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Quantitative Analysis of Autophagy using Advanced 3D Fluorescence Microscopy
Published on: May 3, 2013
Icariin Induces Autophagy-dependent Apoptosis in FaDu Head and Neck Squamous Cell Carcinoma
Kyeong-Rok Kang1, DO Kyung Kim1, Hyangi Lim1
1Department of Oral Physiology, School of Dentistry, Chosun University, Gwangju, Republic of Korea.
Background/Aim:
Icariin (ICA) is a well-known prenylated flavonoid isolated from Epimedium species and has shown potential antitumor effects in multiple malignancies. However, its effects on human head and neck squamous cell carcinoma (HNSCC) have not yet been elucidated. Therefore, the aim of the present study was to evaluate the antitumor effects of ICA in HNSCC and to explore the associated signaling pathways.
Materials And Methods:
FaDu cells were treated with ICA, and its anticancer effects were evaluated using cell viability, apoptosis, autophagy, and protein expression analyses, including MTT assay, flow cytometry, western blotting, caspase activity assay, and immunofluorescence staining. In addition, the antitumor efficacy of ICA was investigated in a FaDu xenograft mouse model through tumor growth assessment and immunohistochemical analysis.
Results:
ICA reduced the viability of FaDu cells and induced marked morphological alterations, including nuclear condensation. It increased the proportion of apoptotic FaDu cells, as determined by flow cytometry. In addition, western blot analysis showed increased cleaved caspase-3 and poly(ADP-ribose) polymerase (PARP), mediated through both intrinsic and extrinsic apoptotic pathways. Furthermore, ICA induced autophagosome formation and modulated the expression of autophagy-related biomarkers, characterized by the upregulation of beclin-1 and LC-3 and the downregulation of p62 in association with suppression of Akt/mTOR signaling pathway. Moreover, pharmacological inhibition of autophagy using 3-methyladenine significantly attenuated ICA-induced apoptosis in FaDu cells. Finally, oral administration of ICA (60 mg/kg/day) suppressed tumor growth in animals xenografted with FaDu cells.
Conclusion:
ICA exerted significant antitumor effects in FaDu cells by inducing autophagy-mediated apoptosis and was associated with inhibition of Akt/mTOR signaling pathway. These results suggest that ICA may serve as a promising therapeutic candidate for HNSCC.
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