The Study Of Ang2-sirna Lentivirus Intervene The Portability Tumor Of Melanoma In Nude Mice

Objective:The model is used by a human malignant melanoma transplanted tumor nude mice model, used Ang2-siRNA lentivirus vector mediated the RNA interference, in order to silence Ang2gene and affect the espression of Ang2gene in model. To research the effect of Ang2-siRNA lentivirus on Ang2gene in transplanted tumor mice, to measure the microvessel density of the transplanted tumor and the tumors’ proliferation and volumes, to evaluate and identify the relationships of Ang2gene expression, microvessel density and tumor proliferation, to provide some basic experimental basis for cancer gene therapy.Methods:1.15male nude mice were randomly divided into PBS control group, Vector control group and RNAi group (n=5).2. Normal cultured and passaged A375cells, made into single cell suspension, and adjust the cell density to5×107cells/ml.3.100μL micro-injector of prepared A375single-cell suspension was inoculated subcutaneously in nude mice right axilla,100μL of cells was inoculated in each mouse, to establish the transplanted tumor nude mice model of human malignant melanoma.4. PBS control group, Vector control group a,RNAi group, three groups of nude mice, respectively, were injected into only PBS, pNL-EGFP-Vector and pNL-EGFP-Ang2-siRNA Lentivirus solution200μL every mouse, and then strengthen intra-abdominal injection of the same solutions500μL every mouse, to carry out interference test research.5. To observe and record the proliferation and volume of transplanted tumor, to analyze the volume difference of transplanted tumor between the groups, and make the tumor growth curve.6. To detect the relative expression levels of Ang2gene in transplanted tumor by Real-time RT-PCR, to contrast Ang2gene expression level differences between the groups, to analyze the relationship of the degree of tumor proliferation. 7. To detect transplanted tumor microvessel density using Immunohistochemical method, analyze differences of microvessel density between groups and the relationship of microvessel density and the degree of tumor proliferation.Results:1. A human malignant melanoma transplanted tumor nude mice model was to establish well.2. Tumor volumes were significantly smaller than the PBS control group and the Vector control group in RNAi group, the difference was statistically significant (P <0.01), but it have no significant differences between the PBS control group and Vector control group (P>0.05).3. The Ang2mRNA expression was significant lower to the PBS control group and Vector control group in RNAi group, it was statistically significant (P<0.01), however, the Ang2mRNA expression level have no significant differences between the PBS control group and Vector control group (P>0.05).4. The transplanted tumor microvessel density was notably lower than the PBS control group and Vector control group in RNAi group, it was statistically significant (P<0.01), however, the microvessel density have no significant differences between the PBS control group and Vector control group (P>0.05).Conclusions:The Ang2-siRNA Lentivirus vector mediated the RNA interference can successfully reduce the level of gene expression of Ang2in human malignant melanoma transplanted tumor in nude mice, reduce the formation of microvessels in transplanted tumors, thereby inhibit the tumor growth and proliferation.