Effects Of Sevoflurane And Isoflurane On Proliferation,apoptosis And Chemosensitivity Of Osteosarcoma Cell Line MG63

Part1 Effects of sevoflurane and isoflurane on proliferation,colony formation and apoptosis of osteosarcoma cell line MG63Objective: To investigate the effects of different concentrations of sevoflurae and isoflurane on proliferation, colony formation and opoptosis of MG63 cells.Methods: MG63 cells were divided into seven groups:group ctrol(group C):MG63 cells was treated by mixed gas of 5%CO2,21%O2,74%N2,group 1.7%Sev(group S1):MG63 cells was treated by mixed gas which contained 1.7%sevoflurane, group 3.4%Sev(group S2):MG63 cells was treated by mixed gas which contained 3.4%sevoflurane, group 5.1%Sev(group S3):MG63 cells was treated by mixed gas which contained 5.1%sevoflurane, group 1.3%Iso(group I1):MG63 cells was treated by mixed gas which contained 1.3%isoflurane, group 2.0%Iso(group I2):MG63 cells was treated by mixed gaswhich contained 2.0%isoflurane, group 2.6%Iso(group I3):MG63 cells was treated by mixed gas which contained 2.6%Isoflurane. The proliferation inhibition rate was measured by MTT assay.The ability of colony formation was detected by the colony formation assay.The opoptosis rates of MG63 cells were detected by flow cytometer respectively after exposure to different concentrations of sevoflurane and isoflurane.Results: 1. The results of MTT assay:compared with group ctronl,the proliferation inhibition rates of the MG63 cells treated by different concentrations of sevoflurane or isoflurane was increased,of which the proliferation inhibition rates of the group 5.1%Sev and group 2.6%Iso were higher than the others(P<0.05).The results displayed different concentrations of sevoflrane and isoflurane could inhibit the proliferation of MG63 cells,the effects of proliferation inhibition of high concentrations of sevoflurane and isoflurane to MG63 cells were more obviously. 2. The results of colony formation assay:the colony formation rates of different concentrations of sevoflurane and isoflurane were lower than the group control(P<0.05).But the effects had nothing to do with the concentrations of sevoflurane and isoflurane. 3. The results of flow cytometry:the opoptosis rates of different concentrations of sevoflurane and isoflurane was not statistically significance compared with the group control.It showed that various concenttations of sevoflurane and isoflurane didn’t promote the apoptosis of MG63 osteosarcoma cells.Conclusion: Different concentrations of sevoflurane and isoflurane inhibit the proliferation of MG63 osteosarcoma cells,the inhibition effects were more obvious by high concentrations of sevoflurane and isoflurane.The ability of colony formation of MG63 cells is inhibited by sevoflurane and isoflurane,but has little to do with concentrations of inhalation anesthetics.Sevoflurane or isoflurane has no effect on the apoptosis rate of MG63 osteosarcoma cells.It shows that the effets of sevoflurane and isoflurane on the proliferation of MG63 cells are independent of the effects of sevoflurane and isoflurane onthe opoptosis rates of MG63 cells.Part2 Effects of sevoflurane and isoflurane on the cisplatin-induced apoptosis to MG63 osteosarcoma cells and the mechanismObjective: To investigate the effects of sevoflurane and isoflurane on the cisplatin-induced apoptosis to MG63 osteosarcoma cells and detecte the expression of the related proteins.Methods: MG63 cells were divided into six groups:group control, MG63 cells was treated by mixed gas containing 5%CO2,21%O2,74%N2;Group sevoflurane, MG63 cells were treated by mixed gas which contained 2.5%sevoflurane;group isoflurane, MG63 cells were treated by mixed gas which contained 2.0%isoflurane;group DDP:MG63 cells were treated by cisplatin with the concentration of 10mg/L for 24 hours;group DDP+2.5%Sev: MG63 cells were treated by cisplatin with the concentration of 10mg/L for 24 hours,within the first 4 hours treated by 2.5%sevoflurane;group DDP+2.0%Iso:MG63 cells were treated by cisplatin with the concentration of 10mg/L for 24 hours,within the first 4 hours treated by 2.0%isoflurane.The viability of MG63 cells were detected by MTT assay.The apoptosis rates of all groups were detected by flow cytometry.The expression levels of p-Akt, NF-κB P65, Hif-1α, Bcl-2, Baxprotein were detected by western blot.Results: 1. Results of MTT assay.Compared with group control,the OD value of each group was lower(P<0.05),Of which the OD values of group DDP,group DDP+2.5%Sev and group DDP+2.0%Iso were significantly lower than the other groups(P<0.05).It showed that sevoflurane,isoflurane and cisplatin decreased the viability of MG63 cells and the effect of cisplatin on the vibility of MG63 cells was more obvious.2. Results of apoptosis rate of MG63 cell:Compared with group control,the apoptosis rate of group DDP,group DDP+2.5%Sev and group DDP+2.0%Iso was significantly increased(P <0.05).It suggested that cisplatin induced the MG63 osteosarcoma cells apoptosis.Compared with group DDP,the apoptosis rate of MG63 cells was decreased,suggesting that sevoflurane and isoflurane attenuated the opoptosis rate of cisplatin-induced to MG63 osteosarcoma cells. 3. Results of western blot assay.Compared with group DDP,the expression level of NF-κB P65, Hif-1α, Bcl-2 of group DDP+2.5%Sev and group DDP+2.0%Iso was increased,while reducing the expression level of Bax(P <0.05).It showed that sevoflurane and isoflurane promoted the activation of NF-κB,improved the expression level of Hif-1α,and changed the expression level of Bcl-2 and Bax.The expression level of p-Akt had no different among the three groups.Conclusion: Sevoflurane and isoflurane decrease the vitality of MG63 cells.And sevoflurane and isoflurane antagonize the toxic effects of cisplatin on MG63 osteosarcoma cells.The apoptosis induced by cisplatin to MG63 osteosarcoma cells is attenuated by sevoflurane and isoflurane.The mechanism of sevoflurane and isoflurane reducing the chemosensitivity of MG63 cells to crisplatin may be related with the activate of NF-κB,the higher expression level of Hif-1α and the changing of the expression of Bcl-2 and Bax.