Transcription profiling of mouse cornea from three strains reveal central corneal thickness is a genetic dependent trait among inbred strains of mice
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ABSTRACT: Central corneal thickness (CCT) exhibits broad variability. We determined the corneal gene expression profile three mouse strains with distinct corneal thickness: C57BLKS/J (88.6 um), SJL/J (123.5 um), and C57BL/6J (100.1 um). Experiment Overall Design: Enucleated eyes from 4 month old mice were dissected in phosphate-buffered saline and a punch of central cornea was collected utilizing a 2-mm biopsy punch. Two central corneal punches (left and right eyes) were pooled from each mouse to form one sample; three samples were analyzed per strain.
Project description:<p>The Ocular Hypertension Treatment Study (OHTS) is an National Eye Institute-sponsored multi-center, randomized, prospective treatment trial designed to determine whether lowering intraocular pressure (IOP) in individuals with ocular hypertension delays or prevents the development of primary open angle glaucoma (POAG). A total of 1,636 individuals with ocular hypertension between 40 and 80 years old were enrolled in the study. In addition to ocular hypertension, subjects in the OHTS were required to have normal optic nerve appearance as determined by the OHTS Optic Disc Reading Center and normal and reliable visual field tests at the time of enrollment by the OHTS Visual Field Reading Center. OHTS subjects were randomly assigned to either an observational group which received close observation or a topical medication group which received medication as needed to achieve a 20% reduction in IOP from their baseline levels. Subjects then were examined at regular intervals for optic disc cupping or visual field defects. Other clinical measures were also obtained from OHTS subjects including central corneal thickness (CCT) and intraocular pressure. The primary outcome monitored for the OHTS was the development of glaucoma in one or both eyes as defined by reproducible visual field abnormality or reproducible optic disc deterioration attributed to POAG by the masked OHTS Endpoint Committee. The OHTS study design has been reported in detail (Gordon, 1999; PMID: <a href="http://www.ncbi.nlm.nih.gov/pubmed/10326953" target="_blank">10326953</a>).</p> <p>The OHTS confirmed that ocular hypertension is a risk factor for developing POAG and showed that lowering IOP reduced the risk for developing POAG (Kass, 2002; PMID: <a href="http://www.ncbi.nlm.nih.gov/pubmed/12049574" target="_blank">12049574</a>). The OHTS also demonstrated that thin CCT is a significant risk factor for the development of POAG (Gordon, 2002; PMID: <a href="http://www.ncbi.nlm.nih.gov/pubmed/12049575" target="_blank">12049575</a>).</p> <p>Blood samples were also collected from 1,077 OHTS participants for an ancillary genetics study. DNA was prepared from these samples and used in a genome-wide association scan (GWAS) designed to identify genetic factors that control the magnitude of quantitative features of glaucoma (baseline IOP, baseline cup-to-disc ratio, and CCT). Genotypes from the GWAS and these clinical data (IOP, cup-to-disc ratio, and CCT) have been provided to dbGaP.</p>
Project description:The precise role of long non-coding RNAs (lncRNAs) as key epigenetic regulators in mediating corneal epithelial wound healing remains elusive. Here, we aim to elucidate the functional contribution of lncRNAs in regulating CEWH. We used Microarray to characterize lncRNA expression profiling during mouse CEWH. The full thickness of the corneal epithelium of C57BL/6 mice, was removed using a 0.5-mm corneal rust ring remover. Subsequently, the entire corneal epithelium was collected from both the injured and fellow eyes for RNA isolation after approximately 48 hours. Microarray analysis revealed dysregulation of numerous lncRNA candidates during CEWH. Out of the 41,655 non-coding RNAs detected by the microarray, 639 were upregulated and 1,006 were downregulated during the process of CEWH.
Project description:To examine whether the upregulation of Ifitm1 is essential for the expansion of stem/early TA cells in response to corneal injury, wild type mouse eyes were topically treated with AAV-shRNA-Ifitm1 or AAV-empty vector (control) and subsequently were subjected to corneal NaOH burn. AAV-shRNA-Ifitm1 treatment downregulated Ifitm1 expression in mouse limbal epithelium . A scRNA-seq assay was conducted using cells isolated from corneal/limbal tissues at three days after corneas were injured. scRNA-seq assay identified stem/early TA cell population using Gpha2, a limbal epithelial stem /eTA cell marker. In AAV-EV treated mouse eyes, NaOH injury increased the numbers of the stem/eTA cells compared to uninjured eyes. Such expansion of stem/eTA cell population following corneal injury was markedly reduced in AAV-shRNA-Ifitm1 treated eyes. This suggests a positive role of Ifitm1 in stem/early TA cell expansion after corneal injury.
Project description:human corneal epithelial cells were isolated from healthy human donor eyes. Cells were cultured with irradiated 3T3 (i3T3) murine fibroblast feeder cells, irradiated human corneal fibroblasts (iHFL) or without feeder layer
Project description:The purpose of the project was to profile protein expression liquid vitreous biopsies from uveal melanoma (UM) patients using mass spectrometry, to identify prognostic biomarkers, signaling pathways, and therapeutic targets. Vitreous biopsies were collected from two cohorts: comparative control eyes with epiretinal membranes (ERM) and test eyes with UM. Samples were analyzed using liquid chromatography-tandem mass spectrometry (LC-MS/MS).
Project description:Transplantation of ex vivo expanded limbal stem cells (LSC) is the main treatment for limbal stem cell deficiency though the clinical problem of donor tissues shortage. Recently, as the development of tissue engineering, embryonic stem cells (ESC) derived corneal epithelial-like cells (ESC-CEC) has become a new direction to this issue.Our group successfully induced ESC into corneal epithelial-like cells, and in the present study we explored various aspects of physiological properties of ESC-CEC. The experiment included three samples: hES, the human embryonic stem cell line H1, RA_SB, the corneal epithelial-like cells derived from hES by differentiation with RA and SB, epithelial_cell, the primary human limbal stem cells from cadaver eyes. hES, the human embryonic stem cell line H1, RA_SB, the corneal epithelial-like cells derived from hES by differentiation with RA and SB, epithelial_cell, the primary human limbal stem cells from cadaver eyes.
Project description:Purpose: Exposure to sulfur mustard (SM) may result in severe ocular injuries. While some of the eyes show a clinical resolution of the injury (defined as clinically non-impaired), part of the eyes develop irreversible late ocular pathologies (defined as clinically impaired) that may lead to corneal blindness. Understanding the pathological mechanisms underlying the development of the late pathology may lead to improved treatment options. Therefore, this study aimed to investigate the mRNA expression profiles of corneas from clinically impaired, clinically non-impaired and naïve eyes. Methods: Rabbit eyes were exposed to SM vapor and a clinical follow-up was carried out up to 4 weeks using a slit lamp microscope. At this time point, corneal tissues from clinically impaired, clinically non-impaired and naïve eyes were processed for RNA sequencing (RNA-seq) and differential expression analyses. The differential expression profiles were further subjected to pathway enrichment analysis using Ingenuity Pathway Analysis (IPA). Real-time PCR was used for RNA-seq validation. Results: The late pathology developed in 54%-80% of the eyes following ocular exposure to SM, clinically manifested by inflammation, corneal opacity and neovascularization. RNA-seq results showed significant differences in mRNA levels of hundreds of genes.xls between clinically impaired, clinically non-impaired and naïve corneas. Pathway enrichment analysis showed common pathways that were activated in all of the exposed eyes, such as Th1 and Th2 activation pathway, in addition to pathways that were activated only in the clinically impaired eyes compared to the clinically non-impaired eyes, such as IL-6 and ERK5 signaling. Conclusions: Corneal mRNA expression profiles for the clinically impaired, clinically non-impaired and naïve eyes generated a comprehensive database that revealed new factors and pathways, which for the first time were shown to be involved in SM-induced late pathology. Our data may contribute to the research on both the pathological mechanisms that are involved in the development of the late pathology and the protective pathways that are activated in the clinically non-impaired eyes and may point out towards novel therapeutic strategies for this severe ocular injury.
Project description:Observational prospective multicenter study to investigate efficacy and safety of endoscopic full thickness resection in the lower GI tract using a novel over-the-scope full thickness resection device.