Effects Of Piper Kadsura Ohwi On IL-1β And TNF-α Expression In Activated Microglia

ObjectiveThis study aimed to explore the activation of microglia cells by different state of aggregation of Aβ1-42at the gene level, and explore the inhibitory effect of extraction from Piper kadsura Ohwi on the release of proinflammatory eytokines IL-1β and TNF-a of microglia cells activated by different state of aggregation of Aβ1-42and to investigate the method of the preparation Aβ1-42oligomers.Methods1. Mice microglia cell line (BV2) was cultured in vitro;2. The consistent soluble oligomeric and fibrillar Aβ1-42were prepared and tested under the transmission electron microscope;3. Microglia cells were divided randomly into4groups:a blank contrast group, without Aβ1-42group, oligomeric Aβ1-42group, fibrillar Aβ1-42group, the cytotoxicity of different extraction from Piper kadsura Ohwi was assessed by Methyl thiazolyl tetrazolium(MTT).4. Microglia cells were divided randomly into5test groups,intervened by oligomeric Aβ1-42, oligomeric Aβ1-42+Piper kadsura Ohwi extracts, fibrillar Aβ1-42and fibrillar Aβ1-42+Piper kadsura Ohwi extracts respectively, besides, a blank contrast group was set without any intervention.48hours later, the expression of IL-1β and TNF-a was assessed by realtime PCR.Results1. Oligomeric Aβ1-42were existed in monomer, spherical or particulate form in diameter at15-30nm without fiber forming, which were observed by the electron microscope after2w incubation at4℃; Fibrillar Aβ1-42were existed in fibrillar crossing, partly oligomeric form after1w incubation at37℃.2. MTT showd that there was no effect on cell survival with the various concentration of Piper kadsura Ohwi extracts (10-80g/L);3. Realtime PCR showed that each of the4test groups had a higher expression of IL-1βand TNF-a than the blank contrast; The oligomeric Aβ1-42+Piper kadsura Ohwi extracts group had a lower expression of IL-1βand TNF-a than the oligomeric Aβ1-42group; likewise, the fibrillar Aβ1-42+Piper kadsura Ohwi extracts group had a lower expression of IL-1β and TNF-a than the fibrillar Aβ1-42group.Conclusion1. Both fibrillar and oligomeric Aβ1-42could activate the microglia at the gene level.2. Their activation of microglia could be inhibited by Piper kadsura Ohwi extracts.3. The preparation of stable oligomer Aβ1-42was succeeded.