The cells were stained with crystal violet, photographed, the dye extracted and absorbance read at 570nm. Scale bars = 2 mm.Ī) Upper panel-mouse mammary epithelial cells, Comma-1D, were incubated in serum-free medium or in conditioned medium (CM) from differentiated (adipocytes) 3T3-L1 cells for 4 days. All of the 6 whole Tg-mammary gland (epithelium and adipocytes) transplants retained their stunted Tg-phenotype in a wt-mouse environment. The #4 wt (left panel) and Tg glands (right panel) of 3–4 weeks old mice were transplanted in contra lateral sites under the skin (between gland #3 and #4) of a wt, 6 week old recipient mouse and processed as in C. The whole mount is representative of 12 individual transplants which were all successful and all showed complete repopulation of the fat pad. Right upper and lower panels, as above, except the donor is Tg. C) Donor wt-mammary epithelium (8 weeks old mouse, gland #4, upper left panel) transplanted into wt mammary gland divested of epithelium, of a 3–4 week old mouse (lower left panel) and analyzed by whole mount 8 weeks later.

B) Quantification of ductal length shown as a ratio between the longest mammary duct measured from the nipple and the length of the fat pad (n=4 glands per group, p<0.001, t-test). All rights reserved.Ī) Whole mounts of mammary glands from wt and DN-RARαG303E transgenic mice isolated at 6, 9 and 12 weeks. Overall, these results support the existence of a novel paracrine loop controlled by adipocyte-RAR that regulates the mammary ductal tree morphogenesis.Ĭopyright © 2010 Elsevier Inc. Moreover, adipocyte-PTHR, another atRA responsive gene, was found to be an up-stream regulator of PTN. Its expression was found to be strongly inhibited by DN-RARα, an inhibition relieved by pharmacological doses of all-trans retinoic acid (atRA) in culture and in vivo. Among them, pleiotrophin (PTN) was identified as the paracrine effectors of epithelial cell migration. Gene expression microarrays revealed a large number of genes regulated by adipocyte-RARs. Co-cultures of primary adipocytes, or in vitro differentiated adipocyte cell line, with mammary epithelium showed that when activated, adipocyte-RARs contribute to generation of secreted proliferative and pro-migratory factors. Transplantation experiments of mammary epithelium and of whole mammary glands implicated a fat-pad dependent paracrine mechanism in the stunted phenotype of the epithelial ductal tree. Transgenic adipocytes had reduced level of RARα, β and γ, which coincided with a severely underdeveloped pubertal and mature ductal tree with profoundly decreased epithelial cell proliferation. We generated a transgenic (Tg)-mouse model expressing a dominant negative-(DN)-RARα, (RARαG303E) under adipocytes-specific promoter to explore the paracrine role of adipocyte retinoic acid receptors (RARs) in mammary morphogenesis.

Liliana model rar