Transforming growth factor-beta 2 modulated extracellular matrix component expression in cultured human optic nerve head astrocytes

Invest Ophthalmol Vis Sci. 2005 Feb;46(2):568-78. doi: 10.1167/iovs.04-0649.

Abstract

Purpose: To study whether glaucomatous extracellular matrix (ECM) modifications in the lamina cribrosa might be induced by TGF-beta 2, the effect of TGF-beta 2 on the expression of collagen types I (Col1 alpha 1), III (Col3 alpha 1), and IV (Col4 alpha 2); fibronectin (FN); tissue transglutaminase (TGM2); connective tissue growth factor (CTGF); and thrombospondin (TSP-1) in cultured human optic nerve head (ONH) astrocytes was investigated.

Methods: Astrocytes were isolated from eyes of five human donors, and cultured monolayers were treated with 1.0 ng/mL TGF-beta 2 for 24 and 48 hours. Expression of Col1 alpha 1, Col3 alpha 1, Col4 alpha 2, FN, TGM2, CTGF, and TSP-1 was examined by semiquantitative RT-PCR and Northern and Western blot analyses. The effect of CTGF silencing on the TGF-beta 2-modulated expression of these genes was investigated by transfection of CTGF small interfering (si)RNA before TGF-beta 2 treatment.

Results: TGF-beta 2 treatment upregulated the expression of Col1 alpha 1, Col4 alpha 2, FN, CTGF, TGM2, and TSP-1 mRNA and protein in cultured astrocytes. Inductions ranged between 1.5- and 4-fold. Expression of Col3 alpha 1 remained unaffected. Transfection of 10 nM CTGF siRNA inhibited the TGF-beta 2-induced upregulation of CTGF, Col4 alpha 2, Col1 alpha 1, TGM2, and FN, whereas TSP-1 expression was not reduced.

Conclusions: TGF-beta 2 is capable of inducing the expression of ECM and basement membrane components in cultured ONH astrocytes via CTGF and upregulated TSP-1, a protein naturally involved in the activation of latent TGF-beta. Therefore, TGF-beta 2 could be a factor in the initiation of the modification of ECM in the glaucomatous ONH. In addition, TSP-1 induction may be a mechanism by which TGF-beta 2 amplifies its own activation.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Aged
  • Aged, 80 and over
  • Astrocytes / drug effects*
  • Astrocytes / metabolism
  • Blotting, Northern
  • Blotting, Western
  • Cells, Cultured
  • Collagen / genetics
  • Collagen / metabolism
  • Connective Tissue Growth Factor
  • Extracellular Matrix Proteins / genetics*
  • Extracellular Matrix Proteins / metabolism
  • Fibronectins / genetics
  • Fibronectins / metabolism
  • GTP-Binding Proteins / genetics
  • GTP-Binding Proteins / metabolism
  • Humans
  • Immediate-Early Proteins / genetics
  • Immediate-Early Proteins / metabolism
  • Intercellular Signaling Peptides and Proteins / genetics
  • Intercellular Signaling Peptides and Proteins / metabolism
  • Middle Aged
  • Optic Disk / drug effects*
  • Optic Disk / metabolism
  • RNA, Messenger / metabolism
  • Reverse Transcriptase Polymerase Chain Reaction
  • Thrombospondin 1 / genetics
  • Thrombospondin 1 / metabolism
  • Transforming Growth Factor beta / pharmacology*
  • Transforming Growth Factor beta2
  • Transglutaminases / genetics
  • Transglutaminases / metabolism
  • Up-Regulation

Substances

  • CCN2 protein, human
  • Extracellular Matrix Proteins
  • Fibronectins
  • Immediate-Early Proteins
  • Intercellular Signaling Peptides and Proteins
  • RNA, Messenger
  • TGFB2 protein, human
  • Thrombospondin 1
  • Transforming Growth Factor beta
  • Transforming Growth Factor beta2
  • Connective Tissue Growth Factor
  • Collagen
  • transglutaminase 2
  • Transglutaminases
  • GTP-Binding Proteins