Researching On Antitumor Effect And Mechanism Of Polysaccharide ? From Bee Pollen Of Carthamus Tinctorius L.on H22 Tumor-Bearing Mice

Objectives:To research the anti-tumor effect and mechanism of Polysaccharide I from Bee Pollen of Carthamus tinctorius L.(PBPCI)in vivo.Methods:Using Bee Pollen of Carthamus tinctorius L as raw material,the total polysaccharide was extracted by water and precipitated with alcohol.After deproteinization,polysaccharide was separated and purified by DEAE-52 cellulose column chromatography and Saphadex-G75 column chromatography to obtain PBPCI.I.Sixty BALB/c female mice were randomly divided into six groups:normal control group,tumor model group,cyclophosphamide(CTX)group and PBPCI low,medium and high dose groups,with 10 mice in each group.Except the normal control group,the other groups were injected with a cell density of 1 × 10~7/ml cells 0.2 ml of H22 hepatoma cells in the right forelimb of the mice to establish H22 liver tumor model..Low,medium and high dose groups were administered with PBPCI 100 mg/kg,200 mg/kg,high,400 mg/kg,and CTX group was subcutaneously injected with CTX 30 mg/kg daily.Normal control group and tumor model group were given equal volume normal saline for 15 days.After the last administration,the mice of each group was fasted for 12 hours(wasn't fasted water).Then taking orbitlal blood,serum was separated.The serum levels of IL-1 a,IL-6,IFN-y,TNF-a IL-2,VEGF were measured by enzyme-linked immunosorbent assay(ELISA).After weighing body weight,mice were sacrificed,the tumor were removed,the spleen and thymus were taken and weighed.The tumor inhibition rate and Immune organ index were calculated.HE staining was used to observe the changes of cell morphology in different groups.Real-time quantitative PCR(RT-qPCR)was used to observe the relative expression of bax,p53,c-myc and bcl-2 genes.The protein expression levels of Bax,P53,C-myc and Bcl-2 were determined by Western blot.Results:PBPC I couldsignificantly inhibt tumor growth Compared with model group,the inhibitory rate of CTX group and PBPC I low,medium and high dose on H22 liver cancer were 67.93%,33.48%,45.69%,57.86%respectively..At the same time,PBPC I low,medium and high dose group could significantly improve the body weight and immune organ index of tumor-bearing mice,and significantly increase the serum content of IFN-?,IL-2 and TNF-? in tumor-bearing mice,as well as reduce the content of IL-1?,IL-6,VEGF in serum.HE staining results showed that with the increase of PBPCI dose,the tumor tissue was loose,the cells were reduced,and some apoptotic features of nuclear condensation,nuclear fragmentation and nuclear fission were observed.RT-qPCR experiments confirmed that PBPCI can down-regulate c-myc and bcl-2 gene transcription and up-regulate bax and p53 gene transcription.At the same time,Western blot experiment showed that the low,middle and high doses of PBPCI could decrease the expression of Bcl-2 and C-myc proteins in tumor tissues,and increase the expression of Bax and P53 proteins.Conclusion:PBPCI has obvious anti-tumor effect on H22 tumor-bearing mice,and its mechanism may promote tumor cell death by protecting immune organs,regulating immune factors,VEGF levels,and regulating the expression levels of Bcl-2,C-myc,Bax and P53.Promote tumor cell apoptosis and inhibit tumor proliferation,proliferation and growth.