COX-2 Inhibitor Celecoxib's Radiosensitizing Effects On SHG-44 Cell Lines And The Preliminary Study Of The Influence On The Expression Of CyclinB1 Gene

Objective To evaluate the COX-2 inhibitor Celecoxib's radiosensitizing effects on SHG-44 cell lines in vitro,characterize the effects on cell cycle redistribution when radiation combined with Celecoxib,and to probe the underlying signal pathway mechanism of Celecoxib's regulation of cell cycle.Methods In this study,human glioma cell SHG-44 was managed in vitro.Group of drug(Group D)was divided according to the different concentration of Celecoxib(0μM,30μM,50μM,100μM),and the group of radiation(Group R)divided according to the different dose of 6MV X-ray(0Gy,2Gy,4Gy,6Gy,8Gy),the other group was formed by Celecoxib combined with radiation(Group D+R).MTT analysis was performed to determine the effects of Celecoxib on SHG-44 cell growth;Celecoxib's radiosensitization was measured by clongenic assay;The cell cycle redistribution was analyzed in different groups by Flow-cytometric analysis;The expression of CyclinB1 mRNA in SHG-44 cells was detected by RT-PCR,and Real Time-PCR was used to determine the relative level of CyclinB1 mRNA.Results1.The outcome of MTT assay suggested that Celecoxib may produce a concentration-dependent and time-dependent inhibition of SHG-44 cell growth.2.By clongenic assay,Celecoxib showed radiosensitizing effect to a certain extent in SHG-44 cells.The survival curve and the related parameters indicated that Celecoxib could enhance the radiosensitivity of SHG-44 cells.The value of D₀,Dq and D_0.01of Group D+R was significantly different from Group R..3.Flow-cytometric analysis showed cell cycle redistribution varied in Group D,Group R and Group D+R.There was more G₂/M phase arrest with 50μmol/L Celecoxib than 0μmol/L and 30μmol/L in both Group D and Group D+R(P＜0.01),while with no significant difference between 30μmol/L and 0μmol/L(P=0.05);SHG-44 cells in Group R showed an enhanced G₂/M phase arrest with 6Gy and 8Gy compared with 0Gy (P＜0.01),but no significant increase with 2Gy and 4Gy(P＞0.05).With 50μmol/L Celecoxib,Group D+R showed more G₂/M phase arrest than Group R(P＜0.05),while with 30μmol/L Celecoxib,there was no significant enhancement(P＞0.05).4.The expression of CyclinB1 gene was proved by RT—PCR in all groups,and Real-Time PCR was performed to determine the level of CyclinB1 mRNA:Compared with 0μmol/L Celecoxib,the expression level of CyclinB1 was downregulated with 30μmol/L,50μmol/L and 100μmol/L Celecoxib in Group D(P＜0.05),but there were no significant differences within the three different concentration(P＞0.05).Compared with Group D,the expression level of CyclinB1 was downregulated significantly with 100μmol/L Celecoxib in Group D+R,while with the other concentration of Celecoxib(0,30,50μmol/L)in Group D+R,there were no significant difference(P＞0.05).Conclusion1.Celecoxib,a COX-2 selected inhibitor,could enhance radiosensitivity of glioma cells SHG-44 in vitro.2.Celecoxib enhanced G₂/M arrest at a certain concentration,and could strengthen the radiation-induced G₂/M arrest in SHG-44 cells.3.One of the underlying mechanisms of radiosensitizing effects of Celecoxib maybe the downregulation of CyclinB1 acted as a key downstream gene in the signal pathway for the G₂/M phase transition.