Calcium plays a critical role in a variety of cell-signalling systems and is increasingly recognised to play a significant part in athe development of cortical cataract. The main aims of this project will be to investigate the mechanisms for achieving calcium homeostasis in the lens and also the sensitivity of these mechanisms to external insult. A further aim will be to elucidate the role of the calcium-activated enzyme, calpain, in the production of lens opacities. Calcium homeostasis will be studied, principally using human lens tissues, but also on athe rat lens in vitro, by a combination of elecrophysiological, radiotracer and fluorimetric dye techniques. Thapsigargin, which selectively inhibits the CaATPase of the endoplasmic reticulum, will be an invaluable tool in these investigations and mathematical modelling techniques will be used to interpret 45Ca flux data from whole human lenses. The sensitivity of calcium permeability (channel) pathways to oxidative insult will also be studied in detail in whole human and rat lenses and in tissue cultured cells. A cation channel antibody will also be used with a combination of immunofluorescence and gold-labelling technique to study the localization of cation channels in lens epithelia and fibre cells. Increases in cytosolic calcium in single lens cells and in whole rat and human lenses will be induced by calcium ionophore (A23187), external membrane SH complexation (pCMPS) and calcium signalling agonists (acetylcholine) and will be quantitatively assayed by the above technique. The effect of these agents on lens transparency will be quantified in whole lens experiments by imaging scattered light. The effect on the lens cytoskeleton (principally actin, tubulin, vimentin and spectrin) will be studied by a combination of DSS-PAGE electrophoresis and immunofluorescence localization techniques. A concomitant study of the colocalization of lens cation channels and cytoskeletal elements will be carried out both in normal and calcium overloaded lens tissues. This study will yield valuable information both on the mechanisms whereby calcium is controlled and also the mechanism responsible for calcium- induced proteolysis and opacification following loss of control. During the course of the project a combined approach will be made that aims both to reduce calcium overload by channel blocking agents and to prevent calcium-induced proteolysis by anti-calpain compounds.

Agency
National Institute of Health (NIH)
Institute
National Eye Institute (NEI)
Type
Research Project (R01)
Project #
1R01EY010558-01
Application #
2164502
Study Section
Visual Sciences A Study Section (VISA)
Project Start
1994-06-01
Project End
1997-05-31
Budget Start
1994-06-01
Budget End
1995-05-31
Support Year
1
Fiscal Year
1994
Total Cost
Indirect Cost
Name
University of East Anglia
Department
Type
DUNS #
City
Norfolk
State
Country
United Kingdom
Zip Code
Hu, Rebecca G; Suzuki-Kerr, Haruna; Webb, Kevin F et al. (2008) Molecular and functional mapping of regional differences in P2Y receptor expression in the rat lens. Exp Eye Res 87:137-46
Marcantonio, J M; Reddan, J R (2004) TGFbeta2 influences alpha5-beta1 integrin distribution in human lens cells. Exp Eye Res 79:437-42
Marcantonio, J M; Syam, P P; Liu, C S C et al. (2003) Epithelial transdifferentiation and cataract in the human lens. Exp Eye Res 77:339-46
Williams, M R; Riach, R A; Collison, D J et al. (2001) Role of the endoplasmic reticulum in shaping calcium dynamics in human lens cells. Invest Ophthalmol Vis Sci 42:1009-17
Collison, D J; Duncan, G (2001) Regional differences in functional receptor distribution and calcium mobilization in the intact human lens. Invest Ophthalmol Vis Sci 42:2355-63
Wormstone, I M; Del Rio-Tsonis, K; McMahon, G et al. (2001) FGF: an autocrine regulator of human lens cell growth independent of added stimuli. Invest Ophthalmol Vis Sci 42:1305-11
Marcantonio, J M; Rakic, J M; Vrensen, G F et al. (2000) Lens cell populations studied in human donor capsular bags with implanted intraocular lenses. Invest Ophthalmol Vis Sci 41:1130-41
Sanderson, J; Marcantonio, J M; Duncan, G (2000) A human lens model of cortical cataract: Ca2+-induced protein loss, vimentin cleavage and opacification. Invest Ophthalmol Vis Sci 41:2255-61
Wormstone, I M; Tamiya, S; Marcantonio, J M et al. (2000) Hepatocyte growth factor function and c-Met expression in human lens epithelial cells. Invest Ophthalmol Vis Sci 41:4216-22
Wormstone, I M; Liu, C S; Rakic, J M et al. (1997) Human lens epithelial cell proliferation in a protein-free medium. Invest Ophthalmol Vis Sci 38:396-404

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