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Title: A test bed to explore and exploit the inter-relationships between stem cell differentiantion and matrix elasticity
Author: Lanniel, Mathieu
ISNI:       0000 0004 2725 182X
Awarding Body: University of Nottingham
Current Institution: University of Nottingham
Date of Award: 2011
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Physical and chemical interactions between cells and the extracellular matrix are essential to a number of cellular functions and have also been shown to influence stem cell lineage specification. The aim of the present study is to investigate the interplay of surface stiffness and chemistry on human mesenchymal stem cell (hMsC) differentiation using two engineered substrates, hydrogen silsesquioxane (HsQ) and polyacrylamide qels, both coated with plasma polymers. For HsQI it is found that the surface stiffness can be controlled by the electron beam dose used for curing. A large range of Younq's modulus values can be achieved (0.06 GPa to 1 GPa) with a constant and low surface roughness when the developing step is omitted. Plasma polymerised allylamine (ppAAm) deposition leads to a general decrease in surface stiffness values but the variation caused by differences in electron beam curing is still apparent. hMsCs are able to adhere to the HsQ surface after ppAAm deposition but no dependence of cell morphology on the surface stiffness are observed. For polyacrylamide qels, a range of stiffness values from 7 kPa to 152 kPa for uncoated polyacrylamide gels is achieved. Gel coating with plasma polymerised- altylamlne, acrylic acid (ppAAc) and trimethylphosphite (ppTMP) is compared to collagen coating. Using x-rav photoelectron spectroscopy (X PS) analysts, it is observed that different surface chemistries are achieved while maintaining gel surface stiffness variations and low surface roughness values. Initial cell attachment to the qels, cell spreading and cytoskeletal organizations are mainly affected by surface stiffness with slight differences observed between the surface chemistries studied. Long term cell survival is related to both surface chemistry and stiffness. To assess the differentiation of the hMsCs1 antibody staining is carried out using markers for osteogenic (Runxz), myogenic (MyoD1) and neurogenic (B-Ill tubulin) cell types, which reveals a dependence of marker protein expression upon both surface stiffness and chemistry. The expression of the osteogenic Runx2 marker is maximal for cells cultured on ppTMP coated gels of 41 kPa surface stiffness. Myogenic MyoD1 expression is maximal on the ppAAc coated gels of intermediate stiffness (10 kPa to 17 kPa). Neurogenic differentiation, indicated by B-III tubulin expression, is seen to be greatest on the ppAAc coated surfaces but shows less dependency on stiffness than the other two markers. The addition of soluble factors in the medium to induce osteogenic behaviour results in the formation of bone nodules and matrix calcification for gel stiffness values higher than 10 kPa, especially on ppAAm coatings. The results indicate that control over differentiation fate of hMSCs can be exerted using not only surface stiffness, a result previously widely reported, but also surface chemistry working in tandem with the influence of compliance. This has great significance in developing stem cell therapies when synthetic surfaces are used as scaffolds, delivery vehicles or culture ware.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available