Granulocyte macrophage colony-stimulating factor expression in human herpetic stromal keratitis: implications for the role of neutrophils in HSK

Rui Duan, Lies Remeijer, Jessica M van Dun, Albert D M E Osterhaus, Georges M G M Verjans

Research output: Contribution to journalArticleResearchpeer-review


PURPOSE: Granulocyte macrophage colony-stimulating factor (GM-CSF) is thought to play a key role in chronic inflammatory diseases by governing the survival and function of infiltrating neutrophils. The objective of this study was to determine the putative role of GM-CSF in the pathogenesis of human herpetic stromal keratitis (HSK).

METHODS: Primary human corneal fibroblast (HCF) cultures and a telomerase-immortalized human corneal epithelial (HCE) cell line representative of native HCE were stimulated with the known HSK-inducing cytokines interferon (IFN)-gamma, interleukin (IL)-1beta, and tumor necrosis factor (TNF)-alpha. Alternatively, the T-cell cytokine IL-17 was added solely or simultaneously. Human neutrophils were incubated with conditioned medium (CM) of the HCF and HCE stimulated with the aforementioned cytokines, or recombinant GM-CSF, and their viability or activation status was determined by flow cytometry. GM-CSF and IL-8 secretion levels in the CM were determined by ELISA. The antibody-dependent cellular cytotoxicity (ADCC) of neutrophils toward herpes simplex virus (HSV)-infected HCFs was determined by flow cytometry. The expression of GM-CSF was determined in HSK and control corneal buttons by real-time RT-PCR and immunohistology.

RESULTS: Compared with IFN-gamma, CM of either cell type stimulated with IL-1beta, or in the case of HCE cells, stimulated with TNF-alpha or IL-17, delayed neutrophil apoptosis significantly. Only in HCFs did IL-17 exhibit a synergistic effect with TNF-alpha. The antiapoptotic activity was attributable in part to the GM-CSF secreted by the activated HCFs and HCE cells. GM-CSF stimulation of neutrophils induced their activation and the secretion of IL-8. GM-CSF did not increase significantly the ADCC reaction of neutrophils toward HSV-infected HCFs. Finally, GM-CSF was expressed in corneas of the patients with HSK but not in control subjects.

CONCLUSIONS: The data suggest that GM-CSF, expressed by cornea-resident cells such as HCFs and HCE cells, may play a role in the immunopathogenesis of HSK by prolonging the survival and modulating the effector function of corneal infiltrating neutrophils.

Original languageEnglish
Pages (from-to)277-84
Number of pages8
JournalInvestigative ophthalmology & visual science
Issue number1
Publication statusPublished - Jan 2007


  • Antibody-Dependent Cell Cytotoxicity
  • Cells, Cultured
  • Corneal Stroma/cytology
  • Dose-Response Relationship, Drug
  • Enzyme-Linked Immunosorbent Assay
  • Epithelium, Corneal/drug effects
  • Fibroblasts/drug effects
  • Granulocyte-Macrophage Colony-Stimulating Factor/genetics
  • Herpesvirus 1, Human/physiology
  • Humans
  • Immunoenzyme Techniques
  • Interferon-gamma/pharmacology
  • Interleukin-1beta/pharmacology
  • Interleukin-8/metabolism
  • Keratitis, Herpetic/metabolism
  • Neutrophil Activation
  • Neutrophils/physiology
  • RNA/isolation & purification
  • RNA, Messenger/metabolism
  • Reverse Transcriptase Polymerase Chain Reaction
  • Tumor Necrosis Factor-alpha/pharmacology


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