Precise and Facile Endotracheal Lung-tumor-implantation Mouse Model Visualized by GFP Expression

In osteosarcoma, lung metastasis is a major cause of cancer-related death, as the 5-year survival rate for patients with metastases is approximately only 20-30%. To develop improved therapeutic strategies against lung-metastatic osteosarcoma, an experimental lung-tumor-implantation mouse model is needed for basic research. In the present study, we developed a precise and facile endotracheal lung-tumor-implantation technique.

For establishment of the lung-tumor-implantation mouse model of metastatic osteosarcoma, 5 mice were used. A 15-mm longitudinal incision was made in the center of the neck to expose the salivary glands. The salivary glands were then split, exposing the trachea covered by the sternohyoid muscles. The trachea was then clearly exposed by cutting the sternohyoid muscles longitudinally. A 22 G gavage needle was tilted slightly toward the left side of the mouse and inserted from the oral cavity into the bronchus, with confirmation of the position of the tip of the gavage needle visualized through the tracheal wall, followed by injection of 0.5% crystal violet to first confirm the accuracy of endotracheal injection in the lung. A 143B-GFP cell suspension (2.0×10⁶ cells/50 μl PBS) was then injected endotracheally in other mice.

The procedure, including anesthesia and suturing, took approximately 10 minutes. The left lobe of the lung, in which crystal violet was injected endotracheally, was stained in 3 out of 3 mice (100%). 143B-GFP-osteosarcoma tumors were detected with GFP fluorescence in the left lobe of the lung in 3 out of 4 mice (75%), 5 weeks after endotracheal injection. One mouse died 4 weeks after 143B-GFP-cell implantation.

This novel technique of establishing tumors in the lung via endotracheal injection of cancer cells is precise and facile and can be used widely, since neither a surgical microscope nor X-ray imaging are needed.