Vai ai contenuti. | Spostati sulla navigazione | Spostati sulla ricerca | Vai al menu | Contatti | Accessibilità

| Crea un account

Serena, Elena (2009) Microscale Tissue Engineering of human skeletal and cardiac muscles for in vitro applications. [Tesi di dottorato]

Full text disponibile come:

[img]
Anteprima
Documento PDF
9Mb

Abstract (inglese)

Recently, engineered tissues have found an alternative application as in vitro models. They will never be implanted directly into patients, but will instead be used to transform the way we study human tissue physiology and pathophysiology in vitro. The development of new drugs and therapies for diseases such as Duchenne Muscular Dystrophy or myocardium failure is greatly slowed and hindered by the lack of adequate in vitro models, which should be functional, representative of human tissue, easy to use and economic. Therefore the requirements are the use of a human cell source; working at the microscale, in order to reproduce with high precision the cell microenvironment and to guide cell differentiation correctly. The microscale gives the additional possibility of developing micrometric array of cells which can be coupled to microfluidic platforms for highthroughput experiments, which are fundamental for multifactorial diseases such as DMD and HF. Moreover the use of easy methodologies and simple techniques allowed the use of the model by researchers with different background and skills. In this scenario, the aim of this thesis is the obtainment of human functional skeletal and cardiac tissues, through the application of innovative microscale techniques to standard cell culture devices. The strategy employed is biomimetic and multidisciplinary: the cell culture microenvironment has been engineered in order to reproduce in vitro the major stimuli that guide muscle cell differentiation in vivo. The coupling of tools and methodologies of the tissue engineering with innovative microscale technologies developed by engineers of the laboratory has led to a precise control of the cell microenvironment. In particular, the chemical-physical properties of the substrates, the topologic organization of cells and the application of exogenous electrical stimuli were optimized during this work. The developed devices were used with cell culture of particular interest, such as human primary dystrophic myoblasts and human embryonic stem cells (hESC). The functional analysis of the obtained skeletal muscle tissue, both dystrophic and healthy, highlighted a more rapid differentiation process of myoblasts cultured with the innovative techniques in comparison to standard cell culture systems. Electrical stimulation was applied to hESC in order to favor the cardiac differentiation pathway and an array of human beating cardiomyocytes has been developed.Taken together these results open new promising prospective for the development of in vitro model for preclinical trials of drugs or therapies for the treatment of pathologies of skeletal and cardiac muscles.

Abstract (italiano)

L’utilizzo di tessuti ingegnerizzati come modelli in vitro è recentemente emerso come applicazione alternativa al loro tradizionale impianto in vivo. Lo sviluppo di nuovi farmaci e terapie, nel caso di malattie quali la Distrofia Muscolare di Duchenne (DMD) o l’infarto miocardico (HF), sono infatti fortemente limitati e rallentati della mancanza di adeguati modelli in vitro: rappresentativi del tessuto umano e delle sue proprietà funzionali, di semplice utilizzo ed accessibili economicamente. Prerequisiti necessari all’ottenimento di un modello che risponda a tali esigenze sono: l’utilizzo di una fonte cellulare umana primaria; l’impiego di tecniche micrometriche, in primo luogo per il fine controllo del microambiente e del conseguente differenziamento cellulare in tessuto funzionale e per la riduzione dei costi di ricerca. Lo sviluppo su microscala permette inoltre di effettuare sperimentazioni multiparametriche, essenziali per patologie multifattoriali quali DMD e HF, con un elevato numero di dati in uscita. Infine, l’utilizzo di metodologie e tecniche semplici permette il trasferimento tra laboratori e ricercatori di diversa formazione. Lo scopo di questa tesi è stato quindi l’ottenimento di tessuti umani funzionali di muscolo scheletrico e cardiaco mediante l’utilizzo di tecniche di microscala, al fine di soddisfare le attuali esigenze di ricerca. È stato utilizzato un approccio biomimetico e multidisciplinare: un’accurata ingegnerizzazione del microambiente cellulare ha permesso di riprodurre in vitro i principali stimoli che in vivo guidano la differenziazione cellulare, così da ottenere un tessuto funzionale e rappresentativo del tessuto naturale. Le metodologie classiche dell’ingegneria dei tessuti sono state accoppiate ad innovative tecnologie di microscala, precedentemente sviluppate dagli ingegneri del laboratorio, per il preciso controllo dell’ambiente cellulare a livello micrometrico. In particolare sono state ottimizzate le proprietà chimico-fisiche del substrato, l’organizzazione topologica delle colture e l’applicazione di stimolazione elettrica esogena. Tali sistemi sono stati utilizzati con colture di particolare interesse quali mioblasti umani distrofici e cellule staminali embrionali umane (hESC). L’analisi funzionale del tessuto scheletrico umano così ottenuto, sia distrofico che sano, ha evidenziato come i sistemi sviluppati siano in grado di indurre un processo differenziativo più rapido rispetto ai tradizionali metodi di coltura. È stato valutato l’effetto di stimolazioni elettriche esogene sul differenziamento cardiomiocitario di hESC ed è stato sviluppato un array di cardiomiociti umani contrattili. Tali risultati aprono promettenti prospettive per lo sviluppo di modelli in vitro che permettano lo screening preclinico di nuovi farmaci o nuove terapie per la cura delle patologie a carico della muscolatura scheletrica e cardiaca.

Statistiche Download - Aggiungi a RefWorks
Tipo di EPrint:Tesi di dottorato
Relatore:Gamba, Piergiorgio
Correlatore:Elvassore, Nicola - Vunjak-Novakovic, Gordana
Dottorato (corsi e scuole):Ciclo 21 > Scuole per il 21simo ciclo > BIOLOGIA E MEDICINA DELLA RIGENERAZIONE > INGEGNERIA DEI TESSUTI E DEI TRAPIANTI
Data di deposito della tesi:01 Febbraio 2009
Anno di Pubblicazione:01 Febbraio 2009
Parole chiave (italiano / inglese):skeletal muscle tissue engineering, cardiac tissue engineering, human embryonic stem cell, Duchenne Muscular Dystrophy, microscale technologies, in vitro model, biomaterials, bioreactors
Settori scientifico-disciplinari MIUR:Area 09 - Ingegneria industriale e dell'informazione > ING-IND/24 Principi di ingegneria chimica
Struttura di riferimento:Dipartimenti > pre 2012 - Dipartimento di Principi e Impianti di Ingegneria Chimica "I. Sorgato"
Codice ID:1392
Depositato il:01 Feb 2009
Simple Metadata
Full Metadata
EndNote Format

Bibliografia

I riferimenti della bibliografia possono essere cercati con Cerca la citazione di AIRE, copiando il titolo dell'articolo (o del libro) e la rivista (se presente) nei campi appositi di "Cerca la Citazione di AIRE".
Le url contenute in alcuni riferimenti sono raggiungibili cliccando sul link alla fine della citazione (Vai!) e tramite Google (Ricerca con Google). Il risultato dipende dalla formattazione della citazione.

1 Bach AD, Beier JP, Stern-Staeter J and Horch RE. Skeletal muscle tissue engineering. Journal of Cell. Mol. Med 2004; 8(4):413-422 Cerca con Google

2 Partridge TA. Invited review: myoblast transfer: a possible therapy for inherited myopathies? Muscle Nerve 1991; 14(3):197-212 Cerca con Google

3 Huard J LY, Fu F. H. Muscle Injuries and Repair: Current Trends in Research. J. Bone Joint Surg. Am. 2002; 84(822-832 Cerca con Google

4 Carrier RL, Papadaki M, Rupnick M, Schoen FJ, Bursac N, Langer R, et al. Cardiac tissue engineering: Cell seeding, cultivation parameters, and tissue construct characterization. Biotechnology and Bioengineering 1999; 64(5):580-589 Cerca con Google

5 Okano T and Matsuda T. Muscular tissue engineering: capillary-incorporated hybrid muscular tissues in vivo tissue culture. Cell Transplantation 1998; 7(5):435-442 Cerca con Google

6 Freed LE and Vunjak-Novakovic G. Culture of organized cell communities. Advanced Drug Delivery Reviews 1998; 33(1-2):15-30 Cerca con Google

7 Hill E, Boontheekul T and Mooney D. Designing Scaffolds to Enhance Transplanted Myoblast Survival and Migration. Tissue Engineering 2006; 12(5):1295-1304 Cerca con Google

8 De Coppi P, Delo D, Farrugia L, Udompanyanan K, Yoo JJ, Nomi M, et al. Angiogenic Gene-Modified Muscle Cells for Enhancement of Tissue Formation. Tissue Engineering 2005; 11(7-8):1034-1044 Cerca con Google

172 Appendix C Cerca con Google

9 Nomi M, Atala A, De Coppi P and Soker S. Principals of neovascularization for tissue engineering. Molecular Aspects of Medicine 2002; 23(6):463-483 Cerca con Google

10 Lewis MC, MacArthur BD, Malda J, Pettet G and Please CP. Heterogeneous proliferation within engineered cartilaginous tissue: the role of oxygen tension. Biotechnology and Bioengineering 2005; 91(5):607-615 Cerca con Google

