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Luteolin Induces Apoptosis in Oral Squamous Cancer Cells
S.-F. Yang1,
W.-E. Yang2,
H.-R. Chang3,
S.-C. Chu4 and
Y.-S. Hsieh2,*
1 Institute of Medicine,
2 Institute of Biochemistry and Biotechnology, Chung Shan Medical University, Taichung 402, Taiwan;
3 Division of Nephrology, Department of Internal Medicine, Chung Shan Medical University Hospital, Taichung 402, Taiwan; and
4 Department of Food Science, Central Taiwan University of Science and Technology, Taichung 406, Taiwan
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Figure 1. The effect of luteolin on viability/number and death of SCC-4 cells. The chemical structure of luteolin (3',4',5,7-tetrahydroxyflavone) (A). Cells were treated with luteolin at a concentration of 0, 20, 40, 60, 80, or 100 µM for 24, 48, and 72 hrs. At the end of treatment, cell viability was determined by MTT assay (B), and dead cells were counted by the trypan blue dye exclusion assay (C), as described in MATERIALS & METHODS. Human gingival fibroblasts and OC2 and SCC-4 cells were treated with luteolin and 7-hydroxyflavanone at a concentration of 0, 20, 40, 60, 80, or 100 µM for 24 hrs. After treatment, cell viability was determined by MTT assay (D). The data are means ± SD of 3 independent experiments (*P < 0.05; **P < 0.01; ***P < 0.001).
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Figure 2. Effects of luteolin on cell-cycle progression and protein levels of cell-cycle regulators. Cells were treated with 0, 20, 40, 60, 80, or 100 µM of luteolin for 24 hrs. The cell-cycle distribution was analyzed by flow cytometry (A). For protein levels, cells were subjected to Western blot for analysis of the expression of CDK2, 4, and 6, cyclin D1, D3, and E (B), Cip1/p21, Cip2/p27, and phospho-Rb (C), together with β-actin as an internal control. The data are means SD of 3 independent experiments (*P < 0.05; **P < 0.01; ***P < 0.001).
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Figure 3. Apoptotic effect of luteolin on SCC-4 cells. After a 48-hour luteolin treatment with 0, 20, 40, 60, 80, and 100 µM, DNA condensation and fragmentation were analyzed by DAPI staining (A) and DNA fragmentation assay (B). Meanwhile, treated cell lysates were subjected to SDS-PAGE, followed by Western blotting with antibodies against Bcl-2 and Bax, cleaved forms of caspase 3, 9, and PARP, with β-actin as an internal control (C). The data are means ± SD of 3 independent experiments (*P < 0.05; **P < 0.01; ***P < 0.001).
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Figure 4. The improved effect of luteolin with paclitaxel on SCC-4 cells, and its anticancer effects in vitro and in vivo. (A) Cells were cultured and treated with paclitaxel (0.3 nM) alone or in combination with luteolin (5, 10 µM), and cell viability was determined at the end of Days 4 and 7 by MTT assay. (B) After subcutaneous implantation of SCC-4 cells, BALB/c nu/nu mice received vehicle (DMSO as control; n = 5), luteolin (3 or 5 mg/kg body weight; n = 5), paclitaxel (1 mg/kg body weight; n = 5), or combination treatment with luteolin (3 mg/kg body weight) and paclitaxel (1 mg/kg body weight; n = 5), then were analyzed for tumor growth. The data are means ± SD of 3 independent experiments (*P < 0.05; **P < 0.01; ***P < 0.001).
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Journal of Dental Research, Vol. 87, No. 4,
401-406 (2008)
DOI: 10.1177/154405910808700413
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