Figure 5.
Estrogen suppresses JNK1 activity but does not alter RANK and c-Fms expression in RAW 264.7 cells. (A) Cells were pretreated with vehicle or 17β-estradiol (10−8 M) (15 min or 24 h) and then stimulated for 5 min with RANKL (80 ng/ml). Cell lysates were examined for JNK1 activity. RANKL-induced JNK1 activity was reduced 0 and 30% following a 15-min and 24-h treatment with estrogen, respectively. (B) Cells were treated for 24 h with vehicle, 17β-estradiol (10−8 M), 4-hydroxytamoxifen (10−7 M), or raloxifene (10−7 M) and then stimulated for 5 min with RANKL (80 ng/ml). Extracts were examined for JNK1 activity and by Western blot analysis. (C) RNA was isolated from cells treated for 24 h with either vehicle or 17β-estradiol (10−8 M) and subjected to RT-PCR by using oligonucleotide primer pairs for murine RANK, c-Fms, and glyceraldehyde-3-phosphate dehydrogenase (GAPDH). The cycle-dependent appearance of specific DNA products was assessed by agarose gel electrophoresis. d, Cells were treated for 24 h with either vehicle or 17β-estradiol (10−8 M), and stimulated for 15 min with RANKL and nuclear extracts subjected to Western blot analysis by using Abs to c-Jun.