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R. J. Walker, J. J. Steinle; Treatment With Salmeterol Restores Insulin Signaling in Retinal Müller Cells. Invest. Ophthalmol. Vis. Sci. 2010;51(13):3727.
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© ARVO (1962-2015); The Authors (2016-present)
To determine the regulation of insulin signaling by beta-adrenergic receptor subtypes on Müller cells in culture.
rMC-1 cells were cultured in high glucose (25mM) conditions until the cells reached 80% confluency. After becoming appropriately confluent, cells were serum-starved for 18-24 hours to eliminate any effects of insulin from the FBS. Following starvation, cells were treated with 10µM isoproterenol for 1, 6, 12, and 24 hours, followed by Western blot and ELISA analysis. For investigation of dominant receptor subtype in Müller cells, cells were treated with either 10uM xamoterol (selective beta-1-adrenergic agonist) or 10uM salmeterol (selective beta-2-adrenergic receptor agonist) to stimulate beta-adrenergic receptors without presence of antagonist. We then compared the effects of each of these drugs cleaved caspase 3 levels, phosphorylation of Akt, and insulin receptor to previous results with isoproterenol (a non-selective beta-adrenergic receptor agonist).
Treatment of cells with 10uM isoproterenol significantly increased the phosphorylation activity of insulin receptor beta and Akt, while significantly decreasing caspase 3 levels at 24 hours. Treatment with a beta-1-adrenergic receptor agonist, xamoterol, significantly decreased caspase 3 levels following 1 and 6 hours of treatment. Treatment with a beta-2-adrenergic receptor agonist, salmeterol, significantly decreased cleaved caspase 3 levels at all timepoints. Both xamoterol and salmeterol significantly increased phosphorylation of insulin receptor beta in cells cultured in high glucose. Phosphorylation of Akt, an anti-apoptotic factor, was also significantly increased following treatment with isoproterenol and salmeterol, but salmeterol produced a stronger response than isoproterenol.
These results indicate that change in glucose environment reduces insulin signaling in cultured rat Muller but treatment with a non-selective- or selective beta- adrenergic receptor agonist can restore normal signaling. These results also suggest that treatment with a beta-2 adrenergic receptor agonist can restore insulin signaling for longer periods of time vs. a non-selective- or beta-1-adrenergic receptor selective drug. Overall, these results are the first to report that beta-adrenergic receptors (specifically beta-2-adrenergic receptors) can regulate insulin signaling in hyperglycemic Müller cells, which may offer a new avenue for therapeutic development.
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