ABSTRACT
Objective: To investigate the antitumour activity of ginsenoside Rh2 against human leukemia KG1α cells through apoptosis and autophagy pathway. Methods: CCK-8 assay was used to screen the most effective ingredient on the proliferations among ginsenoside Rh2 in leukemia KG1α cell line; FCM detected cell apoptosis; Hoechst staining observed the cell morphological changes of apoptosis; Acridine staining detected Rh2 effected on autophagy; Western blotting and RT-PCR detected the expression levels of the proteins closely associated with autophagy and apoptosis. After joining autophagy inhibitors, using CCK-8 to test the proliferation activity of cells, cell apoptosis was measured by FCM. Results: CCK-8 indicated that Rh2 could inhibit the proliferation of KG1α cells significantly with dose- and time-dependent manners; FCM indicated that Rh2 induced apoptosis; Hoechest staining showed that KG1α cells had typical apoptotic morphological changes by treated Rh2; Acridine staining revealed that Rh2 cause increase in the number of acidic autophagy vesicles in cells, causing cell autophagy levels increased; Western blotting and RT-PCR results showed that Rh2 increased the expression of Beclin-1, LC3A, and LC3B, activated Caspase-3 and Bax/Bcl-2 rates, and MAPK, ATK, and ERK signaling pathway; After using autophagy inhibitors (3-MA), autophagy crippled that Rh2 inhibited the proliferation and induced apoptosis in KG1α cells. Conclusion: Ginsenoside Rh2 could significantly enhance autophagy through activated MAPK, ATK, and ERK signaling pathway, and then inhibit the proliferation and induce apoptosis in KG1α cells.
ABSTRACT
AIM: To investigate the autophagy of breast cancer cells induced by baicalein and to explore its mechanism.METHODS: The effects of baicalein on the viability of MCF-7 cells and 4T1 cells were investigated by MTT assay, and the dosage of the drug was determined.The expression levels of microtubule-associated protein 1 light chain 3-II (LC3-II) and LC3-I in the MCF-7 cells and 4T1 cells treated with baicalein at doses of 25, 50 and 100 μmol/L, or combined with autophagy inhibitor 3-methyladenine (3-MA) were determined by Western blot.In order to confirm the role of baicalein in autophagy, the effect of 3-MA on the apoptosis of both MCF-7 cells and 4T1 cells induced by baicalein was analyzed by flow cytometry.The protein levels of p-mTOR, mTOR, p-AKT and AKT were examined by Western blot and the role of AKT-mTOR pathway in the induction of autophagy in breast cancer induced by baicalein was determined by the combination of activators.RESULTS: Baicalein at 50 μmol/L and above doses significantly inhibited the viability of breast cancer cells in a dose-and time-dependent manner.The expression of LC3-II/LC3-I in both MCF-7 cells and 4T1 cells was significantly enhanced after the action of baicalein, and the ratio of LC3-II/LC3-I was significantly decreased after 3-MA addition.The results of flow cytometry showed that, compared with baicalein group, the combination of baicalein and 3-MA promoted the levels of necrosis and apoptosis.Moreover, the protein levels of p-mTOR and p-AKT were significantly decreased and were rescued by EGF, while their total protein levels were not changed.CONCLUSION: Baicalein induces autophagy through AKT-mTOR pathway both in MCF-7 cells and 4T1 cells.