title: Chemokine expression in retinal pigment epithelial ARPE-19 cells in response to coculture with activated T cells.
abstract: OBJECTIVE
  To investigate the effects of T-cell-derived cytokines on gene and protein expression of chemokines in a human RPE cell line (ARPE-19).
  
  
  METHODS
  We used an in vitro coculture system in which the RPE and CD3/CD28-activated T-cells were separated by a membrane. RPE cell expression of chemokine genes was quantified using three different types of microarrays. Protein expression was determined by single and multiplex ELISA and immunoblotting.
  
  
  RESULTS
  Coculture with activated T-cells increased RPE mRNA and protein expression of chemokines CCL2 (MCP-1); CCL5 (RANTES); CCL7 (MCP-3); CCL8 (MCP-2); CXCL1 (GRO-α); IL8 (CXCL8); CXCL9 (MIG); CXCL10 (IP10); CXCL11 (ITAC); and CX3CL1 (fractalkine). CCL7, CXCL9, CXCL10, and CXCL11 were secreted significantly more in the apical direction. Using recombinant human cytokines and neutralizing antibodies, we identified IFNγ and TNFα as the two major T-cell-derived cytokines responsible for the RPE response. For CCL5, CXCL9, CXCL10, CXCL11, CXCL16, and CX3CL1, we observed a synergistic effect of IFNγ and TNFα in combination. CCL20, CXCL1, CXCL6, and IL8 were negatively regulated by IFNγ.
  
  
  CONCLUSIONS
  RPE cells responded to exposure to T-cell-derived cytokines by upregulating expression of multiple chemokines related to microglial, T-cell, and monocyte chemotaxis and activation. This inflammatory stress response may have implications for immune homeostasis in the retina, and for the further understanding of inflammatory ocular diseases such as uveitis and AMD.
authors.affiliation: Eye Research Unit, Department of International Health, Immunology and Microbiology, University of Copenhagen, Copenhagen, Denmark. hjuel@sund.ku.dk
authors.forename: Helene B
authors.initials: HB
authors.lastname: Juel
authors.forename: Carsten
authors.initials: C
authors.lastname: Faber
authors.forename: Maja S
authors.initials: MS
authors.lastname: Udsen
authors.forename: Lasse
authors.initials: L
authors.lastname: Folkersen
authors.forename: Mogens H
authors.initials: MH
authors.lastname: Nissen
mesh_terms: D018106:CD28 Antigens / Q000378:metabolism; D017252:CD3 Complex / Q000378:metabolism; D002460:Cell Line; D018925:Chemokines / Q000235:genetics* / Q000378:metabolism*; D018920:Coculture Techniques; D004797:Enzyme-Linked Immunosorbent Assay; D005786:Gene Expression Regulation / Q000502:physiology*; D055106:Genome-Wide Association Study; D006801:Humans; D015151:Immunoblotting; D007371:Interferon-gamma; D008213:Lymphocyte Activation / Q000502:physiology*; D012333:RNA, Messenger / Q000378:metabolism; D055213:Retinal Pigment Epithelium / Q000166:cytology / Q000378:metabolism*; D013601:T-Lymphocytes / Q000166:cytology / Q000502:physiology*; D014409:Tumor Necrosis Factor-alpha
chemical_list: D018106:CD28 Antigens; D017252:CD3 Complex; D018925:Chemokines; D012333:RNA, Messenger; D014409:Tumor Necrosis Factor-alpha; D007371:Interferon-gamma