Effect Of Icariin On Wnt/?-catenin Pathway In Apoptosis Of Multiple Myeloma Cells

ObjectiveTo observe the effect of icariin on the proliferation and apoptosis of multiple myeloma cell U266,and to explore the relationship between its mechanism and Wnt/?-catenin pathway.MethodsThe multiple myeloma U266 cells were divided into different concentrations of the icariin group,and the inhibition rate of different concentrations of icariin was detected by CCK-8 to explore the optimal concentration.The multiple myeloma U266 cells were divided into control group,lenalidomide group,optimal concentration of icariin group and lenalidomide+optimal concentration of icariin group.The cell proliferation was detected by CCK-8.The Annexin-V/PI double staining was used to detect early apoptosis and qRT-PCR was used to detect the expression of Wnt pathway regulatory molecules?-catenin,GSK3 p and downstream target genes c-Myc and cyclin D1 mRNA in each group.The ?-catenin and GSK3? proteins were detected by Western blot.Resu ts1.CCK-8 detects cell proliferation resultsCompared with the control group,there was no statistical difference in the otyonin OD values between the 24h and 72h concent rations.There was a statistical difference between the 48h 32 u mol/L concentration group and the control group,P<0.05.Compared with the control group,the lenalidomide group and the icariin group had no statistically significant inhibition on the proliferation of U266 cells,but there was a statistical difference between the icariin+lenalidomide group,P<0.05.There was no statistically significant difference between the icariin group and the lenalidomide group.2.Annexin-V/PI double staining detection of early apoptosis resultsThe effect of icariin on the early apoptosis of U266 cells was not obvious in the middle and low concentrations(2?mol/L,4? mol/L,8 u mol/L).After high concentration(16 ? mol/L,32 ? mol/L)treatment with icariin,the early apoptosis rate of U266 cells increased.The early apoptotic rate of the fourth quadrant of the control group,the icariin group,the lenalidomide group,the icariaxin+ lenalidomide group was not obvious,but the rate of apoptosis and necrotic cells in the second quadrant increase.3.Detection of mRNA expression in each group by qRT-PCRRelative expression of ?-catenin mRNA:There was no statistical difference in the lenalidomide group compared with the control group;the icariin group was statistically different,P<0.05;the icariin+lenalidomide group Statistical difference,P<0.05.There was a statistically significant difference between the lenalidomide group and the icariin group,P<0.05.There was a statistically significant difference between the icariin+lenalidomide group and the lenalidomide group,P<0.05;there was no statistical difference compared with the icariin group.Relative expression of GSK3? mRNA:Compared with the control group,there was no statistical difference between the lenalidomide group and the icariin group.The icariin+lenalidomide group had statistical difference,P<0.05.There was no statistical difference between the lenalidomide group and the icariin group.There was a statistically significant difference between the icariin+lenalidomide group and the lenalidomide group,P<0.05;there was no statistical difference compared with the icariin group.Relative expression of c-Myc mRNA:Compared with the control group,there was no statistical difference between the lenalidomide group and the icariin group;the icariin+lenalidomide group had statistical difference,P<0.05.There was no statistical difference between the lenalidomide group and the icariin group.There was a statistically significant difference between the icariin+ lenalidomide group and the lenalidomide group and the icariin group,P<0.05.Relative expression of cyclin D1 mRNA:There was no statistical difference in the lenalidomide group compared with the control group;the icariin group was statistically different,P<0.05.There was a statistically significant difference between the icariin+lenalidomide group,P<0.05.There was a statistically significant difference between the lenalidomide group and the icariin group,P<0.05.There was a statistically significant difference between the icariin+ lenalidomide group and the lenalidomide group,P<0.05;there was no statistical difference compared with the icariin group.4.Detection of the expression of ?-catenin and GSK3 ? protein in each group by Western blot?-catenin protein expression:Compared with the control group,there was no statistical difference in the lenalidomide group;the icariin group,the icariin+lenalidomide group had statistical difference,P<0.05.There was a statistically significant difference in protein expression between the icariin+lenalidomide group and the lenalidomide group,P<0.05.GSK3 ? protein expression:Compared with the control group,the difference between the lenalidomide group and the icariin group was statistically significant,and the expression of icariin+lenalidomide was statistically different,P<0.05.There was a statistically significant difference in the expression of histones between lenalidomide and icariin+lenalidomide,P<0.05.ConclusionIcariin can down-regulate the gene and protein expression of ?-catenin and GSK3 ? related to Wnt pathway,and down-regulate the gene expression of c-Myc and cyclin D1 downstream of Wnt.Icariin can inhibit the proliferation of multiple myeloma cells,promote apoptosis of multiple myeloma cells,and increase the sensitivity of lenalidomide chemotherapy.