TGFβ induced myofibroblast differentiation of rabbit keratocytes requires synergistic TGFβ, PDGF and integrin signaling

James V. Jester, Jiying Huang, Walter M Petroll, Harrison D Cavanagh

Research output: Contribution to journalArticle

133 Citations (Scopus)

Abstract

There is a growing consensus that corneal myofibroblasts are derived from adjacent stromal keratocytes which undergo an orderly phenotypic transition from quiescent keratocyte to activated fibroblast to myofibroblast. Both in vivo and in vitro studies have shown this transition to be dependent, in part, on transforming growth factor beta (TGFβ). In many fibroblastic cells autocrine production of platelet derived growth factor (PDGF) is known to mediate the growth up-regulation by TGFβ. In this study, blocking antibodies to PDGF significantly reduced by 80 % (P < 0.025) the TGFβ1 stimulated cell cycle entry of serum-free cultured rabbit corneal keratocytes. AntiPDGF treatment also markedly reduced the TGFβ1-induced intracellular actin filament re-organization, fibronectin fibril assembly, and focal contact formation as well as reducing by 80 % the expression of α-smooth muscle (α-SM) specific isoform of actin characteristic of myofibroblast differentiation. Although PDGF treatment of quiescent keratocytes produced an activated, fibroblastic cell type, PDGF stimulated keratocytes exhibited the same temporal, myofibroblastic differentiation response to TGFβ1 as did quiescent keratocytes. Furthermore, blocking TGFβ1 induction of myofibroblast differentiation with the Arg-Gly-Asp containing peptide, GRGDdSP, for 3 days followed by allowing progression of myofibroblast differentiation by removing GRGDdSP did not change the temporal response or tyrosine phosphorylation cascade (2-72 hr) leading to myofibroblast differentiation. Nor did PDGF treatment of keratocytes reverse the RGD blockade of TGFβ1 induced myofibroblast differentiation. Overall these cumulative findings indicate that myofibroblast differentiation in the rabbit corneal keratocyte requires synergistic growth factor/integrin signaling involving TGFβ, PDGF, and the fibronectin receptor. Additionally, the similar TGFβ1 temporal response of PDGF-stimulated compared to naïve keratocytes suggests that myofibroblast differentiation does not require transition through a fibroblast phenotype.

Original languageEnglish (US)
Pages (from-to)645-657
Number of pages13
JournalExperimental Eye Research
Volume75
Issue number6
DOIs
StatePublished - 2002

Fingerprint

Myofibroblasts
Platelet-Derived Growth Factor
Integrins
Transforming Growth Factor beta
Rabbits
Corneal Keratocytes
Fibroblasts
Integrin alpha5beta1
Focal Adhesions
Blocking Antibodies
Actin Cytoskeleton
Fibronectins
Smooth Muscle
Tyrosine
Actins
Intercellular Signaling Peptides and Proteins
Cell Cycle
Protein Isoforms
Up-Regulation
Phosphorylation

Keywords

  • Alpha smooth muscle actin
  • Cornea
  • Integrin
  • Keratocyte
  • Myofibroblast
  • Platelet derived growth factor
  • Transforming growth factor beta

ASJC Scopus subject areas

  • Ophthalmology
  • Sensory Systems

Cite this

TGFβ induced myofibroblast differentiation of rabbit keratocytes requires synergistic TGFβ, PDGF and integrin signaling. / Jester, James V.; Huang, Jiying; Petroll, Walter M; Cavanagh, Harrison D.

In: Experimental Eye Research, Vol. 75, No. 6, 2002, p. 645-657.

Research output: Contribution to journalArticle

@article{a6ed15ad5922479ebf39510a2abe7a54,
title = "TGFβ induced myofibroblast differentiation of rabbit keratocytes requires synergistic TGFβ, PDGF and integrin signaling",
abstract = "There is a growing consensus that corneal myofibroblasts are derived from adjacent stromal keratocytes which undergo an orderly phenotypic transition from quiescent keratocyte to activated fibroblast to myofibroblast. Both in vivo and in vitro studies have shown this transition to be dependent, in part, on transforming growth factor beta (TGFβ). In many fibroblastic cells autocrine production of platelet derived growth factor (PDGF) is known to mediate the growth up-regulation by TGFβ. In this study, blocking antibodies to PDGF significantly reduced by 80 {\%} (P < 0.025) the TGFβ1 stimulated cell cycle entry of serum-free cultured rabbit corneal keratocytes. AntiPDGF treatment also markedly reduced the TGFβ1-induced intracellular actin filament re-organization, fibronectin fibril assembly, and focal contact formation as well as reducing by 80 {\%} the expression of α-smooth muscle (α-SM) specific isoform of actin characteristic of myofibroblast differentiation. Although PDGF treatment of quiescent keratocytes produced an activated, fibroblastic cell type, PDGF stimulated keratocytes exhibited the same temporal, myofibroblastic differentiation response to TGFβ1 as did quiescent keratocytes. Furthermore, blocking TGFβ1 induction of myofibroblast differentiation with the Arg-Gly-Asp containing peptide, GRGDdSP, for 3 days followed by allowing progression of myofibroblast differentiation by removing GRGDdSP did not change the temporal response or tyrosine phosphorylation cascade (2-72 hr) leading to myofibroblast differentiation. Nor did PDGF treatment of keratocytes reverse the RGD blockade of TGFβ1 induced myofibroblast differentiation. Overall these cumulative findings indicate that myofibroblast differentiation in the rabbit corneal keratocyte requires synergistic growth factor/integrin signaling involving TGFβ, PDGF, and the fibronectin receptor. Additionally, the similar TGFβ1 temporal response of PDGF-stimulated compared to na{\"i}ve keratocytes suggests that myofibroblast differentiation does not require transition through a fibroblast phenotype.",
keywords = "Alpha smooth muscle actin, Cornea, Integrin, Keratocyte, Myofibroblast, Platelet derived growth factor, Transforming growth factor beta",
author = "Jester, {James V.} and Jiying Huang and Petroll, {Walter M} and Cavanagh, {Harrison D}",
year = "2002",
doi = "10.1006/exer.2002.2066",
language = "English (US)",
volume = "75",
pages = "645--657",
journal = "Experimental Eye Research",
issn = "0014-4835",
publisher = "Academic Press Inc.",
number = "6",

