Spelling suggestions: "subject:"fissue engineering"" "subject:"anissue engineering""
371 |
Development of a prevascularized bone implant / Entwicklung eines prävaskularisierten KnochenimplantatsRücker, Christoph January 2019 (has links) (PDF)
The skeletal system forms the mechanical structure of the body and consists of bone, which is hard connective tissue. The tasks the skeleton and bones take over are of mechanical, metabolic and synthetic nature. Lastly, bones enable the production of blood cells by housing the bone marrow. Bone has a scarless self-healing capacity to a certain degree. Injuries exceeding this capacity caused by trauma, surgical removal of infected or tumoral bone or as a result from treatment-related osteonecrosis, will not heal. Critical size bone defects that will not heal by themselves are still object of comprehensive clinical investigation. The conventional treatments often result in therapies including burdening methods as for example the harvesting of autologous bone material. The aim of this thesis was the creation of a prevascularized bone implant employing minimally invasive methods in order to minimize inconvenience for patients and surgical site morbidity. The basis for the implant was a decellularized, naturally derived vascular scaffold (BioVaSc-TERM®) providing functional vessel structures after reseeding with autologous endothelial cells. The bone compartment was built by the combination of the aforementioned scaffold with synthetic β-tricalcium phosphate. In vitro culture for tissue maturation was performed using bioreactor technology before the testing of the regenerative potential of the implant in large animal experiments in sheep. A tibia defect was treated without the anastomosis of the implant’s innate vasculature to the host’s circulatory system and in a second study, with anastomosis of the vessel system in a mandibular defect. While the non-anastomosed implant revealed a mostly osteoconductive effect, the implants that were anastomosed achieved formation of bony islands evenly distributed over the defect.
In order to prepare preconditions for a rapid approval of an implant making use of this vascularization strategy, the manufacturing of the BioVaSc-TERM® as vascularizing scaffold was adjusted to GMP requirements. / Das Skelett bildet die mechanische Struktur des Körpers und besteht aus Knochen, einem harten Bindegewebe. Knochen übernehmen mechanische, metabolische und synthetische Aufgaben. Schlussendlich ermöglichen Knochen die Synthese von Blutzellen durch die Beherbergung des Knochenmarks. Wird die Heilungskapazität von Knochen durch Trauma, operative Entfernung von infiziertem oder tumorösem Knochen oder als Ergebnis behandlungsbedingter Osteonekrose, überschritten, findet keine vollständige Heilung statt. Knochendefekte, die eine kritische Größe überschreiten, sind daher immer noch Gegenstand umfangreicher, klinischer Forschung. Bei herkömmlichen Behandlungsmethoden können Eingriffe notwendig werden, die den Patienten belasten, wie bei der Gewinnung von autologem Knochenmaterial. Das Ziel der vorliegenden Arbeit war die Herstellung eines prävaskularisierten Implantats unter Verwendung minimalinvasiver Methoden, um die Belastung von Patienten und die Morbidität an der Entnahmestelle, zu verringern. Zur Herstellung eines vaskularisierten Implantats bildete ein dezellularisiertes Darmsegment (Jejunum) porcinen Ursprungs die Grundlage (BioVasc-TERM®). Diese Trägerstruktur stellte ein funktionales Blutgefäßsystem nach Wiederbesiedelung mit autologen Endothelzellen bereit. Der Knochenanteil des Implantats wurde durch die Kombination der genannten Trägerstruktur mit dem synthetischen Knochenersatzmaterial β-Tricalciumphosphat gebildet. In-vitro-Kultivierung in einem Bioreaktor führte zur Reifung des Implantats vor der Testung seines Potenzials zur Knochenregeneration in Großtierversuchen bei Schafen. Ein Tibiadefekt wurde behandelt ohne die Anastomose des implantateigenen Gefäßsystems an den Blutkreislauf und ein Mandibeldefekt wurde mit Gefäßanschluss behandelt. Das Implantat ohne Gefäßanschluss hatte einen osteokonduktiven Effekt, während das anastomosierte Implantat zur Bildung zahlreicher Knocheninseln, gleichmäßig über den Defekt verteilt, führte. Um eine zügige Zulassung eines Implantats, das diese Strategie zur Vaskularisierung von Knochen nutzt, zu ermöglichen, wurde die Herstellung der BioVaSc-TERM® an die Vorgaben der Guten Herstellungspraxis angepasst.
|
372 |
Tissue Engineering of a Vascularized Meniscus Implant / Tissue Engineering eines vaskularisierten Meniskus-ImplantatesKremer, Antje January 2019 (has links) (PDF)
The knee joint is a complex composite joint containing the C-shaped wedge-like menisci composed of fibrocartilage. Due to their complex composition and structure, they provide mechanical resilience to the knee joint protecting the articular cartilage. Because of the limited repair potential, meniscal injuries do not only affect the meniscus itself but also lead to altered joint homeostasis and inevitably to secondary osteoarthritis.
