Figure 1.

Doxorubicin (DXR) induces senescence in human breast cancer cells. A, Three breast cancer cell lines were cultured with the indicated doses of DXR (nmol/L) for 72 h. Medium alone (background) was subtracted. In these experiments, cell viability (%) was determined using the WST‐8 assay. The results are shown as the means of 3 wells. B, Three breast cancer cell lines were cultured with DXR (250 nmol/L for MDA‐MB‐231, 100 nmol/L for BT‐549, and 200 nmol/L for MCF‐7) for 48 h. Using the tumor lysates, immunoblotting analysis was carried out using anti‐γH2AX Ab. β‐Actin was used as a control. C, Similarly, 3 breast cancer cell lines were cultured with DXR for 48 h. Immunoblotting analysis was undertaken using anti‐p21 and anti‐p16 Abs. β‐Actin was used as a control. D, To examine the expression of senescence‐associated β‐Gal, cancer cells were treated with DXR (250 nmol/L for MDA‐MB‐231, 100 nmol/L for BT‐549, and 200 nmol/L for MCF‐7) for 2 d and stained with SPiDER β‐Gal. Confocal imaging was carried out on untreated or DXR‐treated cancer cells. Scale bar = 10 μm. E, Similarly, 3 cancer cell lines were treated with or without DXR for 2 d. After harvesting, cancer cells were cultured without DXR for 2 d. Thereafter, the levels of interleukin (IL)‐6 and IL‐8 in the supernatants were examined by ELISA. **P < .01, ***P < .005