Preparation and Testing of Cells Expressing Fluorescent Proteins for Intravital Imaging of Tumor Microenvironment
Intravital microscopy is widely used for in vivo studies of the mechanisms of carcinogenesis and response to antitumor therapy. For visualization of tumor cells in vivo, cell lines expressing fluorescent proteins are needed. Expression of exogenous proteins can affect cell growth rate and their tumorigenic potential. Therefore, comprehensive analysis of the morphofunctional properties of transduced cells is required for creating appropriate models of tumor microenvironment. In the present study, six lines of mouse tumor cells expressing green and red fluorescent proteins were derived. Analysis of cells morphology, growth kinetics, and response to chemotherapy in vitro revealed no significant differences between wild-type and transduced cell lines. Introduction of fluorescent proteins into the genome of 4T1 (murine breast cancer) and B16-F10 (murine melanoma) cells did not affect tumor growth rate after subcutaneous implantation to mice, while both CT26-GFP and CT26-RFP cells (murine colon cancer) were rejected starting from day 8 after implantation. Elucidation of the mechanisms underlying CT26-GFP/RFP rejection is required to modify transduction technique for creating the models of tumor microenvironment accessible for in vivo visualization. Transduced 4T1 and B16-F10 cell lines can be used for intravital microscopic imaging of tumor cells, neoplastic vasculature, and leukocyte subpopulations.
Key Wordsfluorescent cell lines transduction tumor uptake intravital microscopy
Unable to display preview. Download preview PDF.
- 4.Caysa H, Hoffmann S, Luetzkendorf J, Mueller LP, Unverzagt S, Mäder K, Mueller T. Monitoring of xenograft tumor growth and response to chemotherapy by non-invasive in vivo multispectral fluorescence imaging. PLoS One. 2012;7(10):e47927. doi: https://doi.org/10.1371/journal.pone.0047927.CrossRefGoogle Scholar
- 5.Chishima T, Miyagi Y, Wang X, Yamaoka H, Shimada H, Moossa AR, Hoffman RM. Cancer invasion and micrometastasis visualized in live tissue by green fluorescent protein expression. Cancer Res. 1997;57(10):2042-2047.Google Scholar
- 8.Goto H, Yang B, Petersen D, Pepper KA, Alfaro PA, Kohn DB, Reynolds CP. Transduction of green fluorescent protein increased oxidative stress and enhanced sensitivity to cytotoxic drugs in neuroblastoma cell lines. Mol. Cancer Ther. 2003;2(9):911-917.Google Scholar
- 11.Immunobiology: The Immune System in Health and Disease. Janeway СA, Travers P, Walport M, Shlomchik M, eds. New York, 2001. Russian.Google Scholar
- 18.Taghizadeh RR, Sherley JL. CFP and YFP, but not GFP, provide stable fluorescent marking of rat hepatic adult stem cells. J. Biomed. Biotechnol. 2008;2008. ID 453590. doi: https://doi.org/10.1155/2008/453590.
- 19.Yang M, Baranov E, Jiang P, Sun FX, Li XM, Li L, Hasegawa S, Bouvet M, Al-Tuwaijri M, Chishima T, Shimada H, Moossa AR, Penman S, Hoffman RM. Whole-body optical imaging of green fluorescent protein-expressing tumors and metastases. Proc. Natl Acad. Sci. USA. 2000;97(3):1206-1211.CrossRefGoogle Scholar
- 20.Yuzhakova DV, Shirmanova MV, Serebrovskaya EO, Lukyanov KA, Druzhkova IN, Shakhov BE, Lukyanov SA, Zagaynova EV. CT26 murine colon carcinoma expressing the red fluorescent protein KillerRed as a highly immunogenic tumor model. J. Biomed. Opt. 2015;20(8):ID 88002. doi: https://doi.org/10.1117/1.JBO.20.8.088002.