The release of a neutrophil chemotactic factor from UV-B irradiated rabbit corneas in vitro

Rabbit corneas were isolated, mounted on plastic rings to form a cup and the endothelium was covered with RPMI tissue culture medium. The preparation was then irradiated with 1 J. cm-2 of 300 nm light over 1 hour and then incubated for a further two hours in the dark. The supernatant fluid was assayed for chemotactic activity toward rabbit neutrophils in an in vitro Boyden chamber assay. The results indicated that medium from irradiated corneas had a chemotactic activity that was 42% of that produced by the standard chemoattractant f-met-leu-phe, (10(-9) M) while medium from unexposed corneas and exposed medium alone had less than 3% activity. An in vivo assay using sub-epidermal injection into the back of a rabbit gave qualitatively similar results, only f-met-leu-phe and the medium from irradiated corneas causing neutrophil infiltration of the tissue. A checkerboard analysis confirmed that the activity was chemotactic rather than chemokinetic. Release of a chemotactic factor following UV-B irradiation provides a mechanism for the recruitment of neutrophils, at specific localized areas of the endothelium, that is seen after discrete in vivo irradiation. The results also confirm the importance of corneal inflammatory mediators in the development of tissue damage subsequent to exposure to toxic agents.