br Results br Discussion Abnormally elevated levels of BMP
Discussion Abnormally elevated levels of BMP2 were specifically detected in luminal breast tumors but produced by tumor microenvironment and not by cancer anti fungal themselves as reported in leukemia (Gerber et al., 2013; Laperrousaz et al., 2013). Chronic exposure to BMP2 in the presence of IL-6 promotes the engagement of immature epithelial cells toward transformation, as shown by anchorage-independent growth and in vivo experiments. Consistent with difficulties engrafting established human breast tumors in recipient mice (2.5% for luminal versus 30% for nonluminal tumors) (Cottu et al., 2012), we did not observe large and aggressive cell outgrowth. Nevertheless, we obtained a significant and reproducible engraftment of long-term BMP2-treated-MCF10A cells, and loss of BMPR1B expression resulted in a decrease in engrafted cells. BMP2-treated cells displayed a robust luminal tumor-like phenotype, as confirmed by multiparameter comparison using two independent sets of primary tumors and induce and amplify intrinsic alterations, such as BMPR1B overexpression. However, the observed in vivo lesions did not yet resemble human breast carcinoma, suggesting that our model mimics an early stage of tumor development. Very interestingly, in vivo cells organized close to neovessels that colonized the coinjected matrigel. This was not observed in the xenografts of cells expressing shBMPR1B, confirming that BMPR1B was required for transformation and identifying it as the predominant receptor involved in luminal tumors from the earliest stages to established tumors despite its contradictory role in breast cancer prognosis (Bokobza et al., 2009; Helms et al., 2005). This is also consistent with our demonstration that in chronic phase of chronic myeloid leukemia the BMP/BMPR1B pathway provides key proliferation and expansion signals to leukemic stem cells (Laperrousaz et al., 2013). Aberrant regulation of the BMP pathway could therefore be a general mechanism in early phases of the transformation of immature cells. We showed that the BMPR1B ligands BMP2 and BMP4 are present in the human normal mammary gland, where they occupy different functions in the regulation of mammary stem/progenitor cell fate (Figure 7). BMP2, but not BMP4, is able to commit cell to luminal differentiation and expansion of the luminal progenitor compartment, like in the murine mammary gland (Forsman et al., 2013). Interestingly, luminal progenitors have been reported to be potentially more susceptible to oncogenic events (Lim et al., 2009; Molyneux et al., 2010) due to short telomere length (Kannan et al., 2013). This might confer a more genetically unstable feature to these cells, also specific targets of BMP2 biological cell fate control. Our data indicate that binding of BMP2 to BMPR1B rapidly induces a sustained signaling involving GATA3 (as in development; Forsman et al., 2013) and a change of the FOXA1/FOXC1 balance leading to luminal immature progenitors expansion that further transformed through a BMPR1B-dependent signaling. Therefore, it is possible that under BMP2 signal transformation arises either from a stem/basal cell that first engages toward a luminal progenitor and then proliferates and further progresses or directly emerges from an already genetically altered committed luminal progenitor. Based on our findings, we cannot rule out one of the two hypotheses. Irrespective of the impact of BMP2 on luminal lineage commitment, the IHC analysis of normal and tumor tissue indicated that tumor cells themselves are the target rather than the origin of BMP2 overproduction. The transformation process may therefore either lead to the loss of BMP2 expression in mammary epithelial cells or occur in an epithelial cell that does not produce BMP2. We showed that environmental pollutants such as BPA (Rundle et al., 2000), and its substitutes, BPS or radiation (Sun et al., 2012), are able to shift the balance of secreted BMP molecules in favor of BMP2. This seems to happen more frequently in individuals susceptible of developing breast cancer. Radiation and/or BPA accumulation in the mammary stem cell niche might contribute to increase the local concentrations of BMP2 and IL-6, which in turn may facilitate epithelial cell transformation and progression. Indeed, IL-6 secretion by stromal cells can be induced by BPA (Ben-Jonathan et al., 2009) and BMP2 (Hyzy et al., 2013; Zara et al., 2011), suggesting a possible feedback loop that maintains transforming conditions. This is consistent with the high BMP2 staining detected in tumor endothelial cells also involved in increased angiogenesis (Finkenzeller et al., 2012; Raida et al., 2005), causing a continuous influx of BMP-laden platelets, thereby sustaining local high concentrations of BMPs (Labelle et al., 2011).