PROTEINASE K DECREASES PSEUDOMONAS-AERUGINOSA ADHESION TO WOUNDED CORNEA

The present study was designed to obtain further information regarding the molecular nature of the corneal receptor(s) facilitating Pseudomonas aeruginosa adhesion to cornea. Scarified adult mouse corneas in organ culture were treated for 10 or 60 min with a panel of lipase-free proteases [each at 20 μg ml-1 or 0·22 Units (U) ml-1, activity] including trypsin, chymotrypsin, V8 protease, elastase, subtilisin A, pronase protease and proteinase K. All of these, except proteinase K treatment (20 μg ml-1 for 60 min), either significantly elevated or had no effect (proteinase K 20 μg ml-1 for 10 min) on subsequent bacterial adhesion at 60 min following topical application of the inoculum to the scarified corneal surface. Enzyme treatment times of 10, 30 or 60 min at a higher concentration (50 μg ml-1) of proteinase K, significantly decreased binding at 60 min after bacterial application for each enzyme treatment time. The combined effects of proteases and lipase on bacterial binding also was examined. Eyes treated with proteinase K (20 μg ml-1 for 1 hr) or protease-free lipase (50 000 U ml-1 for 1 hr) alone or in combination, all reduced bacterial binding, but the effect was not additive. Trypsin or lipase alone significantly enhanced or decreased binding, respectively. In contrast, trypsin (20 μg ml-1 for 1 hr) followed by lipase treatment (50 000 U ml-1 for 1 hr) resulted in binding which was not significantly different than phosphate-buffered saline (PBS) control binding, indicating that the trypsin exposed receptor was lipase sensitive. Lastly, sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) of corneal epithelium obtained from trypsin versus proteinase K-treated scarified corneas, revealed that trypsin treatment (20 μg ml-1 for 10 min) generally enhanced corneal epithelial proteins, while similar treatment with proteinase K, but for 30 min, generally decreased corneal epithelial proteins. These findings provide evidence that the binding of P. aeruginosa to the wounded adult ocular surface involves interaction of bacterial adhesins with corneal epithelial host receptor proteins as well as lipids. © 1992 Academic Press Limited.