【作者】 王伟明；

【导师】 李文辉； 崔建国； 秦继勇；

【作者基本信息】 昆明医学院 ， 影像医学与核医学， 2008， 硕士

【摘要】 【研究目的】探讨PI3K/Akt特异性抑制剂Ly294002对云南宣威肺腺癌XWLC-05细胞株的放射增敏作用。【研究方法】1.克隆形成实验测定XWLC-05细胞在不同剂量X线照射下的存活分数,分别用单靶多击数学模型拟合细胞存活曲线,并求出放射生物学参数D₀、D_q、N、SF₂、SER。2.四氮唑兰比色法（MTT法）检测X线4Gy照射24h、48h后不同浓度Ly294002对XWLC-05细胞的抑制率,评价细胞放射敏感性。3.流式细胞术测定不同条件下Ly294002对XWLC-05细胞的周期分布和细胞凋亡情况的影响,评价Ly294002的作用机制。【研究结果】1.克隆形成实验:XWLC-05细胞在对照组各个剂量点的存活分数均高于实验组,对照组D0、Dq、N、SF2的值均大于实验组,SER值为1.05,对照组和实验组存活分数经成组t检验有显著差异（t=2.856,p=0.036）。2.MTT法实验:不同浓度（5、10、20、50μmol/l）Ly294002作用XWLC-05细胞24小时及48小时后,对照组抑制率小于实验组,Ly294002作用24小时后,对照组和实验组抑制率经成组T检验有显著差异（t=-17.489,p=0.003）,Ly294002作用48小时后对照组和实验组抑制率经成组T检验无显著差异（t=-2.201,p＞0.05）。3.流式细胞检测:Ly294002作用12小时后,未照射实验组G₂/M期细胞比例大于对照组,照射实验组G₂/M期细胞比例小于对照组,对照组和实验组G₂/M期细胞经成组t检验无显著差异。Ly294002作用24小时和36小时后,实验组G₂/M期细胞比例均大于对照组,对照组和实验组G₂/M期细胞经成组t检验有显著差异（p＜0.05）。Ly294002作用12小时、24小时和36小时后,各组凋亡结果不明显,经秩和检验,对照组和实验组细胞凋亡结果无明显差异（p＞0.05）。【结论】1.Ly294002对XWLC-05细胞有放射增敏作用。2.不同浓度Ly294002作用XWLC-05细胞24小时后,对细胞的抑制皇浓度依从性,随着Ly294002浓度的增加,抑制率增加;且照射组抑制率大于单纯Ly294002组;提示24小时可能为Ly294002最佳作用时间。3.XWLC-05细胞经照射后出现G2/M期细胞阻滞,经Ly294002作用后G2/M期细胞阻滞增加,凋亡率不明显,提示Ly294002可能通过增加G2/M期细胞数增加XWLC-05细胞的放射敏感性。更多还原

【Abstract】 PurposeTo investigate the radiosensitive effect of PI3K/Akt’s specific inhibitor-Ly294002 on adenocarcinoma of lung cancer cell at xuanwei,yunnan province.Materials and methods1.Clonogenic survival assay was performed to determine survival fraction of XWLC-05 cell at different doses.The parameters D₀,D_q,N,SF₂ and SER from multi-target hit mathematical model were calculated to evaluate radiosensitivity of the cells and were fitted cell survival curve.2.Tetrazolylazo-colorimetry was detected to inhibition rates of XWLC-05 cells as different concentrations of Ly294002 after 24 hours and 48 hours with 4Gy irradiation,Cell radiosensitivity was evaluated.3.Flow cytometry was performed to check the impact on the XWLC-05 cells cycle and apoptosis on different conditions of Ly294002 in order to evaluated the mechanism of Ly294002.Results1.Clonogenic survival assay:The XWLC-05 cells’s survival fraction of the control group in different points were higher than experimental group.The control group’s values of D₀,D_q, SF₂ were higher than the experimental group.The value of SER is 1.05.The survival fraction between the control group and the experimental group were significantly different（p＜0.05）.2.Detection of MTT colorimetric assay:With different concentrations（5,10,20,50μmol/l）, inhibition rate of the control group inhibition rate was less than that of the experimental group after 24 hours and 48 hours.By the data t test group,the inhibiting results processed for 24 hours with Ly294002,between the control group and the experimental group were significantly different（p＜0.05）.The cells are processed for 24 hours with Ly294002 are of statistical significance（p＜0.05）.The inhibition were processed for 24 hours by Ly294002,the control group and the experimental group were of no statistical significance（p＞0.05）.3.Detection of the Flow cytometry assay:Having processed for 24 hours with Ly294002, the G₂/M phase cell ratio of the experimental group of no irradiation was greater than the control group,the G₂/M phase cell ratio of the experimental group of irradiation wasless than the control group（p＞0.05）.Having processed for 24 hours and 36 hours with Ly294002,the G₂/M phase cell ratio of the experimental group of irradiation was more than the control group（p＜0.05）.The result of apoptosis was not obvious after the cells were processed for 12 hours,24 hours and 36 hours with Ly294002,the control group and experimental group apoptosis result there was no significant difference（p＞0.05）.Conclusions1.Ly294002 have an radiosensitive effect on XWLC-05 cell.2.After 24 hours,the role of different concentrations Ly294002 inhibited on XWLC-05 cells was in a dose-compliance.With the increasing concentration of Ly294002,inhibition rate increase.Comparing with the control group,the inhibitory rate of the irradiated group was more greatly increasing;The best time processing with Ly294002 may be 24h.3.The G₂/M phase of XWLC-05 cells was blocked after irradiating,the G₂/M phase cells block were increased by Ly294002,The rate of apoptosis was not obvious,It was suggested that the mechanism of Ly294002 on XWLC-05 cell line’s radio-sensitizing effect may be blocking cells during G₂/M phase.更多还原