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ABSTRACT:
PROVIDER: PRJNA693337 | ENA |
REPOSITORIES: ENA
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Project description:In humans, the lacrimal gland produces the aqueous component of the tear film, which e.g. moistens and nourishes the ocular surface to maintain ophthalmic health. Decreased production of the aqueous component leads to the development of dry eye disease, which affects over 250 million people worldwide. Despite the impact on patients, the availability of primary human material to study underlying disease mechanisms is severely constrained. Here, we report the development of an immortalized human lacrimal gland epithelial cell line that improves accessibility to study the molecular pathogenesis-mechanisms of dry eye disease and link them to causal treatments.
2024-04-17 | GSE239464 | GEO
Project description:Dry eye is a common ocular inflammatory disorder characterized by tear film instability and reduced tear production. There is increasing evidence that homeostasis of the ocular surface is impacted by the intestinal microbiome. We are interested in investigating the potential role of microbially produced small molecules in mediating the interaction between the intestinal microbiota and the ocular surface. One such molecule is butyrate, a short-chain fatty acid (SCFA) produced by certain members of the gut microbiota through fermentation of dietary fiber. We have shown that oral administration of tributyrin, a prodrug form of butyrate, is protective of the ocular surface in mice undergoing desiccating stress. To gain insight into the mechanism, we analyzed gene expression in conjunctival tissue from mice treated with either tributyrgn or vehicle control.
2022-02-07 | GSE195578 | GEO
Project description:Glycosylation of proteins is one of the most common post-translational modification (PTM) and plays important regulatory functions in diverse biological processes such as protein stability or cell signaling. Accordingly, glycoproteins are also a consistent part of the human tear film proteome maintaining the proper function of the ocular surface and forming the first defense barrier of the ocular immune system. Irregularities in the glycoproteomic composition of the tear film might promote development of chronic eye diseases indicating glycoproteins as valuable source for biomarker discovery or drug target identification. The present study aimed to develop a lectin-based affinity method for the enrichment and concentration of tear glycoproteins/glycopeptides and the characterization of their specific N-glycosylation sites by high-resolution mass spectrometry (MS). For method development and evaluation, we accumulated first native glycoproteins from human tear sample pools and assessed the enrichment efficiency of different lectin column systems by 1D gel electrophoresis and specific protein stainings (Coomassie and glycoproteins). The best-performing multi-lectin column system (comprising the four lectins ConA, JAC, WGA and UEA I, termed as 4L) was applied for glycopeptide enrichment from human tear sample digests followed by MS-based detection and localization of their specific N-glycosylation sites. As main result, the present study identified in total 26 N glycosylation sites of 11 N-glycoproteins in tear sample pools of healthy individuals (n=3 biological sample pools). Amongst others, we identified tear film proteins lactotransferrin (N497 and N642, LTF), Ig heavy chain constant α-1 (N144 and 340, IGHA1), prolactin-inducible protein (N105, PIP) as well as extracellular lacritin (N105, LACRT) as highly reliable and significant N glycoproteins, which were already associated with the pathogenesis of various chronic eye diseases such as the dry eye syndrome (DES). In conclusion, the results of the present study will serve as important tear film N-glycoprotein catalogue for future studies focusing the human tear film and ocular surface-related inflammatory diseases.
2023-01-25 | PXD038280 | Pride
Project description:Tears are a biological fluid that has diagnostic potential for ocular diseases. Extracellular vesicles (EVs), wildly detected in various biofluids including tears, are nanoparticles released by living cells and considered as promising detection sources for non-invasive liquid biopsy. Understanding the roles of tears and tear-EVs in ocular diseases such as dry eye can facilitate the studies of clinical diagnosis, which usually entails detecting such liquid objects with a rapid and effective method. In this study, we utilized a mass spectrometry based strategy to analyze peptidome/proteome profiles of tear and EVs for rapid dry eye diagnosis. Nano-sized EVs were isolated from tears of either healthy control (HC) individuals or dry eye syndrome (DES) patients, and the tear compositions were further analyzed by tracking their fingerprints with matrix-assisted laser desorption ionization/time-of-flight mass spectrometry (MALDI-TOF-MS). The fingerprints of tear-EVs could be observed in a dose-dependent manner as well as tears, allowing comparing their discriminant peaks between tears and EVs. By analyzing these peaks, the fingerprints of both tear and tear-EVs were showed to have a capability of distinguishing DES patients from HC donors, and providing an efficient way for screening potential DES biomarkers. The proposed tear and EV fingerprinting approach is expected to be a potential tool in rapid diagnosis of ocular disease and in-depth researches of pathogenesis.
2020-08-20 | PXD020217 | Pride
Project description:Corneal architecture is essential for vision and is greatly perturbed by the absence of tears due to the highly prevalent disorder dry eye. With no regenerative therapies available, pathological alterations of the ocular surface in response to dryness, including persistent epithelial defects and poor wound healing, result in lifelong morbidity. Here, using a mouse model of aqueous-deficient dry eye, we reveal that topical application of the synthetic tear protein lacripep reverses the pathological outcomes of dry eye through restoring the extensive network of corneal nerves that are essential for tear secretion, barrier function, epithelial homeostasis and wound healing. Intriguingly, the restorative effects of lacripep occur despite extensive immune cell infiltration, suggesting tissue reinnervation and regeneration can be achieved under chronic inflammatory conditions. In summary, our data highlight lacripep as a first-in-class regenerative therapy for returning the cornea to a near homeostatic state in individuals who suffer from dry eye.
