放疗对卵巢癌顺铂耐药细胞的影响

Effect of Radiotherapy on Cisplatin-Resistant Ovarian Cancer Cells

目的探究放疗对卵巢癌顺铂(cisplatin,CDDP)耐药细胞耐药性的影响,并探讨相关机制。方法取对数期卵巢癌SKOV3细胞株、SKOV3/CDDP耐药细胞株,二甲基噻唑(dimethythiazole,MTT)法检测SKOV3、SKOV3/CDDP对CDDP敏感性,以及放疗对SKOV3/CDDP的杀伤作用;取对数期SKOV3/CDDP耐药细胞株,采用0.5 Gy×4次低剂量分割放疗干预,设为低剂量分割放疗组,取不处理SKOV3/CDDP耐药细胞株为空白组,平板克隆实验测定2组的克隆形成能力;流式细胞术测定凋亡率;碘化丙啶(propidium iodide,PI)染色测定细胞周期;Western blot检测细胞周期蛋白-D1(cyclin-D1)、骨髓细胞瘤病毒癌基因(cellular-myelocytomatosis viral oucogene,c-myc)、β-链蛋白(β-catenin)、p-β-catenin蛋白表达。结果CDDP对SKOV3、SKOV3/CDDP细胞的半数抑制剂量(half inhibitory concentration,IC 50)分别为2.41、6.15 mg·L^(-1),SKOV3/CDDP对CDDP的耐药指数(resistance index,RI)为2.55倍;不放疗、0.5 Gy、0.5 Gy×4次、2 Gy时,CDDP对SKOV3/CDDP细胞的IC 50分别为6.15、5.17、3.92、4.31 mg·L^(-1),0.5 Gy、0.5 Gy×4次、2 Gy时耐药逆转倍数(reversal fold,RF)分别为1.19倍、1.57倍、1.43倍;低剂量分割放疗组克隆形成率,S、G 2/M占比,cyclin-D1、c-myc蛋白相对表达量,p-β-catenin/β-catenin低于空白组,凋亡率、G 0/G 1占比高于空白组,差异有统计学意义(P<0.05)。结论低剂量分割放疗可抑制卵巢癌CDDP耐药细胞增殖,促进其凋亡,具有化疗增敏作用,推测与抑制Wnt/β-catenin信号通路有关。

Objective To explore the effect of radiotherapy on the drug resistance of ovarian cancer cisplatin(CDDP)resistant cells and explore the related mechanisms.Methods SKOV3 cell line and SKOV3/CDDP-resistant cell line of logarithmic stage ovarian cancer were taken.Dimethylthiazole(MTT)method was used to detect the sensitivity of SKOV3,SKOV3/CDDP to CDDP and the killing effect of radiotherapy on SKOV3/CDDP.The log-phase SKOV3/CDDP-resistant cell line was taken and intervened with 0.5 Gy×4 low-dose split radiotherapy,which was set up as the low-dose split radiotherapy group,and the untreated SKOV3/CDDP-resistant cell line was taken as the blank group,and the plate cloning assay was performed to determine the clone-forming ability of the two groups.Flow cytometry was used to determine the apoptosis rate,propidium iodide(PI)staining was used to determine the cell cycle,and Western blot was used to detect cyclin-D1,cellular-myeloma virus oncogene(c-myc),β-catenin and p-β-catenin protein expression.Results The half inhibitory concentration(IC 50)of CDDP on SKOV3 and SKOV3/CDDP cells was 2.41 and 6.15 mg·L^(-1),respectively,and the resistance index(RI)of SKOV3/CDDP to CDDP was 2.55 times.The IC50 of CDDP on SKOV3/CDDP cells was 6.15,5.17,3.92 and 4.31 mg·L^(-1) at no radiotherapy,0.5 Gy,0.5 Gy×4 times and 2 Gy,respectively;and the reversal fold(RFT)of resistance was 1.19 times,1.57 times,and 1.43 times at 0.5 Gy,0.5 Gy×4 times,and 2 Gy,respectively.The clone formation rate,the proportion of S,G 2/M,the relative expression of cyclin-D1 and c-myc protein,p-β-catenin/β-catenin in the low-dose fractional radiotherapy group were lower than those of the blank group,of which the apoptosis rate,the proportion of G 0/G 1 were higher than those of the blank group(P<0.05).Conclusion Low-dose fractional radiotherapy can inhibit the proliferation of CDDP-resistant ovarian cancer cells,promote their apoptosis,and have chemosensitizing effects,which is presumably related to the inhibition of Wnt/β-catenin signaling pathway.