Figure 4

AAT inhibits RANK and related gene expression in OCP cells. bMM cells were treated with M-CSF for 4 d in the presence or absence of AAT as indicated and stimulated by RANKL (Day 4–7). (A) Cell surface RANK receptors detected by flow cytometry expression. Left panel, the mean fluorescence intensity of cell surface RANK receptor levels at d 4. (Right panel) The representative flow cytometry histogram (n = 5). b Western blot analysis for detection of total RANK protein at d 4. (Upper panel) Representative image of Western blot. (Lower panel) RANK protein band intensities of four individual experiments. band intensity was analyzed with Image J software. (C–J) Relative gene expression (mRNA) levels (fold changes) detected by quantitative real-time PCR. (C) RANK at d 4. (D) RANK at d 7. (E) NFκb at d 4. (F) NFκb at d 7. (G) cFos at d 4. (H) cFos at d 7. (I) NFATc1 at d 4. (J) NFATc1 at d 7. Values are means ± SEM of at least triplicate samples. Data were analyzed using one-way analysis of variance followed by Dunnett’s multiple comparison test. Student t test was used to compare two samples. *P <0.05, **P <0.001, ***P <0.0001.