Single-cell RNA transcriptome defines the signaling networks stimulated by alkali burn
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ABSTRACT:
ORGANISM(S): Mus musculus
PROVIDER: GSE275678 | GEO | 2025/05/19
REPOSITORIES: GEO
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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.
2025-05-19 | GSE277598 | GEO
Project description:Purpose Corneal neovascularization (CNV) impairs corneal transparency and visual acuity. The study aims to deepen our understanding of the molecules involved in CNV induced by alkali burns, facilitate a better grasp of CNV mechanisms, and uncover potential therapeutic targets. Methods Mice were selected for establishing CNV models via alkali burns. On days 3, 7, and 14 after the burns, corneal observations and histological investigations were conducted. An integrated analysis of RNA sequencing (RNA-seq)-based transcriptomics and label-free quantitative proteomics was performed in both normal and burned corneas. Bioinformatics approaches, encompassing GO and KEGG analysis, were applied to discern differentially expressed genes (DEGs) and crucial signaling pathways. Four potentially CNV-related genes were validated using qRT-PCR and Western blot. Results Significant CNV was observed on the seventh day. Forty-one genes were differentially expressed in neovascularized corneas, with 15 upregulated and 26 downregulated at both mRNA and protein levels. Bioinformatics analysis revealed that these DEGs participated in diverse biological processes, encompassing retinol and retinoic acid metabolism, neutrophil chemotaxis, and actin filament assembly, along with significant enrichment pathways like cytochrome P450, tyrosine, and phenylalanine metabolism. The upregulation of lymphocyte cytosolic protein 1 (LCP1) and cysteine and glycine-rich protein 2 (CSRP2) genes and the downregulation of transglutaminase 2 (TGM2) and transforming growth factor-beta-induced (TGFBI) genes were confirmed. Conclusions We analyzed gene expression differences in mouse corneas seven days after alkali burns, finding 41 genes with altered expression. The exact role of these genes in CNV is not fully understood, but exploring angiogenesis-related molecules offers potential for CNV treatment or prevention.
2024-01-26 | PXD047367 | Pride
Project description:In order to find out the vital genes during the corneal neovascularization (CNV), we set up two CNV models, one being induced by suture placement and the other by alkali burn to the corneal center. Young adult Balb/c and C57BL/6 were used. The corneas were checked daily for CNV development and some of the corneas were removed at two pre-determined time points for RNA extraction. Preliminary experiment indicated slightly different dynamics in suture- or chemical burn-induced CNV. Vigorous growth appeared at day 5 and day 6 in S-CNV and CB-CNV, and reached maximum length around day 10 and day 14, respectively, in S-CNV and CB-CNV. To check whether genetic MHC background has any effect on development of CNV, we compared CB-CNV in Balb/c and C57BL/6 mice at day 6. Corneas from Balb/c mice were isolated 5 and 10 days after suture-induced CNV, and 6 and 14 days after chemical burn-induced CNV. Corneas from C57BL/6 mice were isolated 6 days after chemical burn-induced CNV. Each time point had two or three biological replications.
2011-02-08 | E-GEOD-23347 | biostudies-arrayexpress
Project description:In order to find out the vital genes during the corneal neovascularization (CNV), we set up two CNV models, one being induced by suture placement and the other by alkali burn to the corneal center. Young adult Balb/c and C57BL/6 were used. The corneas were checked daily for CNV development and some of the corneas were removed at two pre-determined time points for RNA extraction. Preliminary experiment indicated slightly different dynamics in suture- or chemical burn-induced CNV. Vigorous growth appeared at day 5 and day 6 in S-CNV and CB-CNV, and reached maximum length around day 10 and day 14, respectively, in S-CNV and CB-CNV. To check whether genetic MHC background has any effect on development of CNV, we compared CB-CNV in Balb/c and C57BL/6 mice at day 6.
2011-02-08 | GSE23347 | GEO
Project description:Current drugs that directly target pro-angiogenic factors to inhibit or reverse corneal neovascularization, the major sight-threatening pathology caused by angiogenic stimuli, require multiple rounds of administration and have limited efficacies. Here we report the profiling of anti-angiogenic corneal microRNAs (miRNAs), and a framework that employs discovered miRNAs as biotherapies deliverable by recombinant adeno-associated viruses (rAAVs). By querying differentially expressed miRNAs in neovascularized mouse corneas induced by alkali-burn, we have revealed 39 miRNAs that are predicted to target more than 5,500 differentially expressed corneal mRNAs. Among these corneal miRNAs, we selected miR-204 and assessed its efficacy as a therapeutic miRNA in injured corneas. Our results show that delivery of miR-204 by rAAV is efficacious and safe for mitigating corneal NV. Overall, our work demonstrates the discovery of therapeutic miRNAs in corneal disorders and their translation into viable clinical vectors.
