Use of alpha(v) Integrin Linked to Green Fluorescent Protein in Osteosarcoma Cells and Confocal Microscopy to Image Molecular Dynamics During Lung Metastasis in Nude Mice

摘要

Background: We report here imaging of the behavior of alpha(v) integrin linked to green fluorescent protein (GFP) in human osteosarcoma cells colonizing the lung of nude mice. Materials and Methods: 143B osteosarcoma cells expressing alpha(v) integrin-GFP were generated by transfection of an alpha(v) integrin-GFP fusion-gene vector pCMV-AC-ITGAV-GFP. In order to generate experimental lung metastases, 143B osteosarcoma cells (1x10(6)), stably expressing alpha(v) integrin-GFP, were injected intravenously via the tail vein. The osteosarcoma cells were transplanted orthotopically in the tibia of nude mice in order to generate spontaneous metastases. Lungs were harvested and imaged by confocal microscopy within 1 hour. Results: In the experimental lung-metastasis model, extravasating and deformed osteosarcoma cells expressing alpha(v) integrin-GFP were observed. Pseudopodia of the osteosarcoma cells contained small puncta of alpha(v) integrin-GFP. In early-stage spontaneous lung metastasis, tumor emboli were observed in pulmonary vessels. At high magnification, small alpha(v) integrin-GFP puncta were observed in the tumor embolus. In late-stage spontaneous metastasis, tumor emboli were also observed in pulmonary vessels. Invading cancer cells with strong expression of alpha(v) integrin-GFP were observed at the margin of the tumor emboli. Conclusion: The results of this study demonstrate that molecular dynamics of alpha(v) integrin-GFP can be imaged in lung metastasis, which will allow further understanding of the role of alpha(v) integrin in this process. The results also suggest a general concept for imaging molecular behavior in vivo.