Fig. 1.
Serotonin (5-HT) induces expression of parathyroid hormone-related protein (PTHrP) in normal breast cells. A: PTHrP immunostaining (green fluorescence) in wild-type tryptophan hydroxylase-1 (TPH1+/+) and TPH knockout (TPH1−/−) day 10 lactating mammary glands; nuclei (red). Negative control (no primary antibody) is shown in the inset. B: plasma PTHrP levels in TPH1+/+ (inset) and (TPH1−/−) mice. Bar crossing the ordinate in each graph indicates the lower limit of PTHrP detection. Results are shown for TPH1+/+ mice that were nonlactating (NL) and day 10 lactating (L10). Experimental day 10 lactating TPH1−/− mice were treated with 100 mg/kg 5-hydroxytryptophan (5-HTP) to rescue 5-HT synthesis and killed at time 0, 1 h, and 3 h, as indicated (n = 3 per group). PTHrP mRNA expression was measured in mouse HC11 cells differentiated with lactogenic hormones and treated with control medium (CM) or medium containing 5-HT (2 × 10−4 M) for 48 h (C); and “3D lactogenic” cultures of primary bovine mammary epithelial cells (pBMECs) treated with CM or 5-HT as labeled (D). PCR data were quantified using the 2−ΔΔCT method, with ribosomal S15 used as internal control gene, and comparisons were normalized to control lactogenic cells. Data are represented as means ± SE of log-transformed values of relative expression (n = 4 or 6, ***P < 0.001).