Effect Of Cinobufacin Injection On Glioma U87 Cells

Objective:To study the effect of Cinobufacin injection on proliferation,apoptosis and migration of glioma U87 cells by cell experiment.Research methods:1 Toxicity test of Cinobufacin injection on glioma cells: after the glioma U87 cells are cultured,different concentrations of Cinobufacin injection are added to the culture medium,and then cultured at constant temperature for 24 hours.Add 10 ul CCK-8 solution into the culture medium,continue the culture,after the culture,use the enzyme scale to detect the absorbance value at 450 nm.2 Effects of Cinobufacin injection on the proliferation and apoptosis of glioma cells: take the glioma U87 cells in logarithmic growth period,culture them in constant temperature incubator for 24 hours,then divide them into control group,400ug/ml group and 800ug/ml group.(1)Western blot was used to detect apoptosis related proteins: Western blot was used to detect Caspase-3,Caspase-9,Bax and bcl-2.(2)Flow cytometry was used to detect the apoptosis rate: after the cells were cultured and treated,0.25% trypsin without EDTA was used to digest,collect cells and add reagents.After the reaction,the apoptosis rate was detected by flow cytometry.(3)The growth state of cells in each group was observed under the inverted microscope: after the treatment,the difference of cell morphology and quantity was observed under the inverted microscope.(4)RT-PCR detection of apoptosis related gene expression:RT-PCR detection of Caspase-3,caspase-9,Bax,bcl-2 gene expression.Results:1 CCK-8 was used to detect the toxicity of Cinobufacin to glioma cells.It was found that the effect of Cinobufacin was obvious when the concentration of Cinobufacin was more than 400 ug / ml.2.RT-PCR: after 24 hours of exposure to Cinobufacin solution,the m RNA levels of Bax,caspase-3 and caspase-9 in glioma cells were significantly increased(* * P <0.01),and bcl-2 m RNA was significantly decreased(* * P < 0.01).Flow cytometry was used to detect the apoptosis rate of glioma cells: in the control group,400 ug / ml group and 800 ug / ml group,the apoptosis rate was significantly different(P < 0.05).Western blot showed that the protein content of Caspase-3,caspase-9 and Bax in the control group,400 ug / ml group and 800 ug / ml group increased in turn,and the ratio of Bax to Bcl-2 increased in turn.Under the microscope,observe the effect of Cinobufacin injection on glioma cells: control group,400 ug / ml group,800 ug / ml group,the number of cells decreased in turn,the difference between the control group and 400 ug / ml group was the largest.Conclusion:Cinobufacin injection can obviously inhibit the proliferation of glioma U87 cells,increase the expression of apoptosis gene and promote the apoptosis of glioma U87 cells.