title: Sphingosine 1-phosphate elicits proinflammatory responses in ARPE-19 cells.
abstract: OBJECTIVE
  To investigate the effects of sphingosine 1-phosphate (S1P) on the production of inflammatory mediators and the signaling pathways involved in S1P-mediated production of cytokines by ARPE-19 cells.
  
  
  METHODS
  Expression of S1P receptors was examined using RT-PCR and real-time PCR. ARPE-19 cells were stimulated with S1P or TNF-α, and by coculturing with S1P in the presence or absence of pertussis toxin (PTX) or a series of kinase inhibitors. The induction of inflammatory cytokine production was determined by ELISA. Western blot analysis was used to detect the activation of signaling mediators and S1P(3) receptor.
  
  
  RESULTS
  ARPE-19 cells express all the known receptors for S1P. Moreover, exogenously applied S1P induces ARPE-19 cell secretion of interleukin-8 (IL-8) in a dose- and time-dependent manner, but not IL-6 and monocyte chemotactic protein-1 (MCP-1). S1P-mediated IL-8 secretion is regulated by PTX, extracellular regulated protein kinases 1/2 (ERK1/2), and p38 mitogen-activated protein kinase (MAPK). Interestingly, treatment of ARPE-19 cells with TNF-α increases S1P(3) expression and correlates with the enhancement of S1P-induced IL-8 and IL-6 production.
  
  
  CONCLUSIONS
  This study demonstrates that S1P significantly promoted ARPE-19 cells to secrete inflammatory mediators. extracellular regulated protein kinases 1/2 (ERK1/2), p38 MAPK, and G(i)-dependent pathways are important signaling components in S1P-mediated IL-8 secretion by ARPE-19 cells. Moreover, these results provide evidence that S1P stimulation of RPE cells may play a role in regulating leukocyte function during ocular inflammation.
authors.affiliation: First Affiliated Hospital of Chongqing Medical University, Youyi Road 1, Chongqing Key Laboratory of Ophthalmology, and Chongqing Eye Institute, Chongqing, People's Republic of China.
authors.forename: Yanbin
authors.initials: Y
authors.lastname: Qiao
authors.forename: Ranran
authors.initials: R
authors.lastname: Hu
authors.forename: Qian
authors.initials: Q
authors.lastname: Wang
authors.forename: Jian
authors.initials: J
authors.lastname: Qi
authors.forename: Yan
authors.initials: Y
authors.lastname: Yang
authors.forename: Aize
authors.initials: A
authors.lastname: Kijlstra
authors.forename: Peizeng
authors.initials: P
authors.lastname: Yang
mesh_terms: D015153:Blotting, Western; D002460:Cell Line; D018920:Coculture Techniques; D004797:Enzyme-Linked Immunosorbent Assay; D005786:Gene Expression Regulation / Q000502:physiology; D006801:Humans; D018836:Inflammation Mediators / Q000378:metabolism*; D008246:Lysophospholipids / Q000494:pharmacology*; D020935:MAP Kinase Signaling System / Q000502:physiology; D012333:RNA, Messenger / Q000378:metabolism; D060888:Real-Time Polymerase Chain Reaction; D049349:Receptors, Lysosphingolipid / Q000235:genetics; D055213:Retinal Pigment Epithelium / Q000187:drug effects* / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction / Q000502:physiology*; D013110:Sphingosine / Q000031:analogs & derivatives* / Q000494:pharmacology; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology; D048051:p38 Mitogen-Activated Protein Kinases / Q000378:metabolism
chemical_list: D018836:Inflammation Mediators; D008246:Lysophospholipids; D012333:RNA, Messenger; D049349:Receptors, Lysosphingolipid; D014409:Tumor Necrosis Factor-alpha; C060506:sphingosine 1-phosphate; D048051:p38 Mitogen-Activated Protein Kinases; D013110:Sphingosine