%0 Generic
%A Kaiser, Peter
%D 2009
%F heidok:10125
%K Nanofibrils , Extracellular Matrix
%R 10.11588/heidok.00010125
%T Fabrication and Characterization of Extracellular Matrix Nanofibrils
%U https://archiv.ub.uni-heidelberg.de/volltextserver/10125/
%X All cells in our body are surrounded by Extra Cellular Matrix (ECM), from which they derive biochemical, structural and mechanical signals. One of the main fibrillar ECM protein components is Fibronectin (Fn), which is believed to act as a mechanochemical signal transducer. A current hypothesis is that Fn can undergo structural transitions upon stretching, which can alter Fn binding site accessibility and ultimately lead to an adapted cell response. While this hypothesis has existed for several years, the lack of suitable model systems prevented its proof. The aim of this work was to (i) produce regular arrays of Fn nanofibrils, (ii) control the alignment, diameter and tensile state of those nanofibrils, and (iii) to determine their structural and mechanical properties. During this work, a new method to create regular arrays of Fn nanofibrils was developed. This method allows the control of nanofibril directionality and diameter and can also be used to produce nanofibrils from other ECM proteins, such as Laminin (LM) and Collagen (COL). The method depends both on a protein’s ability to accumulate at the air-buffer interface and its ability to self-associate. The production of nanofibrils from various polymers that share these properties is thus possible. The resulting nanofibrillar arrays can be produced on a variety of mirostructured materials, ranging from Silicon over Poly(dimethylsiloxane) (PDMS) to Polyurethane (PU). The biofunctionality of different ECM nanofibrillar arrays was demonstrated by specific cell adhesion after nanofibril transfer onto non-fouling Polyethyleneglycol (PEG) hydrogels. An investigation of both the molecular structure and the mechanical properties of Fn nanofibrils was performed by Förster Resonance Energy Transfer (FRET) and Atomic Force Microscopy (AFM) experiments. Fn molecules form a surface film after application of Fn into a drop of Phosphate Buffered Saline (PBS). FRET analysis of Fn was performed to determine the degree of Fn molecular unfolding. It could be shown that Fn within surface films only unfolds upon surface dewetting, which coincides with nanofibril formation. The produced nanofibrils show an elongation at break of 200 %. Ruptured nanofibrils retract to 30 % of their original length, but the Fn molecules within nanofibrils do not re-fold completely, as derived from FRET measurements. The pre-strained Fn nanofibrils display a high effective Young’s modulus of E ~ 0.1 - 6 GPa, as determined by AFM experiments. In summary, the production, control and characterization of novel ECM models was accomplished in this work, which can be used to investigate cell adhesive response.