The Effects Of Ginsenoside Rg3 On The Prolifertation,apoptosis,migration And Invasion Of Osteosarcoma Cells And Its Potential Mechanisims

BackgroundsOsteosarcoma(OS)is a bone malignant tumor that originates from transformed progenitor cells with osteogenic differentiation and osteogenesis.Although it is classified as a rare tumor,most patients diagnosed with osteosarcoma are adolescents in their twenties and their 5-year survival rate is not ideal.As one of the oldest Chinese herbal medicines in China,ginseng has been widely used in China and Asia,and ginsenoside Rg3 extracted from ginseng is the most active ingredient.In 2002,Rg3 was approved for small cell lung cancer by China Food and Drug Administration(CFDA).Rg3 owns many anti-cancer activities,such as induce apoptosis,hamper metastasis and angiogenesis.However,the effect of Rg3 and its underlying mechanisms in osteosarcoma are still unknown.ObjectivesTo investigate the effect of ginsenoside Rg3 on the proliferation,apoptosis,migration,invasion in osteosarcoma cells and its underlyingmechanisms.Methods1.MTT assay and crystal violet staining were performed to detect the effect of different concentrations of Rg3 on cell viability in osteosarcoma cells at 24,48,and 72 h.Then,treatment of osteosarcoma cells with appropriate Rg3 concentrations and time period were performed in the following experiments.Colony formation assay was devoted to detect the proliferation of osteosarcoma cells after treated with Rg3.The effect of apoptosis in osteosarcoma cells,which treated with Rg3,was examined by flow cytometer and Hochest 33258 staining assay.The effect of invasion and migration of 143 B and MG63 cells with Rg3 treatment were detected by Trasnwell assay and wound-healing assay,respectively.2.The proteins PCNA,cleaved-Caspase 3,Bax,Bcl-2 were detected by Western blot.RT-qPCR and Western blot were applied to measure the expression of migration,invasion-related proteins in osteosarcoma cells.The signaling pathway,which take part in the regulation of Rg3 was screened by luciferase reporter assay.Then the expression levels of ?-catenin,CyclinD1,c-Myc were further verified by Western blot.Results1.After treated with Rg3,the ability of proliferation of osteosarcoma cells were hampered in dose-and time-dependent manners.And the ability of colony formation also decreased after Rg3 treatment.Flow cytometry andHochest 33258 staining both showed that Rg3 could induce apoptosis.In addition,the greater the concentration of Rg3,the stronger the ability to induce apoptosis.The inhibition of invasion and migration in 143 B and MG63 cells were confirmed by Transwell assay and wound healing,respectively.2.Compared with the control group,Rg3 inhibited the expression of proliferating cell nuclear antigen(PCNA)and Bcl-2,improved the expression of Bax and cleaved-Caspase 3.In addition,the expression levels of MMP2,MMP7,MMP9 and EMT-related proteins were also decreased.Luciferase reporter assay indicated that Rg3 may inhibit the activity of Wnt/?-Catenin signaling pathway.Western blot was further confirmed this inhibition ability,that is Rg3 decreased the expression of ?-catenin,CyclinD1 and c-Myc.Conclusions1.Ginsenoside Rg3 inhibits the ability of proliferation,migration,invasion and induces apoptosis of osteosarcoma cells.2.Ginsenoside Rg3 hampers the ability of proliferation of osteosarcoma cells by regulating the expression of PCNA.3.Ginsenoside Rg3 improves the expression of Bax and cleaved-Caspase 3,decreases Bcl-2 to play the role in inducing apoptosis.4.Ginsenoside Rg3 inhibits the migration and invasion abilities by decreasing the expression levels of MMP2,MMP7,MMP9 and EMT-relatedproteins.5.The anti-cancer effect of Ginsenoside Rg3 in osteosarcoma may be involved in the regulation of Wnt/?-Catenin signaling pathway.