorporated knowledge of a family of auxin importers thought to have an important role in morphogenesis. These results show that auxin importers may have a role in stabilising the patterning of hormones and influence the timing and positioning of hormone peaks. This model highlighted the fact that the idea of a cell’s sensitivity to auxin is important and may have an affect when considering how auxin effects the growth of cells.

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Svitojūtė, Eglė. « Eksperimentinio nefrotoksiškumo modelio in vivo sukūrimas ir jo taikymas kamieninių ląstelių tyrimuose ». Master's thesis, Lithuanian Academic Libraries Network (LABT), 2012. http://vddb.laba.lt/obj/LT-eLABa-0001:E.02~2012~D_20120618_084003-66249.

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Pagrindinis tyrimo tikslas buvo sukurti eksperimentinį nefrotoksiškumo modelį in vivo ir įvertinti jo pritaikomumą ikiklinikiniuose kamieninių ląstelių tyrimuose. Nefrotoksiškumo modeliui parinkti 9–12 sav. amžiaus Wistar linijos žiurkių patinai. Eksperimento metu gyvūnai buvo laikomi individualiuose metaboliniuose narvuose esant pastovioms aplinkos sąlygoms. Inkstų pažeidimui sukelti parinktas gentamicino sulfato injekcinis tirpalas 40mg/1ml, kai jo dozė 80 mg/kg/d i. p. 7 dienas. Sudarytos trys laboratorinių gyvūnų grupės: I – kontrolinė grupė (1.5 ml 0.9 proc. NaCl tirpalo injekcijos i. p. 14 dienų), II – terapinė grupė (gentamicino injekcijos i. p. 14 dienų, dozė 5 mg/kg/d), III – pažeidimo grupė (gentamicino injekcijos i. p. 7 dienas, dozė 80 mg/kg/d). Vertinti žiurkių fiziologiniai, elgesio, biocheminiai šlapimo ir kraujo, frakcinės ekskrecijos, glomerulų filtracijos greičio ir histologiniai inkstų rodikliai. Visi rodikliai lyginti tarp trijų grupių. Laboratoriniams gyvūnams paskirta toksinė gentamicino dozė i. p. 80 mg/kg/d 7 dienas iš eilės sukėlė funkcinius ir morfologinius inkstų pokyčius, t. y. ūminį inkstų funkcijos pažeidimą, kurio pasireiškimą statistiškai patikimai (p<0.05) atspindėjo fiziologiniai, elgesio, biocheminiai kraujo ir šlapimo, frakcinės ekskrecijos, glomerulų filtracijos greičio ir histologiniai inkstų rodikliai lyginant su tais pačiais kontrolinės ir terapinės grupių žiurkių rodikliais. Sukurtas nefrotoksiškumo modelis pritaikytas bandomojo... [toliau žr. visą tekstą]
The main objective of this study was to develop experimental nephrotoxicity model in vivo and to assess its’ applicability for stem cell research. 9–12 weeks old male Wistar rats were chosen for the establishment of this model. During the experiment rats were housed in individual metabolic cages and maintained under standard conditions. Gentamicin sulphate solution for injection (40mg/1ml) was chosen as a toxicant. Toxic dose – genamicin 80 mg/kg/d i. p. for 7 consecutive days. 3 groups were constituted for the study: I group – control group (i. p. injection of saline 1.5 ml for 14 consecutive days), II group – therapeutic group (i. p. injection of genamicin 5mg/kg/d for 14 consecutive days), III group – acute kidney injury group (i. p. injection of genamicin 80mg/kg/d for 7 consecutive days). We evaluated physiological parameters, behavioural parameters, biochemical urine and blood parameters, fractional excretion parameters, glomerular filtration rate and histological kidney parameters. All parameters were compared between groups. Gentamicin administration in a very high dose (80 mg/kg/d i. p. for 7 consecutive days) caused functional and morphological renal changes and induced acute kidney injury, which was marked by statistically significant changes of physiological, behavioural, biochemical, fractional excretion, glomerular filtration rate and histological parameters, comparing with the same parameters of control and therapeutic groups. Nephrotoxicity model was applied... [to full text]

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Hornsby, Jack. « Bladder microstructural and biomechanical modelling : in vivo, in vitro and in silico ». Thesis, University of Oxford, 2016. https://ora.ox.ac.uk/objects/uuid:634324ed-851a-4959-adbd-fb45f05a27b6.

