The role of interferon regulatory factor 8 for retinal tissue homeostasis and development of choroidal neovascularisation
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ABSTRACT:
PROVIDER: PRJNA756533 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR15538635.fastq.gz | Fastqsanger.gz | |||
| SRR15538637.fastq.gz | Fastqsanger.gz | |||
| SRR15538639.fastq.gz | Fastqsanger.gz | |||
| SRR15538641.fastq.gz | Fastqsanger.gz | |||
| SRR15538643.fastq.gz | Fastqsanger.gz |
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Project description:Microglia represent the resident innate immune cells of the retina and are important for retinal development and tissue homeostasis. However, dysfunctional microglia can have a negative impact on the structural and functional integrity of the retina under native and pathological conditions. In this study, we used interferon-regulating factor 8 (Irf8)-deficient mice to determine the transcriptional profile, morphology, and temporal-spatial distribution of microglia lacking Irf8 and to examine the effects on retinal development, tissue homeostasis, and choroidal neovascularization (CNV) formation. We hereby found a severely altered morphological phenotype accompanied by loss of microglia signature genes in IRF8 deficient microglia. An in-depth characterization by funduscopy, fluorescein angiography, optical coherence tomography and electroretinography revealed no major retinal abnormalities. However, in the laser-induced CNV model, Irf8-deficient microglia showed increased activity of biological processes critical for inflammation and cell adhesion and reduced cell number near the lesions, which was associated with significantly increased CNV lesion size. Our results suggest that Irf8 has negligible functions in the steady state but is critical in converting resident microglia to a reactive phenotype under pathologic conditions and thus can suppress retinal inflammation and CNV formation.
2021-08-27 | GSE182504 | GEO
Project description:Consistent with clinical observations that posterior uveitis frequently involves the retinal vasculature and recent recognition of vascular heterogeneity, we hypothesized that retinal vascular endothelium was a cell population of unique molecular phenotype. Donor-matched cultures of primary retinal and choroidal endothelial cells from 6 human cadavers were incubated with either Toxoplasma gondii tachyzoites (10:1, parasite:cell) or Escherichia coli lipopolysaccharide (100 ng/mL); control cultures were simultaneously incubated with medium. Gene expression profiling of endothelial cells was performed using oligonucleotide arrays containing probes designed to detect 8747 human transcripts. After normalization, differential gene expression was assessed by the significance analysis of microarrays, with the false discovery rate set at 5 %. For selected genes, differences in level of expression between retinal and choroidal cells were evaluated by real-time RT-PCR. Graphical descriptive analysis demonstrated strong correlation between gene expression of unstimulated retinal and choroidal endothelial cells, but also highlighted distinctly different patterns of expression that were greater than differences noted between donors or between unstimulated and stimulated cells. Overall, 779 of 8,747 transcripts (8.9 %) were differentially represented. Notably, the 330 transcripts that were present at higher levels in retinal cells included a larger percentage of transcripts encoding molecules involved in the immune response. Differential gene expression was confirmed for 12 transcripts by RT-PCR. Retinal and choroidal vascular endothelial cells display distinctive gene expression profiles. Our findings suggest the possibility of treating posterior uveitis by targeting specific interactions between the retinal endothelial cell and an infiltrating leukocyte. Experiment Overall Design: Retinal and choroidal vascular endothelial cells were isolated from both eyes of 6 human cadaver donors. Endothelial cells of each subtype from each donor were separately pooled and cultured. In a first series of experiments (Experiment 1), donor-matched cultures of retinal and choroidal endothelial cells derived from 3 donors were incubated with either T. gondii tachyzoites or medium alone. In a second set of experiments (Experiment 2), cultured retinal and choroidal endothelial cells derived from the other 3 donors were stimulated with LPS or medium alone. Whenever possible, 2 replicate dishes of endothelial cells were subjected to each set of conditions. Subsequent to incubations, mRNA was extracted from a cell lysate prepared from each dish of cells, and cRNA derived from each mRNA preparation was separately hybridized to one oligonucleotide expression array.
