Dihydroartemisinin Induces Apoptosis In Acute Myeloid Leukemia Cells By Inhibition Of PTEN/AKT Pathway

Objective Acute myeloid leukemia(AML)is the most common type of leukemia in adults.Currently,the commonly used chemotherapy regimens and hematopoietic stem cell transplantation in clinical practice can alleviate and cure some patients,but there is a high rate of drug resistance and high recurrence.The risk of long-term disease-free survival of patients has not been significantly improved,and it is still very important to develop new drugs for the treatment of acute myeloid leukemia.In this study,the acute myeloid leukemia cell lines Kasumi-1,KG-1 and HL60 were used as research objects,and the activity of dihydroartemisinin(DHA)to inhibit the activity and pro-apoptotic ability of acute myeloid leukemia cell lines was observed.Explore its pro-apoptotic mechanism,and provide evidence for the development of drugs with clinical therapeutic value.Methods1.CCK-8 method was used to detect the effect of DHA on the viability of Kasumi-1,KG-1 and HL60 cells.2.PI staining method to observe the inhibitory effect of DHA on Kasumi-1 cells.3.Cell immunohistochemical method was used to detect the expression level of Cleaved-Caspase3 protein in Kasumi-1 cells.4.Flow cytometry was used to detect the apoptosis induction effect of DHA on HL60 cells.5.Add pan-apoptosis inhibitor Z-VAD-FMK to detect its effect on Cleaved-Caspase3 protein expression level and cell viability in Kasumi-1 cells.6.Western Blot method was used to analyze the changes of apoptosis-related proteins and protein expression levels in PTEN / AKT pathway.Results1.The effect of DHA on the growth inhibition of AML cell lines: CCK-8 method found that DHA could inhibit the viability of three AML cells in a dose-dependent manner.PI staining showed that DHA could induce AML cell death in a dose-dependent manner.2.DHA induced apoptosis of AML cells: Cell immunofluorescence detection found that compared with the control group,the expression level of Cleaned-Caspase3 protein was significantly increased in AML cells treated with DHA.Flow cytometry apoptosis detection showed that,compared with the control group the DHA treatment group can induce AML cell apoptosis in a dose-dependent manner,especially in the late stage of apoptosis.Western Blot detection found that after DHA acted on AML cells,the expression levels of pro-apoptosis-related proteins Cleared-caspase3 and c-PARP increased.3.DHA-induced AML cell apoptosis can be recovered by Z-VAD-FMK reagent: Cell immunofluorescence found that pan-apoptotic inhibitor Z-VAD-FMK can attenuate increased expression of Cleaved-caspase3 protein in Kasumi-1 cells after DHA treatment.CCK-8 detection found that the pan-apoptotic inhibitor Z-VAD-FMK can significantly restore the vitality of Kasumi-1 cells after DHA treatment.4.DHA-induced apoptosis of AML cells was accompanied by changes in PTEN and p-AKT protein: Western Blot detection found that compared with the control group,DHA up-regulated the expression level of PTEN protein in Kasumi-1 cells in a dose-dependent manner,down-regulate the expression level of p-AKT protein and reduce the expression ratio of p-AKT / AKT protein.Conclusion1.DHA can inhibit the proliferation of AML cells in vitro by inducing apoptosis.2.PTEN / AKT pathway may be involved in the mechanism of DHA against AML.