The Mechanism Of Killing Effects Against Leukemia Cell By Total Flavones Of Oldenlandia Diffusa Willd(HDE)Combined With Perifosine

Section One:HDE Induced Apoptosis of Lleukemia Cells Line Kasumi-1Objective To observe the inhibition and apoptosis about Kasumi-1 cells induced by total flavones of oldenlandia diffusa willd(HDE),so as to provide guidance for the clinical therapy of leukemia.Methods MTT colorimetric assay was used to examine the growth inhibition on leukemia Kasumi-1 cell by HDE;The cell morphological changes were observed using Hoechst33258 staining;The apoptosis rate was analyzed by Annexin V/PI flow cytometry;The expression of Caspase-3,Caspase-8,Caspase-9,PARP and Bcl-2 family,IAPs family and MAPKs survival signaling pathways were also examined using western-blot analysis.Results Exposure of Kasumi-1 cells treated with(0.125-2mg/mL)of HDE for 24 hours and 48 hours,the IC50 values were 0.07mg/ml and 0.059mg/ml;The apoptosis rate were analyzed by Annexin V/PI flow cytometry showed the early-apoptosis rate of 24 hours were 5.48%?7.53%?10.75%?22.30%?28.00%?32.40.Hoechst33258 staining showed the typical appearance of cell apoptosis after the cells were treated with HDE;Western blot showed an activation of Caspase-3,Caspase-8,Caspase-9,PARP and that down-regulation of MCL-1 proteins was observed in Kasumi-1 cells after treatment with HDE.Conclusions HDE could significantly inhibit the growth of Kasumi-1 cells in a time-and concentration-dependent manner,and HDE also could induces apoptosis in human leukemia Kasumi-1 cells,and the mechanisms associated with mitochondrial signaling pathway.Section Two:HDE Combined with Perifosine Induced Apoptosis of Leukemia Cells Line Kasumi-1Objective To study the synergistic killing effects of HDE combined with Perifosine in leukemia cell line Kasumi-1,and to explore its mechanisms,for the sake of providing new insights for the clinical therapy of leukemia.Methods MTT assay and flow cytometry were used to evaluate cell growth inhibition and apoptosis of HDE and Perifosine respectively or their combination;Hoechst33258 staining was used to observe the morphological changes;The expression of Caspase-3,Caspase-9,PARP,p-p65,p65 which were associated with apoptosis were examined by Western blot.Results HDE combined with Perifosine could significantly inhibit the growth of Kasumi-1 cells,and the apoptosis rate was further confirmed by flow cytometry analysis;Hoechst33258 staining showed more significant morphological changes and more apoptosis in combination treatment;Western blot showed an activation of Caspase-9,-3,PRAP,Cyt-C.HDE combined with Perifosine could down-regulate the expression of p-Akt and p-p65.Conclusions HDE inhibits the growth of Kasumi-1 cells synergistically with Perifosine and that maybe conducted by the PI3k/Akt/NF-kB signaling pathway.