Retinal nerve fiber layer (RNFL) thinning and glaucomatous optic neuropathy in sGCα₁^−/− mice. A

<p>: Quantitative analysis, assessed by SD-OCT (see also <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0060156#pone.0060156.s001" target="_blank">fig. S1</a>), of total retinal thickness (<b>left panel</b>) and RNFL thickness, in young (6-week-old, <b>middle panel</b>) and old (70-week-old, <b>right panel</b>) wild-type (WT, <i>n</i> = 19 and 13, respectively) and soluble guanylate cyclase α₁-deficient (sGCα₁^−/−) mice (<i>n</i> = 15 and 14, respectively; *<i>P</i> = 1.2×10⁻²). <b>B</b>: Representative whole-mount retinas from age-matched young (20-week-old) and old (56-week-old) WT and sGCα₁^−/− mice, reacted with antibodies directed against SMI32, staining retinal nerve fibers yellow. Scale bars: 500 μm. <b>C</b>: Representative confocal images, taken at a similar distance from the optic nerve, of flat-mounted retinas isolated from age-matched 52-week-old WT and sGCα₁^−/− mice that were reacted with antibodies directed against βIII Tubulin, and quantitative analysis of the number of RGCs/high-powered field (<i>n</i> = 8 and 7, respectively; *<i>P</i> = 3.6×10⁻²). A retinal ganglion cell (red) is indicated by an arrow. Scale bars: 20 μm. <b>D</b>: Representative cross sections through the optic nerve of 52-week-old WT and sGCα₁^−/− mice stained with paraphenylenediamine, and quantitative analysis of the calculated number of axons/optic nerve (ON). The arrow indicates an injured area in the optic nerve, characterized by the absence of well-formed myelinated axons (<i>n</i> = 7 and 6, respectively; *<i>P</i> = 4.9×10⁻²). Scale bars: 25 μm.</p

Retinal vascular dysfunction in sGCα₁^−/− mice. A

<p>: Representative trace depicting the diameter in one segment of a retinal arteriole in a wild-type (WT) mouse before, during (arrow), and after injection of the NO-donor compound sodium nitroprusside. Dashed lines indicate the diameter before and after sodium nitroprusside injection. <b>B</b>: Quantitative analysis of the change in diameter (double arrow in fig. 6A) induced by injection of 0.8 mg/kg sodium nitroprusside in WT and sGCα₁^−/− mice. <i>n</i> = 5 mice (3–4 arterioles per mouse were assessed, see also <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0060156#pone.0060156.s003" target="_blank">fig. S3</a>). *<i>P</i> = 4.1×10⁻³.</p

Intraocular pressure (IOP) increases with age in sGCα₁^−/− mice.

<p>IOP, measured serially at 2 time points (19±1 and 37±3 weeks) in eyes from age-matched wild-type (WT, <b>left panel</b>) and soluble guanylate cyclase α₁-deficient (sGCα₁^−/−) mice (<b>right panel</b>). While IOP remained stable in WT mice as they aged from 19 to 37 weeks (14±2 to 14±2 mmHg; <i>n</i> = 25; <i>P</i> = 0.67), IOP increased in sGCα₁^−/− mice (14±2 to 18±3 mmHg; <i>n</i> = 37; *<i>P</i> = 1.9×10⁻⁸).</p

Localization of sGC α₁ and β₁ subunits in the human and murine eye.

<p>Panels <b>A–C</b> depict tissue sections from human eyes. Panels <b>D–F</b> depict tissue sections from mouse eyes. <b>A</b>: Ciliary muscle (CM), stained for α-smooth muscle actin (red), sGCα₁ (green, <b>upper panel</b>), or sGCβ₁ (green, <b>lower panel</b>). Both sGCα₁ and sGCβ₁ co-localized with α-smooth muscle actin in CM (yellow in merged images). Scale bars: 100 μm. <b>B</b>: An arteriole in the iris (IA) and an arteriole in the retina (RA) were stained for α-smooth muscle actin (red), sGCα₁ (green, <b>upper panels</b>), or sGCβ₁ (green, <b>lower panels</b>). Both sGCα₁ and sGCβ₁ co-localized with α-smooth muscle actin in the smooth muscle cell layer of arterioles in the iris and retina (yellow in merged images). ONL: outer nuclear layer, INL: inner nuclear layer. Scale bars: 20 μm. <b>C</b>: sGCα₁ (<b>left panel</b>) and sGCβ₁ (<b>right panel</b>) expression was detected histologically in the outer nuclear layer (ONL), inner nuclear layer (INL), and ganglion cell layer (GCL, white arrow) of the retina. sGCα₁ and sGCβ₁ are visualized by green fluorescence. Scale bars: 20 μm. <b>D</b>: Adjacent sections of a wild-type (WT) murine eye were stained for α-smooth muscle actin (green, <b>left panel</b>) or sGCα₁ (red, <b>right panel</b>). sGCα₁ co-localized with α-smooth muscle actin in ciliary muscle (CM) and in arterioles in the ciliary body (CA). The iridocorneal angle is indicated. Scale bars: 50 μm. <b>E</b>: Adjacent sections of a WT murine eye were stained for α-smooth muscle actin (green, <b>left panel</b>) or sGCα₁ (red, <b>right panel</b>). sGCα₁ co-localized with α-smooth muscle actin in retinal arterioles (RA). Scale bars: 50 μm. <b>F</b>: sGCα₁ (<b>left panel</b>) and sGCβ₁ (<b>right panel</b>) expression was detected histologically in the outer nuclear layer (ONL), inner nuclear layer (INL), and ganglion cell layer (GCL, white arrow) of the mouse retina. sGCα₁ is visualized by brown peroxidase stain and sGCβ₁ is visualized by green fluorescence. Scale bars: 20 μm.</p

Association between <i>GUCY1A3/GUCY1B3</i> single nucleotide polymorphisms (SNPs) and POAG in the Glaucoma Gene and Environment (GLAUGEN) study.

<p>Most significant single nucleotide polymorphisms (SNPs) stratified by gender and type of visual field (VF) loss. 51 SNPs were analyzed within the GUCY1A3/GUCY1B3 locus and 50 kb upstream and downstream of the region. A Bonferroni correction of 3.3×10⁻⁴, correcting for analyzing 51 SNPs and 3 subgroups, was applied to define statistical significance. The top SNP (rs11722059) reached significance in women with paracentral visual field (VF) loss (P value of 3.1×10⁻⁴, bold and italicized). MA: minor allele; OR: multivariable odds ratio associated with each minor allele dose, generated by multiple logistic regression analyses adjusting for age, gender, study site, DNA source, DNA extraction method, and three eigenvectors; CI: confidence interval; Chr: chromosome; N: number.</p