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Journal of Oncology
Volume 2012, Article ID 265487, 8 pages
Research Article

In Vivo Time-Course Imaging of Tumor Angiogenesis in Colorectal Liver Metastases in the Same Living Mice Using Two-Photon Laser Scanning Microscopy

1Department of Gastrointestinal and Pediatric Surgery, Mie University Graduate School of Medicine, 2-174 Edobashi, Tsu, Mie 514-8507, Japan
2Department of Neural Regeneration and Cell Communication, Mie University Graduate School of Medicine, 2-174 Edobashi, Tsu, Mie 514-8507, Japan

Received 27 May 2011; Revised 19 August 2011; Accepted 30 August 2011

Academic Editor: Sundaram Ramakrishnan

Copyright © 2012 Koji Tanaka et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.


In vivo real-time visualization of the process of angiogenesis in secondary tumors in the same living animals presents a major challenge in metastasis research. We developed a technique for intravital imaging of colorectal liver metastasis development in live mice using two-photon laser scanning microscopy (TPLSM). We also developed time-series TPLSM in which intravital TPLSM procedures were performed several times over periods of days to months. Red fluorescent protein-expressing colorectal cancer cells were inoculated into the spleens of green fluorescent protein-expressing mice. First- and second-round intravital TPLSM allowed visualization of viable cancer cells (red) in hepatic sinusoids or the space of Disse. Third-round intravital TPLSM demonstrated liver metastatic colonies consisting of viable cancer cells and surrounding stroma with tumor vessels (green). In vivo time-course imaging of tumor angiogenesis in the same living mice using time-series TPLSM could be an ideal tool for antiangiogenic drug evaluation, reducing the effects of interindividual variation.