title: Proinflammatory cytokines decrease the expression of genes critical for RPE function.
abstract: Proinflammatory cytokines interferon gamma (IFN-γ), tumor necrosis factor alpha (TNF-α), and interleukin-1 beta (IL-1β) secreted by infiltrating lymphocytes or macrophages may play a role in triggering RPE dysfunction associated with age-related macular degeneration (AMD). Binding of these proinflammatory cytokines to their specific receptors residing on the RPE cell surface can activate signaling pathways that, in turn, may dysregulate cellular gene expression. The purpose of the present study was to investigate whether IFN-γ, TNF-α, and IL-1β have an adverse effect on the expression of genes essential for RPE function, employing the RPE cell line ARPE-19 as a model system.
  
  
  
  ARPE-19 cells were cultured for 3-4 months until they exhibited epithelial morphology and expressed mRNAs for visual cycle genes. The differentiated cells were treated with IFN-γ, TNF-α, and/or IL-1β, and gene expression was analyzed with real-time PCR analysis. Western immunoblotting was employed for the detection of proteins.
  
  
  
  Proinflammatory cytokines (IFN-γ + TNF-α + IL-1β) greatly increased the expression of chemokines and cytokines in cultured ARPE-19 cells that exhibited RPE characteristics. However, this response was accompanied by markedly decreased expression of genes important for RPE function, such asCDH1,RPE65,RDH5,RDH10,TYR, andMERTK. This was associated with decreased expression of the genesMITF,TRPM1, andTRPM3, as well as microRNAs miR-204 and miR-211, which are known to regulate RPE-specific gene expression. The decreased expression of the epithelial marker geneCDH1was associated with increased expression of mesenchymal marker genes (CDH2,VIM, andCCND1) and epithelial-mesenchymal transition (EMT) promoting transcription factor genes (ZEB1andSNAI1).
  
  
  
  RPE cells exposed to proinflammatory cytokines IFN-γ, TNF-α, and IL-1β showed decreased expression of key genes involved in the visual cycle, epithelial morphology, and phagocytosis. This adverse effect of proinflammatory cytokines, which could be secreted by infiltrating lymphocytes or macrophages, on the expression of genes indispensable for RPE function may contribute to the RPE dysfunction implicated in AMD pathology.
authors.affiliation: Laboratory of Retinal Cell and Molecular Biology, National Eye Institute, National Institutes of Health, Bethesda, MD.
authors.forename: R Krishnan
authors.initials: RK
authors.lastname: Kutty
authors.affiliation: Laboratory of Retinal Cell and Molecular Biology, National Eye Institute, National Institutes of Health, Bethesda, MD.
authors.forename: William
authors.initials: W
authors.lastname: Samuel
authors.affiliation: Laboratory of Retinal Cell and Molecular Biology, National Eye Institute, National Institutes of Health, Bethesda, MD.
authors.forename: Kaifa
authors.initials: K
authors.lastname: Boyce
authors.affiliation: Laboratory of Retinal Cell and Molecular Biology, National Eye Institute, National Institutes of Health, Bethesda, MD.
authors.forename: Aswini
authors.initials: A
authors.lastname: Cherukuri
authors.affiliation: Laboratory of Retinal Cell and Molecular Biology, National Eye Institute, National Institutes of Health, Bethesda, MD.
authors.forename: Todd
authors.initials: T
authors.lastname: Duncan
authors.affiliation: Laboratory of Retinal Cell and Molecular Biology, National Eye Institute, National Institutes of Health, Bethesda, MD.
authors.forename: Cynthia
authors.initials: C
authors.lastname: Jaworski
authors.affiliation: Radiation Biology Branch, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD.
authors.forename: Chandrasekharam N
authors.initials: CN
authors.lastname: Nagineni
authors.affiliation: Laboratory of Retinal Cell and Molecular Biology, National Eye Institute, National Institutes of Health, Bethesda, MD.
authors.forename: T Michael
authors.initials: TM
authors.lastname: Redmond
mesh_terms: D000429:Alcohol Oxidoreductases / Q000235:genetics; D015153:Blotting, Western; D015820:Cadherins / Q000235:genetics; D002352:Carrier Proteins / Q000235:genetics; D002460:Cell Line; D018925:Chemokines / Q000235:genetics; D016207:Cytokines / Q000235:genetics*; D005136:Eye Proteins / Q000235:genetics*; D005786:Gene Expression Regulation / Q000502:physiology*; D006801:Humans; D007371:Interferon-gamma / Q000494:pharmacology*; D053583:Interleukin-1beta / Q000494:pharmacology*; D051739:Microphthalmia-Associated Transcription Factor / Q000235:genetics; D012333:RNA, Messenger / Q000235:genetics; D060888:Real-Time Polymerase Chain Reaction; D055213:Retinal Pigment Epithelium / Q000187:drug effects* / Q000378:metabolism; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*; D019745:cis-trans-Isomerases / Q000235:genetics; D000099178:Retinoid Isomerohydrolase
chemical_list: D000429:Alcohol Oxidoreductases; D015820:Cadherins; D002352:Carrier Proteins; D018925:Chemokines; D016207:Cytokines; D005136:Eye Proteins; D007371:Interferon-gamma; D053583:Interleukin-1beta; D051739:Microphthalmia-Associated Transcription Factor; D012333:RNA, Messenger; D014409:Tumor Necrosis Factor-alpha; D019745:cis-trans-Isomerases; D000099178:Retinoid Isomerohydrolase; C020932:11-cis-retinal-binding protein; C497234:MITF protein, human; C113992:trans-retinol dehydrogenase