| Hepatic carcinoma is a high incidence rate of malignant disease in China.The prevention and treatment of cancer has always been the key and difficult point of the solution.It has become a focus topic to find anti-tumor drugs that can find out natural and non-toxic sides effects.Our group have preliminary studied the different methods and optimum conditions for the extraction,separation and purification of anthocyanins from black soybean coats.However,the types of anthocyanins contained in black soybean coats,as well as their related properties and mechanisms,require to be more detailed discussion.In order to further investigate the antioxidant properties of anthocyanins from black soybean coats and the molecular mechanisms of inhibition hepatic carcinoma,the following contents are carried out in this article.1.High performance liguid chromatography detection found that compared with the peak time of a variety of standards,the composition of anthocyanins from black soybean coats contains four components,namely,delphindin-3-O-glucoside,cyanidin-3-O-glucoside,petunidin and cyanidin glucoside.Then the different concentrations of the puried samples BSCA were applied on various cells.The changes of antioxidant enzymes index,MDA indicators and the changes of ROS were detected in the cells,the results found that BSCA could promote the activity of SOD,CAT,GSH-Px,et al,but decreased the level of MDA and ROS.The results showed that anthocyanins from black soybean coats showed good antioxidant properties.2.Cell proliferation was detected by MTT and EDU methods in order that screened one of the most obvious cell lines after anthocyanins from black soybean coats treatment cells.The conclusion is that anthocyanins from blacksoybean coats has the most significant inhibitory effect on heatoma Hep G2 cells,and there is a corresponding time-dose correlation.Then we used different concentrations of anthocyanins from black soybean coats to act on cells,respectively so that we could observe the change of cell morphology and nucleus,observation of confocal laser scanning microscopy showed that cell morphology significantly shrinked,the gap of cells increased and the number of cells decreased,the nucleus collapsed and became crescent-shaped,after cells were treated with BSCA.As well as the results of flow cytometry detection showed that anthocyanins from black soybean coats can induce mitochondrial membrance potential decreased,compared with the untreated cells.3.Hep G2 cells and SMMC7721 cells were treated with different concentrations of anthocyanins from black soybean coats,and the expression of JAK2,STAT3,p-JAK2 and p-STAT3 in the JAK2/STAT3 signaling pathway and their gene expression were detected by Western blot and qRT-PCR.At the same time,the expression of PARP,Caspase-3?BcL-2,Bax,p53 and p63.The results showed that the expression levels of p-JAK2,p-STAT3,Caspase-3 and Bcl-2 were reduced along with the extension of concentration and time,on the contrary,the expression of p53 and p63 were increased.It found that anthocyanins from black soybean coats could promote the expression of pro-apoptotic related proteins,moreover,the expression of anti-apoptosis related proteins is expected to be inhibited in a certain extent.Furthermore,after the shRNA interference was applied to JAK2,the expression of JAK2 protein was significantly down-regulated.The anthocyanins from black soybean coats acted on the shJAK2 cell line,which also inhibited JAK2,the inhibitory effect was more obvious.So the anthocyanins from black soybean coats can cause cell apoptosis by inhibiting the JAK2/STAT3 signaling pathway in hepatoma Hep G2 and SMMC7721 cells.The above results provide a theoretical basis for the development ofdrugs with natural non-toxic side effects,and also provide a new idea for the research of target treatment. |
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