目的 探討三七總皂苷（PNS）對(duì)脂多糖（LPS）誘導(dǎo)的RAW264.7細(xì)胞iNOS-NO-NF-κB信號(hào)通路相關(guān)分子表達(dá)及活性的影響。方法 MTT比色法檢測(cè)PNS對(duì)RAW264.7細(xì)胞增殖的抑制作用；不同濃度PNS（25、50、100 μg/mL）干預(yù)體外培養(yǎng)LPS誘導(dǎo)的RAW264.7細(xì)胞24、48 h后，Griess法檢測(cè)NO變化；PNS干預(yù)24 h后，Western blotting檢測(cè)iNOS、NF-κB、p-NF-κB、IKKα、p-IKKα、p-ΙκΒα蛋白表達(dá)量的變化；PNS干預(yù)4 h后，激光共聚焦顯微鏡檢測(cè)NF-κB轉(zhuǎn)位入核情況。結(jié)果 PNS濃度大于150 μg/mL時(shí)，才表現(xiàn)出顯著抑制細(xì)胞增殖的作用。25、50、100 μg/mL PNS作用于RAW264.7細(xì)胞24和48 h后，與模型組比較，NO生成量均顯著降低（P<0.001）。50、100 μg/mL PNS作用于RAW264.7 24 h后，iNOS、NF-κB蛋白表達(dá)量顯著降低，磷酸化蛋白p-NF-κB、p-IKKα、p-ΙκΒα表達(dá)水平也顯著降低（P<0.01、0.001）。與模型組比較，PNS 25、50、100 μg/mL組核因子p65入核熒光強(qiáng)度顯著降低（P<0.01、0.001）。結(jié)論 PNS能夠顯著抑制LPS誘導(dǎo)的RAW264.7細(xì)胞iNOS-NO-NF-κB信號(hào)通路的活性，降低細(xì)胞炎癥反應(yīng)。

Objective To investigate the effect of Panax notoginseng saponins (PNS) on the expression and activity of iNOS-NONF-κB signaling pathway related molecules in lipopolysaccharide (LPS)-stimulated RAW264.7 cells in vitro. Methods RAW264.7 cells were treated with different concentrations (25-100 μg/mL) of PNS for different periods of time (24-48 h). Antiinflammation effects of PNS in LPS-stimulated RAW264.7 macrophages were measured based on the detection of nitric oxide (NO) overproduction (Griess method, DAF-FM DA fluorescence assay and NO₂^- scavenging assay), and the cell proliferation was measured by MTT assay, the expression of iNOS, NF-κB, p-NF-κB, IKKα, p-IKKα, p-IκBα by Western blot, the NF-κB p65 translocation by confocal laser scanning fluorescence microscopy. Results When the concentration of PNS was more than 150 μg/mL, the inhibition of cell proliferation was significant. 25, 50, 100 μg/mL PNS treated RAW264.7 cells for 24 and 48 h, compared with the model group, NO production was significantly reduced (P<0.001). After PNS (25, 50 and 100 μg/mL) was applied to RAW264.7 cells for 24 h, the expression of iNOS, NF-κB proteins and the expression of p-NF-κB, p-IKKα, p-IκBα proteins were all decreased in a dose-dependment manner (P<0.01, 0.001). Conclusion Taken together, PNS exerts anti0inflammatory effects, by inhibiting the activity of iNOS-NO-NF-κB signaling pathway in LPS-treated RAW264.7 macrophages.

R285.5

國(guó)家自然科學(xué)基金資助項(xiàng)目（81760818、81160492）；云南省應(yīng)用基礎(chǔ)研究計(jì)劃項(xiàng)目-中醫(yī)聯(lián)合專項(xiàng)[2017FF11（-013）]