Figure 8.

Overexpression of CrkII results in enhanced collagen deposition, which is associated with delayed ductal outgrowth during puberty. A: Heat map of differentially expressed genes associated with the extracellular matrix, using technical replicates as an additional control. The subset of differentially expressed collagen genes from the extracellular matrix heat map are represented in a collagen signature. B: Collagen type12 alpha1 differential expression was validated using RT-PCR analysis. Data from three independent experiments were collected and expressed as the average fold-change between the T47D control cells and the T47D-CrkII cells. A student's t-test was performed on the data collected from the three experiments to determine significance. C: Comparison of FVB terminal end buds (left panels) to those from MMTV/CrkII transgenic mice (right panels) shows that CrkII terminal end buds have increased collagenous stroma (see black arrow, upper right panel) encompassing the end bud. Trichrome staining revealed that increased collagen surrounds the MMTV/CrkII terminal end buds but not FVB terminal end buds (see black arrows). Immunohistochemistry reveals that Crk is indeed expressed in the terminal end buds of transgenic mice (see black arrow) but not in FVB littermates. All images taken at ×20. Scale bar = 50 μm. D: The percentage of terminal end bud structures encompassed by stroma (“halo-like structures”) in FVB versus MMTV/CrkII mice ± SE, P < 0.05. E: Immunohistochemistry identifies F4/80 positive macrophages (see black arrows) surrounding the terminal end buds of both MMTV/CrkII and FVB mice. All images taken at ×20. Scale bar = 50 μm. F: F4/80 macrophages were scored in a blinded fashion and averaged over the total number of terminal end buds counted, ± SE. The average number of F4/80 macrophages was not significantly different between the wild-type and MMTV/CrkII transgenic mice.