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Robert J. Walker, Nancy M. Anderson, Youde Jiang, Suleiman Bahouth, Jena J. Steinle; Role of β-Adrenergic Receptor Regulation of TNF-α and Insulin Signaling in Retinal Müller Cells. Invest. Ophthalmol. Vis. Sci. 2011;52(13):9527-9533. doi: https://doi.org/10.1167/iovs.11-8631.
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The goal of this study was to determine the relationship of TNF-α and the downregulation of insulin receptor signaling in retinal Müller cells cultured under hyperglycemic conditions and the role of β-adrenergic receptors in regulating these responses.
Retinal Müller cells were cultured in normal (5 mM) or high (25 mM) glucose until 80% confluent and then were reduced to 2% serum for 18 to 24 hours. The cells were then treated with 10 μM salmeterol followed by Western blot analysis or ELISA. For TNF-α inhibitory studies, the cells were treated with 5 ng/mL of TNF-α for 30 minutes or by a 30-minute pretreatment with TNF-α followed by salmeterol for 6 hours. In the TNF-α short hairpin (sh)RNA experiments, the cells were cultured until 90% confluent, followed by transfection with TNF-α shRNA for 18 hours.
TNF-α-only treatments of Müller cells resulted in significant decreases of tyrosine phosphorylation of the insulin receptor and Akt in high-glucose conditions. Salmeterol (10 μM), a β-2-adrenergic receptor agonist, significantly increased phosphorylation of both insulin receptor and Akt. TNF-α shRNA significantly decreased phosphorylation of IRS-1Ser307, which was further decreased after salmeterol+TNF-α shRNA. Both TNF-α shRNA and salmeterol significantly reduced death of the retinal Müller cells.
These studies demonstrate that β-adrenergic receptor agonists in vitro can restore the loss of insulin receptor activity noted in diabetes. By decreasing the levels of TNF-α and decreasing the phosphorylation of IRS-1Ser307 while increasing tyrosine phosphorylation of insulin receptor, these results suggest a possible mechanism by which restoration of β-adrenergic receptor signaling may protect the retina against diabetes-induced damage.
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