fig5

Figure 5. Inhibition of the CXCL12/CXCR4 axis expression by withaferin A in F cells which were pretreated with interferon-γ (10 U/mL for 24 h). (a) Time-dependent inhibition of CXCR4, CXCL12, and vimentin expression by withaferin A. Interferon-γ pretreated floating (F) cells were treated with 5 μmol/L withaferin A for various lengths of time as indicated, and the expression of each protein was measured by Western blot analysis; (b) suppression of cellular invasion by interferon-γ pretreated F cells by neutralizing anti-CXCR4 antibody. Interferon-γ pretreated F cells were incubated with 5 μg/mL control IgG, 5 μg/mL anti-CXCR4 antibody, or 5 μmol/L withaferin A for 24 h. Asterisks denote a statistically significant difference between anti-CXCR4 antibody treatment and control IgG treatment (P < 0.01); (c) knockdown of CXCR4 expression by CXCR4-siRNA. Interferon-γ pretreated F cells were transfected with control-siRNA or CXCR4-siRNA, and the expression of CXCR4 and vimentin were measured by Western blot analysis; (d) suppression of cellular invasion by interferon-γ pretreated F cells by treatment with CXCR4-siRNA. Interferon-γ pretreated F cells were transfected with control-siRNA or CXCR4-siRNA. Interferon-γ pretreated F cells were then harvested, and incubated in a chamber for 24 h. Asterisks denote a statistically significant (P < 0.01) difference between CXCR4-siRNA treatment and control-siRNA treatment