Study Cytotoxicity Effect And Mechanism Of Celecoxib On Human Osteosarcoma Cell Line MG-63

Objective:to sutdy the cytotoxicity effect and mechanism of Celecoxib on human osteosarcoma cell line MG-63.Methods:MG-63 cells were cultured in the environment of 37°C 5%CO2 with modified Eagle medium (MEM)medium.The logarithmically growing MG63 cells were used。1 The suppressive effects of resveratrol on the proliferaition of MG63 cells were evaluated in vitro by MTT colorimetricassay. After cultured for several generations,the cells were digested and transferred into 96-well culture plates. after adherenced,add to Celecoxib,culture 24,48,72hours,add to MTT 20ul,incub- ation 4 hours,abandon to supernate fluid,add to DMSO, to swi ng for 5 min,determin OD490 with analysator,according to for- mula calculate inhibition ratio2 Using light microscope observe the change of morphology: MG63 cells were treated with different concentrations of Cele- coxib for 24,48,72 hours,the change of morphology was obse- rved by inverted phase contrast microscope in different time and photographed.3 Flow cytometric analysis:MG63 cells were treated with Celec- oxib for 48h.Experimental group and control group cells were harvested and washed twice with PBS.Cells were fixed with ice cold 70% ethanol at 4°C. Precipitates were digested with 0.5% pepsin after centrifugation.And then cells were resuspended in 0.5 mL propidium iodide(PI)/RNase A solution.Cells were inc- ubated in the dark at room temperature for 15 min.The fluor- escence emission of stained cells was measured with a flow cyt- ometer.Data were analyzed with Multipcycle software.4 Morphology analysis by transmission electron microscopy: MG63 cells treated with different concentrations of Celecoxib for 48 h, and pre-fixed with 2.5%glutaraldehyde at 4°C.The cells were postfixed for 1 h with 1%osmium tetroxide,then dehy- drated through a graded ethanol series,and embedded in Epon 812.The ultrastructure of cells was analyzed inultrathin sections in a transmission electron microscope after the sections were stained with uranylacetate and lead citrate.Results: 1 MG63 cells were treated with at various Cele- coxib concentrations for 24 h,48 h,and 72 h respectively,the growth of cells was inhibited significantly in a time-and dose- dependent fashion. The biggest inhibitory rate of Celecoxib was 69.17% at the concentration of 200μg/ml for 72 h(P<0.01).2 Elecrton microscope revealed the characteristic apoptosis alet- rations,shrinking cellular morphology,integrity cell membr- ane,karyopycnosis,fragment of nucleus,chromatin condensation and margination,crescent nucleus,cytoplasmiec vacuoles.3 after cells were exposed to Celecoxib with at various Cele- coxib concentrations for 48 h,The quantity of treated cells in G0/G1 phase increased but that in S,G2/M phase decreased. MG63 cells rates in G0/G1 phase were 63.4±0.85% at the con- centration of 200μg/ml whlie MG63 cells rates in G0/G1 phase were 49.5±0.72% in control group.The analysis of cellular DNA content by FCM showed that the apoptosis rates become higer with drug concentration.there was a typical apoptotic peak when drug concentration were 2~200ug/ml.After MG63 cells were exposed to Celecoxib (200μg/ml)for 48h,the apoptosis rates were 21.41±0.85%,which were significantly higher than control (4.04±0.52%)(P<0.01).Conclusion: Celecoxib has cytotoxicity againsthumna osteosaeroma cell line MG-63.The mechnanism includes that Celecoxib can eeffetively inhibit the proliferation of MG-63 cell line,arrest the cell cycle at GO/G1 phase and induce the apopt- osis of MG-63 cell line.The effect of Celecoxib displyas a time-dependent and dose-dependent manner.