Figure 4. Invasion in vivo no longer requires the CSF-1/CSF-1R paracrine loop upon HBEGF expression.

A,B, MTLn3 ErbB1 control (A) and HBEGF (B) primary tumors were tested for their ability to invade in vivo upon inhibition of the paracrine loop of invasion by using 1uM JnJ, a CSF-1R inhibitor. C, PBS-containing control or clodronate-containing liposomes were injected i.v. 48 h and 24 h prior to analysis into the tail veins of animals bearing MTLn3 ErbB1 HBEGF tumors. Four hours prior to analysis, 70kDa Texas Red dextran was injected intravenously, and the uptake of dextran by macrophages in the spleen and tumor was assessed using multiphoton imaging. Representative images of the spleen (left columns) and primary tumor (right columns) are shown. Magenta, collagen fibers; red, macrophages; green, tumor cells. Scale bar, 50 um. D, Needles containing buffer or 25nM EGF were inserted into the primary tumors of animals that were treated with PBS-containing control liposomes (vehicle) or clodronate-containing liposomes (clodronate) and the number of cells invading into the needle was determined. E, 3D in vitro invasion of MTLn3 ErbB1 and MDA-MB 231 transductant cell lines in the absence and presence of BAC macrophages measured as the fraction of carcinoma cells invading ≥ 20um into collagen gel. Data are means and SEM; **p < 0.01, ***p < 0.001 by t- test.