Cleavage of CD14 on Human Gingival Fibroblasts Cocultured with Activated Neutrophils Is Mediated by Human Leukocyte Elastase Resulting in Down-Regulation of Lipopolysaccharide-Induced IL-8 Production

摘要

Activated polymrophonuclear leuckocytes (PMNs) release various types of proteases and express them on the cell surface.The proteases play important roles in PMN-mediated events.In the present study,flow cytometric analysis revealed that CD14 expression onhuman gingival fibroblasts (HGF) was markedly reduced by PMA-activated PMNs in a coculture system.We found that this reduction was caused by both secreted and cells urface proteases produced by activated PMNs.A protease responsible for the reduction was found to be hmuan leukocyte elastase (HLE) secreted from the activated PMNs by use of various proteas inhibitors,although HLE was only partially involved in CD14 reduction caused by cell-bound-molecule(s) finxed PMNs.Analysis with prrified HLE revealed a time- and dose-dependent reduction of CD174 HGF,and complete reduction was observed by 20 #mu#g/ml HLE treatment for 30-60 min,but the other molecules such as CD26,CD59,CD157,and MHC class I on HGF were only slightly reduced.This reduction of CD14 resulted from direct proteolysis by HLE on the cell surface,because HLE reduced CD14 on fixed HGF and also on purified cell membranes.As a result of CD14 proteolysis,IL-8 production by HGF was suppressed when triggered by 10 ng/ml LPS,but not by IL-1#alpha#,indicating that HLE inhibited a CD14-dependent cell activation.These findings suggested that activated PMNs have a potential negative feedback mechanism for HGF function at the inflammatory site,particulatry in peridontal tissues.