PTN in the Mammary Gland Figure 7. PTN effect on MAP kinase pathway activity in developing mammary glands. Phospho-ERK1/2 (A), phospho-p38 (B), and phospho-JNK (C) in whole mammary gland extracts from anti-PTN treated mice (n = 10) versus controls (n = 9). The ratio of phospho- to total protein was obtained from multiplex assays. Ns, not significant; *P,0.05. (D, E) Western blot analysis for pERK1/2. Representative Western blots (mouse m1 to m6) and quantitation of phospho ERK1/2 relative to total ERK1/2 in 3 independent experiments (n = 9 to 10 per group). (F) Representative Western blots of whole cell lysates from MECs treated for 24 hours with either UO126 (500 nM) alone or in combination with anti-PTN blocking antibody and quantitation of phospho ERK1/2 relative to total ERK1/2. Numbers represent the ratio of the quantification of phospho ERK1/2 to total ERK1/2. Data are means 6 SE; *P,0.05. doi:10.1371/journal.pone.0047876.g007 Discussion PTN is expressed and highly regulated during embryonic and postnatal development of different tissues [13]. During adulthood, PTN expression is confined to the nervous system, the iris, the uterus and the basal layer of the tongue epithelium [13], though several reports showed PTN expression also in the mouse mammary gland and in the human breast [39,40,41]. The detection of PTN expression in primary breast cancers and in estrogen receptor negative breast cancer cell lines and its pro- angiogenic and pro-metastatic function [19,22,34,35] make PTN a good candidate for therapeutic targeting. Here we sought to understand PTN function in the normal mammary gland. The mammary gland is one of the few organs that mature after birth and continues development during puberty under hormonal stimuli until adulthood when it becomes quiescent. During pregnancy, the mammary gland goes through several changes such as proliferation, differentiation and apoptosis of the alveolar cells. The multiple cell populations in the mammary gland change in number and ratio during the process of development as well as PLOS ONE | www.plosone.org 8 October 2012 | Volume 7 | Issue 10 | e47876