title: Role of β-adrenergic receptor regulation of TNF-α and insulin signaling in retinal Muller cells.
abstract: OBJECTIVE
  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.
  
  
  METHODS
  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.
  
  
  RESULTS
  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-1(Ser307), which was further decreased after salmeterol+TNF-α shRNA. Both TNF-α shRNA and salmeterol significantly reduced death of the retinal Müller cells.
  
  
  CONCLUSIONS
  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-1(Ser307) 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.
authors: Department of Ophthalmology, Hamilton Eye Institute, University of Tennessee Health Science Center, Memphis, Tennessee 38163, USA.
authors: Robert J
authors: RJ
authors: Walker
authors: Nancy M
authors: NM
authors: Anderson
authors: Youde
authors: Y
authors: Jiang
authors: Suleiman
authors: S
authors: Bahouth
authors: Jena J
authors: JJ
authors: Steinle
mesh_terms: D058666:Adrenergic beta-2 Receptor Agonists / Q000494:pharmacology*; D000420:Albuterol / Q000031:analogs & derivatives* / Q000494:pharmacology; D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D015153:Blotting, Western; D053148:Caspase 3 / Q000378:metabolism; D002478:Cells, Cultured; D015536:Down-Regulation; D004797:Enzyme-Linked Immunosorbent Assay; D006943:Hyperglycemia / Q000378:metabolism; D055504:Insulin Receptor Substrate Proteins / Q000378:metabolism*; D009457:Neuroglia / Q000187:drug effects / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D051381:Rats; D012160:Retina / Q000166:cytology / Q000378:metabolism*; D000068299:Salmeterol Xinafoate; D015398:Signal Transduction / Q000502:physiology*; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism* / Q000494:pharmacology
chemical_list: D058666:Adrenergic beta-2 Receptor Agonists; C526292:IRS1 protein, human; D055504:Insulin Receptor Substrate Proteins; D014409:Tumor Necrosis Factor-alpha; D000068299:Salmeterol Xinafoate; D051057:Proto-Oncogene Proteins c-akt; D053148:Caspase 3; D000420:Albuterol