Nicotine modulation of in vitro human gingival fibroblast beta1 integrin expression.
BACKGROUND
Smoking is clearly a high-risk behavior for the development of periodontal disease, and nicotine is the major vasoactive component in tobacco. Integrins are a large family of homologous transmembrane adhesion proteins serving as the principal receptors on animal cells to bind and communicate with many extracellular matrix proteins such as collagen, fibronectin, and laminin. This study's aim was to evaluate nicotine's effects on beta1 integrin expression as a function of either 1) the generalized effects on RNA/protein synthesis or 2) as a specific modulation of beta1 integrin synthesis.


METHODS
Pooled human gingival fibroblasts (HGFs) were cultured in nicotine concentrations commonly seen in smokers (0.025 to 0.8 microM), and relative incorporation of [35S]methionine into newly synthesized protein or [3H]uridine into newly synthesized RNA was measured. Cultures were harvested at various times, and duplicate cell aliquots were homogenized and fractionated to obtain cell membrane-enriched preparations or solubilized to obtain whole cell lysates. Radiolabeled RNA and proteins were quantitated by trichloroacetic acid (TCA) precipitation and liquid scintillation spectrometry. Beta1 integrin subunits were detected by SDS-PAGE and Western blotting, and the relative intensities of reactive bands were quantitated by scanning densitometry.


RESULTS
After 17 hours of exposure, 0.4 and 0.8 microM nicotine resulted in a dose-dependent increase in beta1 integrin in whole cell lysates, and a decrease in beta1 integrin in the corresponding membrane-enriched fractions. There was also a statistically significant decrease (P < or = 0.05) in radiolabeled proteins in culture. Although there appeared to be a mild, generalized reduction in radiolabeled RNA in nicotine-treated cultures compared to controls, a 1-way analysis of variance showed no statistical significance between values.


CONCLUSIONS
Our results suggest that nicotine may induce an altered compartmentalization process in which beta1 integrin molecules are produced, but are not appropriately transferred to the membrane. Nicotine effects on cellular protein synthesis and its modulation of beta1 integrin expression may impair gingival fibroblast ability to adhere to and communicate with one another and with the extracellular matrix, which could impair wound healing and/or exacerbate periodontal disease.
Snyder
Harold B
HB
U.S. Army Dental Activity, Baumholder, Germany.
Caughman
Gretchen
G
Lewis
Jill
J
Billman
Michael A
MA
Schuster
George
G
D000704:Analysis of Variance; D015153:Blotting, Western; D002478:Cells, Cultured; D011232:Chemical Precipitation; D003720:Densitometry; D004305:Dose-Response Relationship, Drug; D004591:Electrophoresis, Polyacrylamide Gel; D005347:Fibroblasts / Q000187:drug effects* / Q000276:immunology; D005786:Gene Expression Regulation; D005881:Gingiva / Q000166:cytology / Q000187:drug effects* / Q000276:immunology; D006801:Humans; D019012:Integrin beta1 / Q000187:drug effects* / Q000235:genetics; D008715:Methionine / Q000378:metabolism; D009538:Nicotine / Q000008:administration & dosage / Q000494:pharmacology*; D018722:Nicotinic Agonists / Q000008:administration & dosage / Q000494:pharmacology*; D014176:Protein Biosynthesis; D011506:Proteins / Q000187:drug effects; D012313:RNA / Q000187:drug effects / Q000235:genetics; D019275:Radiopharmaceuticals; D013057:Spectrum Analysis; D013462:Sulfur Radioisotopes; D014238:Trichloroacetic Acid; D014316:Tritium; D014529:Uridine / Q000378:metabolism
D019012:Integrin beta1; D018722:Nicotinic Agonists; D011506:Proteins; D019275:Radiopharmaceuticals; D013462:Sulfur Radioisotopes; D014316:Tritium; D014238:Trichloroacetic Acid; D012313:RNA; D009538:Nicotine; D008715:Methionine; D014529:Uridine