2008-06-15 | E-GEOD-7850 | biostudies-arrayexpress
Project description:PURPOSE Diseases that involve choroidal or retinal endothelial vascular cells are leading causes of vision loss: age-related macular degeneration, retinal ischemic vasculopathies and non-infectious posterior uveitis. Proteins differentially expressed by these endothelial cell populations are potential drug targets. We used deep proteomics to define the molecular phenotype of human choroidal and retinal endothelial cells at the protein level. METHODS Choroidal and retinal endothelial cells were separately isolated from 5 human eye pairs by selection on CD31. Total protein was extracted and digested, and peptide fractions were analysed by reverse-phase liquid chromatography tandem mass spectrometry. Peptide sequences were assigned to fragment ion spectra and proteins were inferred using public protein databases. Protein abundance was determined by spectral counting. Protein expression in choroidal versus retinal endothelial cells was compared using edgeR package in R, and annotation enrichment analysis was performed. RESULTS Human choroidal or retinal endothelial cells expressed 5042 non-redundant proteins. Setting the false discovery rate at 5%, 498 proteins (14.4%) of 3454 quantifiable proteins with minimum mean spectral counts of 2.5 were differentially expressed between cell populations. Choroidal and retinal endothelial cells were enriched in angiogenic proteins, and retinal endothelial cells were also enriched in immunologic proteins. CONCLUSIONS This work demonstrates the protein heterogeneity of human choroidal and retinal vascular endothelial cells and provides multiple candidates for further study as novel treatments or drug targets for posterior eye diseases.
2018-03-26 | PXD005972 | Pride
Project description:Consistent with clinical observations that posterior uveitis frequently involves the retinal vasculature and recent recognition of vascular heterogeneity, we hypothesized that retinal vascular endothelium was a cell population of unique molecular phenotype. Donor-matched cultures of primary retinal and choroidal endothelial cells from 6 human cadavers were incubated with either Toxoplasma gondii tachyzoites (10:1, parasite:cell) or Escherichia coli lipopolysaccharide (100 ng/mL); control cultures were simultaneously incubated with medium. Gene expression profiling of endothelial cells was performed using oligonucleotide arrays containing probes designed to detect 8747 human transcripts. After normalization, differential gene expression was assessed by the significance analysis of microarrays, with the false discovery rate set at 5 %. For selected genes, differences in level of expression between retinal and choroidal cells were evaluated by real-time RT-PCR. Graphical descriptive analysis demonstrated strong correlation between gene expression of unstimulated retinal and choroidal endothelial cells, but also highlighted distinctly different patterns of expression that were greater than differences noted between donors or between unstimulated and stimulated cells. Overall, 779 of 8,747 transcripts (8.9 %) were differentially represented. Notably, the 330 transcripts that were present at higher levels in retinal cells included a larger percentage of transcripts encoding molecules involved in the immune response. Differential gene expression was confirmed for 12 transcripts by RT-PCR. Retinal and choroidal vascular endothelial cells display distinctive gene expression profiles. Our findings suggest the possibility of treating posterior uveitis by targeting specific interactions between the retinal endothelial cell and an infiltrating leukocyte. Keywords: Cell type comparison, stress response
2007-05-25 | GSE7850 | GEO
Project description:Retinal pigment epithelial (RPE) cells and choroidal stromal fibroblast (CSF) were isolated from healthy human donor eyes. Cells were cultured and RNA extracted.
2018-11-25 | GSE122885 | GEO
Project description:BackgroundMicroglia cells represent the resident innate immune cells of the retina and are important for retinal development and tissue homeostasis. However, dysfunctional microglia can have a negative impact on the structural and functional integrity of the retina under native and pathological conditions.MethodsIn this study, we examined interferon-regulatory factor 8 (Irf8)-deficient mice to determine the transcriptional profile, morphology, and temporospatial distribution of microglia lacking Irf8 and to explore the effects on retinal development, tissue homeostasis, and formation of choroidal neovascularisation (CNV).ResultsOur study shows that Irf8-deficient MG exhibit a considerable loss of microglial signature genes accompanied by a severely altered MG morphology. An in-depth characterisation by fundus photography, fluorescein angiography, optical coherence tomography and electroretinography revealed no major retinal abnormalities during steady state. However, in the laser-induced CNV model, Irf8-deficient microglia showed an increased activity of biological processes critical for inflammation and cell adhesion and a reduced MG cell density near the lesions, which was associated with significantly increased CNV lesion size.ConclusionsOur results suggest that loss of Irf8 in microglia has negligible effects on retinal homeostasis in the steady state. However, under pathological conditions, Irf8 is crucial for the transformation of resident microglia into a reactive phenotype and thus for the suppression of retinal inflammation and CNV formation.
