Local stimuli are sufficient for induction of bax gene expression and phosphorylation changes in Stat5a/5b and Stat3. (A) Northern blot analysis of steady state levels of bax mRNA in open (O), closed (C), and normal involution day 3 (I3) glands. Loading control, ethidium bromide staining of 18S and 28S RNA. (B) Western blot analysis of steady state levels of bax and WAP protein in open (O) and closed (C) glands compared with spleen (S). Coomassie blue staining of the protein gel demonstrated equal loading of all samples. (C) Analysis of Stat5a and b protein levels, phosphorylation, and heterodimerization by immunoprecipitation and Western blot in open (O) and closed (C) glands at 12, 24, 48, and 72 h. α-5a, anti-Stat5a Ab; α-5b, anti-Stat5b Ab; α-pY, antiphosphotyrosine Ab. (D) Analysis of Stat3 protein levels and phosphorylation by immunoprecipitation and Western blot in open (O) and closed (C) glands at 12, 24, 48, and 72 h. α-3: anti-Stat3 Ab.