Effect of stevia polysaccharide on apoptosis of diffuse large B-cell lymphoma cell line U2940 and its molecular mechanism
Objective To investigate the effect and mechanism of stevia polysaccharide on large B lymphoma cell line U2940.Methods U2940 cells were treated with stevia polysaccharides at the concen-tration of 0-64 μg/ml,and the proliferation activity of U2940 cells was determined by cell counting kit-8(CCK-8).The cytotoxicity of stevia polysaccharides was confirmed by lactate dehydrogenase(LDH)release assay,JC-1 staining was used to observe mitochondrial membrane potential,flow cytometry was used to detect apoptosis rate,and real-time quantitative reverse transcriptase-polymerase chain reaction(RT-qPCR)and Western blotting were used to determine gene transcription and expression level.In addi-tion,the inhibitory effect of stevia polysaccharide on tumor growth and survival time of diffuse large B-cell lymphoma(DLBCL)-bearing mice were also observed.Paired sampl t test andx2 test were used for statisti-cal analysis.Results The proliferation activity of U2940 cells in the experimental group was lower than that in the control group.After 24 h,16μg/ml and 64μg/ml stevia polysaccharide inhibited the prolifera-tion of U2940 cells less than that in the control group(0.24±0.01 vs.0.31±0.01,t=6.31,P＜0.05;0.20±0.03 vs.0.31±0.01,t=4.42,P＜0.05),and the apoptosis of U2940 cells was induced by this drug.The transcription of Ras/Raf/mitogen extracellular kinase(MEK)/extracellular signal-regulated kinase(ERK),phosphatidylinositol 3 kinase(PI3K)/protein kinase B(Akt)/mammalian target of rapamy-cin(mTOR)and nuclear factor-KB(NF-κB)pathway and downstream bcI-2,bcI-XL and MCL-1 genes in the experimental group was lower than that in the control group(0.44±0.02 vs.1.00±0.06,t=6.11,P＜0.05;0.51±0.09 vs.1.00±0.03,t=3.77,P＜0.05;0.39±0.06vs.1.00±0.04,t=2.78,P＜0.05).The expressions of cysteinyl aspartate-specific protease(Caspase)-3,Caspase-8 and Caspase-9,which are directly related to apoptosis,in the experimental group were higher than those in the control group,(3.69±0.08 vs.1.00±0.03,t=6.75,P＜0.05);(2.39±0.10 vs.1.00±0.02,t=6.45,P＜0.05;3.67±0.08 vs.1.00±0.03,t=6.32,P＜0.05);The tumor inhibition experiment showed that compared with the control group,stevia polysaccharide could inhibit the growth of tumor tissue and prolong the survival time of tumorous mice for(54 days vs.14 days,t=4.66,P＜0.05).Conclusion Stevia polysaccharide can inhibit the proliferation and induce apoptosis of U2940 cells by affecting RAS/RAF/MEK/ERK,PI3K/Akt/mTOR and NF-κB signaling pathways,and has a significant anti-DLBCL effect.
Stevia polysaccharidesDiffuse large B-cell lymphomaSignaling pathwayProliferative activityCell apoptosis
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