The meniscus was characterized focusing on its anatomy, structure and meniscal markers such as aggrecan, collagen type I (Col I) and Col II. The components relevant for meniscus tissue engineering, namely cells, Col I scaffolds, biochemical and biomechanical stimuli were studied. Meniscal cells (MCs) were isolated from meniscus, mesenchymal stem cells (MSCs) from bone marrow and dermal microvascular endothelial cells (d-mvECs) from foreskin biopsies. For the human (h) meniscus model, wedge-shape compression of a hMSC-laden Col I gel was successfully established. During three weeks of static culture, the biochemical stimulus transforming growth factor beta-3 (TGF beta-3) led to a compact collagen structure. On day 21, this meniscus model showed high metabolic activity and matrix remodeling as confirmed by matrix metalloproteinases detection. The fibrochondrogenic properties were illustrated by immunohistochemical detection of meniscal markers, significant GAG/DNA increase and increased compressive properties. For further improvement, biomechanical stimulation systems by compression and hydrostatic pressure were designed. As one vascularization approach, direct stimulation with ciclopirox olamine (CPX) significantly increased sprouting of hd-mvEC spheroids even in absence of auxiliary cells such as MSCs. Second, a cell sheet composed of hMSCs and hd-mvECs was fabricated by temperature triggered cell sheet engineering and transferred onto the wedge-shaped meniscus model. Third, a biological vascularized scaffold (BioVaSc-TERM) was re-endothelialized with hd-mvECs providing a viable vascularized network. The vascularized BioVaSc-TERM was suggested as wrapping scaffold of the meniscus model by using two suture techniques, the all-inside-repair (AIR) for the posterior horn, and the outside-in-refixation (OIR) for the anterior horn and the middle part.
This meniscus model for replacing torn menisci is a promising approach to be further optimized regarding vascularization, biochemical and biomechanical stimuli. / Das Knie ist ein komplex zusammengesetztes Gelenk mit zwei C-förmigen Keilen aus Bindegewebsknorpel, die Menisken. Sie sorgen für die mechanische Belastbarkeit des Knies, wodurch der Gelenksknorpel geschützt wird. Aufgrund des limitierten Heilungspotentials beeinträchtigen Meniskusverletzungen nicht nur den Meniskus selbst, sondern schädigen auch das Gelenksgleichgewicht und führen zu sekundärer Osteoarthritis.
Der Meniskus wurde in seiner Anatomie, Struktur und Meniskusmarkern wie Aggrekan, Kollagen I und Kollagen II charakterisiert. Die Komponenten von Meniskus Tissue Engineering, Zellen, Kollagen I Materialien, biochemische und biomechanische Stimuli wurden untersucht. Meniskuszellen (MCs) wurden aus Meniskus isoliert, mesenchymale Stammzellen (MSCs) aus Knochenmark und dermale mikrovaskuläre Endothelzellen (d-mvECs) aus Vorhautbiopsien. Für das humane (h) Meniskus-Modell wurde die keilförmige Kompression eines hMSC-beladenen Kollagen I Gels erfolgreich etabliert. Während drei Wochen statischer Kultur führte der biochemische Stimulus transformierender Wachs-tumsfaktor beta-3 (TGF beta-3) zu einer kompakten Kollagenstruktur. An Tag 21 zeigte dieses Meniskus-Modell eine hohe metabolische Aktivität und Matrixumbau durch die Detektion von Matrix-Metalloproteasen. Der Bindegewebsknorpel wurde durch immunhistochemische Detektion der Meniskusmarker, einem signifikanten GAG/DNA Anstieg und erhöhter Kompressionseigenschaften bestätigt. Für weitere Verbesserungen wurden biomechanische Stimulierungssysteme mittels Kompression und hydrostatischen Druck aufgebaut. Als Vaskularisierungsansatz führte die direkte Stimulierung mit Ciclopirox Olamine (CPX) sogar in Abwesenheit von Helferzellen wie MSCs zu einem erhöhten Sprouting der hd-mvEC Spheroide. Zweitens wurde ein hMSC/hd-mvEC Sheet mithilfe eines Temperatur-abhängigen Verfahrens produziert und auf das keilförmige Meniskus-Modell transferiert. Drittens wurde ein vaskularisiertes Biomaterial (BioVaSc-TERM) mit hd-mvECs besiedelt, wodurch ein vitales Gefäßystem bereitgestellt wurde. Die vaskularisierte BioVaSc-TERM wurde als Hülle des Meniskus-Modells unter der Verwendung von zwei Nahttechniken vorgeschlagen: die All-Inside-Repair (AIR) für das Hinterhorn und die Outside-In-Refixation (OIR) für das Vorderhorn und den mittleren Teil.
