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Originally published online as doi:10.1189/jlb.0801757 on June 16, 2003

Published online before print June 16, 2003
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(Journal of Leukocyte Biology. 2003;74:412-419.)
© 2003 by Society for Leukocyte Biology

Enhancement by neutrophils of collagen degradation by corneal fibroblasts

Qin Li*, Ken Fukuda{dagger}, Ying Lu*, Yoshikuni Nakamura*, Tai-ichiro Chikama*, Naoki Kumagai* and Teruo Nishida*,1

* Departments of Biomolecular Recognition and Ophthalmology and
{dagger} Ocular Pathophysiology, Yamaguchi University School of Medicine, Ube City, Japan

1Correspondence: Department of Biomolecular Recognition and Ophthalmology, Yamaguchi University School of Medicine, 1-1-1 Minami-Kogushi, Ube City, Yamaguchi 755-8505, Japan. E-mail: nishida1{at}yamaguchi-u.ac.jp

Activated corneal fibroblasts and infiltrated leukocytes are thought to contribute to corneal ulceration. The potential roles of neutrophil-fibroblast and cell-matrix interactions in the degradation of stromal collagen associated with corneal ulceration have now been investigated with the use of three-dimensional cultures of rabbit cells in collagen gels. Degradation of collagen fibrils during culture was measured by spectrophotometric determination of released hydroxyproline. Whereas corneal fibroblasts alone degraded collagen fibrils to a small extent, neutrophils did not. However, the addition of neutrophils or neutrophil–conditioned medium (CM) to cultures of corneal fibroblasts resulted in a marked increase in the amount of collagen degraded by the fibroblasts. The effect of CM from neutrophils cultured in collagen gels on collagen degradation by corneal fibroblasts was greater than that of medium conditioned by neutrophils in monolayer culture. Immunoblot as well as reverse transcription and real-time polymerase chain reaction analyses revealed that neutrophil–CM stimulated the synthesis of matrix metalloproteinase (MMP)-1 and MMP-3 by corneal fibroblasts. The stimulatory effect of neutrophils on collagen degradation by corneal fibroblasts was inhibited by the synthetic MMP inhibitor ilomastat and by interleukin-1 (IL-1) receptor antagonist. These results suggest that factors secreted by collagen-stimulated neutrophils augment collagen degradation by corneal fibroblasts through a stimulatory effect on MMP synthesis and that IL-1 released by neutrophils may contribute to this effect.

Key Words: corneal ulcer • collagen gel • IL-1 receptor antagonist • matrix metalloproteinase




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