Exploring the in vitro behaviour of endothelial cells in different cell culture models

  • Lucy Harvey

Student thesis: Master's ThesisMaster of Science by Research (MScR)

Abstract

Maintenance of vascular homeostasis requires a coordinated and regulated response by endothelial cells. ECs form the inner lining of blood and lymphatic vessels and form an interface between the luminal vessel fluid and surrounding tissue. They are hence involved in a large number of physiological processes and implicated in many pathologies. The fundamental biology of ECs must therefore be studied in order to understand the role of ECs in normal physiology and disease. My lab focusses on research into ECs and in the generation of in vitro EC culture models to explore its roles in angiogenesis. I aimed to analyse EC growth and proliferation under different conditions. An additional aim was to optimise a tumour angiogenesis co-culture assay, which analyses the growth of blood vessels towards tumour spheroids. Finally, I set out to study the surface polarity of a number of important cell surface proteins in human umbilical vein endothelial cells by means of surface protein labelling and analysis. A variety of in vitro assays were used across this work, including that for cell proliferation, tube formation and protein identification.

Optimal HUVEC culture conditions were analysed and determined. HUVEC grow and proliferate greatest in Lonza EGM-2 media, however ECs from transgenic fluorescent zebrafish larvae require alternate cell culture conditions. HUVEC also have varying growth abilities on different surfaces, with a nanostructured surface generating low confluency of cells which also developed markers of genetic instability. Optimisation of HUVEC culture conditions led to investigation of their capacity to form vascular tubules in a co-culture with normal human dermal fibroblasts. An optimal vascular density to effectively determine vascular directionality towards tumour spheroids was achieved, with future prospects to assess the angiogenic effects of growth factors and drugs in culture. Failure to develop a novel method to label HUVEC surface proteins for proteomic analysis revealed that further work on this needs to be carried out, however immunofluorescence has shown to be a valid, low-throughput method to label and identify surface proteins
Date of Award25 Jun 2019
Original languageEnglish
Awarding Institution
  • The University of Bristol
SupervisorHarry H Mellor (Supervisor) & Kevin Gaston (Supervisor)

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