POSTN promotes a mesenchymal phenotype in MCF-10A and MCF-7 cells.

<p><b>A.</b> POSTN-overexpressing cells exhibit a mesenchymal-like morphology. <b>B.</b> POSTN promotes cell invasion of human mammary epithelial cells and BCCs as detected by a matrigel-coated transwell invasion assay. <b>C, D.</b> Immunofluorescence analysis revealed that the mesenchymal markers N-cadherin, fibronectin, vimentin and α-SMA in POSTN-expressing cells were increased while the epithelial marker E-cadherin was decreased. <b>E, F.</b> POSTN-expressing cells show increased levels of N-cadherin, fibrnectin, vimentin and α-SMA and decreased E-cadherin. Expression of epithelial and mesenchymal markers was analysed by western blotting.</p

POSTN induces adipogenic and chondrogenic differentiation.

<p><b>A.</b> Following adipogenic differentiation, MCF-10A/POSTN, MCF-7/POSTN cells and hMSCs stained positive with oil red O (top) and fluorescent LipidTox, which stains oil droplets (bottom). <b>B, C.</b> Real-time RT-PCR analysis for the expression of the adipocyte markers <i>PPARγ</i>and <i>ADFP</i> in MCF-10A and MCF-7 cells and their POSTN-overexpressing cells subjected to adipocyte differentiation for 21 days. The data are means ± SD. *P<0.05, **P<0.01. <b>D.</b> Chondrocytic nodules formed by MCF-10A/POSTN cells and hMSCs stained positive with alcian blue 8 GX (left panel). Immunohistochemistry was performed on chondrocyte sections using antibody against collagen II (right panel). MCF-10A/Vector cells, MCF-7/Vector and MCF-7/POSTN cells did not form any chondrocytic nodules under identical conditions.</p

Model of role of POSTN in mammary epithelial neoplasia and metastasis.

<p>POSTN might confer mammary epithelial cells and BCCs with stem cell-like traits and a mesenchymal phenotype, as well as the multipotent potentials of MSCs to promote tumorigenesis and metastasis.</p

POSTN induces osteoblastic and myogenic differentiation.

<p><b>A, B.</b> Following culture in osteoblastic differentiation media for 21 days, MCF-10A/POSTN, MCF-7/POSTN cells and hMSCs were positive for alizarin red S and von Kossa staining. <b>C, D.</b> Relative levels of mRNAs encoding BSP and Runx2 in MCF-10A and MCF-7 cells expressing the vector or POSTN were determined by real-time RT-PCR. <i>Cyclophilin G</i> mRNA was used to normalize the variability in template loading. The data are the means ± SD. *P<0.05, **P<0.01. <b>E.</b> Following myogenic differentiation for 4 weeks, MCF-10A/POSTN cells stained positive for CD56. MCF-7/POSTN cells died under the same myogenic differentiation condition and did not differentiate into myogenic lineages (data not shown). <b>F.</b> Real-time RT-PCR analysis of <i>MyoG</i> and <i>Pax3</i> showing the expression of myogenic markers. The data are the means ± SD. *P<0.05.</p

POSTN-overexpressing cells accelerates the tumor growth and metastatic properties of BCCs.

<p><b>A.</b> 2.5×10⁶ MCF-7/Vector or MCF-7/POSTN cells were injected subcutaneously into 5- to 6-week-old female Balb/c nude mice (n = 4 mice per group). Mice were sacrificed 30 days after injection and examined for the growth of subcutaneous tumors. **P<0.01. <b>B.</b> Tumor weight measurements of 1×10⁶ MDA-MB-231 cells subcutaneously injected into nude mice with 3×10⁶ MCF-10A/Vector cells or MCF-10A/POSTN cells. <b>C.</b> The tumors were measured once or twice per week. Circles, MDA-MB-231 cells; squares, MDA-MB-231 cells plus MCF-10A/Vector cells; triangles, MDA-MB-231 cells plus MCF-10A/POSTN cells; diamonds, MCF-10A/POSTN cells (n = 4 mice per group). **P<0.01. <b>D.</b> Mice subcutaneously implanted with 3×10⁶ MDA-MB-231 mixed with 9×10⁶ MCF-10A/Vector cells or with MCF-10A/POSTN cells (n = 3 mice per group). Twelve weeks after cell injection, the mice were euthanized and the lung metastasis indices for each tumor bearing mouse were determined. *P<0.05. <b>E.</b> Representative haematoxylin-and-eosin-stained sections of lungs of mice bearing the indicated tumors in (D). Scale bar = 500 µm.</p