Dieses Meniskus-Modell ist ein vielversprechender Ansatz für den Meniskusersatz, um in Vaskularisierung, biochemischer und biomechanischer Stimuli weiter optimiert zu werden.
|
373 |
Entwicklung und Charakterisierung von Gelatine-basierten Hydrogelen und PLGA-basierten Janus-Partikeln / Development and characterization of gelatin-based hydrogels and PLGA-based Janus particlesSchönwälder, Sina Maria Siglinde January 2016 (has links) (PDF)
Zusammenfassung
In der Regenerativen Medizin sind polymerbasierte Biomaterialien von großer Bedeutung für
die Entwicklung und Anwendung verbesserter bzw. neuer Therapien. Die Erforschung der
Oberflächeneigenschaften von Biomaterialien, welche als Implantate eingesetzt werden, ist
eine grundlegende Voraussetzung für deren erfolgreichen Einsatz. Die Protein-Oberflächen-
Interaktion geschieht initial, sobald ein Implantat mit Körperflüssigkeiten oder mit Gewebe
in Kontakt kommt, und trägt maßgeblich zur direkten Wechselwirkung von Implantat und
umgebenden Zellen bei. Dieser Prozess wird in der vorliegenden Arbeit an Gelatine untersucht.
Daher bestand ein Ziel darin, stabile, nanometerdünne Gelatineoberflächen herzustellen
und darauf die Adsorption von humanen Plasmaproteinen und bakteriellen Proteinen zu
analysieren.
Die Abscheidung der Gelatinefilme in variabler Schichtdicke auf zuvor mit PPX-Amin modifizierten
Oberflächen wurde unter Verwendung eines Rotationsbeschichters durchgeführt.
Um stabile Hydrogelfilme zu erhalten, wurden die Amingruppen der disaggregierten Gelatinefibrillen
untereinander und mit denen der Amin-Modifizierung durch ein biokompatibles
Diisocyanat quervernetzt. Dieser Prozess lieferte einen reproduzierbaren und chemisch stabilen
Gelatinefilm, welcher durch die substratunabhängige Amin-Modifizierung kovalent auf
unterschiedlichste Oberflächen aufgebracht werden konnte. Die durch den Herstellungsprozess
präzise eingestellte Schichtdicke (Nano- bzw. Mikrometermaßstab) wurde mittels Ellipsometrie
und Rasterkraftmikroskopie ermittelt. Die ebenso bestimmte Rauheit war unabhängig
von der Schichtdicke sehr gering. Gelatinefilme, die auf funktionalisierte und strukturierte
Proben aufgebracht wurden, konnten durch Elektronenmikroskopie dargestellt werden. Mit
Hilfe der Infrarot-Reflexions-Absorptions-Spektroskopie wurden die Gelatinefilme im Hinblick
auf ihre Stabilität chemisch charakterisiert. Zur Quantifizierung der Adsorption humaner
Plasmaproteine (Einzelproteinlösungen) und komplexer Proteingemische aus steril filtrierten
Kulturüberständen des humanpathogenen Bakteriums Pseudomonas aeruginosa wurde die
Quarzkristall-Mikrowaage mit Dissipationsüberwachung eingesetzt. Hiermit konnte nicht
nur die adsorbierte Menge an Proteinen auf dem Gelatinehydrogel bzw. Referenzoberflächen
(Gold, PPX-Amin, Titan), sondern auch die viskoelastischen Eigenschaften des adsorbierten
Proteinfilms bestimmt werden. Allgemein adsorbierte auf dem Gelatinehydrogel eine geringere
Proteinmasse im Vergleich zu den Referenzoberflächen. Circa ein Viertel der adsorbierten
Proteine migrierte in die Poren des gequollenen Gels und veränderte dessen viskoelastische
Eigenschaften. Durch anschließende MALDI-ToF/MS- und MS/MS-Analyse konnten die bakteriellen
Proteine auf den untersuchten Oberflächen identifiziert und untereinander verglichen
werden. Hierbei zeigten sich nur geringfügige Unterschiede in der Proteinzusammensetzung.
