Mechanism Of Wnt And NF-?B Signaling Pathways In LPS-stimulated BV2 Cellular Inflammation

Objective:To study the regulation mechanism of Wnt/?-catenin signaling pathway and NF-?B signaling pathway on lipopolysaccharide(LPS)-induced BV2 cell inflammation and to explore its role in neuroinflammation.Methods:BV2 cells were cultured in a DMEM high glucose-containing complete culture medium containing 5%fetal bovine serum(FBS)and placed in a 37?,5%C02 incubator.When the cells growth well density moderate,divided them randomly into:?blank control group(Control):5%DMEM high-glucose complete culture medium was used without any treatment;?LPS treatment group(LPS):5%DMEM high-glucose complete culture medium was used,then add lug/ml LPS to treat cells for different time(0.5h,1h,2h,4h,6h);?inhibitor group(LPS+XAV939):use 5%DMEM high glucose type complete culture medium,add ImM XAV939 pretreatment cells for 6h.The cells were then treated with 1 ?g/ml LPS for 1 h.RT-PCR was used to detect the expression of inflammatory related factors such as tumor necrosis factor-a(TNF-a)and cyclooxygenase-2(COX-2);?-catenin,phosphorylated nuclear factor-icB(p-NF-?B),phosphorylated nuclear factor-?B inhibitory protein(p-I?B)was detected by Western Blotting;immunofluorescence assay for TNF-a,COX-2,nuclear factor-?B(NF-?B),?-catenin expression and nuclear translocation of NF-?B and ?-catenin.Results:?Compared with the blank control group,LPS-stimulated BV2 cells 0.5 h,1 h,2 h,4 h,and 6 h,expressions of TNF-a,COX-2 genes were up-regulated;immunofluorescence suggested that LPS stimulated BV2 after 1h,the translational levels of TNF-a and COX2 were up-regulated in the LPS-stimulated BV2 cells;?The expression of ?-catenin,p-NF-?B and p-I?B were up-regulated after 0.5h,1h,2h,4h and 6h of LPS-stimulated BV2 cells and significantly upregulate At 1h.Immunofluorescence double staining suggested that the nuclear translocation of NF-?B and ?-catenin was significantly increased compared with the blank group.?Compared with LPS treatment group,the transcription and translation levels of TNF-a and COX-2 in the inhibitor group were significantly down-regulated;the expression levels of ?-catenin,p-NF-?B and p-IKB were down-regulated and the nuclear translocation of NF-?B and ?-catenin was obviously decrease in inhibitor group.Conclusion:?LPS can stimulate BV2 cells to produce inflammatory reaction at different times(0.5h-6h),which is related to activation of Wnt/?-catenin signaling pathway and NF-?B signaling pathway.When LPS stimulation for 1h,the above two Signal pathway activation is most pronounced.?After inhibiting the wnt signaling pathway,the inflammatory response of BV2 cells was significantly down-regulated,and the activation of NF-?B signaling pathway was significantly inhibited-This suggests that XAV939 may downregulate the NF-?B signaling pathway by inhibitingthe activity of wnt signaling pathway,thereby down-regulating the inflammatory response.