CAJ | 학술논문

目的观察替莫唑胺(TMZ)诱导人脑胶质瘤U251耐药细胞株(U251/TR)经放射线照射后细胞周期及P-糖蛋白(P-gp)表达的改变.方法逐步递增剂量诱导方法建立U251/TR细胞株,细胞计数试剂盒(CCK-8)法测定其药物敏感性;倒置显微镜观察亲本U251细胞及U251/TR两种细胞结晶紫染液染色后形态差异;医用直线加速器6 Gy剂量放射线照射两种细胞后观察细胞周期的变化;Western blot分析耐药蛋白细胞膜P-gp表达变化.结果成功诱导建立人脑恶性胶质瘤U251耐药细胞株后,结晶紫染液染色显示显微镜下两种细胞存在形态学差异.CCK-8法检测显示U251/TR对TMZ、阿霉素、环磷酰胺、依托泊苷、硫酸长春新碱化疗药物的耐受指数分别为2.63、2.76、2.15、10.58、3.54.流式细胞术检测发现在同步化之后,经6Gy X线照射后6、24、48、72 h细胞周期各时比较,提示U251/TR细胞株较亲本细胞G2/M期明显减少,G0/G1期和S期明显增多.Western blot结果显示6 Gy放射线照射U251/TR和U251细胞后,P-gp的蛋白表达均下降(相对蛋白表达量分别为0.535±0.002、0.349±0.004),说明6 Gy放射线对两种细胞均有杀伤作用.结论经小剂量6Gy X线辐射照射可抑制甚至杀伤胶质瘤U251细胞及耐TMZ细胞U251/TR,一定剂量放疗后两种细胞放射敏感性均增高.
목적 관찰체막서알(TMZ)유도인뇌효질류U251내약세포주(U251/TR)경방사선조사후세포주기급P-당단백(P-gp)표체적개변.방법 축보체증제량유도방법건립U251/TR세포주,세포계수시제합(CCK-8)법측정기약물민감성;도치현미경관찰친본U251세포급U251/TR량충세포결정자염액염색후형태차이;의용직선가속기6 Gy제량방사선조사량충세포후관찰세포주기적변화;Western blot분석내약단백세포막P-gp표체변화.결과 성공유도건립인뇌악성효질류U251내약세포주후,결정자염액염색현시현미경하량충세포존재형태학차이.CCK-8법검측현시U251/TR대TMZ、아매소、배린선알、의탁박감、류산장춘신감화료약물적내수지수분별위2.63、2.76、2.15、10.58、3.54.류식세포술검측발현재동보화지후,경6Gy X선조사후6、24、48、72 h세포주기각시비교,제시U251/TR세포주교친본세포G2/M기명현감소,G0/G1기화S기명현증다.Western blot결과현시6 Gy방사선조사U251/TR화U251세포후,P-gp적단백표체균하강(상대단백표체량분별위0.535±0.002、0.349±0.004),설명6 Gy방사선대량충세포균유살상작용.결론 경소제량6Gy X선복사조사가억제심지살상효질류U251세포급내TMZ세포U251/TR,일정제량방료후량충세포방사민감성균증고.
Objective To investigate the cell cycle and P-glycoprotein expression changes of drugresistance cell lines of U251 induced by temozolomide (TMZ) after radiation exposure.Methods The U251 cells resistant to TMZ (U251/TR) were maked by increasing concentrations.The muhidrug resistance was detected by cell counting kit-8 (CCK-8).Cell morphology of U251 and U251/TR was observed under optical microscope with crystal violet staining.We induced U251 parents and U251/TR cells used 6 Gy X-rays by linear accelerator.The flow cytometer was used to test the change of cells cycle after X-rays irradiation.Protein expression of P-glycoprotein (P-gp) was detected by using Western blotting.Results We were successful in inducing to establish a constant drug-resistace cell lines U251/TR by TMZ.Inverted microscope to observe morphology of the U251 and U251/TR were different after dyeing.CCK-8 method test showed that:the U251/TR to TMZ,doxorubicin,cyclophosphamide,etoposide,sulfuric acid vincristine chemotherapy drugs resistance index for parents of U251 respectively 2.63,2.76,2.15,10.58,3.54.The flow cytometric test showed the U251/TR cell lines of G2/M phase was decreased significantly than parent cell,but G0/G1 phase and S phase were increased obviously.Western blotting result analysis:the P-gp protein expression was decreased after exposure to 6 Gy X-ray irradiation (The relative quantity of protein expression,respectively was 0.535 ± 0.002,0.349 ± 0.004).Conclusion Small doses X-ray (such as 6 Gy) radiation exposure to inhibit or even killing.Both U251 and U251/TR Cells were increased radiosensitivity after Right Dosage X-ray irradiation.