The Effect Of Piper Kadsura Extract On Aβ Oligomer-induced IL-1β And IL-6 Secretion From Microglia

Objective: To explore the method of preparation and characterization of oligomers, to explore the inflammatory response of microglia induced bβoligomers and Aβfiber, and explore the inhibitory effects of Piper kadsura extract on this inflammatory response.Methods: 1.To get Aβmonomer, dissolve the lyophilized Aβin HFIP; allow HFIP to evaporate in the open tubes in the biological safety cabinet; dissolve the Aβin DMSO, and then dissolve the DMSO-Ap in Ham's F-12; transfer to 4℃and incubate for 24 h; observe the prepaired Aβoligomers under the transmission electron microscope.2. The cultured microglial cells were divided into 7 groups and each group with five parallel holes:①blank control;②Aβ25-35 oligomers;③Aβ25-35 oligomers+Piper kadsura extract;④Aβ25-35 oligomers+DMSO;⑤Aβ25-35 fiber;⑥Aβ25-35 fiber+Piper kadsura extract;⑦Aβ25-35 fiber+DMSO.48 hours later, collect the supernatants to test IL-1βand IL-6 with ELISA.Results:1. Aβ42 and AP25-35 oligomers in diameter at 12-25nm and 10-20nm respectively were observed after 1h incubation at 4℃; After 24h incubation at 4℃, there is no significantly change in diameter of Aβ42 oligomers compared with the Aβ42 oligomers incubated 1h and so do Aβ25-35 oligomers after 48h incubation at 4℃; there is no fiber formed.2. Compared with the control group, the level of IL-1βand IL-6 in the supernatant of Aβoligomers group and Aβfibril group increase significantly (P<0.01) after 48h stimulation of microglial cells; the level of IL-1βand IL-6 in the supernatant of Aβoligomers group is significantly higher than Aβfibril group (P<0.01); the level of IL-1βand IL-6 in the supernatant of Aβoligomers group is significantly higher than Aβoligomers+Piper kadsura extract group(P<0.01); the level of IL-1βin the supernatant of Aβfiber group is significantly higher than Aβfiber+Piper kadsura extract group(P<0.01); the level of IL-6 in the supernatant of Aβfiber group is significantly higher than Aβfiber+Piper kadsura extract group(P<0.05).Conclusion:1. Aβ42 oligomers and AP25-35 oligomers in diameter at 12-25nm and 10-20nm respectively were observed under the transmission electron microscope. 2. Aβoligomers are stronger than the Aβfiber in activation of microglia; Piper kadsura extract can significantly inhibite the Aβoligomers and Aβfiber induced microglial activation.