May 2003
Volume 44, Issue 13
Free
ARVO Annual Meeting Abstract  |   May 2003
Doxycycline Inhibits TGF-ß1 Stimulated MMP-9 in Human Corneal Epithelial Cells via Smad and MAPK Pathways
Author Affiliations & Notes
  • S.C. Pflugfelder
    Ophthalmology-Ocular Surf Ctr, Cullen Eye Institute, Houston, TX, United States
  • H. Kim
    Ophthalmology-Ocular Surf Ctr, Cullen Eye Institute, Houston, TX, United States
  • D. Li
    Ophthalmology-Ocular Surf Ctr, Cullen Eye Institute, Houston, TX, United States
  • Footnotes
    Commercial Relationships  S.C. Pflugfelder, None; H. Kim, None; D. Li, None.
  • Footnotes
    Support  EY11915-05
Investigative Ophthalmology & Visual Science May 2003, Vol.44, 1404. doi:
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      S.C. Pflugfelder, H. Kim, D. Li; Doxycycline Inhibits TGF-ß1 Stimulated MMP-9 in Human Corneal Epithelial Cells via Smad and MAPK Pathways . Invest. Ophthalmol. Vis. Sci. 2003;44(13):1404.

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      © ARVO (1962-2015); The Authors (2016-present)

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Abstract

Abstract: : Purpose: To evaluate the effects of doxycycline on the expression of TGF-ß1 stimulated matrix metalloproteinase 9 (MMP-9) in human corneal epithelial cells and on the activation of Smad2, c-Jun N-terminal kinase (JNK) and extracellular-regulated kinase (ERK) signaling intermediates that are induced by TGF-ß1. Methods: Primary human corneal epithelial cells were cultured from limbal explants in 6-well plates until confluence. The cells were then exposed to serum-free media alone (control), or with different concentrations of TGF-ß1 (0.1, 1,10 ng/ml) with or without TGF-ß1 neutralizing mAb (5 µg/ml), SB202190 (10-40 µM) and doxycycline (5-40 µg/ml) for different lengths of time. The conditioned media were collected from cultures treated for 24 hours for zymography and MMP-9 activity assay. The cells treated for 5-60 min were lysed in RIPA buffer and subjected to Western blot with phospho-specific antibodies against Smad2, JNK or ERK. Results: TGF-ß1 dose-dependently increased the production and activity of MMP-9 by human corneal epithelial cells. This stimulation was abolished by 5 µg/ml of TGF-ß1 neutralizing mAb or 10 µg/ml of doxycycline. TGF-ß1 dose-dependently induced phorsphorylated JNK1/2, ERK and Smad2, as early as 5 min, peaking at 15 or 30 min respectively. Doxycycline markedly inhibited the TGF-ß1 induced activation of JNK1/2, ERK1 and Smad2 with activity equal to SB202190. Conclusions: Doxycycline inhibits TGF-ß1 induced MMP-9 production and activity, perhaps through the Smad and MAPK pathways. This finding may explain the reported efficacy of doxycycline in treating MMP-mediated ocular surface disease.

Keywords: cornea: epithelium • cornea: tears/tear film/dry eye • heading 
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