title: CCN2 is necessary for the function of mouse embryonic fibroblasts.
abstract: CCN2 is expressed by mesenchymal cells undergoing active tissue remodeling, and is characteristically overexpressed in connective tissue pathologies such as fibrosis and cancer. However, the physiological roles and mechanism of action of CCN2 are largely unknown. Here, we probe the contribution of CCN2 to the biology of mouse embryonic fibroblasts (MEFs) using genome-wide mRNA expression profiling, proteomic and functional bioassay analyses. We show that ccn2-/- mouse embryonic fibroblasts (MEFs) have significantly reduced the expression of pro-adhesive, pro-inflammatory and pro-angiogenic genes such as interleukin-6 (IL-6), ceruloplasmin, thrombospondin-1, lipocalin-2 and syndecan 4. Anti-syndecan 4 antibody reduced ERK phosphorylation in ccn2+/+ MEFs. In ccn2+/+ MEFs, the MEK inhibitor U0126 and dominant negative ras reduced expression of IL-6 and lipocalin-2. Overexpressing syndecan 4 in ccn2-/- MEFs restored IL-6 and lipocalin-2 mRNA expression. Syndecan 4 has been shown to mediate cell migration. We found that ccn2+/+ MEFs migrated significantly faster than ccn2-/- MEFs; anti-syndecan 4 antibody and U0126 reduced the migration of ccn2+/+ MEFs to that of ccn2-/- MEFs. These results collectively support the notion that syndecan 4 acts downstream of CCN2 in MEFs, and that reduced syndecan 4 expression contributes to at least part of the ccn2-/- phenotype. Further, these results suggest that CCN2 is required for MEFs to contribute to aspects of tissue remodeling. Consistent with this notion, whereas ccn2+/+ MEFs displayed actin stress fibers and focal adhesions at the cell periphery consistent with a migratory phenotype, ccn2-/- MEFs displayed reduced focal adhesions and actin stress fibers, and a reduced ability to transduce forces across a collagen gel matrix. Collectively, these results suggest that CCN2 supplies essential, non-redundant functions required for fibroblasts to properly participate in features of embryogenesis, and further suggest that CCN2 may play essential roles in adult wound healing, tissue repair and fibrogenesis.
authors.affiliation: Division of Oral Biology and Department of Physiology and Pharmacology, CIHR Group in Skeletal Development and Remodeling, Schulich School of Medicine and Dentistry, University of Western Ontario, Dental Sciences Building, London, ON, Canada N6A 5C1.
authors.forename: Laura
authors.initials: L
authors.lastname: Kennedy
authors.forename: Shangxi
authors.initials: S
authors.lastname: Liu
authors.forename: Xu
authors.initials: X
authors.lastname: Shi-Wen
authors.forename: Yunliang
authors.initials: Y
authors.lastname: Chen
authors.forename: Mark
authors.initials: M
authors.lastname: Eastwood
authors.forename: Mojhgan
authors.initials: M
authors.lastname: Sabetkar
authors.forename: David E
authors.initials: DE
authors.lastname: Carter
authors.forename: Karen M
authors.initials: KM
authors.lastname: Lyons
authors.forename: Carol M
authors.initials: CM
authors.lastname: Black
authors.forename: David J
authors.initials: DJ
authors.lastname: Abraham
authors.forename: Andrew
authors.initials: A
authors.lastname: Leask
mesh_terms: D008841:Actin Cytoskeleton / Q000502:physiology; D000818:Animals; D002465:Cell Movement; D002478:Cells, Cultured; D003199:Computer Systems; D055513:Connective Tissue Growth Factor; D004622:Embryo, Mammalian / Q000166:cytology*; D005109:Extracellular Matrix / Q000502:physiology; D005347:Fibroblasts / Q000378:metabolism / Q000502:physiology*; D022001:Focal Adhesions / Q000502:physiology; D020869:Gene Expression Profiling / Q000379:methods; D017874:Immediate-Early Proteins / Q000235:genetics / Q000378:metabolism* / Q000502:physiology*; D036341:Intercellular Signaling Peptides and Proteins / Q000235:genetics / Q000378:metabolism* / Q000502:physiology*; D051379:Mice; D016133:Polymerase Chain Reaction; D040901:Proteomics; D015398:Signal Transduction; D053671:Syndecan-4 / Q000378:metabolism; D014945:Wound Healing
chemical_list: C526318:CCN2 protein, mouse; D017874:Immediate-Early Proteins; D036341:Intercellular Signaling Peptides and Proteins; D053671:Syndecan-4; D055513:Connective Tissue Growth Factor