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Jin Liang, Iris D Navarro, Pedro Gonzalez, Fan Yuan, W Daniel Stamer; Mechanical and cytokine stimulation of smooth muscle actin expression in human trabecular meshwork cells. Invest. Ophthalmol. Vis. Sci. 2014;55(13):2887.
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The goal of the present study was to determine the impact of mechanical stress and transforming growth factor beta-2 (TGF-β2) on the protein expression of the smooth muscle actin (SMA) in human trabecular meshwork (TM) cells.
Primary cultures of human TM cells isolated from three different donor eye pairs were used as models. Confluent and mature monolayers of TM cells were subjected to either (i) TGF-β2 (0.1-5 ng/ml) for 24 hr or (ii) TGF-β2 (5ng/ml) for 6, 24 and 48 hours or (iii) cyclic mechanical stretch (16% strain at 1 Hertz) using a Flexcell System ±TGF-β2 (5ng/ml) for 24 hours. Relative expression of SMA was determined by Western Blot using anti-SMA specific IgGs normalized to total protein on nitrocellulose blots by Pierce reversible stain and imageJ analysis.
SMA protein levels in human TM cells increased 3.5 fold (p=0.03) after 5ng/ml TGF-β2 treatment, but remained unchanged at lower concentrations (0.1 and 1ng/ml). Maximum effects of TGF-β2 were observed at 48 hours after treatment. By comparison, exposing human TM cells to mechanical stretch increased SMA 2-fold at 6 hours and 3-fold at 24 hours (p<0.05). The combination of mechanical stretch and TGF-β2 treatment synergistically increased SMA production after 24 hours. (n=2)
Results show that human TM cells rapidly upregulate their contractile machinery in response to both mechanical and cytokine stimuli, and when present together responses are exacerbated. Prolonged stimulation may facilitate transdifferentiation of TM cells to accommodate stress in a homeostatic or pathological fashion, such as may occur in glaucoma.
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