Figure 4.

TLR2 and TLR4 both participate in PepO inducing macrophage function enhancement. (a) The PDMs were incubated with PepO, PepO 1–430, and PepO 431–630, respectively, the cell lysates were subjected to His-Tag IP and the interaction of different proteins with TLR2 and TLR4 were identified by TLR2 or TLR4 immunoblot. GAPDH was used as loading control. (b) PepO enhanced the phagocytosis of tlr2^−/- macrophage, though higher dose was required. (c) PepO enhanced the phagocytosis of tlr4^−/- macrophage. (d) tlr2/tlr4^−/- macrophage did not respond to PepO stimulation. (e) PepO failed to reduce the bacterial load of S. pneumoniae in tlr2/tlr4^−/- mice (n = 6 mice/group). (f) The macrophages were treated with Bafilomycin A1 before PepO treatment, and the phagocytosis against S. aureus was assessed. (g) The bactericidal activity against S. aureus of Bafilomycin A1 and PepO-treated macrophages was assessed. (h, i, and j) The relative NOS activity of different macrophages treated with PepO or with Bafilomycin A1 before PepO treatment was assessed. Data are shown as mean + SEM (n = 3) and are representative of three independent experiments. ns: not significant; **p < 0.01. Student’s t-test was employed for statistical analysis.