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- Title
Overexpression and Activation of the α9-Nicotinic Receptor During Tumorigenesis in Human Breast Epithelial Cells.
- Authors
Chia-Hwa Lee; Ching-Shui Huang; Ching-Shyang Chen; Shih-Hsin Tu; Ying-Jan Wang; Yu-Jia Chang; Ka-Wai Tam; Po-Li Wei; Tzu-Chun Cheng; Jan-Show Chu; Li-Ching Chen; Chih-Hsiung Wu; Yuan-Soon Ho
- Abstract
Background Large epidemiological cohort studies in the United States have indicated that active and passive smoking are associated with increased breast cancer risk. However, there was no direct evidence of an effect of tobacco carcinogens on the cellular molecules involved in breast tumorigenesis. Methods Reverse transcription--polymerase chain reaction was used to determine the expression of all of the nicotinic acetylcholine receptor (nAChR) subunits in 50 human breast cancer samples and to determine the expression of the α9-nAChR subunit in 276 surgical and laser capture microdissected breast tumor vs normal tissue pairs. Stable MDA-MB-231 breast cancer cell lines were established in which expression of the α9-nAChR subunit was inhibited using short interfering RNA. MCF-10A normal human breast epithelial cells were established in which the α9-nAChR subunit could be conditionally overexpressed by removal of doxycycline from the culture fluid. Cell proliferation and soft agar assays and tumor growth in nude mice were used as measures of cell transformation. All statistical tests were two-sided. Results In 186 (67.3%) of the 276 paired samples, α9-nAChR mRNA was expressed at (mean 7.84-fold) higher levels in breast cancers than in surrounding normal tissue. Stable expression of α9-nAChR short interfering RNA in MDA-MB-231 cells attenuated nicotine-stimulated proliferation and growth in soft agar and reduced tumor volume when the cells were introduced as xenografts in SCID mice (n = 5 mice per group; mean tumor volume at 6 weeks treatment in mice injected with Si α9 cells = 995.6 mm³, in mice injected with parental cells = 2993.2 mm³, difference = 1997.6 mm³, 95% confidence interval [CI] = 1705 to 2290.2 mm³, P = .009). Long-term treatment of MCF-10A normal breast epithelial cells with either nicotine or its active metabolite, 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone, triggered precancerous transformation as defined by soft agar assay. Inducible overexpression of α9-nAChR in MCF-10A cell xenografts in nude mice substantially increased tumor growth (n = 5 mice per group; DOX+, mean tumor volume without nicotine vs with nicotine = 266.2 vs 501.6 mm³, difference = 235.4 mm³, 95% CI = 112.7 to 358 mm³, P = .009; DOX-, mean tumor volume without nicotine vs with nicotine = 621.2 vs 898.6 mm³, difference = 277.4 mm³, 95% CI = 98.1 to 456.7 mm³, P = .016; mean tumor volume in the presence of nicotine, DOX+ vs DOX- = 501.6 vs 898.6 mm³, difference = 397 mm³, 95% CI = 241.3 to 552.6 mm³, P = .009). Conclusion The α9-nAChR is important for nicotine-induced transformation of normal human breast epithelial cells.
- Publication
JNCI: Journal of the National Cancer Institute, 2010, Vol 102, Issue 17, p1322
- ISSN
0027-8874
- Publication type
Academic Journal
- DOI
10.1093/jnci/djq300