Project description:MicroRNA expression in the mouse eye.MicroRNAs (miRNAs) are key regulators of biological processes. To define miRNA function in the eye, it is essential to determine a high-resolution profile of their spatial and temporal distribution. In this report, we present the first comprehensive survey of miRNA expression in ocular tissues, using both microarray and RNA in situ hybridization (ISH) procedures. We initially determined the expression profiles of miRNAs in the retina, lens, cornea and retinal pigment epithelium of the adult mouse eye by microarray. Each tissue exhibited notably distinct miRNA enrichment patterns and cluster analysis identified groups of miRNAs that showed predominant expression in specific ocular tissues or combinations of them. Next, we performed RNA ISH for over 220 miRNAs, including those showing the highest expression levels by microarray, and generated a high-resolution expression atlas of miRNAs in the developing and adult wild-type mouse eye, which is accessible in the form of a publicly available web database. We found that 122 miRNAs displayed restricted expression domains in the eye at different developmental stages, with the majority of them expressed in one or more cell layers of the neural retina . This analysis revealed miRNAs with differential expression in ocular tissues and provided a detailed atlas of their tissue-specific distribution during development of the murine eye. The combination of the two approaches offers a valuable resource to decipher the contributions of specific miRNAs and miRNA clusters to the development of distinct ocular structures. microRNA profiling of ocular tissues from mouse. In particular we analysed the cornea, lens, Retina Pigment Epithelium (RPE) and retina and compared them against RNA extracted from the entire eye. The purpose of this experiment was to understand which microRNAs are present nd/or show differential expression in the various structures of the eye (cornea, lens, RPE, retina). The samples numbered 1 & 2 (i.e. CORNEA1, CORNEA2 etc ) are biological replicates, prepared from tissues dissecyed from different groups of wild-type animals. RNA extracted from the entire eye (EYE) served as the unique reference sample. For each tissue to be analysed we performed the following hybridizations: - 2 slides for lens (LENS1, LENS2) vs entire eye (EYE) - 2 slides for RPE (RPE1, RPE2) vs entire eye (EYE) - 2 slides for retina (RETINA1, RETINA2) vs entire eye (EYE) - 2 slides for cornea (CORNEA1, CORNEA2) vs entire eye (EYE) - 1 slide for entire eye (EYE) vs entire eye (EYE)

Project description:Dr. Liu's research group is interested in studying the expression and functions of galectin-3, -7 and -12, in particular the roles of these proteins in inflammation and neoplasm. Members of the galectin family are known to participate in cellular homeostasis by modulating cell growth, controlling cell cycle progression, and inducing or inhibiting apoptosis. It is known that some galectins have similar functions. However, it is not fully understood whether they work cooperatively or not. As the outermost barrier of the body, skin is directly and frequently exposed to a prooxidative environment, including solar ultraviolet A (UVA), ultraviolet B (UVB) radiation, and air pollution. Several reports have shown that exposure of cells to UV increase or decrease the levels of galectins. For example, the amounts of galectin-7 mRNA and protein are increased rapidly after UVB irradiation of keratinocytes (Proc. Natl. Acad. Sci. USA 1999; 96:11329-34). Heat shock and subculturing decrease, while alkylating agents and UV-light increase galectin-3 (Cell Physiol Biochem 2000; 10:149-58). To analyze the change of all galectin gene expression profiles after UVB irradiation and to determine the presence or absence of coordinate regulation, we analyzed the gene expression profiles of keratinocytes exposed to UVB. Normal human epidermal keratinocytes (NHEK) were irradiated with 200 J/m2 of UVB. Total RNA will be extracted at 0, 6, 12 and 24 h after irradiation (duplicate) for analysis on the Glyco gene chip. Several reports have shown that exposure to UV light can regulate levels of galectin in skin. This study seeks to analyze the changes in all galectin gene expression profiles post-UVB irradiation to determine the presence or absence of coordinate regulation. In this study, normal human keratinocytes were irradiated with 200J/m2 of UVB. Total RNA was extracted at 0, 6, 12, and 24-hour post irradiation time points, in duplicate. Samples were hybridized and analyzed using the GLYCOv2 array.

