Study of collagen structure in canine myxomatous mitral valve disease
Abstract
Myxomatous mitral valve disease (MMVD) is the single most common acquired
cardiac disease of dogs, and is a disease of significant veterinary importance. It also
bears close similarities to mitral valve prolapse in humans and therefore is a disease
of emerging comparative interest. Realising the importance of collagen fibres in
mitral heart valves and considering the paramount significance of myxomatous
mitral valve disease, a better understanding of the pathogenesis of MMVD is
essential. Thus, this study was designed to investigate the changes in collagen
molecules, including fibril structure, fibril orientation, d-spacing, collagen density,
collagen content, thermal stability, and the status of mature and immature crosslinks.
A combination of biophysical and biochemical tools such as x-ray diffraction,
neutron diffraction, HPLC were utilised in order to fulfil the objectives. Biochemical
assay of hydroxyproline revealed a 10% depletion of collagen in mildly affacted
(grade I and II) leaflets, while a 20% depletion of fibrillar collagen was revealed by
mapping the collagen fibrils onto the anatomy of cardiac leaflets using x-ray data.
Differential scanning calorimetry showed that there were no significant differences
in the onset temperature of denaturation of collagen between the healthy and affected
leaflets. However, in affected areas of leaflets, the enthalpy of denaturation
significantly dropped by 20%. In the affected regions, neutron diffraction results
showed an increase in the immature reducible cross-links though the low number of
the samples can be considered a limiting factor in this regard. However, the HPLC
results showed a 25% decrease in the number of mature cross-links. Additionally, the
recently introduced imaging technologies to biology and medicine such as differential enhancing imaging (DEI) and coherent anti-Stokes Raman scattering
spectroscopy (CARS) were, to the author’s best knowledge, applied for the first time
to this disease. In doing so, this thesis furthers our understanding of the pathogenesis
of MMVD, especially in relation to the collagen. The thesis provides new findings
about MMVD and demonstrates the potential of biophysical tools for studying
similar conditions.