Fallatah, Safaa Mohammed Yousif; (2020) Stem Cell and Material Bioengineering Approaches for Vascular Regenerative Medicine Applications. Doctoral thesis (Ph.D), UCL (University College London).

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Abstract

Cardiovascular disease (CVD) is considered a significant public health problem, and one of the main leading causes of vascular occlusive states, including congestive heart failure (CHF), myocardial infarction (MI) and strokes. CVD is responsible for about 35% of death in the United Kingdom, which accounts for 150,000 deaths each year. Moreover, this incidence is anticipated to increase by a further 20,000 deaths by the year 2022. The major objectives of this research project were to investigate several and possible aspects of treating CVD and developing a CVD model as current in vitro disease models are failing to meet the need for new efficacious drugs and in vivo animal disease models are not fully translatable to humans. Firstly, we investigated whether we could generate populations of vascular cells from a renewable source that would benefit disease modelling and regenerative medicine applications. Human and murine induced pluripotent stem cells (iPSCs) were therefore selected for this part of the study for their ability to differentiate into functional endothelial cells (ECs) and vascular smooth muscle cells (vSMCs). Moreover, we observed a successful derivation of pericytes (PCs) from human iPSCs (hiPSCs). We also determined that early addition of growth factors including vascular endothelial growth factor (VEGF) and Bone Morphogenetic Protein 4 (BMP-4), in combination with early FLK-1+ sorting, derived ECs with greater functionality, as determined by an increased tube forming capacity and nitric oxide (NO) production as well as cell migration following injury. As an alternative to differentiating iPSCs on collagen IV investigated in Chapter 3, we next examined the utility of the natural vessels ECM in promoting iPSCs differentiation by substituting collagen IV for vessel ECM gel. The vessels of donors were subjected to a decellularisation process to remove all cellular material, leaving the ECM scaffold intact. The optimal decellularisation protocol developed herein demonstrated that Glycosaminoglycans (GAGs) and collagen were preserved at the same level as native vessels with efficient DNA material removal. Next, a decellularised vessel extracellular matrix gel (V-gel) was successfully created that is not toxic to the cells. Uncontrolled hiPSC differentiation on V-gel was observed, although, V-gel supported endothelial progenitor cells (EPCs) metabolic activity. To determine whether we could generate a bioengineered vessel for these studies, we next utilised polyhedral oligomeric silsesquioxane poly(caprolactone-urea) urethane (POSS-PCLUU) as a synthetic scaffold on which to grow blood vessel cells. POSS-PCLUU is a biocompatible, non-toxic, versatile and biodegradable nanocomposite polymer material suitable for a range of tissue engineering applications. We postulated that POSS-PCLUU could be utilised to replace damaged vessels as it exhibits the same mechanical strength properties as native blood vessels. However, work herein showed that while this material could support cell growth and proliferation for 12 days, beyond that period, cells grown on POSS-PCLUU started to lose their metabolic activity and functionality. We also looked at another aspect that could be utilised to disease a cardiovascular model. This was by utilising endothelial progenitor cells (EPCs) that circulate in the bloodstream. EPCs isolation and characterisation from healthy donors was achieved. On the other hand, the derivation of EPCs from diabetic patients with cardiovascular complications was not accomplished.

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