rhKAI1 suppress LPS-induced inflammatory response in RAW 264.7 macrophages via blocking of JNK/NF-κB signaling pathway. (A) Relative levels of mRNA expression for IL-1β, IL-6, TNF-α, and COX-2 were expressed as fold of control. (B) In the supernatants, NO concentration was measured by the Griess reaction. (C) The secretion levels of PGE2, IL-1β and IL-6 levels were determined using commercial ELISA kits. Data are expressed as the mean ± SD (n = 4). ***P < 0.001 compared to control. ###P < 0.001 compared to LPS-stimulated cells. (D) The cells subjected to immunofluorescence staining with NF-κB p65 antibody and representative images were acquired using a confocal laser scanning microscopy. Green fluorescence indicates the localization of NF-κB p65, and blue fluorescence by DAPI allows the nuclei. Scale bar = 50 μm. (E) The protein expression of MAPK signaling molecules including ERK, p-ERK, JNK, p-JNK, p38 MAPK and p-p38 MAPK. (F) Bar diagram showing the relative protein density after normalization with actin based on Western blot analysis. *P < 0.05, **P < 0.01, and ***P < 0.001 compared to control. ###P < 0.001 compared to LPS-stimulated cells.
