Current Advancements in Mouse Models of Retinal Disease.

The field of retinal degenerative (RDs) disease study has been in a state of exponential growth from discovering the underlying genetic components of such diseases as age-related macular degeneration (AMD) and retinitis pigmentosa (RP) to the first gene therapy developed and approved for human Leber congenital amaurosis. However, a source for high-fidelity animal models of these complex, multifactorial, and/or polygenic diseases is a need that has yet to be fulfilled. While models for AMD and RP do exist, they often require aging the animals for a year or more, feeding special diets, or introduction of external modulators such as exposure to cigarette smoke. Currently, work is being done to uncover high-fidelity naturally occurring models of these retinal diseases with the hope and intent of providing the vision community the tools it needs to better understand, treat, and, one day, cure the patients suffering from these devastating afflictions.

Chronic Proinflammatory Signaling Accelerates the Rate of Degeneration in a Spontaneous Polygenic Model of Inherited Retinal Dystrophy.

Collectively, retinal neurodegenerative diseases are comprised of numerous subtypes of disorders which result in loss of a varying cell types in the retina. These diseases can range from glaucoma, which results in retinal ganglion cell death, to age-related macular degeneration and retinitis pigmentosa, which result in cell death of the retinal pigment epithelium, photoreceptors, or both. Regardless of the disease, it's been recently found that increased release of proinflammatory cytokines and proliferation of active microglia result in a remarkably proinflammatory microenvironment that assists in the pathogenesis of the disease; however, many of the details of these inflammatory events have yet to be elucidated. In an ongoing study, we have used systems genetics to identify possible models of spontaneous polygenic age-related macular degeneration by mining the BXD family of mice using single nucleotide polymorphism analyses of known genes associated with the human retinal disease. One BXD strain (BXD32) was removed from the study as the rate of degeneration observed in these animals was markedly increased with a resultant loss of most all photoreceptors by 6 months of age. Using functional and anatomical exams including optokinetic nystamography, funduscopy, fluorescein angiography, and optical coherence tomography, along with immunohistochemical analyses, we show that the BXD32 mouse strain exhibits a severe neurodegenerative phenotype accompanied by adverse effects on the retinal vasculature. We also expose the concurrent establishment of a chronic proinflammatory microenvironment including the TNFα secretion and activation of the NF-κB and JAK/STAT pathways with an associated increase in activated macrophages and phagoptosis. We conclude that the induced neuronal death and proinflammatory pathways work synergistically in the disease pathogenesis to enhance the rate of degeneration in this spontaneous polygenic model of inherited retinal dystrophy.

Working memory and hippocampal expression of BDNF, ARC, and P-STAT3 in rats: effects of diet and exercise.

OBJECTIVES: Mounting evidence suggests diet and exercise influence learning and memory (LM). We compared a high-fat, high-sucrose Western diet (WD) to a plant-based, amylose/amylopectin blend, lower-fat diet known as the Daniel Fast (DF) in rats with and without regular aerobic exercise on a task of spatial working memory (WM). METHODS: Rats were randomly assigned to the WD or DF at 6 weeks of age. Exercised rats (WD-E, DF-E) ran on a treadmill 3 times/week for 30 min while the sedentary rats did not (WD-S, DF-S). Rats adhered to these assignments for 12 weeks, inclusive of ab libitum food intake, after which mild food restriction was implemented to encourage responding during WM testing. For nine months, WM performance was assessed once daily, six days per week, after which hippocampal sections were collected for subsequent analysis of brain-derived neurotrophic factor (BDNF), activity-regulated cytoskeletal protein (ARC), and signal transducer and activator of transcription 3 (P-STAT3, Tyr705). RESULTS: DF-E rats exhibited the best DSA performance. Surprisingly, the WD-S group outperformed the WD-E group, but had significantly lower BDNF and ARC relative to the DF-S group, with a similar trend from the WD-E group. P-STAT3 expression was also significantly elevated in the WD-S group compared to both the DF-S and WD-E groups. DISCUSSION: These results support previous research demonstrating negative effects of the WD on spatial LM, demonstrate the plant-based DF regimen combined with chronic aerobic exercise produces measurable WM and neuroprotective benefits, and suggest the need to carefully design exercise prescriptions to avoid over-stressing individuals making concurrent dietary changes.

Proinflammatory Pathways Are Activated in the Human Q344X Rhodopsin Knock-In Mouse Model of Retinitis Pigmentosa.

