Establishment Of Mouse Lymph Node Metastasis Model With Lewis Lung Carcinoma Cells Transduced By VEGF-D

Objective VEGF-D is a mitogen for endothelial cells. It can induce tumor lymphangiogenesis and promote lymphatic spread of tumors. This research is to explore the feasibility of establishing mouse lymph node metastasis model with Lewis lung carcinoma cells transduced by VEGF-D. Methods The mouse VEGF-D cDNA was amplified from total RNA isolated from mouse embryo tissue, then it was inserted into pDrive plasmid (Qiagen) and subcloned into pcDNA3.1(+) (Invitrogen) eukaryotic expression vector. After the full-length sequence of expected was confirmed by emzymatic digestion and dideoxy sequence, we transduced mouse Lewis lung carcinoma cells (LLC) with the pcDNA3.1(+) expression vector containing mouse VEGF-D and neomycin resistance genes. Stable VEGF-D high-expression LLC clones were selected in vitro with G418, which were then combined and injected into muscle, footpad and subcutaneously inoculated separately in hind limb of mice to observe their metastatic properties compared with LLC cells transduced only with null-vector and wild type LLC cells. Results Thefull-length sequence of mouse VEGF-D was confirmed by dideoxy sequenceto be identical to that reported in Gene bank. Stable VEGF-D high-expression LLC clones were obtained by selection in vitro with G418, RT-PCR and western blot analysis. Results of animal experiment indicated that expression of VEGF-D in tumor cells promoted tumor growth, formation of lymphatics was observed within tumors and VEGF-D high-expression tumor cells showed a significantly increased lymph node metastasis rate in mice compared with control groups. Conclusion Our study indicated that transduction of VEGF-D may significantly enhance the lymph node metastasis of tumor cells and be very useful in establishing tumor metastasis animal model.