title: Transforming growth factor-β1 (TGF-β1)-stimulated fibroblast to myofibroblast differentiation is mediated by hyaluronan (HA)-facilitated epidermal growth factor receptor (EGFR) and CD44 co-localization in lipid rafts.
abstract: Fibroblast to myofibroblast differentiation drives effective wound healing and is largely regulated by the cytokine transforming growth factor-β1 (TGF-β1). Myofibroblasts express α-smooth muscle actin and are present in granulation tissue, where they are responsible for wound contraction. Our previous studies show that fibroblast differentiation in response to TGF-β1 is dependent on and mediated by the linear polysaccharide hyaluronan (HA). Both the HA receptor, CD44, and the epidermal growth factor receptor (EGFR) are involved in this differentiation response. The aim of this study was to understand the mechanisms linking HA-, CD44-, and EGFR-regulated TGF-β1-dependent differentiation. CD44 and EGFR co-localization within membrane-bound lipid rafts was necessary for differentiation, and this triggered downstream mitogen-activated protein kinase (MAPK/ERK) and Ca(2+)/calmodulin kinase II (CaMKII) activation. We also found that ERK phosphorylation was upstream of CaMKII phosphorylation, that ERK activation was necessary for CaMKII signaling, and that both kinases were essential for differentiation. In addition, HA synthase-2 (HAS2) siRNA attenuated both ERK and CaMKII signaling and sequestration of CD44 into lipid rafts, preventing differentiation. In summary, the data suggest that HAS2-dependent production of HA facilitates TGF-β1-dependent fibroblast differentiation through promoting CD44 interaction with EGFR held within membrane-bound lipid rafts. This induces MAPK/ERK, followed by CaMKII activation, leading to differentiation. This pathway is synergistic with the classical TGF-β1-dependent SMAD-signaling pathway and may provide a novel opportunity for intervention in wound healing.
authors.affiliation: Institute of Nephrology, School of Medicine and Cardiff Institute of Tissue Engineering and Repair, University of Cardiff, Heath Park, Cardiff CF14 4XN, United Kingdom.
authors.forename: Adam C
authors.initials: AC
authors.lastname: Midgley
authors.forename: Mathew
authors.initials: M
authors.lastname: Rogers
authors.forename: Maurice B
authors.initials: MB
authors.lastname: Hallett
authors.forename: Aled
authors.initials: A
authors.lastname: Clayton
authors.forename: Timothy
authors.initials: T
authors.lastname: Bowen
authors.forename: Aled O
authors.initials: AO
authors.lastname: Phillips
authors.forename: Robert
authors.initials: R
authors.lastname: Steadman
mesh_terms: D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2 / Q000235:genetics / Q000378:metabolism; D002454:Cell Differentiation / Q000502:physiology*; D002461:Cell Line, Transformed; D004789:Enzyme Activation / Q000502:physiology; D066246:ErbB Receptors / Q000235:genetics / Q000378:metabolism*; D048049:Extracellular Signal-Regulated MAP Kinases / Q000235:genetics / Q000378:metabolism; D014453:Glucuronosyltransferase / Q000235:genetics / Q000378:metabolism; D006801:Humans; D018960:Hyaluronan Receptors / Q000235:genetics / Q000378:metabolism*; D000076002:Hyaluronan Synthases; D006820:Hyaluronic Acid / Q000235:genetics / Q000378:metabolism*; D021962:Membrane Microdomains / Q000235:genetics / Q000378:metabolism*; D058628:Myofibroblasts / Q000166:cytology / Q000378:metabolism*; D015398:Signal Transduction / Q000502:physiology; D051785:Smad Proteins / Q000235:genetics / Q000378:metabolism; D053773:Transforming Growth Factor beta1 / Q000235:genetics / Q000378:metabolism*; D014945:Wound Healing / Q000502:physiology
chemical_list: C497483:CD44 protein, human; D018960:Hyaluronan Receptors; D051785:Smad Proteins; C508960:TGFB1 protein, human; D053773:Transforming Growth Factor beta1; D006820:Hyaluronic Acid; D014453:Glucuronosyltransferase; C101754:HAS2 protein, human; D000076002:Hyaluronan Synthases; C512478:EGFR protein, human; D066246:ErbB Receptors; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D048049:Extracellular Signal-Regulated MAP Kinases
keywords: CD44; Epidermal Growth Factor Receptor (EGFR); Hyaluronate; Lipid Raft; Myofibroblast