Macrophage plays a critical role in inducing inflammatory response, activated though TLR4 receptor by endotoxin lipopolysaccharide (LPS). It has been reported that cell inflammation is related to the oxidative stress response. However, it is not clear how the LPS-induced inflammatory signaling pathway is mechanistically related to oxidative stress responses. Here, we examined the impact of LPS-induced inflammation and oxidative stress using RAW 264.7 cells. LPS treatment up to 1 µg/ml did not show any cell toxicity. LPS stimulation causes Akt and ERK1/2 activation. However, treatment with LY294002, an Akt signaling inhibitor, completely inhibited Akt activation but not ERK1/2 activation. Interestingly, We found that LPS induces down-regulation of Nrf2 nuclear expression and translocation, whereas inhibition of Akt activation with LY294002 reverses the effects of LPS on Nrf2, as judged by Western blotting with nuclear fraction protein and immunofluorescence analysis. In addition, to investigate the role of Akt signaling in NF-κB activation-induced inflammatory response, NF-κB nuclear translocation and its target gene expression were analyzed by immunofluorescence assay and qRT-PCR analysis. LPS-induced NF-κB nuclear translocation was suppressed by LY294002 treatment. Furthermore, LY294002 inhibited LPS-induced mRNA expression of inflammatory markers, including IL-1β and MCP-1. In summary, these findings suggest that Akt signaling plays a critical role in the LPS-induced inflammatory and oxidative stress responses in RAW 264.7 cells.

KeywordsAkt, inflammation, lipopolysaccharide, Nrf2, NF-κB