Effects Of ZYY48-induced Autophagy And Apoptosis In Burkitt's Lymphoma CA46 And Acute Myeloid Leukemia HL-60 Cell Lines

Objective This study was focused on the investigation of the effects of Emodin derivative ZYY48 on human Burkitt's lymphoma cell line CA46 and acute myeloid leukemia cell line HL-60.The roles of ZYY48-induced apoptosis and autophagy were highlighted in this study.Methods 1.MTT assay and colony formation assay were applied to access the cell growth inhibition in CA46 and HL-60 cells after ZYY48 treatment,respectively.The cell viability of primary mononuclear bone marrow from acute leukemia,and thrombocytopenia patients,and healthy donors was also measured in this study.2.The ultrastructure changes of cellular morphology in CA46 and HL-60 cells were identified by electron microscope.4.The morphology changes of autophagy in both cell lines were observed by fluorescence microscope after MDC staining.5.The expression level Beclin-1 m RNA was performed by real-time quantitative PCR(RQ-PCR).6.Western blot was employed to measure the expression levels of autophagy related proteins including LC3-I/II and Beclin-1.The same assay was also performed to evaluate the expression changes of apoptosis related proteins,such as PARP,AKT,p-AKT,c-Myc,NF-?B,p-NF-?B and MAPKs.Result 1.ZYY48 dose-and time-dependently inhibited CA46 and HL-60 cell proliferation with the average IC50 values of 1.82±0.13?mol/L and 1.03±0.14?mol/L,respectively.Primary acute leukemia cells were more sensitive to ZYY48 treatment compared with the primary mononuclear cells from thrombocytopenia patients and healthy donors.The average IC50 value was 3.07±0.44?mol/L in leukemia cells,which was 1.93-and 2.50-folds lower than that in healthy donor specimens(5.89±0.47?mol/L)and thrombocytopenia(7.57±0.57?mol/L),respectively.2.ZYY48 inhibited colony formation and induced apoptosis in CA46 and HL-60 cells in a dose-dependent way.4.MDC staining result showed that a large number of different sizes of autophagic vacuoles appeared in cell cytoplasm after ZYY48 treated leukemia cells.5.The Beclin-1 m RNA expression levels in both CA46 and HL-60 cell lines were enhanced as the concentration of 2?mol/L of ZYY48 was added in the system.6.Western blot results showed that when increasing doses of ZYY48,the apoptosis-related proteins and the marker proteins of autophagy were markedly changed.It down-regulated the expression levels,such as Ras,ERK1/2,p-ERK1/2,p-Raf,p38 MAPK,and up-regulated the molecules,such as p-p38 MAPK,p-HSP27,p-ATF-2,LC3-I/II and Beclin-1,which lead to the induction of apoptosis and autophagy in both cell lines.However,the following proteins including AKT,p-AKT,c-Myc,NF-?B and p-NF-?B were not grossly affected.Conclusion Emodin derivative ZYY48 could efficiently inhibit proliferation in human Burkitt's lymphoma cell line CA46 and acute myeloid leukemia cell line HL-60.ZYY48 also induces cell apoptosis and autophagy at the same time,which can kill the cells of CA46 and HL-60.ZYY48 induces CA46 and HL-60 cells apoptosis through triggering activation of ERK/MAPK and p38 MAPK signaling pathway.ZYY48 could efficiently inhibit proliferation in primary leukemic cells,which did not significantly affect the viability of normal cells compared with the leukemia cells.