title: Differential regulation of ENA-78 and GCP-2 gene expression in human corneal keratocytes and epithelial cells.
abstract: OBJECTIVE
  To determine whether interleukin (IL)-1alpha- and tumor necrosis factor (TNF)-alpha-stimulated human corneal epithelial cells (HCECs) and human corneal keratocytes (HCKs) produce the alpha-chemokines epithelial cell-derived neutrophil attractant (ENA)-78 and granulocyte chemotactic protein (GCP)-2.
  
  
  METHODS
  Cultures of HCECs and HCKs were stimulated with either human recombinant IL-1alpha or TNF-alpha. At selected times after stimulation, culture supernatants were harvested and assayed for ENA-78 and GCP-2 by enzyme-linked immunosorbent assay. RNA was extracted from cell cultures to measure steady state levels of intracellular ENA-78 and GCP-2 pre-mRNA and mRNA by the reverse transcription-polymerase chain reaction.
  
  
  RESULTS
  Exposure of HCECs to either IL-1alpha or TNF-alpha stimulated a more than 4.5-fold increase in ENA-78 RNA and protein synthesis without stimulating a significant increase in either GCP-2 RNA synthesis or protein production. Exposure of HCK to IL-1alpha stimulated a 10-fold increase in ENA-78 and GCP-2 RNA synthesis and a more than 300-fold increase in ENA-78 and GCP-2 protein production. In contrast, exposure of keratocytes to TNF-alpha significantly enhanced ENA-78 RNA synthesis, resulting in a more than 68-fold increase in ENA-78 protein synthesis without significantly enhancing either GCP-2 gene expression or protein secretion.
  
  
  CONCLUSIONS
  ENA-78 gene expression is significantly enhanced in both HCECs and HCKs in response to either IL-1alpha or TNF-alpha stimulation. In contrast, GCP-2 synthesis is only inducible in IL-1alpha-stimulated HCKs. The results suggest that GCP-2 gene expression is more tightly regulated in diseased or injured corneal tissue than is ENA-78 gene expression.
authors.affiliation: Department of Microbiology and Immunology, College of Medicine, University of South Alabama, Mobile, Alabama, USA.
authors.forename: Rebecca A
authors.initials: RA
authors.lastname: Fillmore
authors.forename: Shannon E
authors.initials: SE
authors.lastname: Nelson
authors.forename: Robert N
authors.initials: RN
authors.lastname: Lausch
authors.forename: John E
authors.initials: JE
authors.lastname: Oakes
mesh_terms: D002478:Cells, Cultured; D054365:Chemokine CXCL5; D054427:Chemokine CXCL6; D019743:Chemokines, CXC / Q000235:genetics* / Q000378:metabolism; D003319:Corneal Stroma / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D004797:Enzyme-Linked Immunosorbent Assay; D019573:Epithelium, Corneal / Q000187:drug effects* / Q000378:metabolism; D005347:Fibroblasts; D005786:Gene Expression Regulation / Q000187:drug effects*; D006801:Humans; D007375:Interleukin-1 / Q000494:pharmacology*; D016209:Interleukin-8 / Q000031:analogs & derivatives* / Q000235:genetics* / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*
chemical_list: C486263:CXCL5 protein, human; C515503:CXCL6 protein, human; D054365:Chemokine CXCL5; D054427:Chemokine CXCL6; D019743:Chemokines, CXC; D007375:Interleukin-1; D016209:Interleukin-8; D012333:RNA, Messenger; D014409:Tumor Necrosis Factor-alpha