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Lower urinary tract disorders are significant prognostic indicators of institutionalisation and lower quality of life in the elderly and their incidence increases with age. Urodynamics, the gold standard in diagnosis, replicates symptoms to assess functionality through controlled filling and voiding of the bladder but its interpretation is subjective and may be inconclusive; often requiring further testing or leading to inappropriate treatment. Normal filling and voiding biomechanics of the bladder relate directly to the structural composition of the bladder wall. Alterations to tissue composition in aging and pathology have significant impacts on biomechanics but are yet to be fully described. The aim of this thesis was to gain insight into the individual microstructural components of the bladder wall and how they relate to the gross mechanical response. Additionally, representation of these observations in a mathematical model that can be used to improve our understanding of urodynamic data. This aim was achieved through a combination of in situ mechanical testing and the development of a microstructural constitutive model, which was then included within an overall micturition framework to simulate filling and voiding functions, and evaluated with clinical data. Coupled systems of multiphoton microscopy and uniaxial, biaxial and inflation testing were used to correlate extra cellular matrix interactions with the mechanical response of young and aged murine bladder. Wall-layer specific collagen fibre orientation, dispersion and recruitment were quantified and implemented into a novel microstructural constitutive model. The bladder was modelled as a nonlinear elastic, constrainedmixture planar membrane with contribution from smooth muscle and collagen fibres in the detrusor. Collagen recruitment in the detrusor was observed to occur at a finite stretch; correlated with a steep increase in stiffness of the tissue, while collagen of the lamina propria plays a capacitance role. Collagen recruitment was modelled using a triangular probability density function; quantified from sequential microscopy images and fitted to mechanical data. Increased collagen area fraction and changes in dominant fibre orientation were attributed to reduced compliance in aged bladder. This behaviour was captured by the model. The microstructural model was modified to an isotropic thin-walled spherical membrane for the filling phase of a micturition model framework, consisting of a bladder outlet relation and urethral resistance relation. A contractile smooth muscle element was included in the active response. In the first steps towards clinical application the model was applied to male and female 'normal' patient urodynamic data to observe quality of model fit and estimate baseline parameter values. The model simulated key filling and voiding features seen in normal male and female clinical data. Mechanobiological modelling combined with clinically relevant micturition modelling has the potential to quantify bladder dysfunction. Moreover, improved understanding of how the microstructure influences macroscopic mechanics will yield improved understanding of how changes to the bladder impair its functionality. We predict that modelling will become a clinically relevant tool in urodynamics; leading to new options for diagnosis and management of patients with bladder dysfunction.

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Verdon, Renaud. « Infection a cryptosporidium parvum : developpement de modeles in vivo et in vitro ». Paris 7, 1997. http://www.theses.fr/1997PA077339.

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L'infection a cryptosporidium parvum du sujet immunodeprime est une entite de decouverte recente dont la frequence s'est accrue avec l'emergence du syndrome d'immunodeficience acquise. Aucun traitement efficace autre que la correction du deficit immunitaire n'a ete a ce jour trouve. L'objectif de ce travail est le developpement de modeles in vivo et in vitro susceptible de contribuer a la mise au point d'un traitement efficace. L'utilisation d'un modele chez le rat immunodeprime a permis de verifier l'efficacite partielle et insuffisante de la paromomycine. Une des raisons possibles de l'echec de cette molecule est la persistance de l'infection dans des gites peu accessibles au traitement tels que les voies biliaires. Un modele permettant l'obtention d'une cholangite spontanement resolutive chez la souris adulte immunocompetente et d'une cholangite associee a une infection ileale chez la souris immunodeprimee par la dexamethasone est decrit dans ce travail. Ces 2 modeles devraient permettre l'etude d'une part des mecanismes impliques dans l'eradication de l'infection et d'autre part de molecules potentiellement efficaces sur les 2 localisations de l'infection. En raison du faible nombre de molecules candidates, il est apparu que des modeles permettant l'etude de mecanismes precis de l'infection de la cellule hote pourrait permettre de definir de nouvelles cibles therapeutiques. Ainsi, un modele d'attachement des sporozoites aux cellules hotes, avec detection par reaction immuno-enzymatique s'est revele interessant dans l'etude de substances pouvant inhiber une etape initiale de l'invasion cellulaire. De telles substances sont des glycoproteines pouvant interferer avec une activite lectine et des anticorps monoclonaux ou polyclonaux diriges contre des epitopes du sporozoite. Enfin, 2 modeles d'infection cellulaire utilisant la lignee enterocytaire caco-2 et une lignee epitheliale biliaire recemment produite (h69) ont ete developpes avec lecture immuno-enzymatique. Ces 2 modeles qui permettent une lecture rapide, semi-quantitative et objective de l'intensite de l'infection sont particulierement adaptes au screening de substances potentiellement efficaces contre l'infection a c. Parvum.

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Thèses sur le sujet « Modelli in vivo »

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