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Am J Physiol Regul Integr Comp Physiol 274: R104-R111, 1998;
0363-6119/98 $5.00
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Vol. 274, Issue 1, R104-R111, January 1998

Smokeless tobacco-exposed oral keratinocytes increase macromolecular efflux from the in situ oral mucosa

Israel Rubinstein1, Xiao-Pei Gao1, Sergei Pakhlevaniants1, and Dolphine Oda2

1 Department of Medicine, University of Illinois at Chicago, and West Side Department of Veterans Affairs Medical Center, Chicago, Illinois 60612; and 2 Department of Oral Biology, University of Washington, Seattle, Washington 98195

The purpose of this study was to determine whether supernatants of cultured human oral keratinocytes (HOK) exposed to an aqueous extract of smokeless tobacco (STE) increase macromolecular efflux from the oral mucosa in vivo and, if so, whether bradykinin mediates in part this response. Subconfluent monolayers of HOK were incubated with STE or media, and supernatants were collected 24, 48, and 72 h thereafter. Using intravital microscopy, we found that suffusion of supernatants of STE- but not media-exposed HOK elicited significant concentration- and time-dependent increases in efflux of fluorescein isothiocyanate-labeled dextran (mol mass 70 kDa) from the in situ hamster cheek pouch (P < 0.05). These effects were significantly attenuated by HOE-140 and NPC-17647 but not by des-Arg9,[Leu8]-bradykinin. Proteolytic activity was increased in supernatants of STE- but not media-exposed HOK. However, a mixture of leupeptin, Bestatin, and DL-2-mercaptomethyl-3-guanidinoethylthiopropanoic acid had no significant effects on HOK supernatant-induced responses. Collectively, these data suggest that oral keratinocytes modulate smokeless tobacco-induced increase in macromolecular efflux from the in situ oral mucosa in part by elaborating proteases that may account for local bradykinin production.

gingiva; inflammation; bradykinin; proteases; hamster


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