HBD-1 and hBD-2 expression in HaCaT keratinocytes stimulated with nicotine.
OBJECTIVE
The impact of nicotine on the local innate immune response in the oral cavity is unclear. The aim of the present study was to evaluate the possible effects of nicotine on the gene expression of human beta-defensin-1 and -2 in HaCaT keratinocytes.


METHODS
HaCaTs were cultured in six-well plates in Dulbecco's minimum essential medium (DMEM) supplemented with 10% FBS at a density of ×10(6). Cells were pretreated with 10 μg/ml nicotine (12 h), and then stimulated with 50 ng/ml TNF-α (during the following 12 h); or were pretreated with 50 ng/ml TNF-α, and then stimulated with 10 μg/ml nicotine; or were not pretreated but only stimulated with either nicotine or TNF-α, or a combination of both. Total RNA was extracted and analysed by real-time RT-PCR for human beta-defensins-1-, -2-, and interleukins IL-1β- and IL-6-, as well as GAPDH-mRNA. The obtained data were analysed using Tukey's B multiple comparison test for post hoc analysis.


RESULTS
Pretreatment with nicotine caused a significant 2.5-fold inhibition of TNF-α-stimulated hBD-2 mRNA expression compared to TNF-α alone (p = 0.004). Simultaneous treatment with TNF-α and nicotine caused a significant 2-fold inhibition of hBD-2 mRNA compared to TNF-α alone (p = 0.041).


CONCLUSIONS
The present results suggest that the pre-exposition to nicotine seems to reduce a stimulating effect of TNF-α on the gene expression of hBD-2.
Centre for Operative Dentistry and Periodontology, University of Dental Medicine and Oral Health, Danube Private University (DPU), Steiner Landstrasse 124, 3500 Krems, Austria. michael.wolgin@dp-uni.ac.at
Michael
M
Wolgin
Stylianos
S
Liodakis
Axel R
AR
Pries
Andreas
A
Zakrzewicz
Andrej M
AM
Kielbassa
D000704:Analysis of Variance; D002460:Cell Line; D002478:Cells, Cultured; D023082:Defensins / Q000378:metabolism; D020869:Gene Expression Profiling; D024581:Glyceraldehyde-3-Phosphate Dehydrogenase (Phosphorylating) / Q000378:metabolism; D053583:Interleukin-1beta / Q000378:metabolism; D015850:Interleukin-6 / Q000378:metabolism; D015603:Keratinocytes / Q000187:drug effects* / Q000378:metabolism*; D009538:Nicotine / Q000494:pharmacology*; D060888:Real-Time Polymerase Chain Reaction; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*; D023083:beta-Defensins / Q000187:drug effects* / Q000235:genetics*
C406407:DEFB1 protein, human; C106641:DEFB4A protein, human; D023082:Defensins; D053583:Interleukin-1beta; D015850:Interleukin-6; D014409:Tumor Necrosis Factor-alpha; D023083:beta-Defensins; D009538:Nicotine; D024581:Glyceraldehyde-3-Phosphate Dehydrogenase (Phosphorylating)