McKenzie, Andrew (2015) Integrated studies of the interactions of macrophages with synthetic hydrogels. PhD thesis, University of Sheffield.
Abstract
In this project a glycerol methacrylate hydrogel surface was synthesised, which acted as a
biomaterial. This biomaterial was used as a surface on which fibroblast and macrophage
type cells were grown. The hydrogels were synthesised from glycerol methacrylate, a
hydrophilic monomer, cross-linked with glycerol dimethacrylate using chiefly thermal
polymerisation methods. The hydrogels were shown to have similar water contents of
around 75-85% mass and similar physical structures. Polystyrene latex particles were also
synthesised using emulsion polymerisation. Latex particles were synthesised in 3 sizes: 100
nm, 250 nm and 720 nm. These particles were produced easily and in large quantities in the
three sizes and with good control. The particles were incorporated into the hydrogel
structures to give three sets of hydrogels with surfaces of varying roughnesses but identical
surface chemistry. The surfaces were analysed with electron microscopy and white light
interferometry to gain an understanding of the roughness and appearance of the surfaces.
These latex hydrogels were optimised for cell culture and shown to be non-cytotoxic by the
culture of 3T3 fibroblast cells on their surfaces. Normal human dermal fibroblast cells were
also grown on the hydrogels and were shown to have survived on the surface for at least 48
hours with some evidence of proliferation. A simple staining and imaging method using
Giemsa nuclear stain, PicoSirius red cellular collagen stain and upright inverted light
microscopy was demonstrated. Optimisation of the hydrogels for the culture of THP-1
macrophage like cells was demonstrated and ruled out epifluorescence imaging for these
materials due to excessive autofluorescence generated by the hydrogels. It was shown that
it was possible to induce differentiation of the THP-1 cells using PMA and for the cells to
adhere to the hydrogel surface, however an experimental attempt to investigate to what
extent the hydrogel surfaces caused an activation of the macrophages was inconclusive.
Using ELISA it was shown that the detection of very small levels of inflammatory cytokines
was a possible.
Alongside these investigations, a set of water swollen core-shell particles was synthesised
from a protected monomer in water using emulsion polymerisation. These particles were
shown to swell with water on the removal of a protecting acetonide group. The particles
increased in size from around 100-300 nm to 5-6 µm. This water swelling is a key indication
of hydrogel function. The core shell particles were shown to be able to adsorb protein
molecules (lysozyme, albumin and fibrinogen) onto their surfaces and the change of surface
charge, measured by zeta potential was shown. Higher adsorbed protein concentration had
a more marked effect on the elevation of charge on the particles and the particles with a
smaller shell diameter showed the largest change in zeta potential with adsorbed proteins.
Total protein content adsorbed to the particles was measured using the BCA assay. The
protein adsorption showed that these particles may have potential used in a biological
context and could be investigated further in the area of drug or biomolecule uptake and
release.
Metadata
Supervisors: | Rimmer, Stephen and Ingham, Eileen and Southgate, Jenny |
---|---|
Awarding institution: | University of Sheffield |
Academic Units: | The University of Sheffield > Faculty of Science (Sheffield) > Chemistry (Sheffield) The University of Sheffield > Faculty of Science (Sheffield) |
Identification Number/EthosID: | uk.bl.ethos.679833 |
Depositing User: | Mr Andrew McKenzie |
Date Deposited: | 16 Feb 2016 09:41 |
Last Modified: | 03 Oct 2016 13:07 |
Open Archives Initiative ID (OAI ID): | oai:etheses.whiterose.ac.uk:11949 |
Download
Thesis corrected final
Filename: Thesis corrected final.pdf
Licence:
This work is licensed under a Creative Commons Attribution-NonCommercial-NoDerivs 2.5 License
Export
Statistics
You do not need to contact us to get a copy of this thesis. Please use the 'Download' link(s) above to get a copy.
You can contact us about this thesis. If you need to make a general enquiry, please see the Contact us page.