| Objective: To investigate the effects of huaier aqueous extract on the morphology,proliferation,invasion,cell cycle and autophagy of triple negative breast cancer cell lines MDA-MB-231 and BT-549?Methods:1.MDA-MB-231 and BT-549 triple-negative breast cancer cell lines were cultured in vitro.Observed the morphological changes of breast cancer cell treated with different concentrations of huaier aqueous extract(4mg/ml,8mg/ml)for 48 hours under light microscope.2.Observe the effects of different concentrations(2mg/ml,4mg/ml,8mg/ml,16mg/ml)of huaier aqueous extract for different time(24h,48h)on the proliferation of triple negative breast cancer cells by CCK-8 experiment.3.Observe the effect of huaier aqueous extract on invasion and migration of triple negative breast cancer cells by transwell experiment.4.Detect the cell cycle of triple negative breast cancer cells under the effects of huaier aqueous extract at different concentrations.5.Observe the proliferation of triple negative breast cancer cells under the combination of huaier aqueous extract and autophagy inhibitor by CCK-8 experiment.6.Western blot was used to observe the LC-3II/LC-3I expression ratio in triple negative breast cancer cells under different concentrations of huaier aqueous extract.Results:1.Huaier aqueous extract could promote the apoptosis and necrosis of MDA-MB-231 and BT-549 cell lines,inhibited the proliferation of two cell lines,and had a dose-time effect relationship.The IC50 of MDA-MB-231 cells at 24 h and 48 h were 8.92mg/ml and 5.52mg/ml,and of BT-549 cells at 24 h and 48 h were 8.41mg/ml and 4.75mg/ml.Inhibition of different times between the same cells statistically different,there was no significant difference between the two cell lines.2.In the Transwell experiment,we observed that a large number of MDA-MB-231 and BT-549 cells that have been cultured for 24 hours could penetrate the membrane of the chamber with or without Matrigel gel.But the cells cultured by 4mg/ml huaier aqueous extract could hardly penetrate the membrane.3.Cell cycle was detected after the cells cultured 24 h and 48 h by 4mg/ml and 8mg/ml huaier aqueous extract.The proportion of G0/G1 phase of MDA-MB-231 cells showed a downward trend,and the S phase showed an upward trend,but there was no statistical difference overall.The proportion of G0/G1 phase in BT-549 cells showed a decreasing trend and an increase in S phase.Among the experimental groups treated with 8mg/ml huaier aqueous extract for 24 h and 48 h,the proportion of S phase reached 34% and 33%.Compared with 15% of the control group,there is a significant difference.4.MDA-MB-231 and BT-549 cells were cultured with autophagy inhibitors CQ and 3-MA for 48 hours,and the absorbance calculated by the CCK-8 was not significantly different from that of the control group.4mg/ml huaier aqueous extract combined with CQ and 3-MA cultured breast cancer cells for 48 h as the experimental group.Compared with the control group using huaier aqueous extract only,the absorbance of the experimental group with autophagy inhibitor increased slightly,but there was no statistical difference.5.Western Blot was used to detect the LC-3II/LC-3I expression ratio in normal cultured MDA-MB-231 and BT-549 cells and two cells cultured with 4 mg/ml huaier aqueous extract.The LC-3II/LC-3I value of the MDA-MB-231 cells control group was 1.63 and the experimental group was 2.0;the BT-549 cells control group was 1.09 and the experimental group was 1.61.Difference was statistically significant.Conclusions:1.Huaier aqueous extract can inhibit the proliferation,invasion and migration of MDA-MB-231 cells and BT-549 cells.Proliferation inhibition has a time-dose effect relationship.2.Huaier aqueous extract can block the cell cycle of BT-549 cells in S phase,and there is no statistical difference in the blocking effect of the MDA-MB-231 cell cycle.3.The inhibitory effect of huaier aqueous extract on MDA-MB-231 cells and BT-549 cells may be related to enhancing cell autophagy. |
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