Abstract
1/2013
vol. 17
Original paper
DNA damage induced by oridonin involves cell cycle arrest at G2/M phase in human MCF-7 cells
Wspolczesna Onkol 2013; 17 (1): 38–44
Online publish date: 2013/03/15
Aim of the study: To study the mechanisms of inhibition of cell growth and induction of DNA damage in oridonin-treated MCF-7 human breast cancer cells.
Material and methods: The cytotoxicity of oridonin-treated MCF-7 cells was measured by MTT assay. Cell cycle phase distribution was analyzed by flow cytometry. P-ATM, P-CHK2, gH2AX and P-P53 protein expressions were detected by Western blot analysis. The expression of r-h2ax and P-ATM was also detected by immunofluorescence staining. The degree of cellular damage of oridonin-induced MCF-7 human breast cancer cells was confirmed by the comet assay analysis of DNA fragmentation.
Results: Oridonin inhibited cell growth in a time- and dose-dependent manner. The IC50 values at 48 and 72 hours were 78.3 and 31.62 µmol/l, respectively. Oridonin induced G2/M phase arrest in MCF-7 cells. MCF-7 cells treated with oridonin showed significant DNA damage as shown by an increase in olive tail moment (OTM). The protein expression levels of P-ATM, P-CHK2, gH2AX and P-P53 were increased significantly in a dose-dependent manner.
Conclusions: DNA damage provokes p53-mediated G2/M cell cycle arrest in oridonin-induced MCF-7 cells through the mechanism of CHK2 activation by activated ATM protein kinase, which is induced by oridonin.
Material and methods: The cytotoxicity of oridonin-treated MCF-7 cells was measured by MTT assay. Cell cycle phase distribution was analyzed by flow cytometry. P-ATM, P-CHK2, gH2AX and P-P53 protein expressions were detected by Western blot analysis. The expression of r-h2ax and P-ATM was also detected by immunofluorescence staining. The degree of cellular damage of oridonin-induced MCF-7 human breast cancer cells was confirmed by the comet assay analysis of DNA fragmentation.
Results: Oridonin inhibited cell growth in a time- and dose-dependent manner. The IC50 values at 48 and 72 hours were 78.3 and 31.62 µmol/l, respectively. Oridonin induced G2/M phase arrest in MCF-7 cells. MCF-7 cells treated with oridonin showed significant DNA damage as shown by an increase in olive tail moment (OTM). The protein expression levels of P-ATM, P-CHK2, gH2AX and P-P53 were increased significantly in a dose-dependent manner.
Conclusions: DNA damage provokes p53-mediated G2/M cell cycle arrest in oridonin-induced MCF-7 cells through the mechanism of CHK2 activation by activated ATM protein kinase, which is induced by oridonin.
Keywords
oridonin, ATM, CHK2, cell cycle, p53, apoptosis
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