Connective tissue growth factor modulates extracellular matrix production in human subconjunctival fibroblasts and their proliferation and migration in vitro.
OBJECTIVE
We examined the role of connective tissue growth factor (CTGF) in transforming growth factor beta1 (TGFbeta1)-related behavior in cultured human subconjunctival fibroblasts (SCFs), protein production, mRNA expression of CTGF and type I collagen alpha1 chain (colIA1), and cell proliferation and migration. TGFbeta1 is the major factor involved in bleb scarring following filtration surgery.


METHODS
An antisense deoxynucleotide (antisense) (5 microM) for CTGF mRNA was used to block endogenous CTGF expression. Effects of antisense on extracellular matrix (ECM) production and immunolocalization, mRNA expression, and cell proliferation and migration were examined in human SCF cultures with or without TGFbeta1 (5 ng/ml). Cell migration was examined in an in vitro wound model of monolayer fibroblast cultures.


RESULTS
CTGF antisense reduced mRNA expression of CTGF and colIA1 and production of the ECM components type I collagen, and fibronectin much more markedly in cells treated with TGFbeta1 compared with control fibroblasts, and it inhibited the proliferation of cultured SCFs to 71.9% of that of controls after 13 days of culture. CTGF antisense also delayed defect closure in monolayer cell sheets. In the culture, the defect was closed by TGFbeta1 by 36 h, whereas 7.0% of the defect remained at 48 h in the antisense-treated culture.


CONCLUSIONS
These findings indicate that CTGF is involved in ECM production in SCFs activated by exogenous TGFbeta1 in vitro. Inhibition of CTGF expression may be effective in preventing undesirable scar formation during healing following filtration surgery.
Yamanaka
Osamu
O
Department of Ophthalmology, Wakayama Medical University, Wakayama, Japan. yamanaka@wakayama-med.ac.jp
Saika
Shizuya
S
Ikeda
Kazuo
K
Miyazaki
Ken-Ichi
K
Kitano
Ai
A
Ohnishi
Yoshitaka
Y
D002465:Cell Movement / Q000502:physiology; D049109:Cell Proliferation; D002478:Cells, Cultured; D002648:Child; D002675:Child, Preschool; D024042:Collagen Type I / Q000235:genetics / Q000378:metabolism*; D000090602:Collagen Type I, alpha 1 Chain; D003228:Conjunctiva / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D055513:Connective Tissue Growth Factor; D016373:DNA, Antisense / Q000494:pharmacology; D004797:Enzyme-Linked Immunosorbent Assay; D005109:Extracellular Matrix / Q000187:drug effects; D005347:Fibroblasts / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D005353:Fibronectins / Q000378:metabolism*; D006801:Humans; D017874:Immediate-Early Proteins / Q000502:physiology*; D036341:Intercellular Signaling Peptides and Proteins / Q000502:physiology*; D016376:Oligonucleotides, Antisense / Q000494:pharmacology; D012333:RNA, Messenger / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D053773:Transforming Growth Factor beta1 / Q000494:pharmacology
C526317:CCN2 protein, human; D024042:Collagen Type I; D000090602:Collagen Type I, alpha 1 Chain; D016373:DNA, Antisense; D005353:Fibronectins; D017874:Immediate-Early Proteins; D036341:Intercellular Signaling Peptides and Proteins; D016376:Oligonucleotides, Antisense; D012333:RNA, Messenger; D053773:Transforming Growth Factor beta1; D055513:Connective Tissue Growth Factor