Effects On Migration,Proliferation And Mechanisms Of Human Hepatocellular Carcinoma HepG2 Cells Induced By Mehtylnaltrexone

Part ? The inhibition effects of methylnaltrexone on biological behaviors ofliver cancer Hep G2 cellsObjective To observe the proliferation and migration of liver cancer cells Hep G2 influenced by methylnaltrexone(MNTX)Methods The liver cancer cells Hep G2 were cultivated in a period of logarithmic phase.They were divided respectively into three groups with different dose of MNTX(100?mol/L,1000?mol/L)was incubated together with liver cancer Hep G2 cells for 24 h,48h and 72 h respectively.Normal saline in the same volume was used as blank control group.We used CCK8 assay to explore effects of MNTX on cell proliferation.And transwell assay was used to explore influences of MNTX on cell migration.Results CCK8 assay(48h)showed that there was obvious difference between A values of different doses,groups and time.(F group =127.136,P group= 0.000).There was a tendency between the same dose groups with time changing in A values(F time= 230.984,P time= 0.000).And there was significant difference between high-dose group and low-dose group(F dose=18.15,Pdose= 0.000).There was no significant effect of proliferation between low-dose treatment group and blank control group(P =0.112).Transwell assay demonstrated that the high-dose group played a significant role in down-regulating migration of Hep G2 cells at 72 h.Conclusions MNTX could be significantly inhibit proliferation and migration of hepatoma Hep G2 cells.Part ? The mechanisms on inhibiting liver cancer Hep G2 cells growthinduced by methylnaltrexoneObjective To discuss the biological mechanisms on inhibiting liver cancer Hep G2 cells growth induced by methylnaltrexoneMethods Cell culture with different dose of MNTX(100?mol/L,1000?mol/L)was incubated together with liver cancer Hep G2 cells for 72 h.Normal saline in the same volume was used as blank control group.RT-PCR was administrated for detection of the gene expression of VEGF and MMP-9 in Hep G2 cells.The expression of protein of VEGF and MMP-9 gene was detected by western blot.Nude-mouse transplanted tumor model was established to detect inhibition effect of growth by MNTX.Results After 72 hours,the relative expression of VEGF m RNA in control group was 1.00 ± 0.03,which is significantly higher than that of treatment group of 0.38±0.09(t=9.35,P =0.003).And the relative expression of MMP-9 m RNA in control group was 0.96 ± 0.17,which is significantly higher than that of treatment group of 0.75±0.22(t=1.243,P =0.000).After 72 hours,compared with blank control group,the expression of protein of VEGF and MMP-9 in liver cancer cells Hep G2 in high-dose treatment group was significantly lower than that in control group(P < 0.05).Conclusions The results of RT-PCR and western blot showed that compared with control group,MNTX treatment groups could down-regulate the expression of gene and protein of liver cancer cells Hep G2(P < 0.05).And the expression of gene and protein of VEGF and MMP-9 in liver cancer cells Hep G2 in high-dose treatment group was significantly lower than that in low-dose group(P < 0.05).Part ? The growth inhibition effects on nude mice induced bymethylnaltrexoneObjective To establish nude mice transplanted tumor model and explore tumorigenicity of nude mice transplanted of tumor liver cancer Hep G2 cells influenced by methylnaltrexone in vivoMethods The liver cancer cells Hep G2 were cultivated and digested on a period of logarithmic phase.We collected and adapted them with 1*107 cells/ml,200 ul in total.Those reagents were prepared into Hep G2 cell suspension for inoculating nude mice and then they were injected to oxter of nude mice as nude mice transplanted tumor model.We divided the nude mice into two groups,blank control group and mere MNTX group.The mere MNTX group nude mice were injected from 8 days,and nude mice of control group were injected with normal saline in the same volume at the same time.The nude mice were feed for 29 days.Then we observed tumor formulation of each nude mouse.They were peeled completely off the skin for tumor organization,weighed and photographed at 30 th days.Tumor formation rate=(the number of tumor formation nude mice-the number of treatment nude mice)×100%.Results Nude mouse transplanted tumor experiment showed that the general condition of nude mice in MNTX group was significantly greater than that in control group.The weight of nude mice in control group was significantly lower than that in MNTX group(t=2.49,P =0.032).After injection,the weight of tumor in MNTX group was significantly lower than that in control group(t=5.45,P =0.000)Conclusions MNTX could suppress liver cancer Hep G2 cells in vivo with a suppression rate of 59.23%,which accords to the results in vitro.