Figure 4.

Bacteria stimulate FOXO1 nuclear localization through TLR signaling. (A) HL-60 neutrophils were incubated with TLR2 blocking antibody or matched control IgG or TLR4 inhibitor (TAK242) individually or combined. Cells were alternatively incubated with inhibitors to reactive oxygen species (ROS, NAC), NOS (L-NAME), Sirt1(Sirtinol), histone deacetylases (HDAC, Trichostatin A, TSA), or control vehicle alone, DMSO. (A) FOXO1 nuclear localization was determined by immunofluorescence using an antibody specific to FOXO1 and colocalization with DAPI nuclear stain. (B) Quantitation was determined by percent neutrophils with FOXO1 nuclear translocation. The data are representative of two or three independent experiments and expressed as the mean ± SEM from triplicate samples. *Significant difference between neutrophils incubated with bacteria or phosphate-buffered saline (P < 0.05) or ^#treated with inhibitor and vehicle or isotype IgG control (P < 0.05).