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RNA seq analysis of the mouse pNC and Cornea transcriptome

ABSTRACT: Purpose: The goal of this study is to provide a detailed transcriptome expression profile of the mouse cornea and evaluate changes in gene expression during mouse corneal development. Methods: RNA-sequencing is used to profile the mouse transcriptome in the pNC isolated at E10.5 and corneas isolated at E14.5 and E16.5. Samples generated in triplicate for sequencing by Illumina Hiseq 4000. Clean reads are mapped to reference gene and GRCm38 by Bowtie2 and HISAT. Spatiotemporal expression patterns of select genes were validaed by in situ hybrdization. Results: Analysis of the whole transcriptome profile between pNC and embryonic corneas identified 3815 unique differentially expressed genes. Pathway analysis revealed enrichment of differentially expressed genes involved in signal transduction (RA, TGFβ, and Wnt pathways) and transcriptional regulation. Conclusions: Our study for the first time, identifies differential gene expression across progressive stages of mouse corneal development and provides a comprehensive transcriptomic profile of the developing cornea. This serves as a valuable resource for the identification of novel regulatory networks crucial for the advancement of studies in congential defects, stem cell therapy, bioengineering, and adult corneal diseases.

ORGANISM(S): Mus musculus

PROVIDER: GSE121044 | GEO | 2019/01/31

REPOSITORIES: GEO

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Mus musculus cornea experiment

Project description:PURPOSE: To provide a detailed gene expression profile of the normal postnatal mouse cornea. METHODS: Serial analysis of gene expression (SAGE) was performed on postnatal day (PN)9 and adult mouse (6 week) total corneas. The expression of selected genes was analyzed by in situ hybridization. RESULTS: A total of 64,272 PN9 and 62,206 adult tags were sequenced. Mouse corneal transcriptomes are composed of at least 19,544 and 18,509 unique mRNAs, respectively. One third of the unique tags were expressed at both stages, whereas a third was identified exclusively in PN9 or adult corneas. Three hundred thirty-four PN9 and 339 adult tags were enriched more than fivefold over other published nonocular libraries. Abundant transcripts were associated with metabolic functions, redox activities, and barrier integrity. Three members of the Ly-6/uPAR family whose functions are unknown in the cornea constitute more than 1% of the total mRNA. Aquaporin 5, epithelial membrane protein and glutathione-S-transferase (GST) omega-1, and GST alpha-4 mRNAs were preferentially expressed in distinct corneal epithelial layers, providing new markers for stratification. More than 200 tags were differentially expressed, of which 25 mediate transcription. CONCLUSIONS: In addition to providing a detailed profile of expressed genes in the PN9 and mature mouse cornea, the present SAGE data demonstrate dynamic changes in gene expression after eye opening and provide new probes for exploring corneal epithelial cell stratification, development, and function and for exploring the intricate relationship between programmed and environmentally induced gene expression in the cornea. Keywords: other

2004-01-02 | GSE887 | GEO

Single cell RNA-seq of human cornea organoids identifies cell fates of a developing immature cornea

Project description:The cornea is a protective and refractive barrier in the eye crucial for vision. Understanding the human cornea in health, disease and cell-based treatments can be greatly advanced with cornea organoids developed in culture from induced pluripotent stem cells. While a limited number of studies have investigated the single-cell transcriptomic composition of the human cornea, its organoids have not been examined similarly. Here we elucidated the transcriptomic cell fate map of 4-month-old human cornea organoids and human donor corneas. The organoids harbor cell clusters that resemble cells of the corneal epithelium, stroma and endothelium with sub-populations that capture signatures of early developmental states. Unlike the adult cornea where the largest cell population is stromal, the organoids contain large proportions of epithelial and endothelial-like cells. These corneal organoids offer a three-dimensional model to study corneal diseases and integrated responses of different cell types.

