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The formation and maintenance of these barriers is dependent on a series of cellCcell contacts that circumscribe the apical-lateral margin of each cell, known collectively as the apical junction complex (AJC). This complex includes the adherens junction (AJ), which promotes cells integrity by creating a strong adhesive interface between individual cells (Harris and Tepass, 2010 ), and the limited junction (TJ), which forms a physical barrier to the movement between cells of ions, macromolecules, immune cells, and pathogens (Shen and disrupts epithelial morphogenesis in developmental processes, such as gastrulation, tracheal morphogenesis, and ectodermal sheet migration (Jung is not associated with a general failure to form tubular structures. It is instead characterized by Fosfosal changes in the structure of these tubules and the ability of individual cells to properly intercalate with their neighbors (Jung cross-section) or polycarbonate (for en face sections) filter inserts. After 10 d in tradition, epithelia were fixed in 3% glutaraldehyde/0.1 sodium cacodylate/0.05 CaCl2 (pH 7.4), for 1 h at room temperature. Following three rinses with sodium cacodylate buffer, the monolayers were postfixed for 45 min in 1% osmium tetroxide/1.25% potassium ferrocyanide/0.1 sodium cacodylate buffer at space temperature (Russel and Burguet, 1977 ). After washes in deionized water, the cells were stained en bloc with 2% aqueous uranyl acetate for 20 min and dehydrated using increasing concentrations of ethanol (30%, 50%, 75%, 100%, 100%, 10 min each), which was followed by embedment in Polybed 812 epoxy resin (Polysciences, Warrington, PA). For en face sections, the filters were held smooth during embedment with Thermanox coverslips, which were then detached prior to sectioning. The monolayers were sectioned either parallel or perpendicular to the substrate at 70 nm using a diamond knife. Ultrathin sections were collected on 200-mesh copper grids and stained with 4% aqueous uranyl acetate for 15 min, and then Reynolds’ lead citrate for 7 min (Reynolds, 1963 ). Samples were viewed using a LEO EM910 transmission electron microscope operating at 80 kV (LEO Electron Microscopy, Thornwood, NY). Digital images were acquired using a Gatan Orius SC1000 CCD Digital Camera and Digital Micrograph 3.11.0 (Gatan, Pleasanton, CA). SEM.Duplicate cell monolayers were fixed in buffered glutaraldehyde as explained in for correlative SEM. Following aldehyde fixation, the cells were further stabilized using a revised osmium-tannic acid method (Katsumoto ZO-1 protein Polychaetoid regulates embryonic morphogenesis in coordination with Canoe/afadin and Enabled. Mol Biol Cell. 2011;22:2010C2030. [PMC free article] [PubMed] [Google Scholar]Colegio OR, Vehicle Itallie CM, McCrea HJ, Rahner C, Anderson JM. 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