Zudem wurde eine Sekundärionenmassenspektrometrie mit Flugzeitanalyse an reinen Gelatinefilmen
und an mit humanen Plasmaproteinen beladenen Gelatinefilmen durchgeführt.
Durch eine anschließende multivariante Datenanalyse konnte zwischen den untersuchten
Proben eindeutig differenziert werden. Dieser Ansatz ermöglicht es, die Adsorption von
unterschiedlichen Proteinen auf proteinbasierten Oberflächen markierungsfrei zu untersuchen
und kann zur Aufklärung der in vivo-Situation beitragen. Darüber hinaus bietet dieser
Untersuchungsansatz neue Perspektiven für die Gestaltung und das schnelle und effiziente
Screening von unterschiedlichen Proteinzusammensetzungen.
Biomaterialien können jedoch nicht nur als Implantate oder Implantatbeschichtungen eingesetzt
werden. Im Bereich des drug delivery und der Depotarzneimittel sind biologisch
abbaubare Polymere, aufgrund ihrer variablen Eigenschaften, von großem Interesse. Die
Behandlung von bakteriellen und fungalen Pneumonien stellt insbesondere bei Menschen mit
Vorerkrankungen wie Cystische Fibrose oder primäre Ziliendyskinesie eine große Herausforderung
dar. Oral oder intravenös applizierte Wirkstoffe erreichen die Erreger aufgrund der
erhöhten Zähigkeit des Bronchialsekretes oft nicht in ausreichender Konzentration. Daher
besteht ein weiteres Ziel der vorliegenden Arbeit darin, mittels electrohydrodynamic cojetting
mikrometergroße, inhalierbare, wirkstoffbeladene Partikel mit zwei Kompartimenten
(Janus-Partikel) herzustellen und deren Eignung für die therapeutische Anwendung bei
Lungeninfektionen zu untersuchen.
Durch das in dieser Arbeit entwickelte Lösungsmittelsystem können Janus-Partikel aus
biologisch abbaubaren Co-Polymeren der Polymilchsäure (Poly(lactid-co-glycolid), PLGA)
hergestellt und mit verschiedenen Wirkstoffen beladen werden. Darunter befinden sich ein
Antibiotikum (Aztreonam, AZT), ein Antimykotikum (Itraconazol, ICZ), ein Mukolytikum
(Acetylcystein, ACC) und ein Antiphlogistikum (Ibuprofen, IBU). Die Freisetzung der eingelagerten
Wirkstoffe, mit Ausnahme von ICZ, konnte unter physiologischen Bedingungen
mittels Dialyse und anschließender Hochleistungsflüssigkeitschromatographie gemessen werden.
Die Freisetzungsrate wird von der Kettenlänge des Polymers beeinflusst, wobei eine
kürzere Kettenlänge zu einer schnelleren Freisetzung führt. Das in die Partikel eingelagerte
Antimykotikum zeigte in vitro eine gute Wirksamkeit gegen Aspergillus nidulans. Durch das
Einlagern von ICZ in die Partikel ist es möglich diesen schlecht wasserlöslichen Wirkstoff in
eine für Patienten zugängliche und wirksame Applikationsform zu bringen. In Interaktion mit
P. aeruginosa erzielten die mit Antibiotikum beladenen Partikel in vitro bessere Ergebnisse
als der Wirkstoff in Lösung, was sich in einem in vivo-Infektionsmodell mit der Wachsmotte
Galleria mellonella bestätigte. AZT-beladene Partikel hatten gegenüber einer identischen
Wirkstoffmenge in Lösung eine 27,5% bessere Überlebensrate der Wachsmotten zur Folge.
