Design of artificial modular extracellular matrices [Elektronische Ressource] / vorgelegt von: Stefan V. W. Gräter

Stefan Graeter

Sujets

• Biologie

Informations

• Publié par: ruprecht-karls-universitat_heidelberg
• Publié le: 01 janvier 2007
• Nombre de lectures: 15
• Langue: English
• Poids de l'ouvrage: 28 Mo

Extrait

INAUGURAL - DISSERTATION
zur
Erlangung der Doktorwürde
der
Naturwissenschaftlich - Mathematischen Gesamtfakultät
der
Ruprecht - Karls - Universität Heidelberg














Vorgelegt von:
Dipl.-Chem. Stefan V. W. Gräter
aus Esslingen a.N.
Tag der mündlichen Prüfung: 19.07. 2006
















































Design of Artificial Modular Extracellular
Matrices


















Gutachter: Prof. Dr. Joachim P. Spatz
Prof. Dr. Martin Möller













































I. Summary – Zusammenfassung.......................................................................1
II. Introduction and Motivation ..........................................................................5
III. Theory and background..................................................................................9
III.1. Extracellular matrix.......................................................................................9
III.1.1. What is the extracellular matrix? ...........................................................9
III.1.2. Components of the ECM .....................................................................10
III.2. Cell surface receptors...................................................................................14
III.2.1. Transmembrane receptors, and their various functions .......................15
III.2.2. The integrin family of adhesion receptors ...........................................16
III.2.3. L1 of the immunoglobulin family........................................................18
III.3. Interactions between cells and the ECM......................................................19
III.3.1. Mechanical interactions between cells and the ECM ..........................19
III.3.2. Chemical between cells and the ECM..............................22
III.3.3. Structural .............................22
III.4. Design of an artificial extracellular matrix ..................................................24
III.4.1. Approaches used to mimic the extracellular matrix ............................24
III.4.2. Concept of synthetic modular ECMs...................................................30
IV. PEG-based supports with tunable mechanical properties.........................32
IV.1. Tuning the mechanical properties of PEG hydrogels ..................................32
IV.1.1. Crosslinking reaction of PEG-DA .......................................................32
IV.1.2. Crosslinking parameters and mechanical properties............................34
IV.2. Characterizing the mechanical properties of a hydrogel..............................36
IV.2.1. Determination of the mesh size, swelling and gel content...................36
IV.2.2. Measuring elasticity at the nanoscale ..................................................37
IV.3. Experiments, results and discussion ............................................................39
IV.3.1. Crosslinking of PEG ............................................................................39
IV.3.2. Gel content and swelling ratio .............................................................41
IV.3.3. AFM Indentation..................................................................................44
IV.4. Conclusion...................................................................................................45
V. Nanostructured polymeric supports ............................................................46
V.1. Nanostructures.............................................................................................46
V.1.1. Different nano-structuring approaches ................................................46
V.1.2. Block copolymer micelle nanolithography..........................................47
V.1.3. Preparation of gold nanopatterned substrates ......................................51
V.2. Transfer nanolithography.............................................................................53
V.3. Transfer of gold clusters to different polymers............................................54
V.3.1. Linker system: Experiment and characterization.................................55
V.3.2. Transfer of the gold particles: Experiments and results.......................59
V.4. Conclusion...................................................................................................66
VI. Preparation of patterned hydrogel channels...............................................68
VI.1. Decoration of non-planar glass structures....................................................68
VI.2. Transfer lithography for non-planar substrates............................................72
VI.3. Conclusion...................................................................................................74
VII. Chemical modification of PEG hydrogels....................................................76
VII.1. Introduction..................................................................................................77
VII.1.1. Chemical properties of PEG hydrogels................................................77
VII.1.2. Bio-molecules for surface activation ...................................................78
VII.2. Bio-functionalization of gold particles on hydrogels...................................82
VII.2.1. Binding of RGD peptides to the gold particles....................................83
VII.2.2. Binding of proteins to the gold particles via a nickel-NTA complex..84
VII.3. Biofunctionalization of PEG hydrogels.......................................................87
VII.3.1. Copolymerization of carboxyethylacrylate to the PEG-DA ................88
VII.3.2. Binding of peptides to the hydrogel.....................................................90
VII.3.3. Binding of proteins to the hydrogel via a Nickel-NTA complex.........91
VII.4. Conclusion...................................................................................................94
VIII. Cell response to various hydrogel modules .................................................96
VIII.1. Cell culture protocols...................................................................................96
VIII.1.1. Standard culture conditions..................................................................96
VIII.1.2. Cell experiments on hydrogels.............................................................97
VIII.2. Cells on biofunctionalized hydrogels...........................................................98
VIII.3. Conclusion .................................................................................................105
IX. Conclusion and future plans .......................................................................107
X. Materials .......................................................................................................111
XI. Acknowledgments ........................................................................................113
XII. Bibliography .................................................................................................115


























































































I. Summary – Zusammenfassung
Summary
Cellular functions such as cell growth, adhesion and differentiation are essentially
controlled by the surrounding extracellular matrix (ECM). The mechanical, chemical
and structural properties of the ECM are consequently crucial for the selection of cells
at interfaces and the formation of tissues.
The objective of this thesis was to develop an artificial ECM to determine and control
the parameters influencing the crosstalk between cells and their surroundings on a
molecular level. Artificial ECMs which mimic the natural environment of cells enable
precise insights into cell-ECM crosstalk; ultimately, we aim to trigger the crosstalk,
such that specific cell functions are provoked. To this end, a modular ECM system
was developed, consisting of (i) poly(ethylene glycol) (PEG) as the basic material, (ii)
gold nano-particles as the structuring component, and (iii) bioactive molecules which
are immobilized on the basic material and on t