March 2012
Volume 53, Issue 14
Free
ARVO Annual Meeting Abstract  |   March 2012
The Effects of Rho/ROCK Activation in Dexamethasone-Induced Increase of Aqueous-Outflow Resistance
Author Affiliations & Notes
  • Tomokazu Fujimoto
    Ophthalmology & Visual Science, Kumamoto Univ Grad School, Kumamoto, Japan
  • Toshihiro Inoue
    Ophthalmology & Visual Science, Kumamoto Univ Grad School, Kumamoto, Japan
  • Miyuki Inoue-Mochita
    Ophthalmology & Visual Science, Kumamoto Univ Grad School, Kumamoto, Japan
  • Takanori Kameda
    Kobe City Medical Center General Hospital, Kobe, Japan
  • Nanako Kasaoka
    Ophthalmology & Visual Science, Kumamoto Univ Grad School, Kumamoto, Japan
  • Hidenobu Tanihara
    Ophthalmology & Visual Science, Kumamoto Univ Grad School, Kumamoto, Japan
  • Footnotes
    Commercial Relationships  Tomokazu Fujimoto, None; Toshihiro Inoue, None; Miyuki Inoue-Mochita, None; Takanori Kameda, None; Nanako Kasaoka, None; Hidenobu Tanihara, None
  • Footnotes
    Support  The Imai Memorial Fund for Glaucoma Research
Investigative Ophthalmology & Visual Science March 2012, Vol.53, 3227. doi:
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      Tomokazu Fujimoto, Toshihiro Inoue, Miyuki Inoue-Mochita, Takanori Kameda, Nanako Kasaoka, Hidenobu Tanihara; The Effects of Rho/ROCK Activation in Dexamethasone-Induced Increase of Aqueous-Outflow Resistance. Invest. Ophthalmol. Vis. Sci. 2012;53(14):3227.

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

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Abstract

Purpose: : To examine whether the RhoA/Rho kinase (ROCK) pathway is involved in one of the mechanism of dexamethasone (DEX)-induced ocular hypertension.

Methods: : The effect of DEX on the RhoA activation was evaluated by pull-down assay in porcine trabecular meshwork (TM) cells and monkey Schlemm’s canal endothelial (SCE) cells. The effect of selective ROCK inhibitor, Y-27632, on outflow facility was evaluated by perfused porcine anterior organ cultures treated with 100 nM DEX for 7 days. The effects of Y-27632 and DEX on the barrier function of confluent SCE-cell monolayer on micropore filter inserts was evaluated by measurement of transendothelial electrical resistance (TEER). The distributions of junctional proteins and F-actin on SCE cells exposed DEX and Y-27632 were confirmed by immunofluorescence. Collagen type 1 α1 chain (COL1A1) and collagen type4 α1 chain (COL4A1) mRNA expression level was evaluated by quantitative real-time RT-PCR in TM cells treated with DEX and Y-27632.

Results: : Relative RhoA activities were 1.63 ± 0.26 and 1.58 ± 0.06-fold increased 5 minutes after stimulation with 100 nM DEX in TM and SCE cells, respectively. Perfusion with 100 nM of DEX decreased outflow facility by 31.9 ± 14.3% compared with control at 24 hours, and the effect was sustained for 7 days. Combinated treatment with DEX and Y-27632 did not change outflow facility compared with vehicle treated control at 24 hours after drug perfusion. TEER of the SCE-cell monolayer was increased by 100 nM DEX for 3 days (15.42 ± 1.57 ohms·cm2) compared with control (11.43 ± 1.35 ohms·cm2), whereas 10μM Y-27632 diminished the effect of DEX (10.78 ± 0.95 ohms·cm2). ZO-1 was localized on the plasma membrane in SCE cells. A slight increase in intensity of ZO-1 staining was observed in DEX treated SCE cells. Y-27632 disrupted ZO-1 expression in DEX treated SCE cells. Expression of COL4A1 mRNA was 2.1 ± 0.24-fold increased 24 hr after treatment with DEX in TM cells, and Y-27632 inhibited DEX induced COL4A1 expression in TM cells. In contrast, expression of COL1A1 mRNA was not changed by DEX and Y-27632.

Conclusions: : These results suggest that the activation of Rho/ROCK pathway in TM and SCE cells may be involved in the mechanisms of DEX-induced ocular hypertension.

Keywords: outflow: trabecular meshwork • cell adhesions/cell junctions • extracellular matrix 
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