Des Weiteren hatten die Partikel keinen messbaren negativen Einfluss auf die Wachsmotten.
Dreidimensionale Atemwegsschleimhautmodelle, hergestellt mit Methoden des Tissue Engineerings,
bildeten die Basis für Untersuchungen der Partikel in Interaktion mit humanen
Atemwegszellen. Die Untersuchung von Apoptose- und Entzündungsmarkern im Überstand
der 3D-Modelle zeigte diesbezüglich keinen negativen Einfluss der Partikel auf die humanen
Zellen. Diese gut charakterisierten und standardisierten in vitro-Testsysteme machen es
möglich, Medikamentenuntersuchungen an menschlichen Zellen durchzuführen. Hinsichtlich
der histologischen Architektur und funktionellen Eigenschaften der 3D-Modelle konnte eine
hohe in vitro-/in vivo-Korrelation zu menschlichem Gewebe festgestellt werden. Humane
Mucine auf den 3D-Modellen dienten zur Untersuchung der schleimlösenden Wirkung von
ACC-beladenen Partikeln. Standen diese in räumlichem Kontakt zu den Mucinen, wurde deren
Zähigkeit durch das freigesetzte ACC herabgesetzt, was qualitativ mittels histologischen
Methoden bestätigt werden konnte.
Die in dieser Arbeit entwickelten Herstellungsprotokolle dienen als Grundlage und können
für die Synthese ähnlicher Systeme, basierend auf anderen Polymeren und Wirkstoffen,
modifiziert werden. Gelatine und PLGA erwiesen sich als vielseitig einsetzbare Werkstoffe
und bieten eine breite Anwendungsvielfalt in der Regenerativen Medizin, was die erzielten
Resultate bekräftigen. / In the field of regenerative medicine, polymer-based biomaterials are of great importance for the
development and application of improved or new therapies. The research on the surface properties of
biomaterials, which are used as implants, is essential for their successful use. The
protein-surface interaction is the initial step and occurs when an implant comes into contact with
bodily fluids or tissues and significantly increases direct interaction of the implant and the
surrounding cells. This thesis investigates these processes on gelatin. Accordingly, one of the
project’s major goals was to produce stable nanometer-thin gelatin surfaces and analyze the
adsorption of human plasma and bacterial proteins.
The deposition of gelatin films and the assortment of layer thicknesses on PPX-amine modified
surfaces were carried out using a spin coater. To gain hydrogel films with reproducible
properties, the amine groups of the disaggregated gelatin fibrils were cross- linked with each
other and with those of the amine modification by a biocompatible diisocyanate. The result was a
reproducible and chemically stable gelatin film, which could be applied to a wide variety of
surfaces through the substrate-independent amine modification. The manufacturing process precisely
adjusted the layer thickness to the nano- or micrometer scale which could be determined applying
ellipsometry and atomic- force microscopy. The roughness was very low regardless of the layer
thickness. Gelatin films applied to the functionalized and patterned samples could be visualized by
electron microscopy. With the help of infrared reflection absorption spectroscopy, the gelatin
films were chemically characterized in terms of stability. The adsorption of human plasma proteins
(single protein solutions) as well as the complex protein mixtures of sterile filtered supernatants
belonging to Pseudomonas aeruginosa, a human pathogenic bacterium, were quantified by quartz
crystal microbalance with dissipation monitoring. Both the adsorbed amount of proteins on the
gelatin hydrogel or reference surfaces (gold, PPX-amine, titanium) and the viscoelastic properties
of the adsorbed protein film were determined. In general, there was less protein mass adsorbed on
the gelatin hydrogel compared to the reference surfaces. About a quarter of the adsorbed proteins
migrated into the pores of the swollen gel and changed its viscoelastic properties. Subsequent
MALDI-ToF/MS and MS/MS analysis were used to identify and compare the adsorbed bacterial proteins
on the investigated surfaces. Only slight differences were found in the adsorbed protein
composition. A secondary ion mass spectrometry with time-of-flight analysis was performed on pure
gelatin films and gelatin films loaded with human plasma proteins. By subsequent multivariate data
analysis, it was possible to clearly differentiate between the examined samples. Not only does this
approach enable us to screen the adsorption of different proteins on protein-based surfaces without
labeling, but it also contributes to the elucidation of the in vivo-situation. ach provides new
perspectives regarding the design and efficient
screening of different protein compositions. ...
