Tanshinone Ⅰ Inhibits Cancer Cell Growth By Inducing Apoptosis And Autophagy In Vitro

Salvia miltiorrhiza is a traditional Chinese medicine commonly used for promoting blood circulation and removing blood stasis.With the wide application of Salvia miltiorrhiza in cancer treatment,its anti-tumor mechanism has become the focus of modern pharmacological research.Tanshinone I,a kind of lipophilic phenanthroquinone compound extracted from Salvia miltiorrhiza Bunge,is a potent anti-inflammatory,antioxidant,and anti-cancer agent.Our previous research of Salvia miltiorrhiza in another monomer cryptotanshinone(CPT)showed that tanshinone I inhibited the proliferation of breast cancer cells,but the anti-tumor molecular mechanism remains unclear.In the present study,we further explore the anti-tumor effect of tanshinone I and its related molecular mechanism in breast cancer cell lines and liver cancer cell lines,which provides a scientific basis for the extensive application of Salvia miltiorrhiza in clinical treatment of tumor.Objective:To study the anti-tumor effect of tanshinone I and the related molecular mechanism,which provides the theoretical basis for the clinical treatment of tumor.Methods:In this study,three breast cancer cell lines of MCF-7(ER+),MDA-MB-231(ER-)and one liver cancer cell line of HepG2 were used as the research subject.The inhibitory effect of tanshinone I on tumor cells proliferation was detected by MTT.Morphological changes of tumor cells treated with tanshinone I were observed under the microscope.Flow cytometry and western blotting were used to detect the effects of tanshinone I on apoptosis and cycle of tumor cells.The effect of tanshinone I on autophagy of tumor cells was observed by western blotting,fluorescence microscope with acridine orange and transmission electron microscope.Results:We found that μM of tanshinone I significantly inhibited the proliferation of the four tumor cells at 24 h treatment(p<0.05).Moreover,tumor cells shrunk or rounded off and the density decreased significantly after treatment with tanshinone I for 24 h and 48 h.Flow cytometry indicated that tanshinone I induced the apoptosis of tumor cells at 48 h(p<0.05),but no obvious effect was found in the cycle distribution(p>0.05).The accumulation of apoptosis related protein c-Parp(89KD)and autophagy-related protein LC3 Δ were detected by western blotting.Meanwhile,orange-red fluorescence enhancement with acridine orange staining and the formation of autophagic vacuoles were observed under the fluorescence microscope and electron microscope,respectively.MTT showed that the inhibitory effect of tanshinone I on tumor cell growth was attenuated(p<0.05)when tanshinone I was combined with autophagy inhibitor 3-methyladenine(3MA)/chloroquine(CQ).These results suggest that tanshinone I induced autophagic death in tumor cells.Western blotting was used to further explore the molecular mechanism of autophagy induced by tanshinone I in tumor cells.The expression of p-AMPKawas significantly increased after exposure of tumor cells to tanshinone I,suggesting that tanshinone I activated AMPK signaling.Furthermore,tanshinone I induction of autophagy-related protein LC3 Δ and Beclin 1 was markedly attenuated by inclusion of AMPK inhibitor Dorsomorphin 2HCl.These results indicated that tanshinone I induced autophagy of tumor cells by activating AMPK signaling pathway.Conclusion:Tanshinone I can inhibit the proliferation of tumor cells such as breast cancer and liver cancer by inducing apoptosis and autophagy.Tanshinone I can induce autophagy by activating AMPK.