Dataset Information

Alteration in cellular turnover and progenitor cell population in lacrimal glands from thrombospondin 1-/- mice, a model of dry eye.

ABSTRACT: The purpose of this study was to investigate the changes that occur in the lacrimal glands (LGs) in female thrombospondin 1 knockout (TSP1-/-) mice, a mouse model of the autoimmune disease Sjogren's syndrome. The LGs of 4, 12, and 24 week-old female TSP1-/- and C57BL/6J (wild type, WT) mice were used. qPCR was performed to measure cytokine expression. To study the architecture, LG sections were stained with hematoxylin and eosin. Cell proliferation was measured using bromo-deoxyuridine and immunohistochemistry. Amount of CD47 and stem cell markers was analyzed by western blot analysis and location by immunofluorescence microscopy. Expression of stem cell transcription factors was performed using Mouse Stem Cell Transcription Factors RT2 Profiler PCR Array. Cytokine levels significantly increased in LGs of 24 week-old TSP1-/- mice while morphological changes were detected at 12 weeks. Proliferation was decreased in 12 week-old TSP1-/- mice. Three transcription factors were overexpressed and eleven underexpressed in TSP1-/- compared to WT LGs. The amount of CD47, Musashi1, and Sox2 was decreased while the amount of ABCG2 was increased in 12 week-old TSP1-/- mice. We conclude that TSP1 is necessary for maintaining normal LG homeostasis. Absence of TSP1 alters cytokine levels and stem cell transcription factors, LG cellular architecture, decreases cell proliferation, and alters amount of stem cell markers.

SUBMITTER: Shatos MA

PROVIDER: S-EPMC5121046 | biostudies-literature | 2016 Dec

REPOSITORIES: biostudies-literature

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Alteration in cellular turnover and progenitor cell population in lacrimal glands from thrombospondin 1-/- mice, a model of dry eye.

Shatos Marie A MA Hodges Robin R RR Morinaga Masahiro M McNay David E DE Islam Rakibul R Bhattacharya Sumit S Li Dayu D Turpie Bruce B Makarenkova Helen P HP Masli Sharmila S Utheim Tor P TP Dartt Darlene A DA

Experimental eye research 20160930

The purpose of this study was to investigate the changes that occur in the lacrimal glands (LGs) in female thrombospondin 1 knockout (TSP1-/-) mice, a mouse model of the autoimmune disease Sjogren's syndrome. The LGs of 4, 12, and 24 week-old female TSP1-/- and C57BL/6J (wild type, WT) mice were used. qPCR was performed to measure cytokine expression. To study the architecture, LG sections were stained with hematoxylin and eosin. Cell proliferation was measured using bromo- ...[more]

PMID: 27697548

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Project description:Purpose: This work aimed to identify differentially expressed circular RNAs (circRNAs) and elucidate their potential function in aquaporin 5 (AQP5) knockout (AQP5-/-) mice with the primary dry eye phenotype. Methods: A slit lamp examination was performed on AQP5-/- mice to assess corneal epithelial defects using fluorescein sodium staining. Hematoxylin-eosin staining and transmission electron microscopy analysis were performed to identify structural changes in lacrimal gland epithelial cells due to AQP5 deficiency. The expression profiles of circRNA and messenger RNA (mRNA) were determined by a microarray analysis. The selected circRNA was verified by quantitative real-time reverse transcription-polymerase chain reaction (qRT-PCR). Gene Ontology and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses were performed to predict the biological functions and the potential pathways of parental genes involved in lacrimal gland epithelial cell changes. According to the bioinformatics analysis of identified circRNAs, we predicted a circRNA-miRNA-mRNA network of phagosomes. Results: The AQP5-/- mice spontaneously exhibit dry eye symptoms, wherein the AQP5 deficiency changes the structure of lacrimal gland epithelial cells. The analysis revealed that, compared to AQP5+/+ mice, 30 circRNAs in the lacrimal glands of AQP5-/- mice were differentially expressed (fold change ≥ 2.0, p < 0.05). Nine upregulated circRNAs were identified using qRT-PCR, and nine upregulated validated circRNAs, 40 altered microRNAs (miRNAs), and nine upregulated mRNAs were identified through a network analysis. The KEGG analysis showed that these nine target genes were expressed in phagosomes. Conclusion: The AQP5-/- mice have primary and stable dry eye phenotypes from birth. We identified differently expressed circRNAs in the lacrimal glands of AQP5-/- and AQP5+/+ mice, predicting a circRNA-miRNA-mRNA network of phagosomes. CircRNA likely plays an important role in lacrimal gland epithelial cell pathogenesis. Therefore, it is reasonable to use circRNA as a potential therapeutic agent for the treatment of dry eyes.