2022-07-18 | GSE208297 | GEO
Project description:Dry eye syndrome (DES) is a complex ocular condition characterized by an unstable tear film and inadequate tear production, leading to tissue damage. Despite its common occurrence, there is currently no comprehensive in vitro model that accurately reproduce the cellular characteristics of DES. Here we modified a corneal epithelium-on-a-chip (CEpOC) model to recapitulate DES by subjecting HCE-T human corneal epithelial cells to an air-liquid (AL) interface stimulus. We then assessed the effects of AL stimulation both in the presence and absence of diclofenac (DCF). Transcriptomic analysis revealed distinct gene expression changes in response to AL and AL_DCF, affecting pathways related to development, epithelial structure, inflammation, and extracellular matrix remodeling. Both treatments upregulated PIEZO2, linked to corneal damage signaling, while downregulating OCLN, involved in cell-cell junctions. They increased the expression of inflammatory genes (e.g., IL6) and reduced mucin production genes (e.g., MUC16), reflecting dry eye characteristics. TGFB1, crucial for corneal wound healing, was slightly downregulated in AL_DCF, potentially affecting wound healing processes rather than reducing inflammation by DCF. Metabolomic analysis showed increased secretion of metabolites associated with cell damage and inflammation (e.g., methyl-2-oxovaleric acid, 3-methyl-2-oxobutanoic acid, lauroyl-carnitine) in response to AL and even more with AL_DCF, indicating a shift in cellular metabolism. This study showcases the utilization of AL stimulus within the CEpOC as a comprehensive approach to faithfully reproduce the cellular characteristics of DES.
2023-10-11 | GSE244787 | GEO
Project description:“Dry eye” is a multi-factorial inflammatory disease affecting the ocular surface. Tear hyperosmolarity in dry eye contributes to inflammation and cell damage. Recent research efforts on dry eye have been directed towards biomarker discovery for diagnosis, response to treatment and disease mechanisms. This study employed a spontaneously immortalized normal human conjunctival cell line, IOBA-NHC as a model to investigate hyperosmotic stress-induced changes of metabolites and proteins. Global and targeted metabonomic analyses, as well as proteomic analysis were performed on IOBA-NHC cells incubated in serum-free media at 280 mOsm (control), 380 mOsm and 480 mOsm for 24 h. Twenty-one metabolites and seventy-six iTRAQ-identified proteins showed significant changes under at least one hyperosmotic stress treatment as compared to controls. SWATH-based proteomic analysis further confirmed the involvement of inflammatory pathways such as prostaglandin 2 synthesis in IOBA-NHC cells under hyperosmotic stress. This study is the first to identify glycerophosphocholine synthesis and O-linked β-N-acetylglucosamine glycosylation as key activated pathways in ocular surface cells under hyperosmotic stress. These findings extend the current knowledge in metabolite markers of dry eye and provide potential therapeutic targets for its treatment.
2015-08-17 | PXD002661 | Pride
Project description:“Dry eye” is a multi-factorial inflammatory disease affecting the ocular surface. Tear hyperosmolarity in dry eye contributes to inflammation and cell damage. Recent research efforts on dry eye have been directed towards biomarker discovery for diagnosis, response to treatment and disease mechanisms. This study employed a spontaneously immortalized normal human conjunctival cell line, IOBA-NHC as a model to investigate hyperosmotic stress-induced changes of metabolites and proteins. Global and targeted metabonomic analyses, as well as proteomic analysis were performed on IOBA-NHC cells incubated in serum-free media at 280 mOsm (control), 380 mOsm and 480 mOsm for 24 h. Twenty-one metabolites and seventy-six iTRAQ-identified proteins showed significant changes under at least one hyperosmotic stress treatment as compared to controls. SWATH-based proteomic analysis further confirmed the involvement of inflammatory pathways such as prostaglandin 2 synthesis in IOBA-NHC cells under hyperosmotic stress. This study is the first to identify glycerophosphocholine synthesis and O-linked β-N-acetylglucosamine glycosylation as key activated pathways in ocular surface cells under hyperosmotic stress. These findings extend the current knowledge in metabolite markers of dry eye and provide potential therapeutic targets for its treatment.
2015-08-31 | MTBLS214 | MetaboLights
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.
2013-02-08 | GSE44101 | GEO
Project description:In 2005, we determined the glyco-gene expression profile of three normal subjects ( www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE27593). This led to information on the biosynthesis of mucin O-glycans and glycoproteins that may be involved in the protection of the ocular surface. The interaction of the most highly expressed glycoprotein identified by the glyco-gene chip, galectin-3, with other ligands at the ocular surface (i.e., mucin O-glycans), is now under intense study in our laboratory, and has lead to a collaboration with another group in the glycobiology field. Patients with dry eye disease have an alteration of mucin O-glycans at the ocular surface, but the molecular mechanism(s) leading to this alteration remain unknown. In this experiment we (i) pooled three pathological samples per chip to reduce variability, and (ii) used three additional chips to include pooled control samples (normal subjects). Currently, we have RNA from 9 patients with dry eye disease, which have been divided it in 3 groups (D1, D2, and D3)—each group containing RNA pooled from 3 patients. We request three additional chips to include control samples (normal subjects). At this moment, we have RNA from 9 normal subjects, which have been divided it in 3 groups (N1, N2, and N3)—each group containing RNA pooled from 3 normal subjects. RNA was extracted in Trizol, purified using RNeasy column, and dissolved in water were sent to Microarray Core (E). The RNA was amplified, labeled, and hybridized to the GLYCOv3 microarrays. Data Analysis was done by Microarray Core (E).
2011-04-29 | E-GEOD-28941 | biostudies-arrayexpress