Abstract
Purpose :
To maintain tissue transparency, the ocular lens is avascular and operates a microcirculation that delivers nutrients and removes waste products from the lens nucleus. Breakdown of lens structure or function can lead to lens cataract. Globally, lens cataract is the leading cause of preventable blindness, yet the only ‘cure’ is surgical implantation of a synthetic lens. MALDI imaging mass spectrometry (IMS) has been used to map uptake of exogenous molecules to inform development of novel pharmaceutical approaches to delay or prevent the onset of cataract.
Methods :
Bovine lenses were incubated from 5 min-20h in artificial aqueous humour containing stable isotopically-labelled (SIL) lens nutrients (glucose, glutathione) or pharmaceuticals. Axial cryosections (20mm) were analysed by negative and positive mode MALDI-FT-ICR IMS, respectively. Identical samples were microdissected into epithelium, outer cortex, inner cortex and nucleus regions. Gas chromatography mass spectrometry was then used on extracts of these lens regions to validate spatial distribution patterns observed by IMS, and confirm identities of small molecules observed by IMS. Liquid chromatography tandem mass spectrometry-based proteomics and immunohistochemistry were used to detect and localise specific small molecule transport proteins throughout the lens.
Results :
For SIL lens nutrients, MALDI IMS showed signal first appeared in the peripheral epithelium and lens equator. Several members of the solute carrier family of integral membrane proteins (e.g. GLUT1, GLUT3) were also detected by proteomics and immunohistochemistry in this region. In contrast, uptake profiles for pharmaceuticals were less spatially directed than for lens nutrients. Rates of transport of pharmaceuticals throughout the lens were also different between pharmaceuticals with different physicochemical properties.
Conclusions :
A combination of mass spectrometry techniques for protein, metabolite and pharmaceutical detection and mapping can be utilised to aid development of future therapeutic interventions that exploit lens physiological biochemistry to delay the onset of cataract.
This abstract was presented at the 2022 ARVO Annual Meeting, held in Denver, CO, May 1-4, 2022, and virtually.