探究原人参三醇（20S-Protopanaxatriol, PPT）对脂多糖（Lipopolysaccharide, LPS）和三磷酸腺苷（Adenosine Triphosphate, ATP）双诱导激活Nod样受体蛋白3（NOD-Like Receptor Thermal Protein Domain Associated Protein 3, NLRP3）炎症小体的影响，从而发挥改善巨噬细胞炎症的作用。将RAW264.7细胞分为空白对照组、LPS组、LPS+ATP组和PPT给药组（20、40、80 μmol/L）。经LPS+ATP诱导后，一氧化氮合酶（Inducible Nitric Oxide Sythase, iNOS）、环氧合酶2（Cyclooxygenase-2, COX-2）、白细胞介素1β（Interleukin-1β，IL-1β）、肿瘤坏死因子-α（Tumor Necrosis Factor-α, TNF-α）和NLRP3的mRNA水平显著提高。经PPT处理可显著下调iNOS、IL-1β和NLRP3的mRNA水平，而不影响COX-2和TNF-α的mRNA水平。同时，PPT可下调胞质中NLRP3、IL-1β和半胱天冬酶1（Cysteinyl Aspartate Specific Proteinase-1, Cleaved-caspase-1）蛋白的表达量，而不影响核内转录因子κB（Nuclear Factor Kappa-B, NF-κB）蛋白的水平。酶连免疫吸附（Enzyme-Linked Immunosorbnent Assay, ELISA）测定给药组可显著下调IL-1β的表达量，而对TNF-α表达量无显著变化。此外，细胞形态学观察结果显示PPT可显著改善RAW264.7炎症细胞形态，如伪足减少，细胞褶皱平缓，恢复圆形形态。该研究证明PPT可能通过抑制NLRP3炎症小体激活，进一步抑制炎症因子IL-1β的表达水平，从而发挥抗炎作用。为后期探究PPT如何靶向NLRP3炎症小体以及相关药物研发提供理论依据。

The effects of 20(S)-protopanaxatriol (PPT) on NOD-like receptor thermal protein domain-associated protein 3 (NLRP3) inflammasome activation induced by lipopolysaccharide (LPS) and adenosine triphosphate (ATP) were investigated to examine the role of PPT in alleviating macrophage inflammation. RAW264.7 cells were divided into a control group, an LPS group, an LPS+ATP group, and three PPT treatment groups (20, 40, and 80 μmol/L). The mRNA expression levels of inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), Interleukin-1β (IL-1β), tumor necrosis factor-α (TNF-α), and NLRP3 were significantly increased after LPS+ATP treatment. PPT treatment markedly downregulated the mRNA levels of iNOS, IL-1β, and NLRP3; however, there were no significant changes in those of COX-2 and TNF-α. Additionally, the expression levels of NLRP3, IL-1β, and cysteinyl aspartate-specific proteinase-1 in the cell lysate of the PPT treatment group were lower than those in the LPS+ATP group; however, PPT had no effect on the translocation of nuclear factor kappa-B. Enzyme-linked immunosorbent assay revealed that PPT significantly reduced IL-1β expression levels without affecting TNF-α levels. Scanning electron microscopy observation showed that PPT notably improved the inflammatory state of RAW264.7 cells. Specifically, there was a decrease in pseudopodia formation, whereas cell folds were smoothed resulting in rounded cells. This study demonstrated that PPT exhibits immunomodulatory and anti-inflammatory effects. PPT can regulate inflammation in macrophages by inhibiting NLRP3 inflammasome activation and subsequently reducing the expression level of IL-1β. This study provides a theoretical basis for exploring the mechanism by which PPT targets the NLRP3 inflammasome and its potential impact on related drug development.

江苏省高等学校基础科学（自然科学）研究重大项目（23KJA550001）