Project description:Epithelial cells and differentiated fiber cells represent distinct compartments in the ocular lens. While previous studies have revealed proteins that are preferentially expressed in epithelial vs. fiber cells, a comprehensive proteomics library comparing the molecular composition of epithelial vs. fiber cells is essential for understanding lens formation, function, disease and regenerative potential, and for efficient differentiation of pluripotent stem cells for modeling of lens development and pathology in vitro. To compare protein composition between the lens epithelium and fibers, we employed tandem mass spectrometry (2DLC/ MS) analysis of micro-dissected mouse P0.5 lenses. Functional classifications of the top 525 identified proteins into gene ontology categories by molecular process and subcellular localization, were adapted for lens. Expression levels of both epithelial and fiber proteomes were compared with their temporal and spatial mRNA levels using E14.5, E16.5, E18.5, and P0.5 RNA-Seq data sets. During this developmental time window, multiple complex biosynthetic and catabolic processes generate the molecular and structural foundation for lens transparency. As expected, crystallins showed a high correlation between their mRNA and protein levels. Comprehensive data analysis confirmed and/or predicted roles for transcription factors (TFs), RNA-binding proteins, translational apparatus including ribosomal heterogeneity and initiation factors, microtubules, cytoskeletal and membrane proteins in lens formation and maturation. Our data highlighted many proteins with unknown function in the lens that were preferentially enriched in epithelium or fibers, setting the stage for future studies to further dissect the roles of these proteins in fiber cell differentiation vs. epithelial cell maintenance. In conclusion, the present proteomic datasets established reference mouse lens epithelium and fiber cell proteomes, provided quantitative analyses of protein and RNA-Seq data, and probed the major proteome remodeling required to form the mature lens fiber cells.

Project description:Renin-angiotensin system regulates cell proliferation, differentiation, apoptosis and tissue remodeling in skin. Previouly, we reported that angiotensin-converting enzyme (ACE) expression and angiotensin II level were enhanced by repeated UVB-irradiation of hairless mice skin, and ACE inhibitor enalapril maleate accelerated recovery of UVB-induced wrinkle. In present study, we analyzed gene expression with DNA microarray and protein distribution with immunofluorecence method to clarify the process of wrinkle repairing with enalapril maleate in UVB-irradiated hairless mouse. In microarray analysis, we detected up-regulation of various extracullular matrix (ECM) genes and ECM related genes. In immunofluorescence, we confirm that type 1 collagen, fibrillin1, elastin, and dystroglycan 1 were decreased by repeated UVB-irradiation and the distribution of these components were improved by enalapril maleate treatment. In addition, ADAMTS2 and MMP-14, these were perticipate in ECM structure formation and degradation, were increased in enalapril maleate-treated mouse skin. While the functions of these molecules in skin is not clear yet, it is suggested that these molecules participate in improvement of wrinkles induced by UVB-irradiation. Male hairless mice of the SKH-1 strain were purchased from Charles River Laboratories Japan, Inc. (Tokyo, Japan). These animals were approximately 6 weeks old at the start of the experiment. They were fed a commercial diet (CRF-1, Oriental Yeast Co., Ltd, Tokyo, Japan) ad libitum and allowed free access to water. The dorsal region of the mouse was repeatedly irradiated with UVB for 10 weeks. Enalapril maleate treatment was started at 1 week after 10-week irradiation. One hundred microliters of 1% w/v enalapril maleate dissolved in 30 vol % ethanol solution or 30 vol % ethanol (control) was applied 5 times a week for 2 consecutive weeks to the whole dorsal skin of each of the mice. All experimental procedures using mice were approved by the Animal Experiment Committee of Tokyo University of Agriculture and Technology (approval number 24-82). The dosal region of the mouse was repeatedly irradiated with UVB for 10 weeks. Enalapril maleate treatment was started at 1 week after 10-week irradiation. One hundred microliters of 1% w/v enalapril maleate dissolved in 30 vol % ethanol solution or 30 vol % ethanol (control) was applied 5 times a week for 2 or 6 consecutive weeks to the whole dorsal skin of each of the mice. Each of the experimental groups comprised 6 mice. After 2- or 6-week drug treatment, skin samples were collected for microarray analysis.

Dataset Information

UVB irradiation effect on mouse lens epithelium

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure