Peripheral artery disease (PAD) and the consecutive build-up of an
atherosclerotic plaque restricting blood flow to the lower limbs lead to critical
limb ischaemia, one of the most common circulation problems in the world.
Although a small number of interventions (such as surgery or revascularization
treatments) are available, patients with this condition are often too ill for these
procedures, giving a poor prognosis for the disease.
Several strategies to promote neovascularization using different stem cell
populations with angiogenic potential have been proposed as plausible
therapies. Perivascular cells (PCs), key structural components of the wall of
small and large blood vessels have numerous advantages over other cell types
since they are highly abundant, easy to obtain from the stromal vascular
fraction (SVF) of human adipose tissue (an ethically approved source) and have
mesenchymal and angiogenic properties. The work described in this thesis
addressed the hypothesis that PCs isolated from human white adipose tissue
would promote the recovery of blood flow in an ischaemic hindlimb by
increasing blood vessel number and blood perfusion to the foot.
To investigate whether PCs from human white adipose tissue could rapidly
increase neovascularization and, therefore, be used as a possible therapeutic
treatment for PAD and critical limb ischaemia, the initial aim was to validate,
characterise and demonstrate the properties of the murine equivalent of these
cells, in order to establish a direct link between the injected cells and the ones
natively found in the mouse. This was then followed by the use of murine
models of angiogenesis to determine whether transplanted human PCs
stimulate angiogenesis in vivo.
Initial studies using immunohistochemistry, fluorescence-activated cell sorting
(FACS) and in vitro mesodermal differentiation demonstrated that perivascular
cells (namely pericytes and adventitial cells) are present in multiple mouse
organs, can be sorted to purity, and have mesenchymal stem cell (MSC)
properties. These cells had similar characteristics to their human counterparts,
thus validating the mouse as a suitable model for determining whether
transplanted human PCs could stimulate angiogenesis.
Using in vitro and two in vivo (sponge implantation and hindlimb ischaemia)
models, it was shown that human PCs have angiogenic properties being capable
of tube formation and interaction with endothelial cells, as well as promoting
angiogenesis within sponges. Contrary to expectations, PCs did not increase
blood perfusion to the mouse ischaemic hindlimb, despite increasing
microcirculation within the skeletal muscle and myofibre regeneration.
This work showed that PCs obtained from human adipose tissue have important
therapeutic implications in promoting angiogenesis and skeletal muscle
regeneration but failed to increase arteriogenesis which is the key mechanism
allowing the restoration of blood perfusion