The aim of this thesis is to develop automated methods for the analysis of the
spatial patterns, and the functional behaviour of endothelial cells, viewed under
microscopy, with applications to the understanding of atherosclerosis.
Initially, a radial search approach to segmentation was attempted in order to
trace the cell and nuclei boundaries using a maximum likelihood algorithm; it
was found inadequate to detect the weak cell boundaries present in the available
data. A parametric cell shape model was then introduced to fit an equivalent
ellipse to the cell boundary by matching phase-invariant orientation fields of the
image and a candidate cell shape. This approach succeeded on good quality
images, but failed on images with weak cell boundaries. Finally, a support
vector machines based method, relying on a rich set of visual features, and a
small but high quality training dataset, was found to work well on large numbers
of cells even in the presence of strong intensity variations and imaging noise.
Using the segmentation results, several standard shear-stress dependent parameters
of cell morphology were studied, and evidence for similar behaviour
in some cell shape parameters was obtained in in-vivo cells and their nuclei.
Nuclear and cell orientations around immature and mature aortas were broadly
similar, suggesting that the pattern of flow direction near the wall stayed approximately
constant with age. The relation was less strong for the cell and
nuclear length-to-width ratios.
Two novel shape analysis approaches were attempted to find other properties
of cell shape which could be used to annotate or characterise patterns, since a
wide variability in cell and nuclear shapes was observed which did not appear
to fit the standard parameterisations. Although no firm conclusions can yet be
drawn, the work lays the foundation for future studies of cell morphology.
To draw inferences about patterns in the functional response of cells to flow,
which may play a role in the progression of disease, single-cell analysis was performed
using calcium sensitive florescence probes. Calcium transient rates were
found to change with flow, but more importantly, local patterns of synchronisation
in multi-cellular groups were discernable and appear to change with flow.
The patterns suggest a new functional mechanism in flow-mediation of cell-cell
calcium signalling