2023-01-25 | GSE218123 | GEO

Differentially Expressed Wound Healing-Related microRNAs in the Human Diabetic Cornea

Project description:MicroRNAs are powerful gene expression regulators, but their corneal repertoire and potential changes in corneal diseases remain unknown. Our purpose was to identify miRNAs altered in the human diabetic cornea by microarray analysis, and to examine their effects on wound healing in cultured telomerase-immortalized human corneal epithelial cells (HCEC) in vitro. Using microarrays, 29 miRNAs were identified as differentially expressed in diabetic samples. Two miRNA candidates showing the highest fold increased in expression in the diabetic cornea were confirmed by Q-PCR and further characterized. HCEC transfection with h-miR-146a or h-miR-424 significantly retarded wound closure, but their respective antagomirs significantly enhanced wound healing vs. controls. Cells treated with h-miR-146a or h-miR-424 had decreased p-p38 and p-EGFR staining, but these increased over control levels close to the wound edge upon antagomir treatment. In conclusion, several miRNAs with increased expression in human diabetic central corneas were found. Two such miRNAs inhibited cultured corneal epithelial cell wound healing. Dysregulation of miRNA expression in human diabetic cornea may be an important mediator of abnormal wound healing. Total RNA was extracted from age-matched human autopsy normal (n=6) and diabetic (n=6) central corneas, Flash Tag end-labeled, and hybridized to Affymetrix® GeneChip® miRNA Arrays. Select miRNAs associated with diabetic cornea were validated by quantitative RT-PCR (Q-PCR) and by in situ hybridization (ISH) in independent samples.

2013-11-09 | E-GEOD-52233 | biostudies-arrayexpress

The tissue-engineered human cornea as a model to study expression of matrix metalloproteinases during corneal wound healing

Project description:Corneal injuries remain a major cause of consultation in the ophthalmology clinics worldwide. Repair of corneal wounds is a complex mechanism that involves cell death, migration, proliferation, differentiation, and extracellular matrix (ECM) remodeling. In the present study, we used a tissue-engineered, two-layers (epithelium and stroma) human cornea as a biomaterial to study both the cellular and molecular mechanisms of wound healing. Gene profiling on microarrays revealed important alterations in the pattern of genes expressed by tissue-engineered corneas in response to wound healing. Expression of many MMPs-encoding genes was shown by microarray and qPCR analyses to increase in the migrating epithelium of wounded corneas. Many of these enzymes were converted into their enzymatically active form as wound closure proceeded. In addition, expression of MMPs by human corneal epithelial cells (HCECs) was affected both by the stromal fibroblasts and the collagen-enriched ECM they produce. Most of all, results from mass spectrometry analyses provided evidence that a fully stratified epithelium is required for proper synthesis and organization of the ECM on which the epithelial cells adhere. In conclusion, and because of the many characteristics it shares with the native cornea, this human two layers corneal substitute may prove particularly useful to decipher the mechanistic details of corneal wound healing. Primary cultures of human corneal epithelial cells cultivated on BSA (number of replicates: 7), Collagen type I (number of replicates: 2), Collagen type IV (number of replicates: 2), Fibronectin (number of replicates: 2), Tenascin C (number of replicates: 2) and Laminin (number of replicates: 2) matrix. Central, internal and external ring of wounded Tissue-engineered human cornea.

2015-11-25 | E-GEOD-75336 | biostudies-arrayexpress

Differentially Expressed Wound Healing-Related microRNAs in the Human Diabetic Cornea

2013-11-09 | GSE52233 | GEO

Identification of gene expression changes in wild type vs. Pitx2-null cornea.

Project description:To identifycandidate PITX2-dependent genes during corneal development, microarray analysis was used to profile gene expression in corneas taken from flash-frozen e12.5 mouse embryos of control (Pitx2+/-) and temporal knockout Pitx2-deficient embryos.

