Effect About PRL-3 Antibody Combined With Paclitaxel On A549 Cells’ Biologic Effects And Mechanism

Objective: To investigate the effects of liver regeneration phosphatase-3 antibody combined with paclitaxel on the biological characteristics of human lung adenocarcinoma A549 cell line,such as proliferation inhibition,migration,and apoptosis,and its effects on PRL-3,PI3 K,and P-AKT To explore possible mechanisms and provide new ideas and basis for clinical drug treatment of lung cancer.Methods:(1)Culture human lung adenocarcinoma A549 cells: A549 cell line was added to RPMI-1640 medium(containing 10% fetal bovine serum)and cultured in an incubator.A549 cells in logarithmic growth phase were used for subsequent experiments.(2)After 24 hours of treatment of A549 cells with different concentrations of PRL-3 antibodies,the increase inhibition rate and IC50 value of each group were measured: A549 cells in log phase were seeded on 96-well plates,and PRL-3 antibodies(50,100,150,200,250 ng/ml),continue to culture for 24 hours,and then use the CCK-8 method to detect the absorbance value(OD value)of each PRL-3 antibody group and use the formula(cell proliferation inhibition rate% =(Control group OD value-experimental group OD value)/(control group OD value-blank group OD value)× 100%))Calculate the proliferation inhibition rate and half inhibition concentration(IC50)of each group,and select the corresponding concentration of IC50 value as the follow-up Experimental drug concentration.(3)Experimental grouping and processing method: The experiment was divided into 4 groups: control group,PRL-3 antibody group,paclitaxel group,PRL-3 antibody group + paclitaxel group.Treatment methods of each group:(1)Control group: add culture solution.(2)PRL-3 antibody group: PR50 antibody with IC50 concentration was added.(3)Paclitaxel group: IC50 concentration of paclitaxel was added.(4)PRL-3 antibody group + paclitaxel group: PRL-3 antibody + IC50 concentration of paclitaxel was added.Each group was then intervened according to different intervention time(24h,48 h,72h).The CCK-8 method was then used to determine the OD value of each group,calculate the cell proliferation inhibition rate of each group,and compare the differences between the groups;the optimal time point(72h)for the proliferation inhibition rate was selected as the intervention time point for subsequent experiments.(4)Determining the apoptosis of each group: After treating A549 cells according to the above grouping and processing methods for 72 hours,use flow cytometry to determine the apoptosis of each group and compare the differences between the groups.(5)Determining the cell migration capacity of each group: After treating cells for 72 hours according to the same grouping and processing method,use the Transwell method to determine the cell migration capacity of each group and compare the differences between groups.(6)Western Blot detection of PRL-3,PI3 K,p-AKT protein expression levels in each group: grouped and treated according to previous experiments,treated cells for 72 hours,and then determined by Western Blot method in each group The expression levels of PRL-3,PI3 K,and p-AKT proteins were compared,and the differences in PRL-3,PI3 K,and p-AKT protein expressions were compared between groups.Results:(1)The CCK-8 method was used to detect different concentrations of PRL-3 antibodies acting on A549 cells for 24 hours.It was found that with the increase of PRL-3 antibody concentration,the proliferation inhibition rate of A549 cells gradually increased,showing a concentration dependence(P<0.05).The difference was statistically different;(2)CCK-8 method was used to detect the proliferation of PRL-3 antibody(IC50)and PTX group(IC50)in each single and combined drug group at different time(24h,48 h,72h)on A549 cells.Inhibition rate,it was found that the proliferative inhibition rate of A549 cells gradually increased with time,and was time-dependent(P<0.05),and the difference was statistically different;and it was found that the combination of PRL-3 antibody and paclitaxel group acted on A549 The cell proliferation inhibition rate was better than that of the single drug group(P<0.05),and the difference was statistically different;(3)The apoptosis rate of each group was detected by flow cytometry,and the apoptosis of A549 cells treated by the combined drug group was obtained The rate(50.80±0.80%)was better than the apoptosis rate of the PRL-3 antibody group(31.39±0.92%)and the apoptosis rate of the PTX group(32.18±0.93%)(P<0.05),and the differences were statistically different;(4)Transwell method was used to detect the cell migration rate of each group,and PRL-3 antibody and Taxus were obtained.The migration rate(52.60±0.40%)of A549 cells in the combined drug group was significantly lower than that of the PRL-3 antibody group(101.20±1.20%)and the PTX group(99.60±1.20%)(P<0.05).Statistical differences;(5)Western Blot assay showed that PRL-3 antibody(164 ng/ml)could cause down-regulation of PRL-3(0.38±0.06)protein expression after 72 hours of action on A549 cells,and PI3K(0.35±0.01)protein expression down-regulation,P-AKT(0.33±0.05)protein expression down-regulation;PTX(40 ug /ml)after 72 hours on A549 cells can cause down-regulation of PRL-3(0.34±0.04)protein expression,PI3K(0.31±0.01)protein expression was down-regulated,P-AKT(0.29±0.05)protein expression was down-regulated;and PRL-3 antibody and paclitaxel combined with A549 cells for 72 hours caused down-regulation of PRL-3(0.11±0.04)protein expression The expression of PI3K(0.15±0.03)protein was down-regulated,and the expression of P-AKT(0.14±0.07)protein was down-regulated.The expression level was significantly different from that of the single drug group(P<0.05),and the difference was statistically significant.Conclusion:(1)Prl-3 antibody can inhibit the proliferation and migration of lung cancer A549 cells and induce apoptosis in a concentration dependent manner.(2)PRL-3 antibody combined with PTX can significantly inhibit the proliferation and migration of lung cancer A549 cells,and induce apoptosis.The combination of two drugs on lung cancer A549 cells is stronger than the single drug group;(3)PRL-3 antibody combination The effect of PTX on the proliferation,migration and apoptosis of lung cancer A549 cells may be related to its down-regulation of PRL-3,PI3 k,and P-AKT proteins.