The Effects And Mechanisms Of Emodin On Inducing Leukemic U937 Cells Apoptosis And Growth Inhibition

【Objective】To investigate the effects of Emodin on proliferation inhibition and apoptosis induction in human myeloid leukemia cell line U937; on related proteins and mRNA expressions; and on the protein regulation of PI3-K/Akt signal pathway.【Methods】U937 cells were exposed to Emodin at different dosages. Proliferation inhibition was detected by both MTT assay and clone formation assay. The ability of Emodin to induce apoptosis of U937 cells was examined by acridine orange/ethidium bromide (AO/EB) double staining,MitoCapture apoptosis detection,flow cytometry and DNA fragmentation. The activation of caspase-3 was detected by flow cytometry. The expression of Bcl-2,Bax,c-myc,h-TERT,pim-2,TGF-β1,survivin,P21,Mcl-1and wtp53 gene was detected by RT-PCR; protein expressions of Bcl-2,Bax,h-TERT,cyclinD1,procaspase-3,PARP,Akt,pAkt,NF-κB(p65)and pNF-κB(pp65)were detected by Western-blot.【Results】Emodin remarkably inhibited the U937 cells proliferation, with an IC50 value of 30μM after 72 hours of treatment. Staining of cells with AO-EB revealed that Emodin induced nuclear chromatin condensation and nuclear fragmentation. Concomitantly, apoptosis was induced in U937 cells, as measured by detections of changed mitochondrial transmenbrane potential,sub-G1 apoptotic peak and DNA fragmentation. These showed that Emodin induced apoptosis in U937 cells in dose-dependent manners. Compared with the control group, the percentage of the treated cells in G0/G1 phase increased, while that in S phase decreased. Moreover, the activated caspase-3 in treated U937 cells increased. RT-PCR and/or Western-Blot showed the expressions of Bcl-2,c-myc,h-TERT,pim-2,Survivin,procaspase-3,cyclinD1,pAkt,NF-κB(p65) and pNF-κB(pp65) in treated U937 cells decreased, but the expressions of Bax,wtp53,P21 and TGFβ1 increased.【Conclusion】Emodin could efficiently induce proliferation inhibition and apoptosis in U937 cells. Multiple pathways,such as Bcl-2,Bax,c-myc,h-TERT,pim-2,TGF-β1,survivin,P21,wtp53,cyclinD1,procaspase-3,PARP,pAkt, NF-κB(p65),pNF-κB(pp65) may involved in these processes.