|
374 |
In vitro and in vivo bone formation - assessment and applicationChen, Jinbiao, Prince of Wales Clinical School, UNSW January 2006 (has links)
Background: Bone-grafting materials are required in orthopaedic surgery to treat bone defects. Bone formation assessment is required for the development of new strategies and approaches and for quality assurance and quality control of currently available materials. Approaches to the assessment of bone formation are yet to be systematically established, quantified and standardized. Aims: the overall aim of this study was to establish a set of comprehensive quantitative approaches for the assessment of bone formation and to evaluate the role of osteoblastic cells, growth factors, and scaffolds on this process. Materials & methods: both in vitro and in vivo parameters for osteoblast phenotype and bone formation were tested in osteosarcoma cell lines, Saos-2 and U2OS cells, mesenchymal cell line, C2C12 cells, primary adipose derived stromal cells (ADSCs), platelet rich plasma (PRP), and morselized bone grafts. The in vitro parameters used were measurement of alkaline phosphatase (ALP) activity, detection of bone nodules and biomineralization, and quantification of immunocytochemistry and conventional RT-PCR of osteoblast genotyping. In vivo parameters involved ectopic bone formation in nude mice and nude rats and a tibial defect model in nude rats. Histomorphometric and quantitative immunohistochemical analyses were also performed. Results: The in vitro characterization and ectopic bone formation capabiltity of Saos-2 and U2OS cells have been established. Saos-2 cell line, which presents many osteoblast genotype and phenotype, is a stable positive control for both in vitro and in vivo bone formation assessments. The measurement of ALP activity in both solid and liquid phases has been standardized. Both the genotype and phenotype of osteoblast lineage cells has been quantitatively assessed during the capability testing of ADSCs and PRP. Quantitative assessment of new bone formation and related protein markers in vivo has been successfully established through the testing of the biological properties of gamma irradiated morselized bone grafts. Conclusion: A comprehensive knowledge of the assessment of bone regeneration and formation in vitro and in vivo has been integrated and developed through years of study. A whole set of in vitro and in vivo approaches for the assessment of bone formation has been modified and standardized to best suit the different clinical applications. This thesis provides an outline of both in vitro and in vivo bone formation assessment and their clinical applications.
|
375 |
Design, Development, and Optimisation of a Culture Vessel System for Tissue Engineering ApplicationsDamen, Bas Stefaan, bsdamen@hotmail.com January 2003 (has links)
A Tissue Engineering (TE) approach to heart valve replacement has the aim of producing an implant that is identical to healthy tissue in morphology, function and immune recognition. The aim is to harvest tissue from a patient, establish cells in culture from this tissue and then use these cells to grow a new tissue in a desired shape for the implant. The scaffold material that supports the growth of cells into a desired shape may be composed of a biodegradable polymer that degrades over time, so that the final engineered implant is composed entirely of living tissue. The approach used at Swinburne University was to induce the desired mechanical and functional properties of tissue and is to be developed in an environment subjected to flow stresses that mimicked the haemodynamic forces that natural tissue experiences. The full attainment of natural biomechanical and morphological properties of a TE structure has not as yet been demonstrated.
In this thesis a review of Tissue Engineering of Heart Valves (TEHVs) is presented followed by an assessment of biocompatible materials currently used for TEHVs. The thrust of the work was the design and development of a Bioreactor (BR) system, capable of simulating the corresponding haemodynamic forces in vitro so that long-term cultivation of TEHVs and/or other structures can be mimicked. A full description of the developed BR and the verification of its functionality under various physiological conditions using Laser Doppler Anemometry (LDA) are given. An analysis of the fluid flow and shear stress forces in and around a heart valve scaffold is also provided.
