The Role Of MiR-21 In The Inhibition Of Osteosarcoma Cells By Curcumin

PART1 CURCUMIN AFFECTS THE PROLIFERATION OF OSTEOSARCOMA CELL LINES MG-63 AND HOSObjective: To investigate whether curcumin could affect osteosarcoma cell proliferation.Methods: Osteosarcoma cells(MG-63 and HOS)were treated with curcumin at different concentrations(0.1%DMSO,2.5?M,5?M,10?M).CCK-8,colony formation experiments,and flow cytometry were used to determine cell proliferation.Western blot assay was performed to detect the expression of BAX and BCL-2.Results: CCK-8 results showed that the curcumin-treated group(5?M,10?M)had lower absorbance at 48 h and 72 h than the control group(0.1%DMSO).Compared with the control group(0.1%DMSO),the number of colony formed in the curcumin-treated group(10 ?M)was significantly reduced.Flow cytometry indicated that the apoptosis of the cells was increased in curcumin-treated cells.Western blot results showed that curcumin up-regulated the expression of BAX protein and down-regulated the expression of BCL-2 protein.Conclusion: In summary,the results showed that curcumin has an inhibitory effect on osteosarcoma cells MG-63 and HOS.PART2 CURCUMIN AFFECTS THE EXPRESSION OF MIR-21,RECK AND WNT /?-CANTENIN SIGNALING PATHWAY PROTEINS IN OSTEOSARCOMA CELLSObjective: To investigate whether curcumin could affect the expression of mi R-21,the target gene of mi R-21 RECK and Wnt / ?-cantenin signaling pathway related proteins in osteosarcoma cells MG-63 and HOS,exploring the mechanism of curcumin's antitumor effect.Methods: Osteosarcoma cells were treated with curcumin at different concentrations(0.1%DMSO,2.5?M,5?M,10?M)for 48 hours.RT-q PCR was used to detect the expression of mi R-21 and RECK.Western blot was used to detect the expression of wnt pathway proteins ?-catenin,GSK-3?and c-myc.Results: Compared with the control group(0.1% DMSO),the expression of mi R-21 and wnt pathway protein ?-catenin,GSK-3?,c-myc decreased with the increase of curcumin concentration.The expression of RECK was increased in cells treated with curcumin.Conclusion: Curcumin could down-regulate the expression of mi R-21 and wnt/?-catenin signaling pathway protein in osteosarcoma cells,and up-regulated the expression of RECK,indicating that curcumin may play an anti-tumor effect by affecting mi R-21/RECK and wnt/?-cantenin signaling pathways.PART3 TRANSFECTION OF HSA-MIR-21-5P-MIMICS COULD REVERSE THE EFFECT OF CURCUMIN ON OSTEOSARCOMA CELLSObject: To investigate whether up-regulation of mi R-21 could reverse the inhibitory effect of curcumin on osteosarcoma cells,and whether affect the expression of RECK and wnt/?-catenin pathway proteins.Methods: Osteosarcoma cells were transfected with hsa-mi R-21-5p-mimics to up-regulate mi R-21,and RT-q PCR was used to verify the transfection efficiency.CCK-8 and Western blot were performed to detected the effect of up-regulation mi R-21 on curcumin inhibition.Western blot was used to detect the expression of RECK and wnt pathway proteins after transfection.Results: RT-qPCR results showed that the expression of mi R-21 was significantly higher and RECK was lower in the osteosarcoma cell transfection group than the control group,indicating that the cell transfection was successful.Compared with the control group(curcumin group),the absorbance value of the CCK-8 experiment in the experimental group(mimics + curcumin group)was increased,the expression of BAX protein was down-regulated,the expression of BCL-2 was up-regulated,and wnt/?-catenin pathway proteins,?-catenin,GSK-3? and c-myc expression was increased.Conclusion: Up-regulating mi R-21 could reverse the inhibitory effect of curcumin on osteosarcoma cells and affect the expression of RECK and wnt /?-catenin pathway proteins,indicating that curcumin may affect its target gene RECK and wnt / ?-cantenin pathway through mi R-21 to exert its inhibitory effect on cell proliferation.