title: Expression of insulin-like growth factor 2 receptor in corneal keratocytes during differentiation and in response to wound healing.
abstract: OBJECTIVE
  Insulin-like growth factor 2 receptor (IGF2R) associates with ligands that influence wound healing outcomes. However, the expression pattern of IGF2R and its role in the cornea is unknown.
  
  
  METHODS
  Human keratocytes were isolated from donor corneas. Fibroblasts (fibroblast growth factor 2 [FGF2]-treated) or myofibroblasts (TGF-β1-treated) were analyzed for IGF2R and α-smooth muscle actin (α-SMA) expression by Western blotting and immunolocalization. Mouse corneas were wounded in vivo and porcine corneas ex vivo. The IGF2R and α-SMA protein expression were visualized and quantified by immunohistochemistry. The IGF2R gene expression in human corneal fibroblasts was knocked-down with targeted lentiviral shRNA.
  
  
  RESULTS
  The IGF2R is expressed in epithelial and stromal cells of normal human, mouse, and porcine corneas. The IGF2R increases (11.2 ± 0.4-fold) in the epithelial and (11.7 ± 0.9-fold) stromal layers of in vivo wounded mouse corneas. Double-staining with α-SMA- and IGF2R-specific antibodies reveals that IGF2R protein expression is increased in stromal myofibroblasts in the wounded cornea relative to keratocytes in the normal cornea (11.2 ± 0.8-fold). Human primary stromal keratocytes incubated with FGF2 or TGF-β1 in vitro demonstrate increased expression (2.0 ± 0.4-fold) of IGF2R in myofibroblasts relative to fibroblasts. Conversion of IGF2R shRNA-lentiviral particle transduced corneal fibroblasts to myofibroblasts reveals a dependence on IGF2R expression, as only 40% ± 10% of cells transduced converted to myofibroblasts compared to 86% ± 3% in control cells.
  
  
  CONCLUSIONS
  The IGF2R protein expression is increased during corneal wound healing and IGF2R regulates human corneal fibroblast to myofibroblast differentiation.
authors.affiliation: Department of Biochemistry, Medical College of Wisconsin, Milwaukee, Wisconsin, United States.
authors.forename: Richard N
authors.initials: RN
authors.lastname: Bohnsack
authors.affiliation: Department of Biochemistry, Medical College of Wisconsin, Milwaukee, Wisconsin, United States.
authors.forename: Debra J
authors.initials: DJ
authors.lastname: Warejcka
authors.affiliation: Icahn School of Medicine at Mount Sinai, New York, New York, United States.
authors.forename: Lingyan
authors.initials: L
authors.lastname: Wang
authors.affiliation: Icahn School of Medicine at Mount Sinai, New York, New York, United States.
authors.forename: Stephanie R
authors.initials: SR
authors.lastname: Gillespie
authors.affiliation: Icahn School of Medicine at Mount Sinai, New York, New York, United States.
authors.forename: Audrey M
authors.initials: AM
authors.lastname: Bernstein
authors.affiliation: Department of Biochemistry, Medical College of Wisconsin, Milwaukee, Wisconsin, United States Department of Ophthalmology, Medical College of Wisconsin, Milwaukee, Wisconsin, United States.
authors.forename: Sally S
authors.initials: SS
authors.lastname: Twining
authors.affiliation: Department of Biochemistry, Medical College of Wisconsin, Milwaukee, Wisconsin, United States.
authors.forename: Nancy M
authors.initials: NM
authors.lastname: Dahms
mesh_terms: D000199:Actins / Q000378:metabolism; D000818:Animals; D015153:Blotting, Western; D002454:Cell Differentiation / Q000187:drug effects / Q000502:physiology; D002478:Cells, Cultured; D060527:Corneal Keratocytes / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D004195:Disease Models, Animal; D005786:Gene Expression Regulation / Q000502:physiology; D006801:Humans; D007150:Immunohistochemistry; D007335:Insulin-Like Growth Factor II / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D058628:Myofibroblasts / Q000187:drug effects / Q000378:metabolism; D013552:Swine; D016212:Transforming Growth Factor beta / Q000494:pharmacology; D014945:Wound Healing / Q000502:physiology*
chemical_list: C541116:ACTA2 protein, human; D000199:Actins; C035170:IGF2 protein, human; D016212:Transforming Growth Factor beta; D007335:Insulin-Like Growth Factor II
keywords: IGF2R; TGF-β1; cornea; myofibroblasts; wound healing