High-dose Aspirin Attenuates LPS-induced Pro-inflammatory Cytokines By Inhibiting Activation Of NF-κB And P-38MAPKs In Murine RAW264.7Cells

Studies have shown that various diseases were caused or worsened by inflammation response. Additionally, more clinical studies have proved that aspirin plays an important role in preventing metabolite diseases and neurodegenerative diseases, which are all related with the anti-inflammation of aspirin. The mechanism of aspirin has been explored for a long time, but is still not clear and controversy. Therefore, the mechanism studies of aspirin on anti-inflammation will contribute to the development of new drugs and clinical interventions, avoiding or decreasing the side-effects of aspirin. Activating transcription factor3(ATF-3) is a member of the ATF/cAMP responsive element binding protein (CREB) family member of basic leucine zipper-type transcription factors and expressed inducing by various intra or extra damage stress stimulus, while ATF-3is expressed at a low concentration in most normal cells. ATF-3activates or suppresses gene expression depending on the different cell type and stimulus. ATF-3plays a wide role on the regulation of the cell cycle, immune response, cancer formation and apoptosis. Recent studies have suggested that ATF-3is the hub of immune adaptive response, negatively regulating the pro-inflammatory cytokines IL-6and TNF-a expression. Therefore, we suggested that the anti-inflammatory function of aspirin may relate with ATF-3induction. In this study, we used lipopolysaccharide (LPS) to stimulate immune response in RAW264.7macrophage cells. The levels of ATF-3, IL-6, TNF-a and IL-1β mRNA were detected by RT-PCR, while the changes of IL-6and TNF-a proteins in cell culture medium were measured by ELISA. ATF-3, TNF-a, NF-κB and MAPK-associated proteins were examined by Western blot. SiRNA knock down technology was used to detect whether ATF-3down-regulation contributes to the immune reaction in RAW264.7cells. Our results showed that aspirin suppress the expression of pro-inflammatory cytokines, while stimulate ATF-3induction. However, the knock down of ATF-3did not reverse the expression of pro-inflammatory cytokines IL-6 and TNF-a suppressed by aspirin, which indicated that the anti-inflammatory mechanism of aspirin was independent on ATF-3induction. High-dose aspirin performed anti-inflammatory reaction via suppressing NF-κB and p-38MAPK signal transduction.