Nicotine increases the collagen-degrading ability of human gingival fibroblasts.
OBJECTIVE
The objective of this study was to determine the effects that nicotine and the combination of nicotine and Porphyromonas gingivalis supernatant have on human gingival fibroblast-mediated collagen degradation.


METHODS
Human gingival fibroblasts were cultured with 25-500 microg/ml of nicotine in collagen-coated six-well plates. On days 1-5, the conditioned media was collected for zymography and western blot analyses of matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs). The cells were then removed and the collagen cleavage visualized by Coomassie blue staining. To examine the combined effect, 250 microg/ml of nicotine and 10% v/v culture supernatant of P. gingivalis ATCC 33277 were added to the human gingival fibroblasts. The mRNA levels of multiple MMPs and TIMPs were monitored.


RESULTS
Nicotine increased the human gingival fibroblast-mediated collagen cleavage. The MMP-14 and MMP-2 produced by the nicotine-treated human gingival fibroblasts more readily underwent zymogen activation. Nicotine treatment resulted in TIMP-2 redistribution to the cell surface. The mRNAs of multiple MMPs and TIMPs were unaltered by nicotine. An additive collagen cleavage effect was observed when the human gingival fibroblasts were treated with both nicotine and P. gingivalis.


CONCLUSIONS
Nicotine increased human gingival fibroblast-mediated collagen degradation, in part through the activation of membrane-associated MMPs. Nicotine and P. gingivalis had an additive effect on human gingival fibroblast-mediated collagen degradation.
Zhou
J
J
Department of Oral Biology, Indiana University School of Dentistry, Indianapolis, IN, USA. jzhou2@iupui.edu
Olson
B L
BL
Windsor
L J
LJ
D015153:Blotting, Western / Q000379:methods; D002478:Cells, Cultured / Q000187:drug effects; D003094:Collagen / Q000187:drug effects; D005347:Fibroblasts / Q000187:drug effects* / Q000201:enzymology; D005731:Ganglionic Stimulants / Q000009:adverse effects*; D005881:Gingiva / Q000166:cytology / Q000187:drug effects*; D006801:Humans; D020782:Matrix Metalloproteinases / Q000187:drug effects / Q000378:metabolism; D009538:Nicotine / Q000009:adverse effects*; D016966:Porphyromonas gingivalis / Q000378:metabolism*; D012333:RNA, Messenger / Q000187:drug effects / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods; D019714:Tissue Inhibitor of Metalloproteinases / Q000187:drug effects / Q000378:metabolism
D005731:Ganglionic Stimulants; D012333:RNA, Messenger; D019714:Tissue Inhibitor of Metalloproteinases; D009538:Nicotine; D003094:Collagen; D020782:Matrix Metalloproteinases