Distribution of lipids in the human lens and the affect of ageing

The lens is a transparent, biconvex structure located between the iris and the vitreous humour. It is bathed by aqueous humour anterior to the lens, which plays a role in nutrient transport into the lens as it is avascular. The outermost layer of the lens consists of a single layer of epithelial cells. These cells elongate and differentiate into lenticular fibres which are laid down in concentric layers to form the mass of the lens. Mature fibre cells lose all their organelles to maintain lens transparency, thus there is no lipid or protein turnover in the lens nucleus. The lens is first formed in utero and continues to grow throughout a human’s lifespan. The lens formed before birth is referred to as the lens core, where the cells that were laid down immediately after birth (infantile nucleus) are referred to as the inner region. Tissue formed during childhood years make up the first layers of lens cortex outside the nucleus, while cells laid down after middle age are referred to as the outer region. The formation of the barrier between the inner cortex and nucleus at middle age reduces the diffusion of antioxidants and water into the nucleus, as a consequence of the occlusion of membrane pores. Alterations to the membrane lipid composition has the ability to alter the water permeability of aquaporin-0 and also render the membrane more adhesive to α-crystallins, which may reduce water permeability and also cause greater pore occlusion. As a consequence, the lens is more susceptible to protein precipitation and oxidation, leading to age-related nuclear cataract. Experimental evidence suggests that the lipid profile of the lens core changes dramatically around the age of 40, coinciding with the formation of the barrier region. It has been observed that lens sphingomyelin and dihydrosphingomyelin form an annular distribution within the barrier region in an adult lens, while ceramide and dihydroceramide levels that were negligible in the core of young lenses have a sharp increase in abundance after the age of 40. These studies indicate that the change in lipid profile may be related the barrier formation with middle age. In this thesis, methods to optimise mass spectrometric imaging of human lens tissue were developed. It was found that sublimation of matrix was the fastest way to apply a layer of homogenous matrix that was optimal for matrix-assisted laser desorption ionisation (MALDI) imaging. Sublimation conditions were fined tuned for 2-Mercaptobenzothiazole to be used as the matrix in positive ion mode and 2,5-diaminonapthalene to be used as the matrix in negative ion mode