Finally, preliminary results related to a fabricated aortic TEHV-scaffold and the developed cell culture systems are presented and discussed. Attempts to establish viable cell lines from ovine cardiac tissue are also reported.
|
376 |
Feasibility study of selective laser sintering of biopolymer scaffolds for tissue engineeringLee, Siu-hang, Sherman, January 2006 (has links)
Thesis (M. Phil.)--University of Hong Kong, 2007. / Title proper from title frame. Also available in printed format.
|
377 |
Innovative Methods to Determine Material Properties of Cartilaginous Tissues and Application for Tissue EngineeringYuan, Tai-Yi 21 July 2011 (has links)
Low back pain is one of the major health concerns in the US. It affects up to 80% of the population at some time during their lives. It not only causes discomfort to patients and affects their physical ability but also has a huge economic impact on society. Although the cause of low back pain is still poorly understood, it is implicated that degeneration of the intervertebral disc is the primary factor. Currently, researchers are trying to use tissue engineering approaches to develop new treatments capable of removing the degenerated disk and replacing it with a biological substitute. However, to create such a biological substitute, we need to first understand the structure-function relationship of the tissue. Only when we understand the function of the tissue, can we begin creating biological substitutes. While culturing a biological substitute, we also need methods to determine how the substitute responds to its environment. At present, there are many different types of bioreactors developed for cartilaginous tissues. However, there is a lack of a system that can detect the chemical, electrical and mechanical response noninvasively with control feedback in real-time. It is hard to provide the optimal culture environment to the substitute without knowing its response in real-time. The objective of this dissertation is to develop new methods to investigate the transport property, oxygen consumption rate and mechano-electrochemical and mechanical properties of the tissue. Because cells are responsible for the tissue health, it is necessary to understand how they can obtain nutrients under different environments, e.g. under different loading condition. In addition, with the use of a bioreactor with the capability of detecting the real-time response combined with a feedback control system, we can provide the most favorable conditions for tissue or biological substitutes to grow. The new measurement methods developed in this dissertation can contribute to further understanding the function of the tissue. The methods outlined in this dissertation can also provide new tools for future tissue engineering applications. Moreover, the findings in this dissertation can provide information for developing a more comprehensive theoretical model to elucidate the etiology of disc degeneration.
|
378 |
In vitro and in vivo studies of tissue engineering in reconstructive plastic surgeryHuss, Fredrik R.M. January 2005 (has links)
To correct, improve, and maintain tissues, and their functions, are common denominators in tissue engineering and reconstructive plastic surgery. This can be achieved by using autolo-gous tissues as in flaps or transplants. However, often autologous tissue is not useable. This is one of the reasons for the increasing interest among plastic surgeons for tissue engineering, and it has led to fruitful cross-fertilizations between the fields. Tissue engineering is defined as an interdisciplinary field that applies the principles of engineering and life sciences for development of biologic substitutes designed to maintain, restore, or improve tissue functions. These methods have already dramatically improved the possibilities to treat a number of medical conditions, and can arbitrarily be divided into two main principles: > Methods where autologous cells are cultured in vitro and transplanted by means of a cell suspension, a graft, or in a 3-D biodegradable matrix as carrier. > Methods where the tissue of interest is stimulated and given the right prerequisites to regenerate the tissue in vivo/situ with the assistance of implantation of specially designed materials, or application of substances that regulate cell functions - guided tissue regeneration. We have shown that human mammary epithelial cells and adipocytes could be isolated from tissue biopsies and that the cells kept their proliferative ability. When co-cultured in a 3-D matrix, patterns of ductal structures of epithelial cells embedded in clusters of adipocytes, mimicking the in vivo architecture of human breast tissue, were seen. This indicated that human autologous breast tissue can be regenerated in vitro. The adipose tissue is also generally used to correct soft tissue defects e.g. by autologous fat transplantation. Alas 30-70% of the transplanted fat is commonly resorbed. Preadipocytes are believed to be hardier and also able to replicate, and hence, are probably more useful for fat transplantation. We showed that by using cell culture techniques, significantly more pre-adipocytes could survive and proliferate in vitro compared to two clinically used techniques of fat graft handling. Theoretically, a biopsy of fat could generate enough preadipocytes to seed a biodegradable matrix that is implanted to correct a defect. The cells in the matrix will replicate at a rate that parallels the vascular development, the matrix subsequently degrades and the cell-matrix complex is replaced by regenerated, vascularized adipose tissue. We further evaluated different biodegradable scaffolds usable for tissue engineering of soft tissues. A macroporous gelatin sphere showed several appealing characteristics. A number of primary human ecto- and mesodermal cells were proven to thrive on the gelatin spheres when cultured in spinner flasks. As the spheres are biodegradable, it follows that the cells can be cultured and expanded on the same substrate that functions as a transplantation vehicle and scaffold for tissue engineering of soft tissues. To evaluate the in vivo behavior of cells and gelatin spheres, an animal study was performed where human fibroblasts and preadipocytes were cultured on the spheres and injected intra-dermally. Cell-seeded spheres were compared with injections of empty spheres and cell suspensions. The pre-seeded spheres showed a near complete regeneration of the soft tissues with neoangiogenesis. Some tissue regeneration was seen also in the ‘naked’ spheres but no effect was shown by cell injections. In a human pilot-study, intradermally injected spheres were compared with hyaluronan. Volume-stability was inferior to hyaluronan but a near complete regeneration of the dermis was proven, indicating that the volume-effect is permanent in contrast to hyaluronan which eventually will be resorbed. Further studies are needed to fully evaluate the effect of the macroporous gelatin spheres, with or without cellular pre-seeding, as a matrix for guided tissue regeneration. However, we believe that the prospect to use these spheres as an injectable, 3D, biodegradable matrix will greatly enhance our possibilities to regenerate tissues through guided tissue regeneration. / On the day of the defence date the status of article V was In Press.
