Involvement of nicotinic acetylcholine receptor in the proliferation of mouse induced pluripotent stem cells

Aims: As the clinical use of induced pluripotent stem (iPS) cells may have the potential to overcome current obstacles in stem cell-based therapy, the molecular mechanisms that regulate the proliferation of iPS cells are of great interest. However, to our knowledge, no previous studies have examined whether stimulation with nicotinic acetylcholine receptor (nAchR) enhances the growth of iPS cells. In the present study, we examined the involvement of nAchR in the proliferation of mouse iPS cells. Main methods: We performed immunofluorescence staining to determine whether mouse iPS cells could express nAchRs. Mouse iPS cells were treated with nicotine for 24 h under feeder-free conditions in the presence of leukemia inhibitory factor (LIF). The DNA synthesis was examined by the BrdU incorporation assay. Intracellular calcium levels were measured using Fluo-4-acetoxymethyl (a cell-permeable calcium indicator). In addition, we examined the involvement of the CaMK Pi pathway in nicotine-enhanced proliferation of mouse iPS cells. Key findings: The fluorescence images revealed that alpha(4)-nAchR and alpha(7)-nAchR are expressed on mouse iPS cells. Treatment of the cells with 300 nM nicotine significantly increases DNA synthesis. This is significantly inhibited by pretreatment with antagonists of alpha(4)-nAchR and alpha(7)-nAchR or a CaMK Pi inhibitor. In addition, treatment with nicotine increases the intracellular Ca2+ level dose-dependently in mouse iPS cells. Treatment with nicotine significantly enhances CaMK Pi phosphorylation. Significance: The present study indicates that stimulation of alpha(4)-nAchR and alpha(7)-nAchR may lead to a significant increase in the rate of mouse iPS cell proliferation through enhancement of the CaMK Pi signaling pathway. (c) 2012 Elsevier Inc. All rights reserved.