For decades, research has been focussed on finding a way to produce artificial blood as a resolution for the insufficient amount of blood components provided by donation and to provide a more transportable alternative with a longer shelf life. Red blood cells (RBCs) are the most common cell type in blood and are responsible for oxygen transport throughout the human body. It is therefore extremely important to find an alternative oxygen carrier whether these are tissue engineered RBCs or a chemically defined oxygen delivery system The study conducted for this thesis was part of a larger project called Redontap, and was aimed to develop a bioreactor for the manufacture of RBCs. During this research to produce RBCs from adult stem cells in vitro, the main goal was to upscale haematopoietic and erythroid cultures. Understanding the biological signals and their temporal magnitude involved in the division, maturation and migration of the CD34+ haematopoietic stem cells (HSCs) and their differentiated progeny would allow for a controlled continuous production of mature blood cells. The differentiation of HSCs into different blood cell types occurs within different bone marrow niches and so mimicry of the erythrocyte niche is likely to result in maximisation of the rate of red blood cell development. Published research provides evidence that peripheral blood mononuclear cells (PBMCs), including CD34- cells, will be advantageous for erythroid maturation. For this thesis, CD34+ cells were expanded within a population of PBMCs on a stromal layer to recreate a niche-like environment. This approach was also utilised with umbilical cord blood isolated MNCs (UBMCs) to compare HSC expansion potential and subsequently efficiency in erythroid maturation was analysed. Whereas the cell output was limited, differentiated cells proved positive for a range of RBC surface markers and haemoglobin content. As part of the aim for upscaling cell culture by translating static cultures to bioreactor processes, bioreactors with volumes varying between 250mL-3L were analysed for cell retention and viability to achieve high cell densities whilst refreshing culture medium, monitoring culture parameters (e.g. pH, dissolved oxygen), and introducing an hypoxia environment for mimicking the in vivo stem cell niche. In general, this research was focussed on improving dynamic culture conditions for generating higher numbers of cultured erythrocytes than so far has been achieved.
| Identifer | oai:union.ndltd.org:bl.uk/oai:ethos.bl.uk:634454 |
| Date | January 2014 |
| Creators | van Veen, Hendrik Theun |
| Publisher | University of Liverpool |
| Source Sets | Ethos UK |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
| Source | http://livrepository.liverpool.ac.uk/2004970/ |
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