FIGURE 8:

MCF-7 CXCR4ΔCTD cells migrate toward blood vessels and metastasize to the lymph nodes. (a) Intravital images from a time series of GFP-MCF-7 CXCR4ΔCTD cells orthotopically implanted in the fourth mammary fat pad of athymic nude mice 2 wk before imaging. Differentiated HL60 cells were labeled with DiI Cy5 (blue) and injected into the vasculature via a catheter in the femoral vein. Host vasculature was labeled with 30 μl of 20 mg/ml rhodamine dextran (70 kDa), a skin flap was made to expose the mammary fat pad, and images were acquired 2 h after injection of labeled HL60 cells with an LSM 510 META inverted confocal microscope with a 40×/1.3 Plan Apochromat objective. An asterisk is placed over the area of reference to act as a landmark to identify the direction of cell movement. Differentiated HL60 cells (average of two cells in the vasculature adjacent to the tumor) labeled with DiI Cy5 (blue) are indicated by arrows. (b) The 12 most-displaced trajectories of single GFP-MCF-7 CXCR4ΔCTD cells (pink spheres) in the migrating leading edge were tracked over time with Bitplane Imaris. The arrows represent displacement of tracked cells, and colored lines are dragon tails that represent displaced trajectory of the cells tracked over time (Supplemental Movie S7). (c) Differentiated HL60 cells (average of two cells migrated in the tissue toward the tumor cells) labeled with DiI Cy5 (blue) are indicated by arrows. The dHL60 cells migrated toward the leading edge of the migrating GFP-MCF-7 CXCR4ΔCTD tumor cells over the indicated time periods. (d) GFP-MCF-7 CXCR4ΔCTD cell localization in lymph node metastasis. GFP⁺ tumor cells were detected in draining lymph nodes (i) but not contralateral lymph nodes (ii) of the tumor-bearing mouse. Bars, 150 μm.