HBD-1 and hBD-2 expression in HaCaT keratinocytes stimulated with nicotine

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.

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.

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).

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.