Figure 4

Colocalization of PRLr and PRL in THP-1 monocytes activated with LPS. A, fluorescent immunocytochemistry (FI) to detect PRLr and PRL was performed with THP-1 monocytes activated with LPS 1μg/ml and harvested after: 1 h, a, b and c; 4 h, d, e and f; 8 h, g, h and i. Representative results for PRLr, a, d and g; PRL, b, e and h; as well as, merged by double immunocytochemistry for PRLr and PRL are shown (1250X). The nucleus was counterstained with DAPI and an overlay was performed. B, cell signal intensity obtained with FI for PRLr (black) and PRL (white) was quantified using densitometry and compared between 1, 4 and 48 h after stimulation with LPS. Experiments were performed in triplicate, and significance is defined as p<0.001**; p<0.0001*** vs. control or between 1 and 4 h p<0.0001†††. C, Western blot analysis of PRLr; and D, PRL with culture supernatant and nuclear extracts after stimulation with LPS are shown. As positive controls, total MCF-7 extracts and hrPRL were used; and β-Actin was revealed as an endogenous control.