Journal: Oncology Reports
Article Title: Reactive oxygen species induced by icaritin promote DNA strand breaks and apoptosis in human cervical cancer cells
doi: 10.3892/or.2018.6864
Figure Lengend Snippet: Western blot analysis. HeLa and SiHa cells with or without pre-incubation (1 h) with 5 µM NAC were treated by icaritin for 24 h. Multiple SDS-PAGE gels were prepared for each sample, and each gel was used for the detection of one protein. Thus, one loading control, β-actin, was used for all proteins of each sample. (A) Detection of MMP-9. (B) Detection of cell cycle-associated proteins. (C) Detection of apoptosis-associated proteins. (D) Quantification of apoptosis-associated proteins in the 0 µM, 2× IC 50 and 2× IC 50 + NAC groups. Relative protein levels were quantified from western blots using ImageJ and presented as ratios of each protein band relative to the loading control. (*P<0.05, **P<0.01, ***P<0.001 vs. 0 µM). NAC, N-acetyl cysteine, MMP-9, matrix metalloproteinase-9; CDK1, cyclin-dependent kinase 1; CDC25C, cell division cycle 25C; XIAP, X-linked inhibitor of apoptosis protein; Bax, apoptosis regulator Bax; Bcl-2, B-cell lymphoma-2.
Article Snippet: The membranes were blocked in Tris-buffered saline with Tween-20 [10 mM Tris (pH 7.5), 100 mM NaCl, 0.1% Tween-20; PBST) containing 5% (w/v) nonfat milk for 2 h at room temperature, followed by incubation with primary antibodies overnight at 4°C [B-cell lymphoma 2 (Bcl-2; 1:500; cat. no. sc-7382) and apoptosis regulator Bax (Bax; 1:500; cat. no. sc-7480) from Santa Cruz Biotechnology, Inc.; cleaved caspase 3 (1:500; cat. no. ab2302), cleaved caspase 9 (1:200; cat. no. ab2324) and X-linked inhibitor of apoptosis protein (XIAP; 1:1,000; cat. no. ab28151) from Abcam (Cambridge, UK); cyclin-dependent kinase 1 (CDK1)-pT14 (1:1,000; cat. no. bs-3091R), cell division cycle 25C (CDC25C)-pS216 (1:1,000; cat. no. bs-3096R), β-actin (1:5,000; cat. no. bs-0061R) and matrix metalloproteinase 9 (MMP9; 1:1,000; cat. no. bs-4593R) from BIOSS].
Techniques: Western Blot, Incubation, SDS Page
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