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Julie Schaub, Arina Korneva, Elizabeth Cone Cone-Kimball, Mary Ellen Pease, Sarah Quillen, Ian F Pitha, Harry A Quigley; Amyloid precursor protein transport block evaluation as a biomarker for experimental mouse glaucoma. Invest. Ophthalmol. Vis. Sci. 2020;61(7):2001.
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© ARVO (1962-2015); The Authors (2016-present)
To develop a quantifiable marker of axonal transport blockade, which occurs early in glaucoma and is associated with eventual retinal ganglion axonal loss.
Experimental microbead glaucoma (Cone et al. 2012) was induced in one eye for 3 days in CD1 (n=59) mice. A subset of mice were pre-treated with glyceraldehyde (Kimball et al. 2014, n=17), losartan (n=10), or irbesartan (n=9) treatments. Oral treatment with losartan (40-60mg/kg/day) and irbesartan (50mg/kg/day) was began 12 days prior to bead injection. Eyes were cryosectioned and immunostained with DAPI and anti-amyloid precursor protein (APP), then imaged with a LSM710 confocal microscope. Pixel brightness histograms were made in ImageJ (FIJI) and analyzed with MATLAB. The mean intensity of the brightest pixels (brighter than 97.5th percentile of controls) and fraction of brightest pixels (brightest pixels/total pixels) for the pre-optic nerve head (pre-ONH) and unmyelinated optic nerve (UON) were compared between groups.
With 3 day IOP glaucoma, visible clumps of APP within the pre-ONH and UON (Figure 1) led to significantly greater pixel intensity and fraction in glaucoma than control (non-glaucoma) eyes (p<0.05 for all; Figure 2). There was an increase in intensity and pixel fraction in glaucoma + glyceraldehyde eyes compared to control (non-glaucoma) + glyceraldehyde-treated eyes in the UON region (p<0.05). Losartan treatment reduced transport block in the pre-ONH of glaucoma eyes compared to glaucoma only and glaucoma + irbesartan (pixel fraction, p<0.05). The effects in this new marker are consistent with past findings that glyceraldehyde increased glaucoma axonal loss vs glaucoma only (44.6 ± 32.4% vs, 23.0 ± 30.9%, p<0.01), while losartan lowered axonal loss in glaucoma eyes contrasted with glaucoma only (6.2 ± 2.5% vs 25.4 ± 26.4%, p<0.01).
This APP transport block analysis allows an early look at retinal ganglion cell degeneration process in experimental glaucoma.
This is a 2020 ARVO Annual Meeting abstract.
Figure 1. APP (green) and DAPI (blue) expression in mouse optic nerve with the pre-ONH (top) and UON (bottom) outlined in white, with increase of APP over control (A) in glaucoma (B). Scale Bar: 100µm.
Figure 2. Fraction and mean intensity of brightest pixels above 97.5th percentile of controls for (A) glyceraldehyde and (B) oral sartan pretreatment of 3 day bead glaucoma mice. *p<0.05 paired test; #p<0.05 unpaired test
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