Effects of annexin-A1 mimetic peptide Ac2-26 on inflammatory factors released from microglia

Abstract

Objective Objective To observe the effect of Annexin-A1 mimetic peptide Ac2-26 on microglia released tumor necrosis factor-a(TNF-a), interleukin-1b(IL-1b), interleukin-6(IL-6)and nitric oxide(NO), and to explore its impact on expression of Toll Like Receptor-2(TLR2). Methods Rat primary microglial cells were randomly divided into 4 groups by different treatments: Aβ1-42 group(stimulated by Aβ1-42), Aβ1-42 plus scramble control(co-stimulated by Aβ1-42 and scramble control), Aβ1-42plus Ac2-26(co-stimulated by Aβ1-42 and Ac2-26)and blank control group.After above treatments for 24 hour, the microglial cells were collected.Microglial cell viability was measured by CCK-8 assay kits.The inverted phase contrast microscope was applied to observe the morphological changes of microglia.The concentrations of TNF-a, IL-1β, IL-6 and NO in the supernatant were measured by commercial kits, and the level of TLR2 protein expression was detected by Western blot assay. Results The Aβ1-42 and Ac2-26 had no effect on the microglial cell viability.After activation with Aβ1-42, the microglia showed an obvious morphological change.The microglia cell soma showed obvious changes, from resting ramified state into the activated amoeboid macrophage-like appearance.The Aβ1-42 group, Aβ1-42 + scramble control group, Aβ1-42 + Ac2-26 group and blank control group showed that the supernatant levels of TNF-a were(125.17±7.13), (118.78±7.28), (24.02±2.62), (20.89±1.82)ng/L, respectively, the supernatant levels of IL-1β were(117.61±8.73), (108.90±6.53), (27.06±3.32), (24.58±3.45)ng/L, respectively, and the supernatant levels of IL-6 were(108.6 7±5.86), (102.67±4.85), (25.94±2.83), (19.68±2.66)ng/L, respectively.The supernatant levels of TNF-a, IL-1β and IL-6 were significantly lower in the Aβ1-42 + Ac2-26 group than in Aβ1-42 group and Aβ1-42 + scramble control groups, and had no significant difference compared with blank control group; and the supernatant level of NO showed the similar result as the above.The supernatant levels of NO were lower in Aβ1-42 + Ac2-26 group(20.27±2.53)mmol/L than in Aβ1-42 group(75.68±6.14)mmol/L and in Aβ1-42 + scramble control group(69.25±4.50)mmol/L, and had no statistical significance compared with blank control group(16.39±2.96)mmol/L.The protein expression level of TLR2 was significantly downregulated in the Aβ1-42 + Ac2-26 group than in Aβ1-42 group and Aβ1-42 + scramble control group.However it showed no significant difference compared with blank control group. Conclusions Ac2-26 can effectively inhibit the released TNF-a, IL-1β, IL-6 and NO from microglia induced by Aβ1-42, mainly through downregulation TLR2 .Ac2-26 may serve as a novel targeted drug for treatment of Alzheimer's Disease. Key words: Alzheimer Disease; Microglia; Ac2-26; Inflammation; Toll Like Receptor-2