HARVARD DEPARTMENT OF OPHTHALMOLOGY
Category : Basic and Translational Research Candidate : Tessa Fitch Poster #: B7
Resveratrol Protects RPE Against TNFα-Induced Inflammation in Age Related Macular Degeneration
Tessa C Fitch , Scott Frank , Leo A . Kim , Daisy Y . Shu
Purpose : Age-related macular degeneration ( AMD ) is a leading cause of irreversible blindness and vision loss worldwide . Damage to retinal pigment epithelial cells ( RPE ) from inflammatory cytokines plays a key role in the pathogenesis of AMD . This study examined the effects of tumor necrosis factor-α ( TNFα ), a pro-inflammatory cytokine that is heavily involved in AMD , on RPE health and function . Resveratrol is a natural organic compound known to have very potent anti-inflammatory effects . In this study , we investigated the capacity of resveratrol to block TNFα-driven inflammation and metabolic dysfunction in RPE .
Methods : Matured H-RPE cells were treated with TNFα ( 10 ng / ml ) and / or treated with resveratrol ( 50 μM ) for 24 hours to 5 days . Glycolytic and oxidative ( OXPHOS ) metabolic profiles were determined with the Seahorse XFe96 Bioanalyzer . Gene expression of metabolic markers was assessed using qPCR . IL-6 secretion was quantified by using the Human IL-6 ELISA Kit . The EVOS m700 microscope was used for imaging morphology and fluorescence . Data and statistical analysis were performed using GraphPad Prism , Excel , ImageJ , and BioRender .
Results : Resveratrol suppressed TNFα-induced upregulation of IL-6 , IL-8 , TLR2 , and MCP-1 . Maximal respiration , spare respiratory capacity , and glycolysis levels were enhanced with resveratrol on the Seahorse XFe96 . Co-treatment with resveratrol and TNFα further enhances oxygen consumption rate . Cells treated with TNFα showed an elongated morphology with loss of the cuboidal cobblestone morphology . Concurrent treatment with resveratrol rescued morphology with cells appearing similar to control . Cells with resveratrol alone maintained the regular morphology . Resveratrol suppressed TNFα-induced upregulation of cytoplasmic reactive oxygen species ( ROS ) down to basal levels on the CellROX assay .
Conclusions : RPE cells are profoundly affected by the proinflammatory effects of TNFα . TNFα not only disrupts the structural epithelial morphology of H-RPE cells , it also causes dysfunction of metabolic pathways and mitochondria . Treatment with the natural organic compound , resveratrol , efficiently blocks TNFα-induced proinflammatory activation , bioenergetic reprogramming and ROS upregulation of RPE . These results reveal a critical interplay between inflammation , metabolic dysfunction and oxidative stress in RPE , identifying resveratrol as a potent drug to treat AMD .