Figure 4
pH e effect on prolactin receptor signaling is specific. (A) SKBR3 cells were stimulated with vehicle, prolactin or EGF for 30 min at different pHe as indicated. Stat5 and Erk phosphorylation were examined (n = 3). (B) SKBR3 cells were treated with vehicle or 10 nM OSM for 15 min at pHe 7.4 or 6.8. Representative Stat3 and Erk activation were shown (n = 2). (C) SKBR3 cells were treated with vehicle, prolactin or EGF for 1 hour at pHe 7.4 or 6.8. Quantitative PCR of c-jun and CISH transcripts were conducted and representative results are shown in bar graphs (n = 3). (D) 32D-hPrlR cells were treated with 2 nM prolactin at pHe range of 7.4 to 6.6 (n = 3). 32D-hGHR cells were treated with 2 nM growth hormone at similar conditions (n = 5). Representative pYStat5 and total Stat5 blots are shown. Stat5 activation at each pHe were analyzed and compared. (E), 10 nM GH-induced phosphorylation of Jak2, Jak1 and Stat5 at various zinc concentration were compared at normal and acidic tumor pHe in T47D cells (n = 4). One-way ANOVA was followed by Bonferroni’s post hoc test. 32D-hGHR, human growth hormone receptor cells; 32D-hPrlR cells, mouse promyeloid 32D cells stably transfected with human prolactin receptor; ANOVA, analysis of variance; EGF, epidermal growth factor; GH, human growth hormone; pHe, extracellular pH.