title: Hypoxia increases the abundance but not the assembly of extracellular fibronectin during epithelial cell transdifferentiation.
abstract: Increased production and assembly of extracellular matrix proteins during transdifferentiation of epithelial cells to a mesenchymal phenotype contributes to diseases such as renal and pulmonary fibrosis. TGF-β and hypoxia, two cues that initiate injury-induced fibrosis, caused human kidney cells to develop a mesenchymal phenotype, including increased fibronectin expression and secretion. However, upon hypoxia, assembled extracellular fibronectin fibrils were mostly absent, whereas treatment with TGF-β led to abundant fibrils. Fibrillogenesis required cell-generated force and tension. TGF-β, but not hypoxia, increased cell contractility, as determined by phosphorylation of myosin light chain and quantifying force and tension generated by cells plated on engineered elastomeric microposts. Additionally, TGF-β, but not hypoxia, increased the activation of integrins. However, experimentally activating integrins markedly increased the levels of phosphorylated myosin light chain and fibronectin fibril assembly upon hypoxia. Our findings show that deficient integrin activation and subsequent lack of cell contractility are mechanisms that mediate a lack of fibrillogenesis upon hypoxia and they challenge current views on oxygen deprivation being sufficient for fibrosis.
authors.affiliation: Department of Cell and Tissue Biology, University of California, San Francisco, CA 94143, USA Department of Biomedical Engineering, Boston University and Wyss Institute for Biologically Inspired Engineering, Harvard University, Boston, MA 02215, USA.
authors.forename: Manish K
authors.initials: MK
authors.lastname: Rana
authors.affiliation: Department of Cell and Tissue Biology, University of California, San Francisco, CA 94143, USA.
authors.forename: Jyoti
authors.initials: J
authors.lastname: Srivastava
authors.affiliation: Department of Biomedical Engineering, Boston University and Wyss Institute for Biologically Inspired Engineering, Harvard University, Boston, MA 02215, USA.
authors.forename: Michael
authors.initials: M
authors.lastname: Yang
authors.affiliation: Department of Biomedical Engineering, Boston University and Wyss Institute for Biologically Inspired Engineering, Harvard University, Boston, MA 02215, USA.
authors.forename: Christopher S
authors.initials: CS
authors.lastname: Chen
authors.affiliation: Department of Cell and Tissue Biology, University of California, San Francisco, CA 94143, USA diane.barber@ucsf.edu.
authors.forename: Diane L
authors.initials: DL
authors.lastname: Barber
mesh_terms: D015153:Blotting, Western; D054338:Cell Transdifferentiation; D002478:Cells, Cultured; D004847:Epithelial Cells / Q000166:cytology* / Q000378:metabolism; D005109:Extracellular Matrix / Q000378:metabolism*; D005353:Fibronectins / Q000378:metabolism*; D005355:Fibrosis / Q000378:metabolism / Q000473:pathology*; D006801:Humans; D000860:Hypoxia / Q000503:physiopathology*; D016023:Integrins / Q000378:metabolism; D007687:Kidney Tubules, Proximal / Q000166:cytology* / Q000378:metabolism; D016212:Transforming Growth Factor beta / Q000378:metabolism
chemical_list: D005353:Fibronectins; D016023:Integrins; D016212:Transforming Growth Factor beta
keywords: Extracellular matrix; Fibronectin; Fibrosis; Hypoxia; TGF-β