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Thus, the cell clone, 9F-11A, showing enhanced formation of the cell-in-cell structures, was generated from the HCT116 cell lines by the chemical mutagenesis method. On the cell-attachment dishes, the 9F-11A cells often showed different cell shapes as compared to the parent cells (Fig.?3A). Some 9F-11A cells extended slender http://www.selleck.cn/products/MLN8237.html pseudopodia, the lengths of which were sometimes twice to three times the sizes of the cell bodies themselves, whereas the parent cells rarely showed these features. As the altered regulation of actin polymerization was expected in the 9F-11A cells, the cell-in-cell assay using cytochalasin B (CytB), which is an inhibitor of actin polymerization and has been shown to be involved in the process of formation of the cell-in-cell structures (Lugini et?al. 2006; Takeuchi et?al. 2010), was carried out. The rate of the cell-in-cell structures was significantly lower in the CytB-treated cells than in the cells without the treatment of CytB (Fig.?3B; http://www.selleckchem.com/products/CP-690550.html Student's t-test, P?=?0.002 in the samples of 0.2 and 0.8?��g/mL of CytB). This result shows that the actin polymerization pathway is involved in the process of formation of the cell-in-cell structures in the 9F-11A cells. In contrast, similar to the case for the parent cell line, formation of the cell-in-cell structures in 9F-11A seemed to be suppressed on adhesive-coated dishes. Then, we examined the culture condition that would prevent the formation of cell-in-cell structures in 9F-11A, because the presence of cell-in-cell structures at a high frequency can cause http://www.selleckchem.com/products/XL184.html genomic instability of the host cells, and we needed to subculture the 9F-11A cells under a little stimulation for the formation of cell-in-cell structures as possible. As expected, on the adhesive-coated dishes, the number of cell-in-cell structures in the 9F-11A cells decreased to less than one-fourth of that in the suspension culture condition (Fig.?3C; Student's t-test, P?