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Seno et al. Cancer Drug Resist 2019;2:335-50 I http://dx.doi.org/10.20517/cdr.2019.01 Page 345

A B

C

D E

2+
2+
Figure 4. Potential involvement of Ca /Mg -dependent endonuclease in DNA fragmentation. A: We observed no obvious reduction
in full-length ICAD levels during apoptosis of miPS-LLCcm. miPS-LLCcm cells were treated with 100 nmol/L of daunorubicin for the
indicated periods and analyzed by western blotting analyses; B: detection of processed caspases in attached or floating cells; C: apoptotic
cells in the presence of BAPTA-AM were detected by annexin V staining. Cells were pretreated with 10 μmol/L of BAPTA-AM for 1 h,
2+
treated with 100 nmol/L daunorubicin, and analyzed by FACS; D: Ca chelator did not affect caspase levels during apoptosis of miPS-
2+
LLCcm cells; E: suppression of DNA fragmentation by treatment with a Ca chelator. DNA fragmentation in the cells treated with
daunorubicin in the presence of 10 μmol/L of BAPTA-AM was analyzed
glioblastoma multiform . Considering the significant role of the CSC niche, in vitro screening of cancer
[5]
drugs should be performed under the conditions that mimic the CSC niche. Our miPS-CSCs have been
confirmed to exhibit the potential to create an endothelial niche-like environment in vitro [10,11] , in which
[11]
soluble factors, including Notch ligands, were found to promote self-renewal and differentiation of CSCs .
Thus, we performed the experiments using purified miPS-CSCs in the presence of CM obtained from bulk
miPS-CSCs.