Skip to main content
. 1998 Jul 7;95(14):8375–8380. doi: 10.1073/pnas.95.14.8375

Figure 3.

Figure 3

Anandamide interferes with prolactin action. (a) Dose-related effect of prolactin mAb (Pierce) on EFM-19 cell proliferation in the presence or absence of 1 μM anandamide (AEA). (b) Effect of human (h) prolactin (50 ng/ml) on EFM-19 cell proliferation and its counteraction by low doses of anandamide with or without 0.5 μM SR 141716A. (c) Effect on the levels of the long form (100 kDa) of prolactin receptor of 3-day treatment of EFM-19 cells with anandamide (2.5 μM) in the absence (lanes B and E) or presence (lanes C and F) of SR 141716A (0.5 μM); lanes A and D are from untreated cells. (d) Effect on the levels of the brca1 gene product (220 kDa) of 3-day treatment of EFM-19 cells with anandamide (2.5 μM) in the absence (lane B) or presence (lane C) of SR141716A (0.5 μM). In a, the difference observed between the two sets of data was never statistically significant except for 0 μg/ml prolactin antibody. In a and b, data are mean ± SD (n = 3) and are expressed as described in Fig. 1 a, c, and d. ∗, P < 0.05 vs. h-prolactin + [AEA] = 0; ∗∗, P < 0.05 vs. h-prolactin + [AEA] = 0.5 μM. In a, control experiments were performed by using BSA or a NO synthase III polyclonal antibody instead of prolactin mAb, with no effect on proliferation. In c and d, Western immunoblotting was performed with a monoclonal anti-prolactin receptor antibody (c, lanes A–C), polyclonal anti-phosphotyrosine antibody (c, lanes D–F), or a polyclonal anti-brca1 protein antibody (d, lanes A–C). Proteins immunoprecipitated with a monoclonal anti-prolactin receptor antibody or total proteins (50 μg) were used in c and d, respectively. Control experiments (not shown) did not exhibit the bands at 220 or 100 kDa and were carried out with: (i) no proteins, (ii) no first antibody, and (iii) using, as the first antibody, various antibodies other than the ones mentioned above. The mobility of molecular weight markers is shown. Data are representative of at least three separate experiments. Similar data were obtained with MCF-7 cells. Photographs were taken from films exposed with the enhanced chemiluminescence methodology.