Adipose tissue engineering [Elektronische Ressource] : development of a 3-D model system of adipogenesis / vorgelegt von Claudia Fischbach

Claudia Fischbach

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• Gesundheit

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• Publié par: universitat_regensburg
• Publié le: 01 janvier 2004
• Nombre de lectures: 15
• Langue: Deutsch
• Poids de l'ouvrage: 12 Mo

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Adipose Tissue Engineering
Development of a 3-D Model System of
Adipogenesis
Dissertation zur Erlangung des Doktorgrades der Naturwissenschaften
(Dr. rer. nat.)
der Fakultät für Chemie und Pharmazie
der Universität Regensburg
vorgelegt von
Claudia Fischbach
aus Ichenhausen
im Juli 2003Diese Doktorarbeit entstand in der Zeit von Oktober 1998 bis Juni 2003 am Lehrstuhl für
Pharmazeutische Technologie an der Universität Regensburg
Die Arbeit wurde angeleitet von Prof. Dr. Achim Göpferich.
Promotionsgesuch eingereicht am: 9. Juli 2003
Datum der mündlichen Prüfung: 28. Juli 2003
Prüfungsausschuß: Vorsitzender: Prof. Dr. S. Elz
Erstgutachter: Prof. Dr. A. Göpferich
Zweitgutachter: PD Dr. J. Seufert.
Drittprüfer: Prof. Dr. G. FranzMeiner Familie in Liebe und Dankbarkeit gewidmet.
Wir sollen heiter Raum um Raum durchschreiten,
An keinem wie an einer Heimat hängen,
Der Weltgeist will nicht fesseln uns und engen,
Er will uns Stuf´ um Stufe heben, weiten.
(Hermann Hesse, Stufen)Adipose Tissue Engineering
Table of Contents
Chapter 1 Introduction and Goals of the Thesis 7
Chapter 2 Evaluation of Culture Conditions: Influence of Cell Culture Media and
Adipogenic Factors 35
Chapter 3 3-D in vitro-Model of Adipogenesis - Comparison of Culture
Conditions 61
Chapter 4 Analysis of Differential Areas within 3-D Tissue-Engineered Fat
Constructs 89
Chapter 5 Tissue Engineering Allows for Development of 3T3-L1 Cells into Fat
Pads in vitro and in vivo 111
Chapter 6 Does UV Irradiation Affect Polymer Properties Relevant to Tissue
Engineering? 133
Chapter 7 Adipogenesis on Different Polymeric Materials 155
Chapter 8 Summary and Conclusions 177
Appendix List of Abbreviations 187
Primer Sequences and PCR Conditions 189
Chemicals and Instruments 190
Curriculum Vitae 194
List of Publications 195
Acknowledgements 198
- 5 -Adipose Tissue Engineering
Chapter 1
Introduction
and
Goals of the ThesisChapter 1 Introduction and Goals of the Thesis
Tissue engineering
Background:
The loss or failure of an organ or tissue represents one of the most devastating and
costly problems in medicine [1-3]. Current strategies that have evolved to deal with these
defects include organ transplantation, surgical reconstruction, and replacement with
mechanical devices [3-5]. Despite incorporating significant advances, the applicability of
these strategies is substantially limited, especially in the field of transplantation. The
shortcomings associated with organ transplantation include a critical shortage of donor organs
and permanent immunosuppressive medication with its increased risk of adverse side effects
[5-7]. Surgical reconstruction equally suffers from scarcity of available donor tissue.
Furthermore, it often matches the particular reconstructive need only imperfectly and may
entail donor site morbidity [5,8]. Finally, infection, poor biocompatibility, and limited
durability complicate the use of artificial prostheses [3-5]. In order to overcome these
drawbacks, the interdisciplinary field of tissue engineering (TE) emerged. Motivated by the
challenge to develop biological substitutes that restore, maintain, or improve tissue functions,
TE combines the principles of engineering and the life sciences towards the production of
tissue substitutes [1,2,9].
Concept:
The most common approach to guiding tissue regeneration is the use of cell-matrix
constructs [5,7,10] (Fig. 1). This concept stemmed from biological observations reporting that
A) every tissue undergoes constant remodeling, B) isolated cells can reorganize themselves
into the desired tissue when placed in appropriate cell culture conditions, and C) appropriate
histological reorganization only occurs when cells are delivered within a template guiding
restructuring and allowing for diffusion of nutrients and oxygen to the cells [1]. Accordingly,
autologous cells isolated and expanded from a small tissue biopsy are seeded onto scaffolds,
which function to direct the development of the new tissue by providing structural support
and an appropriate three-dimensional (3-D) environment [1,6,8]. Usually, the scaffolds are
fashioned from either synthetic or natural biodegradable and biocompatible polymers that are
gradually be replaced by regenerated tissue, minimizing the inflammatory response [6,8].
Once the cells are adhered onto matrices and proliferate, cellular reorganization is triggered
by treatment with the appropriate tissue-inducing substances, e.g. hormones and growth
- 9 -Chapter 1 Introduction and Goals of the Thesis
factors [1,7]. Subsequent to the formation of the desired tissue, the cell-matrix constructs can
be used for transplantation purposes (Fig. 1). Furthermore, it has become increasingly
recognized that the tissue-like constructs supply valuable 3-D model systems for both basic
research in vitro and in vivo (Fig. 1). Compared to conventional two-dimensional (2-D) cell
culture, the engineered tissue equivalents may better reflect in vivo conditions and are,
therefore, thought to substantially contribute to the understanding of tissue-inherent functions
and processes.
tissue-inducing
substances
isolated,
expanded
cells Transplantationwell plate with
cell-polymer constructs
3-D matrix
In vitro
In vivoCell seeding Tissue formation Basic research
Fig. 1: Tissue engineering
Cell-matrix constructs for clinical application and basic research
Adipose tissue – Challenges for tissue engineering
The traditional role attributed to adipose tissue is the storage of triacylglycerol in times
of nutrient excess and the mobilization of energy during periods of caloric deprivation
[11,12]. However, over the past decades it has become increasingly acknowledged as a major
secretory and endocrine organ involved in a range of functions beyond simple fat storage [13-
15]. Due to its usefulness for plastic and reconstructive surgery and its impact on
metabolically related disorders, adipose tissue has gained substantial clinical and research
interest.
Plastic and reconstructive surgery:
In plastic and reconstructive surgery, autologous fat grafts serve as an appropriate filling
material in the reconstruction of soft tissue defects [16,17]. In particular, adipose tissue offers
the potential to act as a natural bulking material to treat congenital or acquired (traumatic and
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