11 Figallo E, Flaibani M, Zavan B, Abatangelo G and Elvassore N. Micropatterned Biopolymer 3D Scaffold for Static and Dynamic Culture of Human Fibroblasts. Biotechnol Prog 2007; 23(1):210-6 Cerca con Google

12 Cha JM, Park S-N, Park G-O, Kim JK and Suh H. Construction of Functional Soft Tissues From Premodulated Smooth Muscle Cells Using a Bioreactor System. Artificial Organs 2006; 30(9):704-707 Cerca con Google

13 Marolt D, Augst A, Freed LE, Vepari C, Fajardo R, Patel N, et al. Bone and cartilage tissue constructs grown using human bone marrow stromal cells, silk scaffolds and rotating bioreactors. Biomaterials 2006; 27(36):6138-6149 Cerca con Google

14 Papadaki M, Bursac N, Langer R, Merok J, Vunjak-Novakovic G and Freed LE. Tissue engineering of functional cardiac muscle: molecular, structural, and electrophysiological studies. Am J Physiol Heart Circ Physiol 2001; 280(1):H168-178 Cerca con Google

15 Radisic M, Park H, Shing H, Consil T, Schoen FJ, Langer R, et al. Functional assembly of engineered myocardium by electrical stimulation of cardiac myocytes cultured on scaffolds. Proc Natl Acad Sci U S A 2004; 101(52):18129-34 Cerca con Google

16 Kuo CK, Li W-J, Mauck RL and Tuan RS. Cartilage tissue engineering: its potential and uses. Current Opinion in Rheumatology 2006; 18(1):64-73 Cerca con Google

17 Mansbridge J. Commercial considerations in tissue engineering. Journal of Anatomy 2006; 209(4):527-532 Cerca con Google

18 Freed LE, Hollander AP, Martin I, Barry JR, Langer R and Vunjak-Novakovic G. Chondrogenesis in a Cell-Polymer-Bioreactor System. Experimental Cell Research 1998; 240(1):58-65 Cerca con Google

19 Gooch KJ, Blunk T, Courter DL, Sieminski AL, Bursac PM, Vunjak-Novakovic G, et al. IGF-I and Mechanical Environment Interact to Modulate Engineered Cartilage Development. Biochemical and Biophysical Research Communications 2001; 286(5):909-915 Cerca con Google

20 Vunjak-Novakovic G, Obradovic B, Martin I, Bursac PM, Langer R and Freed LE. Dynamic Cell Seeding of Polymer Scaffolds for Cartilage Tissue Engineering. Biotechnol. Prog. 1998; 14(2):193-202 Cerca con Google

21 Bancroft GN, Sikavitsas VI and Mikos AG. Technical Note: Design of a Flow Perfusion Bioreactor System for Bone Tissue-Engineering Applications. Tissue Engineering 2003; 9(3):549-554 Cerca con Google

22 Cartmell SH, Porter BD, Garcia AJ and Guldberg RE. Effects of Medium Perfusion Rate on Cell-Seeded Three-Dimensional Bone Constructs in Vitro. Tissue Engineering 2003; 9(6):1197-1203 Cerca con Google

23 Bilodeau K and Mantovani D. Bioreactors for Tissue Engineering: Focus on Mechanical Constraints. A Comparative Review. Tissue Engineering 2006; 12(8):2367-2383 Cerca con Google

24 Torgan C, Burge S, Collinsworth A, Truskey G and Kraus W. Differentiation of mammalian skeletal muscle cells cultured on microcarrier beads in a rotating cell culture system. Med Biol Eng Comput 2000; 38(5):583-590 Cerca con Google

25 Carrier R, Rupnick M, Langer R, Schoen F, Freed L and Vunjak-Novakovic G. Perfusion Improves Tissue Architecture of Engineered Cardiac Muscle. Tissue Engineering 2002; 8(2):175-188 Cerca con Google

Perfusion bioreactor 173 Cerca con Google

26 Cha J, Park S-N, Park G-O, Kim J and Suh H. Construction of Functional Soft Tissues From Premodulated Smooth Muscle Cells Using a Bioreactor System. Artificial Organs 2006; 30(9):704-707 Cerca con Google

27 Asakura A, Komaki M and Rudnicki M. Muscle satellite cells are multipotential stem cells that exhibit myogenic, osteogenic, and adipogenic differentiation. Differentiation 2001; 68(4-5):245-253 Cerca con Google

28 Conconi MT, Coppi PD, Bellini S, Zara G, Sabatti M, Marzaro M, et al. Homologous muscle acellular matrix seeded with autologous myoblasts as a tissue-engineering approach to abdominal wall-defect repair. Biomaterials 2005; 26(15):2567-2574 Cerca con Google

