Study On The Effect Of Medium Frequency Alternating Microcurrent Combined With Drugs On Lung Adenocarcinoma A549 Cells

Objective:This study is to explore the effect of alternating micro-current at intermediate frequency(ACIF)on lung adenocarcinoma A549 cells,and explore whether the ACIF has a proliferation inhibition effect on A549 cells,and select the optimal inhibition parameters.ACIF will be combined with paclitaxel(TAX)and Kanglaite to intervene in A549 cells,to explore effects of cells proliferation,cell cycle and apoptosis of A549 cells.The purpose of this experiment is to explore whether ACIF which is a novel non-invasive anti-tumor physical method is feasible for the treatment of non-small cell lung cancer.Methods:1.Effects of ACIF on A549 cells1.1 Effects of different frequencies ACIF on A549 cellsUsing A549 cells as the the experimental subjects,according to the single variable method,currents of 30 kHz,50 kHz,100 kHz,150 kHz,200 kHz,300 kHz,and 400 kHz are applied to the cells at a current intensity of 70 mA and 30 min.After 3 days of continuous action,the survival rates of A549 cells are detected by MTT,and the effects of different frequencies ACIF on cells are evaluated,and the optimal experimental current frequency parameter is determined.1.2 Effects of different intensity ACIF on A549 cellsUsing A549 cells as the the experimental subjects,according to the single variable method,currents of 30 mA,50 mA,70 mA,90 mA,110 mA,130 mA,and 150 mA are applied to the cells at a current parameter of 150 kHz and 30 min.After 3 days of continuous action,the survival rates of A549 cells are detected by MTT,and the effects of different intensities ACIF on cells are evaluated,and the optimal experimental current intensity parameter is determined.1.3 Effects of different time ACIF on A549 cellsUsing A549 cells as the the experimental subjects,according to the single variable method,currents of 15 min,30 min,45 min,and 60 min are applied to the cells at a current parameter of150 kHz and 90 mA.After 3 days of continuous action,the survival rates of A549 cells are detected by MTT,and the effects of different ACIF time on cells are evaluated,and the optimal experimental current time parameter is determined.2.Effects of ACIF combined with kanglaite on A549 cellsAccording to the results of previous experiments,the optimal parameters of the ACIF were determined to be 150 kHz,90 mA,and 30 min.To explore the synergistic effect of ACIF combined with kanglaite,A549 cells are exposed to ACIF stimulation,kanglaite stimulation,ACIF combined with kanglaite stimulation and natural growth.After experiment stimulation,the survival rates of each group are detected by MTT,the apoptosis/death and cycle status of each group are analyzed by flow cytometry.3.Effects of ACIF combined with taxol on A549 cellsAccording to the results of previous experiments,the optimal parameters of the ACIF were determined to be 150 kHz,90 mA,and 30 min.To explore the synergistic effect of ACIF combined with taxol,A549 cells are exposed to ACIF stimulation,taxol stimulation,ACIF combined with taxol stimulation and natural growth.After experiment stimulation,the survival rates of each group are detected by MTT,the apoptosis/death and cycle status of each group are analyzed by flow cytometry.Results:1.The current frequency showed a proliferation inhibition effect on A549 cells.The survival rate of A549 cells decrease first and then increase with the increase of current frequency(30kHz-400 kHz).At the current frequency of 150 kHz,A549 cells have the lowest relative survival rate.2.The current intensity inhibited the proliferation of A549 cells.The survival rate of A549 cells decrease first and then increase with the increase of current frequency(30 mA-150 mA).At 90 mA current intensity,A549 cells have the lowest relative survival rate.3.With the increase of the time of ACIF(15-60min),the survival rate of A549 cells gradually decrease,and the proliferation of A549 cells is most sensitive to the time of 30min.4.cell survival rate is reduced to(78.02±0.73)%(P<0.01)after the ACIF(150 kHz,90 mA,30 min)applied to A549 cells.ACIF promotes the apoptosis of A549 cells,the proportion of early and late withered cells is(17.70±0.75)%(P<0.01);the ACIF could block the A549 cell cycle in G0/G1 phase.5.the survival rate of A549 cells is(38.75±0.67)%(P<0.01)after ACIF(150 kHz,90 mA,30 min)combined with Kanglaite(IC50).-The combined group increase the apoptotic rate of A549 cells,and the proportion of early and late withered cells is 58.50±0.84%(P<0.01).The ACIF combined with Kanglaite have little effect on A549 cell cycle.6.the survival rate of A549 cells is(32.87±0.94)%(P<0.01)after ACIF(150 kHz,90 mA,30 min)combined with taxol(IC50).The combined group increase the apoptosis rate of A549 cells,and it is obviously on the late apoptotic cells.The proportation of early and late withered cells is(63.30±1.11)%(P<0.01).the combined group can block the A549 cell cycle in G2/M and S phase.Conclusion:ACIF has a certain proliferation inhibition effect on A549 cells.ACIF combined with Kanglaite injection has synergistic effect,which can enhance the cell proliferation inhibition and promote cell apoptosis,but it is not obvious effect on the cell cycle arrest.The mechanism may be directly mediate The inner apoptotic signaling pathway.ACIF combined with taxol can increase the apoptosis rate of A549 cells and block it in S,G2/M phase.The ACIF combined with taxol has synergistic effects which has effects on cell proliferation,cell apoptosis and cell cycle.ACIF is a new method of tumor physiotherapy,developed by China’s independent innovation.It has anti-tumor effect.Many experiments in vitro have confirmed its therapeutic effectiveness on tumor cells,but there are still problems in vivo testing and clinical application.