The Effect Of Puerarin Combined With 5-fluorouracil On Hepatocellular Carcinoma SMMC7721 Cells

Part I Effects of puerarin exposure on the cells growth and apoptosis of the hepatocellular carcinoma cell line SMMC7721Backgroup and Objective:Recently, some research has showed that puerarin may have anti-carcinogenic effects, but no research has showed that puerarin may have anti-hepatocellular carcinoma effects; study in the part I investigated the effects of puerarin against hepatocellular carcinoma cell line SMMC7721, whether combination effect surpass the effect of 5-FU or puerarin alone against hepatocellular carcinoma SMMC7721.Methods:The human hepatocellular carcinoma cell line SMMC7721 was used in this research. Firstly, the cells were cultured. Then, the growth inhibition of puerarin or 5-FU alone or combined on SMMC7721 cells was determined by measuring CCK-8 dye absorbance in living cells. The median-effect principle was used to evaluate the combined effects. Apoptotic morphological features and percentage were detected by Hoechst 33258 staining and annexin V/PI apoptosis kit, respectively.Results:The mean (SD) growth inhibition rate reached 95.56 (0.81)% at a concentration of 6400 μM,75.91(3.54)% at 640 μM of 5-FU. The combination index (CI) values were< 1 when the fractions affected were between 0.2555 and 0.7420. Bright chromatin condensation and nuclear fragmentation could be found in the treatment groups, while the nucleus stained a weak homogeneous blue in the control cells. Puerarin or 5-FU alone significantly promoted apoptosis compared to the control group. Percentage of apoptotic cells increase markedly in combination group than the groups alone in vitro. Test results on the molecular level showed that the expression of Bax protein in combined group was the highest, while the Bcl-2 is the lowest.Conclusions:Our results demonstrated that puerarin has the activity against hepatocellular carcinoma cell line SMMC7721. Puerarin inhibited the growth and promoted the apoptosis of hepatocellular carcinoma cell SMMC7721.Part 2 Effect of puerarin combination with 5-FU on tumor development in vivoObjective:The part two study whether that puerarin or 5-FU had the activities against hepatocellular carcinoma cell line SMMC7721, respectively. Puerarin combination with 5-fluorouracil (5-FU) may be an important regimen in liver cancer therapy. Our aim was mainly to determine whether combination effect surpass the effect of 5-FU or puerarin alone against hepatocellular carcinoma SMMC7721 in vivo.Methods:The SMMC7721 cell tumor xenograft model was established in nude mice. Puerarin or 5-FU alone or combined were injected into the nude mice, and the tumor growth inhibition rates were evaluated by monitoring tumor volume and weight. In situ apoptosis detection kit was used to detect apoptosis by the TUNEL assay, morphological changes were observed by HE staining, and side effects were observed.Results:Percentage of apoptotic cells increase markedly in combination group than the groups alone in vivo. The tumor volume inhibition rates were 70.58 and 76.26% in puerarin or 5-FU group alone. The tumor weight inhibition rates were 46.20 and 49.86%. The volume and weight tumor inhibition rates were 93.11,75.21% in combination group. In addition, no marked differences were observed in serum marker among the groups.Conclusion:There is a marked increase in the growth inhibition rate and apoptotic percentage in puerarin combination with 5-FU, without liver and renal toxicity occurring. These results suggest that there is a synergistic effect of puerarin combination with 5-FU on hepatocellular carcinoma SMMC7721.