Fraction of MHCII and EpCAM expression characterizes distal lung epithelial cells for alveolar type 2 cell isolation. Gene expression in 10-4A cells was also compared to expression in healthy primary distal lung fibroblasts and primary ATII cells. Scale bar = 50 μm. 4. Cell Biol. Data are expressed as fold expression of the housekeeping gene hydroxymethylbilane synthase (Hmbs). Thoracic Soc. Pulmonary fibrosis in vivo displays increased p21 expression reduced by 5-HT2B receptor antagonists in vitro - A potential pathway affecting proliferation. Similarly, in vivo studies have shown that IFN activities with biological properties identical to those described above can be detected in the serum of fish following natural or experimental viral infection, injection with poly I:C or DNA vaccination with viral G proteins (Dorson et al., 1975; Kim et al., 2000). doi: 10.1083/jcb.200704042, Wuyts, W. A., Agostini, C., Antoniou, K. M., Bouros, D., Chambers, R. C., Cottin, V., et al. Int. The aim of our work was to generate a representative cell line to be used for in vitro fibrosis research. doi: 10.1164/rccm.2009-040GL, Ran, F. A., Hsu, P. D., Wright, J., Agarwala, V., Scott, D. A., and Zhang, F. (2013). Ann. 378, 1811–1823. bioRxiv [Preprint]. Rheumatol. Two-way conversion between lipogenic and myogenic fibroblastic phenotypes marks the progression and resolution of lung fibrosis. (2018). The Filamentous Actin Network in MRC-5 Fibroblast Cells - In this section, the single human fetal lung fibroblast (MRC-5) cell that is featured was resident in a culture that was stained with Alexa Fluor 568 conjugated to phalloidin, which binds to the intracellular filamentous actin network. Primary fibroblast data were obtained from seven different animals, 10-4A data from five different passages. The animal study was reviewed and approved by the Regierungspräsidium Tübingen. Curr. (D) Immunofluorescence staining of 10-4A cells (left) or primary fibroblasts seeded on PDMS with administration of 5 ng/ml TGF-β1 directly after adherence (d0) or 7 days post-seeding. Thus, Dicer partner proteins in both flies and mammals can change where Dicer cleaves some pre-miRNAs, generating miRNA isoforms with distinct lengths, seed sequences, and target specificities compared with those produced by Dicer alone or Dicer bound to alternative dsRBD protein partners. Proc. 49, 999–1008. (2004). doi: 10.14573/altex.1511131, Lederer, D. J., and Martinez, F. J. 19:32. doi: 10.1186/s12931-018-0730-2, Shi, Y., Dong, Y., Duan, Y., Jiang, X., Chen, C., and Deng, L. (2013). (2016). Res. doi: 10.1371/journal.pone.0000108, O’Reilly, S. (2013). Podoplanin in inflammation and cancer. Its mechanism of action may be via down regulation of cell surface C-kit expression and IL-1 receptors (301). 3). Fibroblasts represent a heterogeneous population of cells, which may differ in phenotype and function not only between anatomical sites but also even within a single tissue where immature cells (often called … In the bottom panel, a photograph of the 28S rRNA band is shown to illustrate that equivalent amounts of RNA were present in each gel lane. The editor and reviewers' affiliations are the latest provided on their Loop research profiles and may not reflect their situation at the time of review. Table 1. Expansion of functional personalized cells with specific transgene combinations. Bioinformatics 28, 1530–1532. Only the region of each autoradiogram that contained a mRNA hybridization signal is shown. J. Clin. Human Lung Fibroblast Cells with Texas Red-X, Oregon Green 488, and Hoechst 33258 - In order to label the intermediate filaments in a log phase adherent MRC-5 culture, the fixed and permeabilized cells were blocked and treated with mouse anti-vimentin (porcine eye lens) primary antibodies followed by goat anti-mouse secondary antibodies (IgG) conjugated to Texas Red-X. 7:3. doi: 10.3389/fbioe.2019.00003, Gabazza, E. C., Kasper, M., Ohta, K., Keane, M., D’Alessandro-Gabazza, C., Fujimoto, H., et al. The nuclei were counterstained with Hoechst 33258. Serum-starved cells were either left untreated or treated with FGF-1 for the indicated time periods. A 20 bp single guide RNA (sgRNA) (AAACAGGAGT ATGACGAAGC), binding at the end of the coding region of the Acta2 gene was designed by using the R&D Benchling software3. A confluent monolayer yields 40000 cells/cm2. Overall, the combination of mechanical and chemical stimulation induced a robust and persistent expression of myofibroblast markers in 10-4A cells and primary fibroblasts. doi: 10.14814/phy2.13281, Bagnato, G., and Harari, S. (2015). In addition, this activity is species-specific with respect to the cells being protected but non-specific with regard to the challenge virus, i.e. doi: 10.1056/NEJMra1705751. 186, 866–876. # 926-32213, RRID:AB_621848), IRDye® 680RD Donkey anti-Mouse (cat. Finally, we generated a reporter cell line (10-4ABFP) to express nuclear blue fluorescent protein (BFP) under the promotor of the myofibroblast marker alpha smooth muscle actin (Acta2) using CRISPR/Cas9 technology. It is well established that fish fibroblast or epithelial cell lines (Gravel and Marlsberger, 1965; DeSena and Rio, 1975) and isolated leukocytes (Snegaroff, 1993; Rogel-Gaillard et al., 1993; Congleton and Sun, 1996) can secrete IFN in response to virus infection and poly I This IFN activity is acid (pH 2) stable, relatively temperature resistant but destroyed by trypsin treatment, all properties typical of a type I IFN. Eur. Am. 41, 1207–1218. Pictures were taken with an iMIC Digital Microscope (FEI Munich GmbH). Human fibroblast lines taken from biopsies of patients have been used to explain the pathogenesis of several diseases. 307, L386–L394. Thorax 66, 651–657. (2012). Smallpox vaccines have been shown to be highly effective in preventing infection and in decreasing the severity of illness to those that were exposed without protection. 128, 74–84. (2007). Initiated in September 1966 by J. P. Jacobs, the cell line … Regarding the fibroblast subpopulations stated by Xie et al. To this end, we isolated primary cells from the distal lung of Sprague-Dawley rats and immortalized them using a recently described technology (Kuehn et al., 2016). # A11039; Thermo Fisher Scientific, RRID:AB_142924); Alexa Fluor® 568 goat anti-rabbit (1:300, cat. Opin. (2003). (B) Dose-response curves of BFP expression in 10-4ABFP cells in response to 2 days exposure with increasing doses of TGF-β1, TSLP, IL-4, and IL-33. doi: 10.1165/rcmb.2012-0050OC, Huaux, F., Liu, T., McGarry, B., Ullenbruch, M., and Phan, S. H. (2003). A., Matthes, S. A., Tsui, J. L., et al. (E) Western Blot for αSMA in 10-4A cells and primary fibroblasts cultured over 14 days on soft (PDMS) and stiff (Plastic) matrices, respectively. Based on HEK 293 cells and equipped with flippase recognition target sites containing a murine leukemia virus (MLV)-green fluorescent protein (GFP) vector, these cells allow the efficient Flp recombinase-mediated cassette exchange of MLV vectors; thus, after cassette exchange, the tagged retrovirus producer cell clone is capable of producing vectors containing the transgene of choice at levels similar to those observed for the mother producer cell line [115, 116].

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