2016-12-01 | GSE76047 | GEO

Quantitative metabolomic analysis of the human cornea and aqueous humor

Project description:INTRODUCTION: Cornea is the outermost part of the eye supplied mostly by aqueous humor (AH). Therefore, the comparison of the metabolomic compositions of AH and cornea may help to determine which compounds are produced inside the cornea, and which penetrate into cornea from AH for intra-corneal consumption. Keratoconus (KC) is the most common form of the cornea dystrophy, and the analysis of KC corneas can unravel the metabolomic changes occurring in AH and cornea of KC patients. OBJECTIVES: The work is aimed at the determination of concentrations of a wide range of metabolites in the human cornea and AH, the comparison of the metabolomic profiles of cornea and AH, and the comparison of the metabolomic compositions of samples taken from KC patients and normal donors (post-mortem). METHODS: The quantitative metabolomic profiling was carried out with the use of two independent methods—high-frequency 1H NMR spectroscopy and HPLC with high-resolution ESI-MS detection. RESULTS: The concentrations of 71 most abundant metabolites in cornea and AH from keratoconus patients and from human cadavers have been measured. It is found that the concentrations of purines and organic acids in cornea are significantly higher than in AH. The KC corneas are characterized by the enhanced levels of acetate and citrate, and also by low values of GSH/GSSG ratios. CONCLUSION: A significant difference in the metabolomic compositions of the human AH and cornea has been revealed. The concentrations of glucose and some amino acids in cornea are significantly lower than in AH, indicating their fast consumption inside the cornea. The high levels of organic acids, purines and GSH in cornea should be attributed to their production in the cornea. The enhanced levels of acetate and citrate as well as the low values of GSH/GSSG ratios in KC corneas are the indicators of the oxidative stress.

2018-06-20 | MTBLS444 | MetaboLights

Identification of genes associated with lens regeneration from the cornea in Xenopus laevis tadpoles

Project description:Surgical removal of the lens from larval Xenopus laevis results in a rapid transdifferention of central corneal cells to form a new lens. The trigger for this process is understood to be an induction event arising from the unprecedented contact between the cornea and the vitreous humour that occurs following lens removal. The identity of this trigger is unknown. Here, we have used a functional transgenic approach to show that BMP signalling is required for lens regeneration and a microarray approach to identify genes that are upregulated specifically during this process. Analysis of the array data strongly implicates Wnt signalling and Pitx transcription factors in this process. Pluripotency genes, in contrast, are not upregulated, supporting the idea that corneal cells transdifferentiate without returning to a stem cell state. Furthermore, several genes from the array were expressed in the forming lens during embryogenesis. One of these, nipsnap1, is a known direct target of BMP signalling. We suggest that, as with tail regeneration, activation of multiple developmental signalling pathways could drive lens regeneration from the cornea. Three biological replicates of each of three sample types were analysed. For each replicate, lens tissue (L) was derived from 5 lenses dissected from stage 50 tadpoles. Similarly, 7-8 corneas dissected from stage 50 tadpoles from which the lens had been removed 3 days previously were pooled for each biologocal replicate (R). These samples should contain tissue that is changing from cornea to lens, to regenerate the missing lens. Finally, 7-8 corneas from eyes of tadpoles at the same stage that had the cornea lifetd but the lens left in place 3 days earlier were pooled for each biological replicate (sham operated corneas, designated S).

2011-09-13 | E-GEOD-28014 | biostudies-arrayexpress

The tissue-engineered human cornea as a model to study expression of matrix metalloproteinases during corneal wound healing

2015-11-25 | GSE75336 | GEO

Expression profiling of wildtype mouse cornea in development, adult, and aging

Project description:While the mouse cornea has been well characterized morphologically, the transcriptional changes have not been described in detail. To characterize the genes, pathways, and transcriptional regulators involved in mouse cornea development and aging, we isolated whole cornea from wildtype CB6 mice at several developmental timepoints and every 6 months in the adult. Corneal epithelium and stroma were isolated at one timepoint to provide insights into the genes that are unique to each tissue. Total RNA was purified from whole mouse cornea at 12 time points over the life of the mouse and from corneal epithelium and stroma at a single timepoint.

2013-01-01 | E-GEOD-43155 | biostudies-arrayexpress

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