2018-10-26 | GSE89537 | GEO
Project description:Current drugs that directly target pro-angiogenic factors to inhibit or reverse corneal neovascularization, the major sight-threatening pathology caused by angiogenic stimuli, require multiple rounds of administration and have limited efficacies. Here we report the profiling of anti-angiogenic corneal microRNAs (miRNAs), and a framework that employs discovered miRNAs as biotherapies deliverable by recombinant adeno-associated viruses (rAAVs). By querying differentially expressed miRNAs in neovascularized mouse corneas induced by alkali-burn, we have revealed 39 miRNAs that are predicted to target more than 5,500 differentially expressed corneal mRNAs. Among these corneal miRNAs, we selected miR-204 and assessed its efficacy as a therapeutic miRNA in injured corneas. Our results show that delivery of miR-204 by rAAV is efficacious and safe for mitigating corneal NV. Overall, our work demonstrates the discovery of therapeutic miRNAs in corneal disorders and their translation into viable clinical vectors.
2018-10-26 | GSE89536 | GEO
Project description:For decades, it has been well accepted that corneal epithelial stem cells and their immediate progeny, the early transit amplifying (eTA) cells, reside in the limbal epithelial basal layer. Activation of quiescent stem/eTA cells is required for proper re-epithelialization during wound healing. The molecular profile of activated stem/eTA cells remains unclear because of difficulties in obtaining discrete cell populations for analyses. Single cell RNA sequencing (scRNA-seq) technology can profile the transcriptome at a single cell level, providing information on how stem/eTA cell activation is regulated. Using this technology, we report that ACE2, a key component in the renin-angiotensin system (RAS), functions as a negative regulator of stem/eTA activation. Methods: Mouse corneal epithelium was exposed to 1M NaOH for 30s or mechanically removed with a diamond burr. Corneas were processed for scRNA-seq and data was analyzed using R with a Seurat package. Limbal epithelial cell proliferation was assessed using BrdU incorporation. RT-qPCR, western blotting and immunostaining were conducted to determine the change of gene expression. Results: ACE2 was predominantly expressed in the stem cell-enriched limbal basal epithelium. scRNA-seq combined with GO analysis suggested that ACE2 was involved in limbal stem/eTA cell proliferation. Interestingly, immunostaining and RT-qPCR indicated that ACE2 expression was reduced following corneal injuries. Reduction in ACE2 promoted proliferation in human limbal epithelial cell culture as well as in mouse limbal epithelium after corneal epithelial debridement. Significantly, the negative effect of ACE2 on proliferation was not reversed following treatment with the angiotensin II receptor blocker losartan, indicating that the function of ACE2 in limbal epithelium is independent of RAS. scRNA-seq also revealed that reduction of ACE2 caused activation of the TGFA/EGFR pathway, which reduced expression of Lcn2. Lcn2 is a negative regulator of proliferation in a variety of cells. Inhibition of EGFR or overexpression of Lcn2 reversed the increased proliferation in limbal epithelial cells lacking ACE2. Conclusion: Our findings strongly support the idea that in response to corneal injury, ACE2 is downregulated, which results in the activation of stem/eTA cell proliferation via a novel TGFA/EGFR/Lcn2 signaling pathway in an angiotensin-independent way.
2024-07-15 | GSE263994 | GEO
Project description:We report conditional ablation of lymphatic Vegfr3 alleviated inflammatory process in alkali injured iris. Prox1CreERT2 mice were crossed with Vegfr3fl/fl to selectively target gene expression in Prox1+ lymphatic cells. Injury models were established at 6 weeks postnatally, and Vegfr3 gene depletion was conducted via Tamoxifen injection on the day 13th-17th after corneal alkali injury. Mice were sacrificed 28th days post-injury.
2024-05-29 | GSE240269 | GEO
Project description:Single-cell RNA transcriptome defines the signaling networks stimulated by alkali burn
| PRJNA1152563 | ENA
Project description:Alkali stress is an important means of inactivating undesirable pathogens in a wide range of situations, ranging from environmental cleaning of food processing environments, to the phagolysosomal killing of cells engulfed by mammalian phagocytes. Unfortunately, L. monocytogenes can launch an alkaline tolerance response (AlTR), significantly increasing persistence of the pathogen in such environments. This study compared the transcriptome patterns of alkali stressed and non alkali stressed L. monocytogenes 10403S cells, to elucidate the mechanisms by which this important pathogen adapts and/or grows during short or long-term alkali stress. Transcription profiles associated with alkali shock (AS) responses were obtained by DNA microarray analysis of mid-exponential cells suspended in pH 9 media for 15, 30 or 60 min. Transcription profiles associated with alkali adaptation (AA) were obtained by DNA microarray analysis of cells grown to mid-exponential phase in pH 9 media . Comparison of AS and AA transcription profiles with profiles from control (pH 7.0) cells identified over 2,000 genes that were differentially expressed under alkaline conditions. Rapid (15min) changes in expression included upregulation of genes encoding for multiple metabolic pathways, (including those associated with Na+/H+ antiporters), ABC transporters of functional compatible solutes such as carnitine, motility and virulence-associated genes as well as the σB controlled stress resistance network. Slower (30min and more) responses to AS and adaptation during growth in alkaline conditions (AA), included modest changes in mRNA concentrations, and genes involved in proton export. Keywords: Time course study of gene expression response to alkaline shock and adaptation
2007-07-25 | GSE7854 | GEO