| Objective:To observe the inhibition effects of Vaccinium uliginosum(VU) extract on proliferation of H22hepatocellular carcinoma implanted mouse, and to illustrate its inhibition mechanism by means of assay of IL-2and TNF-a in serum of H22hepatocellular carcinoma implanted mouse, and expression of apaptosis related cyclin E and p27kip1.Methods:First, H22hepatocellular carcinoma implanted mouse model were established.60male KM mices,18-22g, were divided into6groups randomly as Normal (blank),5-Fu, Tumor model, VU high dossage(Group A), VU medium dossage(Group B), VU low dossage(Group C),10mices in each group, respectively. All the mouses in each group were treated with relevant drugs for10days, except Normal group. Body weight of mouses in each group were recorded per day. All the mice were forbidden to eat but allowed to drink for24h after the last drug treatment. Then,to weight all the mice. Taking every mouse’s eyeball for blood and separate the serum. The weights of tumor tissue, spleen and thymus were detected and the suppression ratio, index of spleen and thymus were calculated. IL-2and TNF-a in serum were assayed by ELISA, apoptosis were observed by DNA Ladder agarose gel electrophoresis, and the expression of cyclin E and p27kipl in tumor cell were detected by western blotting.Results:1. H22hepatocellular carcinoma implanted tumor mouse model were established.2. The mean weights of H22hepatocellular carcinoma implanted tumor are0.4134±0.14g,0.6106±0.18g,0.6832g,0.3565±0.11g,1.3736±0.59g, in Group A, B, C,5-Fu and model, respectively. The means of tumor tissue weight of Group A, B,5-Fu were significantly lower than that of model group, P<0.05, respectively. The inhibition ratios were,69.9%,55.6%and74.1%in Group A, B and5-Fu, respectively. The body weight of Group of A, B,5-Fu and model were8.2±1.9g,7.7±3.1g,8.4±2.2g,7.9±2.6g,8.6±2.7g, respectively, and there were no significant differences (P>0.05). The indexes of thymus were1.42±0.07,0.875±0.16,0.922±0.08,0.787±0.11,0.874±0.09in Group A, B, C,5-Fu and model, there was significant difference, Group A vs Group model (P<0.05), while ther was no significant difference between Group5-Fu and model(P>0.05). The indexes of spleen are15.68±0.97g、9.94±0.81g,9.64±0.75g,8.91±0.87g,9.71±0.38g, in Group A, B, C,5-Fu and model. There was significant difference between Group A and Group model (P<0.05), while there was no significant difference, Group5-Fu vs Group model (P>0.05).3. DNA ladder assay were carried out by1%agarose gel electrophoresis. The DNA Ladder phenomenon could be obviousely observed in sample extracted form Group A, while those from Group5-Fu and model were not.4. Cyclin E and p27kipl assay were carried out by Western bloting analysis. The relative ritioes of Cyclin E vs β-actin(inner standard) in Group5-Fu, A, B, C and model were0.3184±0.0047,0.6110±0.0284,0.5668±0.0285,0.5830±0.0451, respectively. The expression of cyclin E in Group A and5-Fu were significant lower than that in Group model (P<0.05). The relative ritioes of p27kip1in Group5-Fu, A, B, C and model were0.4781±0.0081,0.7113±0.0160,0.4344±0.0123,0.2297±0.0188,0.2343±0.0029, respectively. The expression of p27kip1in Group A is significant higher than that in Group model (P<0.05).5. The contents of IL-2in serum of Group blank,5-Fu, A, B, C and model were7.18±0.38ng/ml,3.63±2.07ng/ml,5.73±1.48ng/ml,6.03±1.56ng/ml,5.68±1.72ng/ml and5.73±2.07ng/ml, respectively. The level of IL-2in Group5-Fu is significantly lower than that in Group blank(P<0.05).But,there were no significant differences among other groups (P>0.05).6. The contents of TNF-a in serum in Group blank,5-Fu, A, B, C and model were1.33±0.87ng/ml,2.03±1.41ng/ml,1.83±1.08ng/ml,2.43±1.08 ng/ml,2.63±1.24ng/ml and2.53±0.66ng/ml, respectively. There were no significant differences (P>0.05).Conclusion:The extract of Vaccinium uliginosum could inhibit the proliferation of H22hepatocellular carcinoma, increase the index of thymus and spleen in mice with inplanted H22tumor. It also could be induced H22hepatocellular carcinoma cell apoptosis, it might be by means of down-regulating cyclin E level and up-regulating p27kip1level in tumor cell. The effects of Vaccinium uliginosum extract on IL-2and TNF-a in serum have not being observed in H22hepatocellular carcinoma inplanted KM mice. |
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