Gene regulation by Sox9 in the conjunctival epithelium
Ontology highlight
ABSTRACT:
ORGANISM(S): Mus musculus
SUBMITTER: David Beebe
PROVIDER: E-GEOD-54796 | biostudies-arrayexpress |
REPOSITORIES: biostudies-arrayexpress
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Project description:Wild type or Fgfr2 fx/fx; Le-Cre-positive (Fgfr2-/-) conjunctival epithelium of E14.5 mouse embryos was laser micodissected from three embryos of each genotype. Total RNA was purified and reverse transcribed and amplified using a NuGEN kit. cDNA was biotinylated and hybridized to Illumina MouseWG-6 v2.0 bead arrays. Three wild type and three Ffr2 knockout embryos were used. cDNA obtained from each was used for microarray analysis. Statistical analysis was performed using Illumina Genome Studio software.
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Project description:Wild type or Fgfr2 fx/fx; Le-Cre-positive (Fgfr2-/-) conjunctival epithelium of E14.5 mouse embryos was laser micodissected from three embryos of each genotype. Total RNA was purified and reverse transcribed and amplified using a NuGEN kit. cDNA was biotinylated and hybridized to Illumina MouseWG-6 v2.0 bead arrays.
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Project description:Trachoma is a poorly understood immuno-fibrogenic disease process, initiated by Chlamydia trachomatis (Ct). Differences in conjunctival gene expression profiles between Ethiopians with trachomatous trichiasis (with (TTI) and without (TT) inflammation) and controls (NC) were investigated to identify relevant host responses. Tarsal conjunctival swab samples were collected for RNA isolation and Ct PCR. Transcriptome-wide microarray experiments were conducted on 42 samples (TTI 13, TT 15, NC14). Specific results were confirmed using multiplex quantitative RT-PCR for 16 mRNA targets in an independent collection of case-control samples: 386 case-control pairs (TTI 244, TT 142, NC 386). The gene expression profiles of cases were consistent with: squamous metaplasia (Keratins, SPRR), pro-inflammatory cytokine production (IL-1β, CXCL5, S100A7) and tissue remodelling (MMP7, MMP9, MMP12, HAS3). There was no difference in the level of IFNG between cases and controls. However, cases had increased INDO, NOS2A, and IL13RA2 and reduced IL13. Ct was detected in 1/772. Cases show evidence of ongoing inflammation and tissue remodelling, which were more marked where clinical inflammation was also present. Significantly, these processes appear to be active in the absence of current Ct infection. There was limited evidence of a TH1 response (INDO, NOS2A) and no association between a TH2 response and cases. The epithelium appears actively involved in late cicatricial stages of trachoma through production of pro-inflammatory factors (IL-1β, CXCL5, S100A7). Longitudinal studies are needed to investigate which aetiological factors and pathways are associated with progressive scarring and whether simply controlling chlamydial infection will halt progression in people with established cicatricial disease. Case - control study. Conjunctival swab samples were collected from people with trachomatous trichiasis with (TTI) and without (TT) clinically visable inflammation compared to normal controls (C). Total RNA extracted and analysed on the Illumina WG-6 platrom. Additional extensive qRT-PCR work done on a large set of case-control pairs.
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Project description:Goblet cell numbers decrease within the conjunctival epithelium in drying and cicatrizing ocular surface diseases. Factors regulating goblet cell differentiation in conjunctival epithelium are unknown. Recent data indicate that the transcription factor SAM-pointed domain epithelial-specific transcription factor (Spdef) is essential for goblet cell differentiation in tracheobronchial and gastrointestinal epithelium of mice. Using Spdef -/- mice, we determined that Spdef is required for conjunctival goblet cell differentiation and that Spdef -/- mice, which lack conjunctival goblet cells, have significantly increased corneal surface fluorescein staining and tear volume, a phenotype consistent with dry eye. Microarray analysis of conjunctival epithelium in Spdef -/- mice identified 43 signficantly upregulated genes and 110 signficantly downregulated genes in the conjunctival epithelium of Spdef -/- mice compared to that of Spdef +/+ control mice (3 fold change, p<0.01). Downregulated genes of particular interested included goblet cell-specific genes (Muc5ac, Tff1, Gcnt3). Upregulated genes included epithelial cell differentiation/keratinization genes (Sprr2h, Tgm1) and pro-inflammatory genes (Il1-α, Il-1β, Tnf-α), all of which are upregulated in dry eye. Interestingly, four Wnt pathway genes were downregulated. Conjunctival epithelium of Spdef +/+ and Spdef -/- mice was collected by laser capture microdissection for RNA extraction and hybridization on Affymetrix microarrays to determine if gene expression patterns in the conjunctival epithelium of Spdef -/- mice is altered compared to that of Spdef +/+ mice.
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Project description:Wild type or Pax6 fx/fx; Le-Cre-positive surface ectoderm from E10.25 mouse embryos was laser micodissected from three embryos of each genotype. Total RNA was purified and reverse transcribed and amplified using a NuGEN kit. cDNA was biotinylated and hybridized to Illumina Mouse6 bead arrays. Three wild type and three knockout embryos were used. Each array used the cDNA obtained from the two prospective lens tissues from one embryo.
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