Project description:Purpose: Identify the differentially expressed circular RNA (circRNA) and elucidate their potential function in AQP5 knockout (AQP5-/-) mice with primary dry eye phenotype. Methods: A slit lamp examination was performed on AQP5 knockout mice to assess corneal epithelial defect by fluorescein sodium staining. Hematoxylin-eosin staining and transmission electron microscope analysis were performed to access the structure of lacrimal gland epithelial cells. The expression profiles of circRNA and messenger RNA (mRNA) were determined by microarray analysis. The selected circRNA was verified by quantitative real-time reverse transcription polymerase chain reaction (qRT-PCR). Gene Ontology (GO) and the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis were used to predict biological functions and potential pathways of parental genes involved in lacrimal gland epithelial cell changes. According to the bioinformatics analysis of identified circRNA, we can predict a circRNA-miRNA-mRNA network. Results: AQP5-/- mice exhibit spontaneous dry eye symptoms. AQP5 deficiency obviously changes the structure of lacrimal gland epithelial cells. Analysis showed that compared to AQP5+/+ mice, 30 circRNAs in the lacrimal glands of AQP5‑/- mice had differential expression (fold change ≥ 2.0, P < 0.05). Nine upregulated circRNAs were identified by qRT-PCR; nine upregulated validated circRNAs, 40 altered microRNAs (miRNAs), and nine upregulated mRNAs were involved in the network analysis. KEGG analysis showed these nine target genes were expressed in phagosomes. Conclusions: AQP5-/- mice have primary and stable dry eye phenotypes from birth. The study identified different expressed circRNAs in lacrimal glands between AQP5-/- and AQP5+/+ mice, predicting a circRNA-miRNA-mRNA network of phagosomes. CircRNA likely plays an important role in lacrimal gland epithelial cell pathogenesis. Therefore, it is reasonable to use circRNA as a potential therapeutic target for dry eyes.

Project description:Dry eye disease can be a consequence of lacrimal gland insufficiency in Sjögren's Syndrome or increased tear film evaporation despite normal lacrimal gland function. To determine if there is a correlation between severity effects in these models and underlying pathophysiological responses, we compared the time dependent changes in each of these parameters that occur during a 6 week period. Dry eye was induced in 6-week-old female C57BL/6 mice by exposing them to an Intelligently Controlled Environmental System (ICES). Sixty mice were housed in ICES for 1, 2, 4 and 6 weeks respectively. Twelve were raised in normal environment and received subcutaneous injections of scopolamine hydrobromide (SCOP) 3 times daily for 5 days. Another sixty mice were housed in a normal environment and received no treatment. Corneal fluorescein staining along with corneal MMP-9 and caspase-3 level measurements were performed in parallel with the TUNEL assay. Interleukin-17(IL-17), IL-23, IL-6, IL-1, TNF-α, IFN-γ and TGF-β2 levels were estimated by real-time PCR measurements of conjunctival and lacrimal gland samples (LGs). Immunohistochemistry of excised LGs along with flow cytometry in cervical lymph nodes evaluated immune cell infiltration. Light and transmission electron microscopy studies evaluated LGs cytoarchitectural changes. ICES induced corneal epithelial destruction and apoptosis peaked at 2 weeks and kept stable in the following 4 weeks. In the ICES group, lacrimal gland proinflammatory cytokine level increases were much lower than those in the SCOP group. In accord with the lower proinflammatory cytokine levels, in the ICES group, lacrimal gland cytosolic vesicular density and size exceeded that in the SCOP group. ICES and SCOP induced murine dry eye effects became progressively more severe over a two week period. Subsequently, the disease process stabilized for the next four weeks. ICES induced local effects in the ocular surface, but failed to elicit lacrimal gland inflammation and cytoarchitectural changes, which accounts for less dry eye severity in the ICES model than that in the SCOP model.

Dataset Information

Alteration in cellular turnover and progenitor cell population in lacrimal glands from thrombospondin 1-/- mice, a model of dry eye.

Publications

Alteration in cellular turnover and progenitor cell population in lacrimal glands from thrombospondin 1<sup>-/-</sup> mice, a model of dry eye.

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