The Effect Of Insulin On Chemotherapeutic Drug Sensitivity In Human Lung Adenocarcinoma Cells A549 In Vitro

Objective To investigate the effect and mechanism of insulin, as a metabolic promoter, on chemotherapeutic drug DDP sensitivity in human lung adenocarcinoma cells A549, and then offer evidence for clinical application.Methods Human lung adenocarcinoma cells A549 were cultured and inoculated into the wells. The chemotherapeutic drug was DDP. Insulin was used as inductor. Cell activity and cell metabolism were assessed by MTT assay to reflect the cytotoxity of the anti-tumor agent and the effects combined with insulin. Cell cycle of every groups were examined by floweytometry (FCM). The expressions of CyclinA and CyclinD1 were detected by Western Blot to analyze the chemosensitizationmechanisms of insulin.Results1.In the test of finding the revulsive concentration of insulin, we found thatinsulin promoted the cell metabolism with the concentration 2～32mU/ml. Concentration groups of 4mu/ml,8mu/ml and 16mu/ml had significant differences compared with other groups(P<0.01).2.In the test of finding the best inducement time of insulin, insulin with concentration of 8mu/ml were added.The result showed that the best inducement time of insulin were 8～16h and they had significant differences compared with other groups(P<0.01).3.In the test of finding the chemotherapeutic drug concentration, the analysis showed that inhibitory rate had a linear correlation with DDP. Based on the result of MTT assay, we selected the concentration of 30% inhibitory rate(IC3o) as the concentration of results:DDP IC30=36.87μg/ml.4.In the test of insulin enhancing the chemosensitization on human lung adenocarcinoma cells A549, insulin(8mu/ml) added before adding DDP 8～16h enhanced the chemocytotoxity on Human lung adenocarcinoma cells A549 as indicated by MTT colorimetry. The best suitable concentration of insulin were 2.0～16.0mu/ml, which can promote the sensibility of DDP.5.The percents of S phase of insulin from 4h to 20h were different and the increased S cell cycle were observed in phase synchronized A549 cell treated with 8mu/ml of insulin. Time groups of 12 h had significant differences compared with other groups. Insulin can increase the S phase arrest induced by DDP.6.Insulin treatment enhanced the expression of CyclinD1, while DDP treatment enhanced the expression of CyclinA.Conclusion1.As a reversible metabolic promotor , proliferation of human lung adenocarcinoma cells A549 can be induced by insulin in vitro, at the condition of insulin(4.0～16.0mu/ml, 8～16h). Moreover, an increased S cell cycle is observed in phase synchronized A549 cell treated with 8mu/ml of insulin. The increased expression of cyclinD1 plays an important role in the enhancement.2.Insulin could sensitize A549 to the anticancer activity of DDP in vitro. The key of improving the chemotherapeutic effect is the selection of the inducing occasion of resulsant and concentration. It is the best inducing state when insulin is used before adding chemotherapeutic drug 8～16h,concentration 2.0～16.0mu/ml. The mechanisms may be related to increasing the S phase arrest and the effect can be synergistic with DDP.3.The increased expression of cyclinA plays an important role in the S phase arrest induced by DDP.4.It is possible to increase the growth and metabolism of A549 first so as to enhance the chemosensibility, and then administer chemo- therapeutic agents, thus improving their therapeutic effects.