Parthenolide Promotes Mitochondria-mediated Apoptosis Through Induction Of Oxidative Stress In Gastrointestinal Stromal Tumor Cells

OBJECTIVE:To explore the effect of parthenolide(PTL)-induced oxidative stress in GIST-T1 cell line to promote mitochondrial-mediated apoptosis of GIST cells,and provide an experimental basis for the clinical application of parthenolide in the treatment of GIST.Methods:The human GIST-T1 cell line was selected for in vitro culture for 48 hours,and GIST-T1 cells were cultured for 48 hours using PTLs of 0μM,4μM,8μM,16μM,32μM and 6μM,and the survival rate of each group of cells was calculated using the CCK-8 experiment,and Obtained half of the inhibitory concentration(IC50=20p.M).The experiment was divided into three groups:IC50,IC50/2 and IC50/4.The control group was cultured with McCoy 5A medium containing 0.2%DMSO.The scratching method was used to detect the proliferation and migration ability of GIST-T1 cells in each group after 48 hours of treatment with parthenolide.Using 2’,7’-dichlorofluorescein diacetate(DCFH-DA)method to detect the level of reactive oxygen species(ROS)in each group of cells.Flow cytometry and TUNEL staining were used to detect the apoptosis level of GIST-T1 cells in each group after 48 hours incubation of parthenolide.Western blotting was used to evaluate the expression of Bax and Bcl-2 and the expression of cytochrome c and cleaved caspase(Caspase-3,-8 and-9)after incubation of parthenolide for 48 hours.JC-1 method was used to determine the mitochondrial membrane potential after 48 hours incubation of parthenolide,and the ultrastructure was further examined using transmission electron microscopy(TEM).Results:1.The results of CCK-8 and scratch method showed that parthenolide inhibited the proliferation and migration of GIST-T1 in a dose-dependent manner,and the inhibitory effect increased with the increase of drug concentration(P<0.01).2.After 48 hours of treatment with parthenolide,the reactive oxygen level in GIST-T1 cells increased,and it was proportional to the parthenolide concentration(P<0.01).3.The results of flow cytometry and TUNEL staining showed that parthenolide induced apoptosis,the higher the concentration,the higher the apoptosis rate(P<0.01).4.The results of JC-1,transmission electron microscopy(TEM)and Western blotting showed that the mitochondria of GIST-T1 cells treated with parthenolide induced apoptosis.5.Western blot results showed that parthenolide increased the Bax/Bcl-2 ratio(P<0.05)in a dose-dependent manner,while parthenolactone activated cytochrome c and caspase(Caspase)activity,Further enhance the expression of endogenous apoptosis factors Caspase-3 and 9(P<0.01),but no effect on exogenous apoptosis factors Caspase-8.Conclusion:1.Parthenolide inhibits the proliferation and migration of gastrointestinal stromal tumor cells GIST-T1 in a dose-dependent manner.2.Parthenolide affects the change of mitochondrial membrane potential of tumor cells,allowing cells to enter the apoptosis.3.Parthenolide can induce tumor cell apoptosis through oxidative stress-related pathways,thereby inhibiting tumor cell proliferation and migration.