Research of dihydroartemisinin on anti-colorectal cancer by regulating MAPK/PI3K/Akt signaling pathway
Objective:To investigate the anticancer activity of dihydroartemisinin(DHA)on RKO cells in vivo and its mechanism in vitro.Methods:40 RKO tumor-bearing mouse models were established in specific pathogen-free male nude mice and randomly divided into model group,cisplatin group,high dose DHA(200 mg/kg)group,and low dose DHA(100 mg/kg)group,with 10 mice in each group.The body weight,tumor volume,tumor mass,tumor inhibition rate and serum,tumor necrosis factor-α(TNF-α)level of each group were compared.Hematoxylin-eosin(H&E)staining was used to observe tumor histopathological changes in nude mice of each group.MTT assay was used to detect the effects of DHA at different concentrations(25,50,100 μmol/L)on the proliferation of RKO cells within 0,24,48,and 72 h.Flow cytometry and Hoechst 33258 were used to detect the effects of DHA(0.50,95,150 μmol/L)on the cell cycle and apoptosis of RKO cells after 48 h intervention.Wound healing assay was used to observe the effect of 95 µmol/L DHA on cell migration.Real Time Quantitative PCR(RT-qPCR)was used to detect the effect of DHA on the mRNA expression levels of Caspase-3,Caspase-9,Bcl-2 and Bax.Western blot was used to detect the effect of DHA on the expression levels of Caspase-3,Caspase-9,Bcl-2,Bax,p38-MAPK,p-p38-MAPK,PI3K,p-PI3K,Akt,p-Akt and MMP-9.Results:Tumor volume and mass were significantly reduced and the serum level of TNF-α was lowered by 200 mg/kg DHA,and the tumor inhibition rate was 41.45％.The inhibitory effect on RKO cell proliferation was enhanced with increasing concentration of DHA.RKO cell cycle was arrested in the G2/M phase by(50,95,150 μmol/L)DHA,and the effect of promoting RKO cell apoptosis was enhanced with the increase of DHA concentration.The migration ability of RKO cells was significantly reduced after intervention with 95 μmoUL DHA for 12 and 24 h(P＜0.01).The mRNA expression levels of Caspase-3 and Caspase-9 in RKO cells were significantly up-regulated by 95 µmol/L DHA(P＜0.01),and Bax/Bcl-2 mRNA expression ratio was also significantly up-regulated(P＜0.05).The protein expressions of cleaved-Caspase-9/Caspase-9 and Bax/Bcl-2 in RKO cells were increased by 95 μmol/L DHA(P＜0.01),while the protein expression levels of MMP-9,p-P38 MAPK,p-PI3K,p-Akt and Akt were decreased(P＜0.01).Conclusion:DHA has a good inhibitory effect on colorectal cancer,and its mechanism may be to trigger the endogenous apoptosis of RKO cells to prevent their proliferation and migration through inhibiting the MAPK/PI3K/Akt signaling pathway.
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