The Study Of Inducing Apoptosis Of Tumor Cell And Inhibition Of Tumor Angiogenesis By Celastrus Orbiculatus

Objectives1. To study the effect of celastrus orbiculatus terpenoid on mouse melanoma B16BL6 cell proliferation and apoptosis in vivro and to establish its foundation in aspect of antitumor activity to melanoma.2. To evaluate the effect of celastrus orbiculatus terpenoid on human hepatoma 7721 cell proliferation and the influence of celastrus orbiculatus terpenoid to homogeneity and alloplasm adhesion in the dose of noncytotoxicity, and further study the mechanism of celastrus orbiculatus terpenoid in adhesion ,invasion and migration.3. To evaluate the effect of celastrus orbiculatus terpenoid on human umbilical vain endothelial cells proliferation and apoptosis and to study its possible mechanism of antiangiogenesis.Methods1. MTT assay was performed to study the effect of celastrus orbiculatus terpenoid on mouse melanoma B16BL6 cell, human hepatoma 7721 cells and human umbilical vain endothelial cells proliferation, respectively.2. Flow cytometer was used to test the effect of celastrus orbiculatus terpenoid on mouse melanoma B16BL6 cell and human umbilical vain endothelial cells apoptosis.3. To detect the adhesion ability of between tumor cell, tumor cell and extracellular matrix and tumor and HUVECs by the variation of optical density.4. Flow cytometer was used to dedect the change of MMP-2 protein in the human hepatoma 7721 cells and Bcl-2 protein in the human umbilical vain endothelial cells.5. Indirect enzyme-linked immunosorbent assay (ELISA) was used to detect the change of the vascular endothetial growth factor. Results1. The results of MTT assays demonstrated that different dose of celastrus orbiculatus terpenoid suppressed the mouse melanoma B16BL6 cell,human hepatoma 7721 cell and HUVECs proliferation .They had the dose-effect relationship.2. The results of flow cytometer assay showed that celastrus orbiculatus terpenoid could significantly induce the apoptosis of the mouse melanoma B16BL6 and HUVECs in the dose and time-dependent manner.3. Choose the noncytotoxicity dose of celastrus orbiculatus terpenoid to start experiments .The result showed that different dose of celastrus orbiculatus terpenoid could promote homogeneity adhesion between hepatoma 7721 cells.4. The experiment also indicated that different dose of celastrus orbiculatus terpenoid could obviously inhibit human hepatoma 7721 cell attaching to the matrigel and had a positive correlation between the adhesion inhibition ratio and the dose of celastrus orbiculatus terpenoid.5. The findings also showed that celastrus orbiculatus terpenoid could obviously suppress the human hepatoma 7721 cell attaching the HUVECs in a dose-dependent manner.6. The findings of flow cytometer showed that celastrus orbiculatus terpenoid could down regulate the expression of MMP-2 protein and it had the variability compared to the control(P<0.05). When the dose of celastrus orbiculatus terpenoid gradually increased, the value of FI gradually decreased in a dose-manner.7. Results from flow cytometer showed that different dose of celastrus orbiculatus terpenoid could down-regulated the expression of Bcl-2 protein and it had the special relationship of vascular endothelial cell apoptosis.8. The results of ELISA showed that compared with the control ,celastrus orbiculatus terpenoid could remarkably decrease the content of cytokine VEGF and had significant difference (P<0.05).And it also showed that the content of cytokine VEGF gradually decreased when the dose gradually increased and it was in a dose-dependent manner. Conclusion1. Celastrus orbiculatus terpenoid can inhibit the proliferation of mouse melanoma B16BL6 cell and induce its apoptosis and it has an obvious antitumor activity in the mouse melanoma and it,s deserved to study and research extremely.2. Celastrus orbiculatus terpenoid can promote tumor cell homogeneity adhesion and suppress the tumor cell alloplasm adhesion and it has the obvious antitumor activity in tumor adhesion, mirgration and angiogenesis, and its mechanism is possible concerned with downregulation expression of MMP-2 and VEGF.3. Celastrus orbiculatus terpenoid can inhibit the HUVECs proliferation and induce its apoptosis, which is possible concerned with downregultion oncogene protein of Bcl-2.4. The results of this experiment demonstrated that in rencent celastrus orbiculatus antineoplasmic activitythe is celastrus orbiculatus terpenoid, it has very good antitumor activity in repressing tumor adhesion, invasion and angiogenesis, which is possible concerned with P53-VEGF-VEGFR pathway and deserved to study and research extremely it is possible become an ideal targeted therapy drug of antiangiogenesis .