Cornea lenticule viability and structural integrity after refractive lenticule extraction (ReLEx) and cryopreservation

Molecular Vision173437-3449MVEP

Novel Combination BMP7 and HGF Gene Therapy Instigates Selective Myofibroblast Apoptosis and Reduces Corneal Haze In Vivo.

PurposeWe tested the potential of bone morphogenic protein 7 (BMP7) and hepatocyte growth factor (HGF) combination gene therapy to treat preformed corneal fibrosis using established rabbit in vivo and human in vitro models.MethodsEighteen New Zealand White rabbits were used. Corneal fibrosis was produced by alkali injury. Twenty-four hours after scar formation, cornea received topically either balanced salt solution (BSS; n = 6), polyethylenimine-conjugated gold nanoparticle (PEI2-GNP)-naked plasmid (n = 6) or PEI2-GNP plasmids expressing BMP7 and HGF genes (n = 6). Donor human corneas were used to obtain primary human corneal fibroblasts and myofibroblasts for mechanistic studies. Gene therapy effects on corneal fibrosis and ocular safety were evaluated by slit-lamp microscope, stereo microscopes, quantitative real-time PCR, immunofluorescence, TUNEL, modified MacDonald-Shadduck scoring system, and Draize tests.ResultsPEI2-GNP-mediated BMP7+HGF gene therapy significantly decreased corneal fibrosis in live rabbits in vivo (Fantes scale was 0.6 in BMP7+HGF-treated eyes compared to 3.3 in -therapy group; P 104 gene copies per microgram DNA of BMP7 and HGF genes. The recombinant HGF rendered apoptosis in corneal myofibroblasts but not in fibroblasts. Localized topical BMP7+HGF therapy showed no ocular toxicity.ConclusionsLocalized topical BMP7+HGF gene therapy treats corneal fibrosis and restores transparency in vivo mitigating excessive healing and rendering selective apoptosis in myofibroblasts

Evolution of Melanopsin Photoreceptors: Discovery and Characterization of a New Melanopsin in Nonmammalian Vertebrates

In mammals, the melanopsin gene (Opn4) encodes a sensory photopigment that underpins newly discovered inner retinal photoreceptors. Since its first discovery in Xenopus laevis and subsequent description in humans and mice, melanopsin genes have been described in all vertebrate classes. Until now, all of these sequences have been considered representatives of a single orthologous gene (albeit with duplications in the teleost fish). Here, we describe the discovery and functional characterisation of a new melanopsin gene in fish, bird, and amphibian genomes, demonstrating that, in fact, the vertebrates have evolved two quite separate melanopsins. On the basis of sequence similarity, chromosomal localisation, and phylogeny, we identify our new melanopsins as the true orthologs of the melanopsin gene previously described in mammals and term this grouping Opn4m. By contrast, the previously published melanopsin genes in nonmammalian vertebrates represent a separate branch of the melanopsin family which we term Opn4x. RT-PCR analysis in chicken, zebrafish, and Xenopus identifies expression of both Opn4m and Opn4x genes in tissues known to be photosensitive (eye, brain, and skin). In the day-14 chicken eye, Opn4m mRNA is found in a subset of cells in the outer nuclear, inner nuclear, and ganglion cell layers, the vast majority of which also express Opn4x. Importantly, we show that a representative of the new melanopsins (chicken Opn4m) encodes a photosensory pigment capable of activating G protein signalling cascades in a light- and retinaldehyde-dependent manner under heterologous expression in Neuro-2a cells. A comprehensive in silico analysis of vertebrate genomes indicates that while most vertebrate species have both Opn4m and Opn4x genes, the latter is absent from eutherian and, possibly, marsupial mammals, lost in the course of their evolution as a result of chromosomal reorganisation. Thus, our findings show for the first time that nonmammalian vertebrates retain two quite separate melanopsin genes, while mammals have just one. These data raise important questions regarding the functional differences between Opn4x and Opn4m pigments, the associated adaptive advantages for most vertebrate species in retaining both melanopsins, and the implications for mammalian biology of lacking Opn4x

