Carbon Monoxide Affect Inflammatory Response Of Macrophage In Obese

Objective:Obesity is considered as a chronic inflammatory state with increased level of FFA, and it is closely relatedto the activation of inflammatory response of macrophage. CO is an anti-infalmmatory molecule, and inthis work, we added CO to the FFA treated macrophage to validated this role. The inflammation relatedproteins (NF-KB, GRP78) and the cytokines (TNF-α,IL-6) were deteced to reveal the mechanism of CO onFFA triggered infalmmation with macrophage Preliminarily, and this work may help to improve thepathogenic research and the treatment of obesity and related diseases.Methods:1. FFA treated RAW264.7macrophage was used as cell model.2. The viability of macrophage treated by different concentrations of FFA was measured with MTTmethod.3. Macrophage was treated with different dose of FFA for various time and the secreted TNF-α and IL-6were deteced to determine the optimal concentration for FFA.4. Macrophage was treated with different dose of CORM-2(the donor of CO), and the levels of TNF-αand IL-6were measured by ELISA to determine the optimal concentration for CO.5. Cell in different conditons, the control group, the FFA treated group, the CO treated group and the FFAplus CO group, were treated for24hours, and the protein levels of p-NF-κB and GRP78were detecedby western blot. The secreted inflammatory factors, TNF-α and IL-6, were determined with ELISA.Results:1. Comparing with control group, FAA of different cencentration（25、50、100、200μM）didn’t inhibitthe viability of RAW264.7cells.2. FFA had dose-and time-dependent effect on the secretion of TNF-α and IL-6, and this two cytokinesstarted to increase at12hour and24hours respectively after treatment. The most significant changes forboth the cytokine occured at24hours. Hence, the optimal FFA concentration and the measuring time forTNF-α and IL-6were chosen as100μM and24hour respectively.3. CORM-2had inhibitory and dose-dependent effect on100μM FFA triggered inflammatory response onmacrophage, manifesting by the level of TNF-α and IL-6. There’s no significant difference between thegroups treated with CORM-2of100μM and200μM, so the optimal dose for CORM-2was set as100μM.4. Comparing with the control group, p-NF-κB and GRP78significantly elevated in the FFA treated group,and these effects were significantly inhibited by CORM-2.Conclusions:1. FAA could induce dose-and time-dependent inflammatory response on macrophage.2. CO could inhibit the FFA-induced inflammatory response remarkably, and the mechanisms may berelated to the inhibition of NF-kB phosphorylation and the down-regulation of GRP78.