Retinitis pigmentosa (RP) is a hereditary disease of the retina that results in complete blindness. Currently, there are very few treatments for the disease and those that exist work only for the recessively inherited forms. To better understand the pathogenesis of RP, multiple mouse models have been generated bearing mutations found in human patients including the human Q344X rhodopsin knock-in mouse. In recent years, the immune system was shown to play an increasingly important role in RP degeneration. By way of electroretinography, optical coherence tomography, funduscopy, fluorescein angiography, and fluorescent immunohistochemistry, we show degenerative and vascular phenotypes, microglial activation, photoreceptor phagocytosis, and upregulation of proinflammatory pathway proteins in the retinas of the human Q344X rhodopsin knock-in mouse. We also show that an FDA-approved pharmacological agent indicated for the treatment of rheumatoid arthritis is able to halt activation of pro-inflammatory signaling in cultured retinal cells, setting the stage for pre-clinical trials using these mice to inhibit proinflammatory signaling in an attempt to preserve vision. We conclude from this work that pro- and autoinflammatory upregulation likely act to enhance the progression of the degenerative phenotype of rhodopsin Q344X-mediated RP and that inhibition of these pathways may lead to longer-lasting vision in not only the Q344X rhodopsin knock-in mice, but humans as well.

Enhanced Corneal Penetration of a Poorly Permeable Drug Using Bioadhesive Multiple Microemulsion Technology.

Corneal penetration is a key rate limiting step in the bioavailability of topical ophthalmic formulations that incorporate poorly permeable drugs. Recent advances have greatly aided the ocular delivery of such drugs using colloidal drug delivery systems. Ribavirin, a poorly permeable BCS class-III drug, was incorporated in bioadhesive multiple W/O/W microemulsion (ME) to improve its corneal permeability. The drug-loaded ME was evaluated regarding its physical stability, droplet size, PDI, zeta potential, ultrastructure, viscosity, bioadhesion, in vitro release, transcorneal permeability, cytotoxicity, safety and ocular tolerance. Our ME possessed excellent physical stability, as it successfully passed several cycles of centrifugation and freeze-thaw tests. The formulation has a transparent appearance due to its tiny droplet size (10 nm). TEM confirmed ME droplet size and revealed its multilayered structure. In spite of the high aqueous solubility and the low permeability of ribavirin, this unique formulation was capable of sustaining its release for up to 24 h and improving its corneal permeability by 3-fold. The in vitro safety of our ME was proved by its high percentage cell viability, while its in vivo safety was confirmed by the absence of any sign of toxicity or irritation after either a single dose or 14 days of daily dosing. Our ME could serve as a vehicle for enhanced ocular delivery of drugs with different physicochemical properties, including those with low permeability.

Systems Genetics of Optic Nerve Axon Necrosis During Glaucoma.

In this study, we identify genomic regions that modulate the number of necrotic axons in optic nerves of a family of mice, some of which have severe glaucoma, and define a set of high priority positional candidate genes that modulate retinal ganglion cell (RGC) axonal degeneration. A large cohort of the BXD family were aged to greater than 13 months of age. Optic nerves from 74 strains and the DBA/2J (D2) parent were harvested, sectioned, and stained with p-phenylenediamine. Numbers of necrotic axons per optic nerve cross-section were counted from 1 to 10 replicates per genotype. Strain means and standard errors were uploaded into GeneNetwork 2 for mapping and systems genetics analyses (Trait 18614). The number of necrotic axons per nerve ranged from only a few hundred to more than 4,000. Using conventional interval mapping as well as linear mixed model mapping, we identified a single locus on chromosome 12 between 109 and 112.5 Mb with a likelihood ratio statistic (LRS) of ~18.5 (p genome-wide ~0.1). Axon necrosis is not linked to locations of major known glaucoma genes in this family, including Gpnmb, Tyrp1, Cdh11, Pou6f2, and Cacna2d1. This indicates that although these genes contribute to pigmentary dispersion or elevated IOP, none directly modulates axon necrosis. Of 156 positional candidates, eight genes-CDC42 binding protein kinase beta (Cdc42bpb); eukaryotic translation initiation factor 5 (Eif5); BCL2-associated athanogene 5 (Bag5); apoptogenic 1, mitochondrial (Apopt1); kinesin light chain 1 (Klc1); X-ray repair cross complementing 3 (Xrcc3); protein phosphatase 1, regulatory subunit 13B (Ppp1r13b); and transmembrane protein 179 (Tmem179)-passed stringent criteria and are high priority candidates. Several candidates are linked to mitochondria and/or axons, strengthening their plausible role as modulators of ON necrosis. Additional studies are required to validate and/or eliminate plausible candidates. Surprisingly, IOP and ON necrosis are inversely correlated across the BXD family in mice >13 months of age and these two traits share few genes among their top ocular and retinal correlates. These data suggest that the two traits are independently modulated or that a more complex and multifaceted approach is required to reveal their association.