Growth Inhibition Of Hepatocarcinoma Hep3B Cells By β - Elemene Milk And Its Mechanism

β-elemene is a novel non-cytotoxic antitumor drug,which is isolated from the traditional Chinese medicinal herb Rhizoma zedoariae.It is reported thatβ-elemene has anticancer effects in vitro and in vivo and has substantial clinical activity against various tumors,Meanwhile no bone marrow suppression and drug resistance have been observed in the clinical studies;on the contrary,patient immunity was improved during the therapy withβ-elemene.The present study was to investigate the anti-cancer effect ofβ-elemene on Human hepatocellular carcinoma Hep3B cell and human peripheral blood mononuclear cell（PBMC）,as well as reveal the underlying molecular mechanisms ofβ-elemene on Hep3B cell,which could provide with the evidences for its clinical use.The effects ofβ-elemene on cell proliferation were studied by using MTT and colony formation assay（CF）in vitro,meanwhile the 5-fluorouracil and the PBMC were chosen as the control respectively.DNA damage induced byβ-elemene on Hep3B cell was measured by single cell gel electrophoresis（SCGE）.Apoptosis was detected on Hep3B cell by AO/EB staining,DNA ladder and flow cytometric analysis.And the apoptosis-related mRNA level of Hep3B cell was determined by RT-PCR.The results of the anticancer effects ofβ-elemene in vitro showed that:β-elemene exhibited significant anti-proliferative effects on Hep3B cell(IC₅₀:about 26.7±0.3μg/ml for 48 h),and the IC₅₀on PBMC was about 55.3±4.3μg/ml respectively.The selectivity index（SI）ofβ-elemene was 2.07±0.16,which surpassed the one of 5-fluorouracil（0.85±0.07）.The results of the effects ofβ-elemene on apoptosis in Hep3B cell showed:Hep3B cells were stained with AO/EB and the morphological changes demonstrated thatβ-elemene could induce apoptosis.DNA ladder,a hallmark of apoptosis,was observed on an agarose gel inβ-elemene-treated Hep3B cells.Flow cytometric analysis confirmed thatβ-elemene induced apoptosis in Hep3B cell in a dose- and time-dependent manner.The noticeable cell cycle arrest in G₀/G₁ phase was showed byβ-elemene also.The ability ofβ-elemene to enhance the accumulation of intracellular reactive oxygen species and the loss of mitochondrial membrane potential（△Φm）was examined with 2`7`-dichlorofluorescin diacetate（DCFH-DA）and Rhodamine123 by FCM.As well,β-elemene could increase the pro-Caspase3 mRNA level by RT-PCR,which further demonstrated thatβ-elemene could induce the apoptosis in Hep3B cells.In conclusion,β-elemene exerted the anti-cancer activity on Hep3B cell in vitro and could induce the apoptosis probably involved activation of mitochondrial cell signal pathways and the caspase cascade.