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Fig. 7 | Breast Cancer Research

Fig. 7

From: Nuclear basic fibroblast growth factor regulates triple-negative breast cancer chemo-resistance

Fig. 7

Nuclear basic fibroblast growth factor (bFGF) drives DNA repair and DNA-dependent protein kinase catalytic subunit (DNA-PKcs) expression. a Left panel: BT549 cells transfected with bFGF shRNA or control (ctrl) shRNA were challenged with doxorubicin (Dox) (0.25 μg/ml) for 2 h. Fresh medium was added after chemotherapy removal. DNA damage at sequential time points after chemotherapy treatment was analyzed by neutral comet assay. Representative images are shown for each time point. Cells scored as comet tail-positive are indicated with red arrows in the 48-h time frame. Right panel: percentage of cells with comet tails at indicated time points was quantified with a fluorescence microscope. Error bars represent SD, n = 3 fields (each field containing >50 cells). Significance of data points at 24 and 48 h was determined relative to data reported at 0 h for the indicated cell population (*p <0.05, **p <0.01, ***p <0.001, two-tailed Student’s t test). b SUM159 cells expressing control shRNA, bFGF shRNA, or bFGF shRNA plus indicated addback constructs (as in Fig. 4a) were challenged with doxorubicin (0.25 μg/ml) for 2 h. Fresh medium was added after chemotherapy removal. DNA damage at sequential time points after chemotherapy treatment was analyzed by neutral comet assay. Percentage of cells with comet tails at the indicated time points was quantified with a fluorescence microscope. Error bars represent SD, n = 3 fields (each field containing >50 cells). Significance of data points at 24 and 48 h was determined relative to data reported at 0 h for the indicated cell population (*p <0.05, **p <0.01, ***p <0.001, two-tailed Student’s t test). c SUM159 and BT549 cells transfected with bFGF shRNA or ctrl shRNA were treated with doxorubicin as in Fig. 1a. Nuclear protein from chemo-residual cells was extracted. Equivalent amounts were immunoblotted with DNA-PKCS and lamin A antibodies. Protein bands were quantified, and the relative ratio of DNA-PKCS to loading control is shown for each lane. d Left panel: bFGF shRNA-transfected SUM159 cells expressing indicated addback constructs were treated with doxorubicin as in Fig. 1a. Nuclear protein from chemo-residual cells was extracted. Equivalent amounts were immunoblotted with DNA-PKCS and Lamin A antibodies. Right panel: protein bands from three independent trials were quantified and the relative ratio of DNA-PKCS to loading control is shown for each line. Error bars represent SD, n = 3, **p <0.01, two-tailed Student’s t test

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