|
379 |
Bioactive Poly(ethylene glycol)-based Hydrogels for Angiogenesis in Tissue EngineeringJanuary 2011 (has links)
Because engineered tissue constructs are inherently limited by their lack of microvascularization, which is essential to provide oxygen for cell survival, this thesis presents rationally designed materials and cell culture techniques capable of supporting functional tubule formation and stabilization. Combining a synthetic scaffold material with cells and their cell-secreted signals instigated tubule formation throughout the scaffold. Poly(ethylene glycol) (PEG) based hydrogels, biocompatible polymers which resist protein adsorption and subsequent nonspecific cellular adhesion, were modified to induce desired cell characteristics. Human umbilical vein endothelial cells were used as a reproducible and readily available cell type. Several tubule-stabilization signals, including platelet derived growth factor-BB (PDGF-BB) and ephrinA1, were covalently immobilized via conjugation to PEG to enable prolonged bioactive signaling and controlled local delivery. All hydrogels were further tested in a mouse cornea micropocket angiogenesis assay, a naturally avascular tissue for easy imaging in a reproducible and quantifiable assay. Hydrogels containing soluble growth factors induced vessel formation in the hydrogel, and the resulting vessel morphology was modulated using different growth factor concentrations. Immobilized PDGF-BB led to tubule formation in two dimensions, three dimensions, and in the mouse cornea while immobilized ephrinA1 stimulated secretion of extracellular matrix proteins laminin and collagen IV to stabilize the newly formed tubules. Finally, a co-culture of endothelial and pericyte cells encapsulated into hydrogels formed tubules that anastomosed to the host vasculature and contained red blood cells. PEG-based hydrogels represent a promising technique to induce microvascular formation in engineered constructs, leading to stable and functional vessel formation using covalently immobilized growth factors and encapsulated cells. These materials can be used for replacement of damaged or diseased tissues as the current supply of cadaveric donations cannot meet the demand of tissues for the 110,000 people awaiting an organ in the US.
|
380 |
Coating Collagen Modules with Fibronectin Increases in vivo HUVEC Survival and Vessel Formation through the Suppression of ApoptosisCooper, Thomas 13 January 2010 (has links)
Modular tissue engineering is a novel approach to creating scalable, self-assembling three-dimensional tissue constructs with inherent vascularisation. Under initial methods, the subcutaneous implantation of human umbilical vein endothelial cell (HUVEC)-covered collagen modules in immunocompromised mice resulted in significant host inflammation and limited HUVEC survival. Subsequently, a minimally-invasive injection technique was developed to minimize surgery-related inflammation, and cell death was attributed to extensive apoptosis within 72 hours of implantation. In confirmation of in vitro results, coating collagen modules with fibronectin (Fn) was shown in vivo to reduce short-term HUVEC apoptosis by nearly 40%, while increasing long-term HUVEC survival by 30% to 45%. Consequently, a 100% increase in the number of HUVEC-lined vessels was observed with Fn-coated modules, as compared to collagen-only modules, at 7 and 14 days post-implantation. Furthermore, vessels appeared to be perfused with host erythrocytes by day 7, and vessel maturation and stabilization was evident by day 14.
|
Page generated in 0.074 seconds