| S-EPMC8454118 | biostudies-literature
Project description:Background: To identify and functionally annotate cell type-specific gene expression in the human retinal pigment epithelium (RPE), a key tissue involved in age-related macular degeneration and retinitis pigmentosa. Methodology: RPE, photoreceptor and choroidal cells were isolated from selected freshly frozen healthy human donor eyes using laser microdissection. RNA isolation, amplification and hybridization to 44k microarrays was carried out according to Agilent specifications. Bioinformatics was carried out using Rosetta Resolver, David and Ingenuity software. Principal Findings: Our previous 22k analysis of the RPE transcriptome showed that the RPE has high levels of protein synthesis, strong energy demands, is exposed to high levels of oxidative stress and a variable degree of inflammation. We currently use a complementary new strategy aimed at the identification and functional annotation of RPE-specific expressed transcripts. This strategy takes advantage of the multilayered cellular structure of the retina and overcomes a number of limitations of previous studies. In triplicate, we compared the transcriptomes of RPE, photoreceptor and choroidal cells and we deduced RPE specific expression. We identified at least 114 entries with RPE-specific gene expression. Thirty-nine of these 114 genes also show high expression in the RPE, comparison with the literature showed that 85% of these 39 were previously identified to be expressed in the RPE. In the group of 114 RPE specific genes there was an overrepresentation of genes involved in (membrane) transport, vision and ophthalmic disease. More fundamentally, we found RPE-specific involvement in the RAR-activation, retinol metabolism and GABA receptor signaling pathways. Conclusions: In this study we provide a further specification and understanding of the RPE transcriptome by identifying and analyzing genes that are specifically expressed in the RPE. Six RPE samples, three phot samples, three choroid samples
2010-06-08 | E-GEOD-20191 | biostudies-arrayexpress
Project description:We investigated diurnal changes in gene expression in the retina and choroid at the onset of experimental myopia. We induced visual form deprivation myopia in young chicks, and isolated retinal and choroidal tissues separately in form- deprived and contralateral control eyes every 4 hours over a single 24-hour period.
2023-12-20 | GSE227724 | GEO
Project description:Neovascular age-related macular degeneration represents the most common cause of blindness in the western world. Alterations of the outer Blood-retina barrier integrity and a localized inflammatory microenvironment lead to sprouting of choroidal neovascularization in intimate contact with surrounding myeloid cells and ultimately lead to visual impairment. The discovery of novel targets interfering with angiogenesis and inflammation is vital for the future treatments in AMD patients. To identify novel potential targets in the local phagocytes of the retina, microglia, we performed a comprehensive RNA-seq analysis in the mouse model of laser-induced choroidal neovascularization (mCNV). Here, we identified the angiogenic factor Osteopontin (Opn), also known as "secreted phosphoprotein 1” (Spp1), to be one of the most highly expressed genes in retinal microglia in the course of CNV formation. We could confirm the presence of SPP1 at the lesion site in recruited retinal microglia of Cx3cr1CreER:Rosa26-Tomato reporter mice using immunohistochemistry and in whole retinal tissue lysates by ELISA compared to controls highlighting a massive local production of SPP1. Inhibition of SPP1 by intravitreal injection of anti-SPP1 antibody significantly increased the lesion size compared to IgG-treated control eyes. In line with the results in rodents, we found an increased SPP1 mRNA expression in surgically extracted human choroidal neovascular (hCNV) membranes by the quantitative RNA-seq approach of massive analysis of cDNA ends (MACE) and found numerous IBA1+SPP1+ myeloid cells in human CNV membranes. Taken together, these results highlight the importance of SPP1 in the formation of CNV and potentially offer new opportunities for therapeutic intervention by inhibiting the SPP1 pathway.
2021-02-16 | GSE160011 | GEO
Project description:Following laboratory and clinical findings implicating circadian biology in the pathogenesis of myopia (nearsightedness), we examined gene expression in two crucial tissues controlling post-natal refractive development, the retina and choroid. Inducing unilateral visual form deprivation myopia in young chicks, a widely studied and validated model, we isolated retinal and choroidal tissues every 4 hours over a single day from myopic and contralateral control eyes during a period time when myopia progresses rapidly.
2024-07-01 | GSE261232 | GEO