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E. Clerin, S. Mohand-Saïd, C. Jaillard, T. Cronin, J. Bennett, J.-A. Sahel, T. Léveillard; A High Content Screening Platform for the Measurement of Trophic Factors in Rodent Models of Retinitis Pigmentosa. Invest. Ophthalmol. Vis. Sci. 2009;50(13):1936.
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© ARVO (1962-2015); The Authors (2016-present)
We have developed an automated cell counting platform to validate the activity of the Rod-derived Cone Viability Factors, as therapeutic agents for the treatment of patients suffering from retinitis pigmentosa. The automated cone counting platform for retinal explants is used both for mouse and rat models of rod-cone degeneration. The specific objective was to validate this novel method by comparison to the semi-manual stereological counting used previously (Mohand-Saïd et al., 1998).
Seven retinas from rd1 mouse (C3H) and their wild type isogenic controls (C3H+/+) were dissected from 15 to 90 days post-natal (PN), orientated and fixated with 4% paraformaldehyde. For each mouse, one eye was used for automated counting and the contralateral eye for the semi-manual stereological method. For both methods the cones were labelled with peanut agglutinin (PNA) coupled to Texas-Red. The novel method involves an inverted microscope linked to a computer driven motorized platform. Following automated acquisition in the depth of the tissue, cone density was measured using a special application of the Metamorph software.
The decrease in cone density following rod loss in the rd1 mouse was found to be equivalent using either the stereological method or the novel automated platform. The decrease in cone density from 35 to 90 days post-natal was also quantified in an additional rod-cone degenerative model, the rd10. Regionalisation in the cone density was demonstrated for the rd7 mouse model following labelling with polyclonal antibodies generated against short-wave opsin. We also used this platform to quantity the loss of cones mouse models carrying an inactivation of the genes encoding the Rod derived Cone Viability Factors.
We demonstrate the usefulness of this platform for preclinical studies aimed at preventing the loss of central vision using neurotrophic factors.
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