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Tsugiaki Utsunomiya, Akihiro Ishibazawa, Taiji Nagaoka, Kazuomi Hanada, Akitoshi Yoshida; Effects of mechanical stimulation of fluid shear stress in cultured corneal epithelial cells. Invest. Ophthalmol. Vis. Sci. 2016;57(12):4902. doi: https://doi.org/.
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© ARVO (1962-2015); The Authors (2016-present)
Blinking can generate mechanical stimulation of fluid shear stress on the corneal epithelial cells because blinking results in movement of the tear fluid covering the corneal surface. The mechanical stress can vary in ocular-surface disorders such as dry eye and cause corneal epithelial damage. The stress also can affect some signaling cascades, but the details of that activity have not been determined. We investigated the effects of fluid shear stress on cultured human corneal epithelial cells.
Using a parallel-plate type of flow chamber and peristaltic pump as described previously (Ishibazawa et al., Invest Ophthalmol Vis Sci, 2011), the confluent monolayer of human corneal epithelial cells was exposed to shear stresses of 0, 1.2, and 12 dyne/cm2 for 24 hours. We evaluated the mRNA expressions of transforming growth factor beta (TGF-β) and matrix metalloproteinase (MMP) using real-time reverse transcription polymerase chain reaction and the levels of these proteins in culture supernatant using an enzyme-linked immunosorbent assay. We also evaluated the phosphorylation of SMAD2 using Western blotting.
The expression of TGFβ1 in cells exposed to shear stress increased significantly compared to that in static cells (1.2 dyne/cm2, 1.47±0.13-fold, p = 0.0034; 12 dyne/cm2, 2.28±0.25-fold, p < 0.0001). The expression of MMP9 in cells exposed to high shear stress also increased significantly compared to that in static cells (1.2 dyne/cm2, 2.30±0.86-fold, p = 0.3853; 12 dyne/cm2, 6.78±2.69-fold, p = 0.0003). The levels of TGFβ1 in culture supernatant increased or decreased from baseline (0 dyne/cm2, +212±38 pg/ml; 1.2 dyne/cm2, +5±39 pg/ml; 12 dyne/cm2, -73±36 pg/ml). The phosphorylation of SMAD2 in cells exposed to shear stress also increased significantly.
Shear stress affected the signaling cascade including TGFβ and MMP in cultured human corneal epithelial cells. The mechanical stress caused by blinking may involve these signaling cascades, which are related to cellular proliferation, migration, and inflammation.
This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.
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