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Mounting evidence has shown that side population (SP) cells are enriched for cancer stem cells (CSCs) responsible for cancer malignancy.In this study,SP technology was used to isolate a small subpopulation of SP cells in human gallbladder cancer cell line GBC-SD,and SP cells which had superior potential for proliferation in vitro and tumorigenesis in vivo were identified.Importantly,the abundance of GBC-SD SP cells was increased by a transforming growth factor-β (TGF-β)-induced epithelial-mesenchymal transition (EMT),and this effect was accompanied with a strong up-regulation of ABCG2 mRNA expression,and a decreased sensitivity to mitoxantrone.SP cells were restored upon the removal of TGF-β and the reversion of the cells to an epithelial phenotype,and smad3-specific siRNA reduced SP abundance in response to TGF-β.In conclusion,TGF-β-induced EMT by smad-dependent signaling pathway promotes cancer development and anti-cancer drug resistant phenotype by augmenting the abundance of GBC-SD SP cells,and a better understanding of mechanisms involved in TGF-β-induced EMT may provide a novel strategy for preventing cancer progression.