Study On The Transfection Of Mouse Interleukin-12 Gene Inhibiting The Angiogenesis Of Hepatocarcinoma Of Nude Mouse

Objective: To study the effect of mouse interleukin-12 (mIL-112) gene mediated by adenovirus (AdvmIL-12) on the growth of hepatocarcinoma H22 cell in vitro, and the inhibition of AdvmIL-12 on the angiogenesis of nude mouse hepatocarcinoma in vivo. Methods: (1) AdvmIL-12 was constructed and transfected into H22 cell. The transfection efficiency of AdvmIL-12 on H22 cell was observed by fluorescence microscope. And the content of mIL-12 protein was detected by ELISA in the supernatant. (2) The effect of AdvmIL-12 on the growth of hepatocarcinoma H22 cell in vitro was observed by inverted phase contrast microscope, counting cells, modified MTT and flow cytometry. (3) AdvmIL-12 was transfected into H22 cells in the experimental group. The control group was the H22 cells transfected adenovirus and the H22 cells which were not transfect anything. Then they were inoculated into the subrenal capsule of nude mouse. Then the growth of three kinds of H22 cells and the angiogenesis of hepatocarcinoma were observed. The expression of gene of vascular endothelial growth factor (VEGF) and angiopoietin-2 (Ang-2) were detected by reverse transcription-polymerase chain reaction (RT-PCR).Results: (1) AdvmIL-12 was constructed successfully. When multiplicity of infection (MOI) was 100 and time of infection (TOI) was 2 hours, the transfection efficiency of recombinant adenovirus was (89.71±22.05) ng.48h-1.106cells-1. (2) In vitro the dimension, form, growth, proliferation and the cycle of cells in the experimental and control groups had no difference(P>0.05). (3) In vivo, the volum of hepatocarcinoma tissue was smaller, hemorrhage was lesser in the experimental group. The brightness of VEGF and Ang-2 gene in the experimental group were weaker than that of the control group (p<0.05). The luminance ratio of VEGF (0.8867±0.1924) gene in the experimental group were lower than that of the vector control group (1.1744±0.1189) and the blank control group (1.1874±0.1752) (P<0.05).The luminance ratio of Ang-2 (0.7878±0.1471) gene in the experimental group were lower than that of the vector control group (1.0319±0.1574) and the blank control group (1.0361±0.1536) (P<0.05).Conclusions: (1) AdvmIL-12 was constructed correctly, and its transfection efficiency was higher, and it could express mIL-12 protein highly. (2) In vitro IL-12 can't inhibit the growth of H22 cells. (3) In vivo IL-12 can inhibit the growth of H22 cells and expression of VEGF and Ang-2 gene, accordingly it can inhibit the angiogenesis of hepatocarcinoma and the growth of hepatocarcinoma cells.