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Project 1: The effect of Nrf2 activators on mesenchymal stem cell viability in response to light curable dimethacrylate resins and Project 2: 3D architectural influences on mesenchymal stem cells co-cultured with H400 epithelial oral cell line using hydrogels.

Nicolae, Laura Cristina (2012)
M.Res. thesis, University of Birmingham.

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Abstract

Project 1:
Introduction: Dental restorative materials are useful in the repair of diseased or damaged teeth. These materials can cause cell damage through oxidative stress and the generation of free radicals which can be limited by Nrf2 activators including oleanolic acid and tBHQ (tert-Butylhydroquinone).
Objective: The main purpose of this study was to determine whether Nrf2 activators could protect mesenchymal stem cells (MSCs) from dental resin cytotoxicity.
Methods: MSC viability following exposure to resins over a 4 day period was determined by trypan blue counting and analysis of gene expression of NQO1 (Nad(P)H dehydrogenase, quinone 1), Gclc (glutamate-cysteine ligase, catalytic subunit), and Hmox1 (haemoxygenase (decycling) 1) using the semi-quantitative reverse transcriptase polymerase chain reaction (sqRT-PCR). Resistance to stress was examined by Instron and Vickers Hardness testing.
Results: The addition of 50-\( \mu\)M tBHQ to culture medium increased viable MSC numbers, compared with controls; however cell death occurred in the presence of supplemented resins. Highest Nrf2 activated gene expression occurred in50-\( \mu\)M tBHQ containing resin. The addition of tBHQ or oleanolic acid to resins affected their resistance to stress.
Conclusion: In the presence of dental resins, supplementation of culture medium with Nrf2 activators demonstrates decreased cytotoxicity.

Project 2:
Introduction: Cells cultured in 2D environments behave differently compared with 3D cultures possibly due to less complex interactions/signalling.
Objective: The main purpose of this study was to determine how different 3D culture environments influenced the behaviour and differentiation of bone marrow stromal cells (BMSCs) when co-cultured with the H400 oral epithelial cell line.
Methods: The viability of BMSCs encapsulated in collagen or alginate gels digested with collagenase or 4% sodium citrate respectively was determined using trypan blue cell counting. Gene expression analysis for osteogenic markers in BMSCs and cytokeratin markers in H400 cells were examined using the semi-quantitative reverse transcriptase polymerase chain reaction (sqRT-PCR).
Results: Fewer viable cells were present in 3D compared with 2D environment. BMSCs encapsulated in alginate and collagen gels exhibited increased expression of osteopontin. There was no difference in the expression of cytokeratin markers for encapsulated H400 cells compared with 2D cultured H400 cells.
Conclusion: BMSC encapsulation in 3D matrices may influence these cells to differentiate along particular lineages providing information for future research for clinical purposes.

Type of Work:M.Res. thesis.
Supervisor(s):Palin, William and Cooper, Paul and Shelton, Richard M.
School/Faculty:Colleges (2008 onwards) > College of Medical & Dental Sciences
Department:Biomedical Research, School of Dentistry
Subjects:QH301 Biology
QH426 Genetics
RK Dentistry
Institution:University of Birmingham
ID Code:3756
This unpublished thesis/dissertation is copyright of the author and/or third parties. The intellectual property rights of the author or third parties in respect of this work are as defined by The Copyright Designs and Patents Act 1988 or as modified by any successor legislation. Any use made of information contained in this thesis/dissertation must be in accordance with that legislation and must be properly acknowledged. Further distribution or reproduction in any format is prohibited without the permission of the copyright holder.
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