目的 探討檳榔堿對脂多糖（LPS）誘導小鼠小膠質細胞BV2炎癥反應的改善作用及機制。方法 取對數(shù)生長期的BV2細胞分為空白組和LPS（0.01、0.1、1、10、20 μg/mL）組，CCK-8法檢測細胞活力，分光光度法檢測一氧化氮（NO）含量。另取細胞分為空白組、模型組、檳榔堿（10、20、40 μmol/L）組。CCK-8法檢測細胞活力；分光光度法檢測NO含量；酶聯(lián)免疫吸附法（ELISA）檢測上清中腫瘤壞死因子-α（TNF-α）、白細胞介素6（IL-6）、白細胞介素1β（IL-1β）和抗炎因子白細胞介素10（IL-10）水平；實時熒光定量PCR（qPCR）法檢測細胞中TNF-α、IL-6、IL-1β、一氧化氮合酶（iNOS）mRNA表達表達；Western blotting法檢測Toll樣受體4（TLR4）、p-p65、p65、環(huán)氧化酶2（COX2）、iNOS、胞內磷脂酰肌醇激酶（PI3K）、蛋白激酶B（Akt）、p-Akt蛋白表達水平。結果 0.01～20 μg/mL LPS誘導對BV2小膠質細胞的細胞活力無顯著影響，1 μg/mL LPS誘導顯著上調了細胞中NO含量；檳榔堿在10～40 μmol/L內對細胞活力無顯著影響，且均可緩解LPS誘導的BV2小膠質細胞損傷。與模型組相比，檳榔堿組可降低細胞內NO含量，下調炎癥因子TNF-α、IL-6、IL-1β mRNA相對表達量及其血清水平；抑制TLR4、p-P65、iNOS、COX2、PI3K、p-Akt蛋白表達量。結論 檳榔堿對LPS誘導的BV2小膠質細胞炎癥模型具有顯著改善作用，其作用機制可能與抑制NO生成、下調iNOS表達、調控TLR4/核因子-κB（NF-κB）、PI3K/Akt信號通路有關。

Objective To investigate the effects of arecoline on lipopolysaccharide (LPS) induced BV2 inflammatory response in mouse microglia and its mechanisms. Method BV2 cells were divided into blank group and LPS group (0.01, 0.1, 1, 10, 20 μg/mL). The cell viability was detected by CCK-8 method, and the content of NO was detected by spectrophotometry. Cells were divided into blank group, model group and arecoline group (10, 20, 40 μmol/L). Cell viability was detected by CCK-8 method. NO content was detected by spectrophotometry. The levels of TNF-α, IL-6, IL-1β, and anti-inflammatory factor IL-10 in supernatant were detected by enzyme-linked immunosorbent assay (ELISA). Real-time fluorescence quantitative PCR (qPCR) was used to detect the mRNA expression of TNF-α, IL-6, IL-1β, iNOS in cells. The expression levels of TLR4, p-P65, P65, COX2, iNOS, PI3K, Akt, and p-Akt were detected by Western blotting. Result In the concentration range (0.01-20 μg/mL), LPS induction had NO significant effect on the cell viability of BV2 microglia, and 1 μg/mL LPS induction significantly increased the content of NO in the cells. Arecaline had no significant effect on cell viability in the range of 10 to 40 μmol/L, and could alleviate LPS-induced damage of BV2 microglia. Compared with the model group, arecoline group could decrease the intracellular NO content, down-regulate the mRNA expression of inflammatory factors TNF-α, IL-6, IL-1β and their serum levels. The expression levels of TLR4, p-P65, iNOS, COX2, PI3K and p-Akt were inhibited. Conclusion Arecarecine can significantly improve the LPS-induced inflammation model of BV2 microglia, and its mechanism may be related to inhibiting NO production, down-regulating the expression of iNOS, and regulating the TLR4/NF-κB and PI3K/Akt signaling pathways.

R285;R971

國家重點研發(fā)計劃（2022YFD1600303）；中國農(nóng)業(yè)科學院農(nóng)產(chǎn)品加工研究所創(chuàng)新工程（CAAS-ASTIP-2023-IFST）；三亞中國農(nóng)業(yè)科學院國家南繁研究院“南繁專項”（YYLH05）；三亞中國農(nóng)業(yè)科學院國家南繁研究院“南繁專項”（ZDXM2302）