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Cimetta, Elisa (2009) Design and development of microscale technologies and microfluidic platforms for the in vitro culture of stem cell. [Tesi di dottorato]

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Abstract (inglese)

The impelling needs related to the processes of drug and therapy development for the cure of diseases such as Duchenne Muscular Dystrophy or myocardial infarction, led to an increasing demand for the development of innovative methods and strategies. It is also clear how stem cells could represent a fundamental source for the production of artificial human tissues to be employed in such processes. This perspective, would require both sophisticated tools for the control of stem cells differentiation and their integration within procedures apt to satisfy the fundamental requirements for obtaining tissue-model on which perform pharmacological or therapeutic studies. Among the fundamental requisites are: micronization, versatility, low-cost and highthroughput.
The main aims of this thesis have been the design, development and fabrication of microscale technologies capable of both reproducing a biomimetic stimulation inspired to the in vivo cell microenvironment and responding to the above mentioned technological requirements.
We performed a semi-quantitative analysis of the characteristic times of microscale phenomena that lead to the generation of operative diagrams that would prove useful in the design and development phases of experimental strategies. We developed microfluidic microbioreactors that have been used for biological studies involving cell cultures. In particular, we investigated the role and effect of concentration gradients on the fundamental Wnt signaling pathway.
We developed techniques for obtaining a topological control at the microscale of cell cultures on hydrogels with tunable mechanical properties. These techniques were successfully applied and interfaced with relevant biological systems such as primary human myoblasts from dystrophic patients and human embryonic stem cells-derived cardiomyocytes.
We finally proposed a prototype of an integrated microfluidic platform capable of coupling the topological stimulations to the control of the soluble microenvironment on cell cultures.
The obtained results open new and interesting perspectives for both de efficient development of drugs or therapies for defined diseases and for gaining deeper insights into the complexity of biological systems.

Abstract (italiano)

Le impellenti necessità legate allo sviluppo di nuovi farmaci e di terapie innovative per la cura di malattie dell’apparato muscolare, quali ad esempio la Distrofia Muscolare di Duchenne o l’infarto miocardico, hanno portato alla crescente domanda di nuovi metodi e tecnologie. E’ chiaro inoltre come le cellule staminali possano costituire una risorsa fondamentale per la generazione di tessuti umani artificiali da impiegare in tali processi. Tale prospettiva, richiederebbe da un lato strumenti sofisticati per il controllo e il differenziamento delle cellule staminali e dell’altro, l’integrazione di questi all’interno di procedure in grado di soddisfare i requisiti fondamentali dei modelli sui quali operare per lo sviluppo di nuovi farmaci o strategie terapeutiche. Tra i requisiti fondamentali da rispettare si citano quindi: micronizzazione, versatilità, basso costo e highthroughput.
Obiettivo fondamentale di questa tesi sono stati la progettazione, sviluppo e fabbricazione di tecnologie su scala micrometrica in grado di riprodurre una stimolazione biomimetica ispirata al microambiente cellulare in vivo e, nello stesso tempo, di rispondere ai requisiti tecnologici descritti sopra.
E’ stata effettuata un’analisi semi quantitativa basata sull’analisi dei tempi caratteristi dei fenomeni su microscala, che ha portato alla produzione di diagrammi operativi da impiegarsi nelle fasi di progettazione e sviluppo di tali strategie sperimentali. Sono stati sviluppati microbioreattori all’interno di piattaforme microfluidiche applicate poi allo studio di sistemi cellulari. In particolare, sono stati effettuati studi biologi sull’effetto di gradienti di concentrazione all’interno dell’importante signaling cellulare del Wnt.
E’ stata realizzata una tecnica per l’organizzazione topologica su microscala di colture cellulari su substrati in hydrogel dalle proprietà meccaniche definite. Le tecnologie sviluppate sono state impiegate per casi studio dall’elevato valore scientifico e sono state interfacciate con colture di particolare interesse quali mioblasti umani distrofici e cellule cardiache derivate da staminali embrionali umane.

Infine, è stato proposto un prototipo di piattaforma microfluidica in grado di accoppiare le stimolazioni di tipo topologico al controllo dell’ambiente solubile su colture cellulari.
I risultati ottenuti aprono nuove ed interessanti prospettive sia per lo sviluppo di nuovi farmaci che di strategie terapeutiche volti alla cura di patologie ed allo studio approfondito della complessità dei sistemi biologici.

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Tipo di EPrint:Tesi di dottorato
Relatore:Elvassore, Nicola
Correlatore:Vunjak-Novakovic, Gordana
Dottorato (corsi e scuole):Ciclo 21 > Scuole per il 21simo ciclo > INGEGNERIA INDUSTRIALE > INGEGNERIA CHIMICA
Data di deposito della tesi:02 Febbraio 2009
Anno di Pubblicazione:01 Febbraio 2009
Parole chiave (italiano / inglese):Microfluidics, Stem Cells, Microbioreactor, Microscale, Skeletal Muscle Tissue Engineering, Cardiac Tissue Engineering, Computational Modeling
Settori scientifico-disciplinari MIUR:Area 09 - Ingegneria industriale e dell'informazione > ING-IND/24 Principi di ingegneria chimica
Struttura di riferimento:Dipartimenti > Dipartimento di Principi e Impianti di Ingegneria Chimica "I. Sorgato"
Codice ID:1911
Depositato il:02 Feb 2009
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