}

TY - JOUR

T1 - TGFβ induced myofibroblast differentiation of rabbit keratocytes requires synergistic TGFβ, PDGF and integrin signaling

AU - Jester, James V.

AU - Huang, Jiying

AU - Petroll, Walter M

AU - Cavanagh, Harrison D

PY - 2002

Y1 - 2002

N2 - There is a growing consensus that corneal myofibroblasts are derived from adjacent stromal keratocytes which undergo an orderly phenotypic transition from quiescent keratocyte to activated fibroblast to myofibroblast. Both in vivo and in vitro studies have shown this transition to be dependent, in part, on transforming growth factor beta (TGFβ). In many fibroblastic cells autocrine production of platelet derived growth factor (PDGF) is known to mediate the growth up-regulation by TGFβ. In this study, blocking antibodies to PDGF significantly reduced by 80 % (P < 0.025) the TGFβ1 stimulated cell cycle entry of serum-free cultured rabbit corneal keratocytes. AntiPDGF treatment also markedly reduced the TGFβ1-induced intracellular actin filament re-organization, fibronectin fibril assembly, and focal contact formation as well as reducing by 80 % the expression of α-smooth muscle (α-SM) specific isoform of actin characteristic of myofibroblast differentiation. Although PDGF treatment of quiescent keratocytes produced an activated, fibroblastic cell type, PDGF stimulated keratocytes exhibited the same temporal, myofibroblastic differentiation response to TGFβ1 as did quiescent keratocytes. Furthermore, blocking TGFβ1 induction of myofibroblast differentiation with the Arg-Gly-Asp containing peptide, GRGDdSP, for 3 days followed by allowing progression of myofibroblast differentiation by removing GRGDdSP did not change the temporal response or tyrosine phosphorylation cascade (2-72 hr) leading to myofibroblast differentiation. Nor did PDGF treatment of keratocytes reverse the RGD blockade of TGFβ1 induced myofibroblast differentiation. Overall these cumulative findings indicate that myofibroblast differentiation in the rabbit corneal keratocyte requires synergistic growth factor/integrin signaling involving TGFβ, PDGF, and the fibronectin receptor. Additionally, the similar TGFβ1 temporal response of PDGF-stimulated compared to naïve keratocytes suggests that myofibroblast differentiation does not require transition through a fibroblast phenotype.

AB - There is a growing consensus that corneal myofibroblasts are derived from adjacent stromal keratocytes which undergo an orderly phenotypic transition from quiescent keratocyte to activated fibroblast to myofibroblast. Both in vivo and in vitro studies have shown this transition to be dependent, in part, on transforming growth factor beta (TGFβ). In many fibroblastic cells autocrine production of platelet derived growth factor (PDGF) is known to mediate the growth up-regulation by TGFβ. In this study, blocking antibodies to PDGF significantly reduced by 80 % (P < 0.025) the TGFβ1 stimulated cell cycle entry of serum-free cultured rabbit corneal keratocytes. AntiPDGF treatment also markedly reduced the TGFβ1-induced intracellular actin filament re-organization, fibronectin fibril assembly, and focal contact formation as well as reducing by 80 % the expression of α-smooth muscle (α-SM) specific isoform of actin characteristic of myofibroblast differentiation. Although PDGF treatment of quiescent keratocytes produced an activated, fibroblastic cell type, PDGF stimulated keratocytes exhibited the same temporal, myofibroblastic differentiation response to TGFβ1 as did quiescent keratocytes. Furthermore, blocking TGFβ1 induction of myofibroblast differentiation with the Arg-Gly-Asp containing peptide, GRGDdSP, for 3 days followed by allowing progression of myofibroblast differentiation by removing GRGDdSP did not change the temporal response or tyrosine phosphorylation cascade (2-72 hr) leading to myofibroblast differentiation. Nor did PDGF treatment of keratocytes reverse the RGD blockade of TGFβ1 induced myofibroblast differentiation. Overall these cumulative findings indicate that myofibroblast differentiation in the rabbit corneal keratocyte requires synergistic growth factor/integrin signaling involving TGFβ, PDGF, and the fibronectin receptor. Additionally, the similar TGFβ1 temporal response of PDGF-stimulated compared to naïve keratocytes suggests that myofibroblast differentiation does not require transition through a fibroblast phenotype.

KW - Alpha smooth muscle actin

KW - Cornea

KW - Integrin

KW - Keratocyte

KW - Myofibroblast

KW - Platelet derived growth factor

KW - Transforming growth factor beta

UR - http://www.scopus.com/inward/record.url?scp=0036882153&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0036882153&partnerID=8YFLogxK

U2 - 10.1006/exer.2002.2066

DO - 10.1006/exer.2002.2066

M3 - Article

C2 - 12470966

AN - SCOPUS:0036882153

VL - 75

SP - 645

EP - 657

JO - Experimental Eye Research

JF - Experimental Eye Research

SN - 0014-4835

IS - 6

ER -