29 De Coppi P, Bellini S, Conconi MT, Sabatti M, Simonato E, Gamba PG, et al. Myoblast-Acellular Skeletal Muscle Matrix Constructs Guarantee a Long-Term Repair of Experimental Full-Thickness Abdominal Wall Defects. Tissue Engineering 2006; 12(7):1929-1936 Cerca con Google

30 Partridge TA. Cells that participate in regeneration of skeletal muscle. Gene Therapy 2002; 9(11):752-753 Cerca con Google

31 Boldrin L, Elvassore N, Malerba A, Flaibani M, Cimetta E, Piccoli M, et al. Satellite Cells Delivered by Micro-Patterned Scaffolds: A New Strategy for Cell Transplantation in Muscle Diseases. Tissue Eng 2006; Cerca con Google

32 Portner R, Nagel-Heyer S, Goepfert C, Adamietz P and Meenen N. Bioreactor Design for Tissue Engineering. Journal of Bioscience and Bioengineering 2005; 100(3):235-245 Cerca con Google

33 Bird RB, Stewart WE and Lightfoot EN. Transport Phenomena. Second John Wiley & Sons, inc.; 2002 Cerca con Google

34 Bacabac RG, Smit T-H, Cowin SC, Loon JJWAV, Nieuwstadt FTM, Heethaar R, et al. Dynamic shear stress in parallel-plate flow chambers. Journal of Biomechanics 2005; 38(159-167 Cerca con Google

35 Gosgnach W, Messika-Zeitoun D, Gonzalez W, Philipe M and Michel J-B. Shear stress induces iNOS expression in cultured smooth muscle cells: role of oxidative stress. Am J Physiol Cell Physiol 2000; 279(6):C1880-1888 Cerca con Google

36 Fournier RL. Basic Transport Phenomena in Biomedical Engineering. Lillington, NC: Edwaed Brothers; 1998 Cerca con Google

37 Chow DC, Wenning LA, Miller WM and Papoutsakis ET. Modeling pO(2) distributions in the bone marrow hematopoietic compartment. II. Modified Kroghian models. Biophys J. 2001; 81(2):685-96 Cerca con Google

38 Salathe EP and Gorman AD. Modelling oxygen concentration in skeletal muscle. Mathematical computing modelling 1997; 26(4 Cerca con Google

39 Richardson RS. Oxygen transport and utilization: an integration of the muscle system. Advances in physiology education 2003; 27(4):183-191 Cerca con Google

40 Perry RH and Green DW. Perry’s chemical engineers’ handbook. Seventh Mc Graw-Hill International; 1998 Cerca con Google

41 Brandrup J, Immergat EH and Grulke EA. Polymer handbook. IV Wiley Interscience publication; 1999 Cerca con Google

42 Cussler EL. Diffusion: Mass Transfer in Fluid System. Second Cambridge University Press; 1997 Cerca con Google

43 Zhao F, Pathi P, Grayson W, Xing Q, Locke BR and Ma T. Effects of Oxygen Transport on 3-D Human Mesenchymal Stem Cell Metabolic Activity in Perfusion and Static Cultures: Experiments and Mathematical Model. Biotechnol. Prog. 2005; 21(4):1269-1280 Cerca con Google

174 Appendix C Cerca con Google

44 Sen A, Kallos MS and Behie LA. Expansion of mammalian neural stem cells in bioreactors: effect of power input and medium viscosity. Developmental Brain Research 2002; 134(103-113 Cerca con Google

45 Reutelingsperger CPM, Van Gool RGJ, Heijnen V, Frederik P and T. L. The rotating disc as a device to study the adhesive properties of endothelial cells under differential shear stresses. J. Matr Sci.: Mater. In Medicine 1993; 5(6-7):361-367 Cerca con Google

46 Desai TA. Micro- and nanoscale structures for tissue engineering constructs. Medical Engineering & Physics 2000; 22(9):595-606 Cerca con Google

47 Coletti F, Macchietto S and Elvassore N. Mathematical modeling of three-dimensional cell cultures in perfusion bioreactors. Industrial & Engineering Chemistry Research 2006; 45(24):8158-8169 Cerca con Google

48 Hill E, Boontheekul T and Mooney D. Regulating activation of transplanted cells controls tissue regeneration. Proc Natl Acad Sci U S A 2006; 103(8):2494-2499 Cerca con Google

49 Deasy B M LYaHJ. Tissue engineering with muscle-derived stem cells. Current Opinion in Biotechnology 2004; 15(419–423 Cerca con Google

50 Urish K KY, Huard J. Initial failure in myoblast transplantation therapy has led the way toward the isolation of muscle stem cells: potential for tissue regeneration. Current Topics in Developmental Biology 2005; 68(263-280 Cerca con Google

Download statistics

Solo per lo Staff dell Archivio: Modifica questo record