Hyperosmotic stress and osmo-gene adaptation during early induction of refractive errors

Abstract

Abstract Purpose: Why is myopia a common risk factor for most sight threatening disorders? Our earlier biometric, ultrastructural and elemental analyses of the chick form deprivation model have provided evidence of severe physiological, oxidative and hyperosmotic stress. More recently prolonged hyperosmotic stress has been shown to lead to chronic inflammation in a number of diseases (Brocker etal 2013). We hypothesized that perturbation of axial growth during induction of refractive errors would also be accompanied by hyperosmosis and osmoadaptative gene changes, that should be demonstratable with elemental microanalysis (EDX) and RNA seq respectively. Methods: Chicks were raised with ±10D lenses, or no lens. Following biometric measurements at 1, 2, and 3 days, 8 chicks per lens group were euthanized. RNA was extracted from the retina/RPE/choroid of 4. Four were used for scanning electron-microscopy and EDX. Libraries were sequenced on the Illumina HiSeq1500. Counts per million were imported into GSEA and expression of KEGG and Reactome pathways during myopia/hyperopia induction compared to age-matched no lens chicks (FDR cut-off <.25). Results: Refractive compensation (RC) to -10D defocus continued for 72hrs whereas RC to +10D was in near completion after 24hours. EDX shows sodium and chloride ion distributions were greatly upregulated in outer retina by -10D over the 72hrs but only at the retino-vitreal border in +10D at 72hrs. Potassium profiles in RC to +10D remained upregulated across the retina for 72 hrs with concurrent up-regulation of reactome potassium channel pathways at 72hrs in RNAseq data. Consistent with altered osmotic and oxidative stress, implicated pathways during refractive compensation included those related to synthesis of small molecule osmolytes, structural remodelling, inflammation, and metabolism. Conclusions: The EDX results demonstrate that RC to optical defocus is accompanied by hyperosmotic shifts in ion distribution profiles across the entire posterior eye, while concurrent changes in gene expression profiles were seen in metabolic and ion solute processes. These pathways have previously been associated with osmoadaptation and more severe disease states such as ARM and diabetes. The findings suggest the need for further experimental considerations of hyperosmotic changes as risk factors for severe visual impairments and for development of therapeutics