title: Increased SPARC accumulation during corneal repair.
abstract: Keratocytes can become fibroblasts and myofibroblasts during corneal injury and wound healing. We used the in vitro bovine keratocyte repair model system, which involves culturing collagenase-isolated keratocytes in serum-free media and then adding serum or serum plus TGF-beta to the culture media to induce the fibroblast and myofibroblast phenotypes, respectively, to evaluate the synthesis of secreted products by the cells. Serum and serum plus TGF-beta rapidly induced the fibroblast morphology and alpha smooth muscle actin, a marker of myofibroblasts. Keratocytes cultured in serum and serum plus TGF-beta also increased the synthesis of several high molecular weight products (approximately 100kD and larger) and the accumulation of a 43kD protein shown to be osteonectin/SPARC by both sequencing tryptic peptides from the protein and by reaction with antisera to osteonectin/SPARC. Immunohistochemical staining of mouse corneas with antisera to SPARC seven days post-wounding also demonstrated an increased accumulation of SPARC in the regions undergoing repair. These results indicate SPARC accumulation is a marker for stromal repair.
authors.affiliation: The Department of Biochemistry and Molecular Biology, College of Medicine, University of South Florida, Tampa, FL, USA.
authors.forename: Bridgette L
authors.initials: BL
authors.lastname: Berryhill
authors.forename: Bradley
authors.initials: B
authors.lastname: Kane
authors.forename: Brian M
authors.initials: BM
authors.lastname: Stramer
authors.forename: M Elizabeth
authors.initials: ME
authors.lastname: Fini
authors.forename: John R
authors.initials: JR
authors.lastname: Hassell
mesh_terms: D000818:Animals; D015415:Biomarkers / Q000032:analysis; D015153:Blotting, Western / Q000379:methods; D002417:Cattle; D002478:Cells, Cultured; D003315:Cornea / Q000378:metabolism* / Q000473:pathology; D003320:Corneal Ulcer / Q000378:metabolism* / Q000473:pathology; D004591:Electrophoresis, Polyacrylamide Gel; D005109:Extracellular Matrix; D007150:Immunohistochemistry / Q000379:methods; D015603:Keratinocytes / Q000737:chemistry; D008233:Lymphotoxin-alpha / Q000494:pharmacology; D051379:Mice; D015676:Osteonectin / Q000032:analysis*; D012333:RNA, Messenger / Q000032:analysis; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014945:Wound Healing
chemical_list: D015415:Biomarkers; D008233:Lymphotoxin-alpha; D015676:Osteonectin; D012333:RNA, Messenger