Graded organisation of fibronectin to tune cell behaviour

T Technology (General)

Grigoriou, Eleni ; Salmeron-Sanchez, Manuel

University:University of Glasgow

Department:School of Engineering

关键词: ECM, stem cell differentiation, osteogenic differentiation, BMP-2.;   

Others  :  http://theses.gla.ac.uk/8523/1/2017GrigoriouPhD.pdf

来源: University of Glasgow

【 摘 要 】

Cells are in constant and dynamic interactions with the extracellular environment. They receive several inputs involved in the regulation of cell behaviour. Fibronectin, an abundant protein of the ECM, contains multiple binding domains and binds to cell receptors, growth factors and other ECM proteins. FN undergoes conformational changes through cell-generated contractile forces which consequently affects cell response.Tissue engineering aims at engineering biomaterials that recreate the in vivo ECM. In addition to biomaterials, stem cells have emerged as a promising source due to their inherent differentiation potential. In this work, the role of poly acrylates in controlling human mesenchymal stem cell behaviour (hMSCs) was explored. Particularly, a series of copolymers with specific ratio of ethyl(acrylate), EA, and methyl(acrylate), MA, were used. It is known that poly(ethyl)acrylate, PEA, triggers a network-like conformation of FN upon adsorption, whereas poly(methyl)acrylate, PMA, elicits a globular conformation. It was found that a different degree of FN organisation can be obtained dependent on the EA/MA ratio, with the network being more connected with increased EA ratio. This differential conformation was shown to affect the availability of critical binding sites. This system was further used to study hMSCs response in terms of adhesion and osteogenic differentiation. All surfaces support cell growth and focal adhesion formation. However, increased cell size and spreading was promoted on surfaces with higher EA concentration. Next, the potential of the surfaces after sequential adsorption of FN and the growth factor BMP-2 to drive osteogenic commitment was explored. Enhanced expression of the osteogenic markers RUNX2 and OCN was found with higher concentration of EA whereas the opposite was observed with ALP expression. Another part of this work involved investigating cell migration on PEA and PMA. Higher cell speed was found on PEA where FN adopts a more extended conformation. Moreover, the protein composition of focal adhesions was evaluated by proteomic analysis. The findings of this work give further insights into how the surface with well-defined chemical properties can modulate FN conformation and how these changes affect cellular processes.