Meibum Lipid Composition in Asians with Dry Eye Disease

10.1371/journal.pone.0024339PLoS ONE610

Effect of Fibrin Glue on the Biomechanical Properties of Human Descemet's Membrane

10.1371/journal.pone.0037456PLoS ONE75

Aberrant DNA Methylation of Matrix Remodeling and Cell Adhesion Related Genes in Pterygium

10.1371/journal.pone.0014687PLoS ONE62

Nanomedicine Approaches for Corneal Diseases

Corneal diseases are the third leading cause of blindness globally. Topical nonsteroidal anti-inflammatory drugs (NSAIDs), steroids, antibiotics and tissue transplantation are currently used to treat corneal pathological conditions. However, barrier properties of the ocular surface necessitate high concentration of the drugs applied in the eye repeatedly. This often results in poor efficacy and several side-effects. Nanoparticle-based molecular medicine seeks to overcome these limitations by enhancing the permeability and pharmacological properties of the drugs. The promise of nanomedicine approaches for treating corneal defects and restoring vision without side effects in preclinical animal studies has been demonstrated. Numerous polymeric, metallic and hybrid nanoparticles capable of transporting genes into desired corneal cells to intercept pathologic pathways and processes leading to blindness have been identified. This review provides an overview of corneal diseases, nanovector properties and their applications in drug-delivery and corneal disease management

Update on animal models of diabetic retinopathy: from molecular approaches to mice and higher mammals

Diabetic retinopathy (DR) is the most common microvascular complication of diabetes and one of the major causes of blindness worldwide. The pathogenesis of DR has been investigated using several animal models of diabetes. These models have been generated by pharmacological induction, feeding a galactose diet, and spontaneously by selective inbreeding or genetic modification. Among the available animal models, rodents have been studied most extensively owing to their short generation time and the inherited hyperglycemia and/or obesity that affect certain strains. In particular, mice have proven useful for studying DR and evaluating novel therapies because of their amenability to genetic manipulation. Mouse models suitable for replicating the early, non-proliferative stages of the retinopathy have been characterized, but no animal model has yet been found to demonstrate all of the vascular and neural complications that are associated with the advanced, proliferative stages of DR that occur in humans. In this review, we summarize commonly used animal models of DR, and briefly outline the in vivo imaging techniques used for characterization of DR in these models. Through highlighting the ocular pathological findings, clinical implications, advantages and disadvantages of these models, we provide essential information for planning experimental studies of DR that will lead to new strategies for its prevention and treatment

Dynamics of the expression of intermediate filaments vimentin and desmin during myofibroblast differentiation after corneal injury

Previous studies have suggested that abnormal corneal wound healing in patients after photorefractive keratectomy (PRK) is associated with the appearance of myofibroblasts in the stroma between two and four weeks after surgery. The purpose of this study was to examine potential myofibroblast progenitor cells that might express other filament markers prior to completion of the differentiation pathway that yields a-smooth muscle actin (SMA)-expressing myofibroblasts associated with haze localized beneath the epithelial basement membrane after PRK. Twenty-four female rabbits that had -9 diopter PRK were sacrificed at I week, 2 weeks, 3 weeks or 4 weeks after surgery. Corneal rims were collected, frozen at -80 degrees C, and analyzed by immunocytochemistry using anti-vimentin, anti-desmin, and anti-SMA antibodies. Double immunostaining was performed for the co-localization of SMA with vimentin or desmin with SMA. An increase in vimentin expression in stromal cells is noted as early as 1 week after PRK in the rabbit cornea. As the healing response continues at two or three weeks after surgery, many stromal cells expressing vimentin also begin to express desmin and SMA. By 4 weeks after the surgery most, if not all, myofibroblasts express vimentin, desmin and SMA. Generalized least squares regression analysis showed that there was strong evidence that each of the marker groups differed in expression over time compared to the other two (p < 0.01). Intermediate filaments - vimentin and desmin co-exist in myofibroblasts along with SMA and may play an important role in corneal remodeling after photorefractive keratectomy. The earliest precursors of myofibroblasts destined to express SMA and desmin are detectible by staining for vimentin at I week after surgery. (C) 2009 Elsevier Ltd. All rights reserved.[EY10056][EY015638