May 2006
Volume 47, Issue 13
ARVO Annual Meeting Abstract  |   May 2006
Relative Contribution of LIX/CXCL5, KC/CXCL1, and MIP–2/CXCL2 in LPS Keratitis
Author Affiliations & Notes
  • M. Lin
    Ophthalmology, Case Western Reserve Univ, Cleveland, OH
  • E.C. Carlson
    Ophthalmology, Case Western Reserve Univ, Cleveland, OH
  • E. Diaconu
    Ophthalmology, Case Western Reserve Univ, Cleveland, OH
  • E. Pearlman
    Ophthalmology, Case Western Reserve Univ, Cleveland, OH
  • Footnotes
    Commercial Relationships  M. Lin, None; E.C. Carlson, None; E. Diaconu, None; E. Pearlman, None.
  • Footnotes
    Support  NIH Grant EY10320, Research Prevent Blindness Foundation, Ohio Lions Eye Research Foundation
Investigative Ophthalmology & Visual Science May 2006, Vol.47, 4361. doi:
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      M. Lin, E.C. Carlson, E. Diaconu, E. Pearlman; Relative Contribution of LIX/CXCL5, KC/CXCL1, and MIP–2/CXCL2 in LPS Keratitis . Invest. Ophthalmol. Vis. Sci. 2006;47(13):4361.

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      © ARVO (1962-2015); The Authors (2016-present)

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Purpose: : Neutrophils infiltrating the cornea are crucial to the pathogenesis of keratitis as evidenced by clinical cases and animal models where neutrophil infiltration is hindered resulting in decreased haze, neovascularization, and corneal edema. CXC chemokines are associated with the regulation of neutrophil recruitment, migration, and activation at sites of infection/injury; the relative contribution of CXC chemokines to these functions remains unclear. Preliminary data revealed that corneal fibroblasts are only capable of producing two of the three known ELR+ CXC chemokines,LIX and KC. Therefore, these studies have been concentrated towards determining the contribution of LIX and KC in neutrophil infiltration to the murine cornea and subsequent effects on corneal inflammation.

Methods: : Murine corneal fibroblasts (MK–T1 cells) were stimulated with LPS and assayed at different time points for LIX,KC and MIP–2 mRNA expression by qRT–PCR and for protein by ELISA. C57BL/6 corneas were injected intrastromally with LPS and LIX, KC,and MIP–2 expression was examined by ELISA. An in vivo assay using siRNA to knockdown gene expression was developed by injecting an adenoviral eGFP construct in conjunction with eGFP siRNA into the corneal stroma and efficiency of GFP expression monitored by fluorescence microscopy. LIX siRNA was injected into the corneal stroma prior to LPS stimulation in GFP chimeric or normal C57Bl/6 mice. Cell infiltration in chimerics was monitored in vivo by fluorescence microscopy while corneas of C57Bl/6 mice were excised after 24hrs and assayed for chemokines by ELISA.

Results: : Murine corneal fibroblasts displayed differential mRNA expression of LIX,KC, and MIP–2. However, only LIX and KC protein was detected. In vivo mouse experiments also demonstrated similar patterns of chemokine production in the corneal stroma. eGFP expression in the corneal stroma was significantly decreased in the presence of eGFP siRNA compared to control siRNA. While LIX siRNA selectively decreased LIX production in vivo in response to LPS, this decreased LIX expression did not affect cellular infiltration into the murine cornea.

Conclusions: : These findings demonstrate the selective production of chemokines,LIX and KC,by corneal fibroblasts adding to the notion of a nonmyeloid and myeloid pattern of chemokine production which could reflect their contribution to the inflammatory process. In addition, while the exact contribution of LIX to neutrophil function and corneal inflammation remains to be clarified, these results indicate the potential importance of KC in initial neutrophil recruitment during corneal inflammation.

Keywords: cytokines/chemokines • cornea: stroma and keratocytes • inflammation 

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