The Preliminary Study Of Radiosensitizing Effect Of Ly294002 On XWLC-05 Cell Lines

[Objective]To investigate the radiosensitive effect and the mechanism of LY294002 on lung cancer by applying the specific inhibitor LY294002 on the signaling pathway PI3K/Akt to XWLC-05 cell lines.[Materials and Methods]1.Research object:human lung adenocarcinoma cell lines XWLC-05 from Xuanwei,Yunnan Province,cultivated commonly with 1640 culture medium containing 10%fetal bovine serum.2.Set up the radiotherapy plus drug groups(4Gy X-ray + Ly294002 with different concentrations of 0,5,10,20,50 umol /L).deal the cells for 24h,48h,72h, 96h individually.Calculate the cell survival rate by testing absorbance A at 490nm wave length with MTT method.Deduct the optimum time and concentration of Ly294002 on XWLC-05 cell line when there' s radiosensitive effect.3.Cell cloning experiments:In the control group(1640 medium)and experimental group(1640 medium including Ly294002 of 50 umol/L),cells were cultured for 24h before they were exposed to different doses of X-rays(0 Gy, 1 Gy,2 Gy,4 Gy,6 Gy,8 Gy).14 days later,count the number of cloned cells(cells more than 50),calculate the cell survival rate,use Microsoft Excel to draw the dose-survival fraction curve,and use Constrained Nonlinear Regressin to calculate the value of D₀,D_q,SF₂ and radiation sensitivity ratio (SER). 4.In the control group(radiotherapy)and experimental group(Drug plus radiation),cells were cultured for 24h individually with X-rays of different doses(0 Gy,1 Gy,2 Gy,4 Gy,6 Gy and 8 Gy),before the cell cycle and apoptosis were measured by Flow Cytometry.5.Set up control group,drug group(Ly294002 of 50 umol/h),and radiotherapy group(4 GyX-ray)and radiotherapy plus drug group(46y X-ray + Ly294002of 50 umol/L).In these groups,cells were cultured for 24h before the cell cycle and apoptosis measured by Flow Cytometry.6.Process the data with SPSS13.00 software.T tests were used to analyze means of two groups,ANOVA was used to analyze means of multiple groups,and LSD for comparison between means.[Results]1.The optimum concentration of Ly294002 is 50umol/L as a radiatio-sensitizing agent.The influence of Ly294002 of 50umol/L on the survival of cells was more significant than that of 0,5,10,20 umol/L.The difference is of statistical significance(P<0.05).2.There is no relationship between time duration and the radiothe-rapy sensitizing effect of Ly294002 on XWLC-05.The differences of the survival rate of the cells are of no statistical significance(P<0.05)under the intervening of LY294002 with 24H,48H,72H,96H individually.3.In the cloning experiment of Ly294002 of 50 umol / L on the cells for 24h, the results showed that the Ly294002 on XWLC-05 cell line has radio-sensitizing effect,SER=1.41.4.Ly294002 only doesn't have any impact on cell cycle and apoptosis.Only when it is combined with radiotherapy,it can make effect on the cell cycle and apoptosis,and the difference is of statistical significance(P<0.05).5.The radio-sensitizing effect of Ly294002 on XWLC-05 cell line m-ainly exists in blocking cells in G2/M phase,and the difference is of statistical significance(P<0.05).Ly294002 has little effect on the XWLC-05 cell line during S-phase and cell apoptosis,and the difference is of no statistical significance(P>0.05).[Conclusion]1.Ly294002 has radio-sensitizing effect on XWLC-05 cell line.The best concentration is 50 umol/L.There is no relationship between time duration and the radiothe-rapy sensitizing effect of Ly294002 on XWLC-05.2.Ly294002 alone does not affect the cell cycle and cell apoptosis.However, when it is combined with radiotherapy,it could influence the cell cycle and apoptosis.3.The mechanism of Ly294002 on XWLC-05 cell line's radio-sensitizing effect may be blocking cells during G₂/M phase.While there is little effect on XWLC-05 cell line during S-phase and cell apoptosis.