目的 优化牛膝微沉淀自组装纳米粒（Achyranthis Bidentatae Radix micro-precipitation self-assembled nanoparticles，ABR-MP-SAN）的制备工艺，将其与牛膝传统水煎自组装纳米粒（Achyranthis Bidentatae Radix decoction self-assembled nanoparticles，ABR-D-SAN）进行比较，并基于网络药理学与动物实验评价其抗急性痛风性关节炎（acute gouty arthritis，AGA）的作用与机制。方法 采用微沉淀法制备ABR-MP-SAN，以单因素实验结合响应面法优化提取时间、磁力搅拌时间、生药质量浓度等工艺参数。对优化后的ABR-MP-SAN与ABR-D-SAN的表征及多糖、甾酮含量进行比较。利用网络药理学预测ABR-MP-SAN抗AGA的核心成分、关键靶点及信号通路，并进行分子对接验证。将C57BL/6小鼠分为空白组、模型组、秋水仙碱（1 mg/kg）组、ABR-D-SAN（1.82 g/kg）组和ABR-MP-SAN（1.82 g/kg）组，通过踝关节腔处注射尿酸钠混悬液诱导AGA模型，ig给药7 d后，以检测关节肿胀度、步态评分、血清炎症因子、踝关节组织病理变化与白细胞介素-17A（interleukin-17A，IL-17A）、肿瘤坏死因子-α（tumor necrosis factor-α，TNF-α）、核因子-κB p65（nuclear factor-κB p65，NF-κB p65）蛋白表达来评价药效与机制。结果 ABR-MP-SAN最优工艺为牛膝加10倍量水回流提取60 min，滤渣加10倍量75%乙醇提取60 min，合并2次提取液，400 r/min、25 ℃条件下磁力搅拌25 min，减压除醇后浓缩至0.2 g/mL。所得ABR-MP-SAN呈规则球形，其稳定性、均匀性、多糖和甾酮含量都高于ABR-D-SAN。网络药理学显示核心成分为山柰酚、黄芩素、β-蜕皮甾酮等，关键靶点为IL-1β、TNF、IL-6，主要富集于IL-17、TNF和NF-κB信号通路，分子对接表明核心成分与靶点间具有较强亲和力。动物实验结果显示，与空白组相比，模型组小鼠关节肿胀度、步态评分和血清IL-6、IL-1β、TNF-α水平均明显上升（P＜0.001），踝关节出现明显病理损伤，IL-17A、TNF-α、NF-κB p65阳性表达显著升高（P＜0.01、0.001）；与模型组相比，给药组中上述情况均得到改善，且ABR-MP-SAN组效果明显优于ABR-D-SAN组。结论 微沉淀法制备得到的ABR-MP-SAN粒径小、稳定性好且分布均匀，其抗AGA疗效好，机制可能与抑制IL-17/NF-κB和TNF/NF-κB信号通路、减轻炎症反应有关。

Objective To optimize the preparation process of Achyranthis Bidentatae Radix micro-precipitation self-assembled nanoparticles (ABR-MP-SAN), compare them with ABR decoction self-assembled nanoparticles (ABR-D-SAN), and evaluate their anti-acute gouty arthritis (AGA) effects and mechanisms via network pharmacology and animal experiments. Methods ABR-MP-SAN were prepared using a micro-precipitation method. Single-factor experiments combined with response surface methodology were employed to optimize process parameters including extraction time, magnetic stirring time, and crude drug concentration. The optimized ABR-MP-SAN were compared with ABR-D-SAN in terms of characterization, polysaccharide content, and ecdysteroid content. Network pharmacology was used to predict core components, key targets, and signaling pathways of ABR-MP-SAN against AGA, followed by molecular docking validation. C57BL/6 mice were divided into blank group, model group, colchicine (1 mg/kg) group, ABR-D-SAN (1.82 g/kg) group, and ABR-MP-SAN (1.82 g/kg) group. AGA was induced by intra-articular injection of monosodium urate crystals into the ankle joint cavity. After 7 d of intragastric administration, therapeutic efficacy and mechanisms were evaluated by joint swelling degree, gait score, serum inflammatory cytokines, histopathological changes of ankle joint, and protein expression of interleukin-17A (IL-17A), tumor necrosis factor-α (TNF-α), and nuclear factor-κB p65 (NF-κB p65). Results The optimal process for ABR-MP-SAN was ABR was reflux-extracted with 10 volumes of water for 60 min, the residue was extracted with 10 volumes of 75% ethanol for 60 min, the two extracts were combined, magnetically stirred at 400 r/min and 25 ℃ for 25 min, then concentrated to 0.2 g/mL after ethanol removal under reduced pressure. The obtained ABR-MP-SAN exhibited regular spherical shape, with higher stability, uniformity, polysaccharide content, and ecdysteroid content all higher than those of ABR-D-SAN. Network pharmacology identified core components including kaempferol, baicalein, and β-ecdysterone; key targets including IL-1β, TNF, and IL-6; major pathways enriched in IL-17, TNF, and NF-κB signaling pathways. Molecular docking demonstrated strong binding affinity between core components and targets. Animal experiments showed that compared with the blank group, the model group exhibited significantly increased joint swelling degree, gait score, serum IL-6, IL-1β, and TNF-α levels (P < 0.001), obvious pathological injury of ankle joint, and markedly elevated positive expression of IL-17A, TNF-α, and NF-κB p65 (P < 0.01, 0.001). Compared with the model group, all treatment groups showed improvements, with the ABR-MP-SAN group exhibiting significantly better efficacy than the ABR-D-SAN group. Conclusion ABR-MP-SAN prepared by the micro-precipitation method possess small particle size, good stability, and uniform distribution. They demonstrate potent anti-AGA effects, and the mechanism may involve inhibition of IL-17/NF-κB and TNF/NF-κB signaling pathways to alleviate inflammatory responses.

R283.6

云南省傣医药与彝医药重点实验室开放课题（2024SS24077）；云南民族药二次开发关键技术集成及示范项目（202303AC100025）