目的 制备木芙蓉叶有效组分水凝胶微针，系统考察其理化性质，并评价其对急性乳腺炎模型大鼠的治疗效果与作用机制。方法 采用化学交联法结合模具法制备木芙蓉叶有效组分水凝胶微针。以微针成型性、溶胀度、刺孔率为评价指标，通过单因素实验筛选关键影响因素，结合Box-Behnken设计-响应面法（BBD-RSM）优化交联温度、干燥方式、基质［聚甲基乙烯基醚共聚马来酸（S-97）、聚乙二醇10000（PEG 10000）、无水碳酸钠（Na₂CO₃）］用量等工艺参数。利用扫描电子显微镜（SEM）和体视显微镜表征微针的外观形态与结构特征；采用高效液相色谱法（HPLC）测定微针中芦丁、异槲皮苷、山柰酚的含量；通过铝箔穿刺实验、离体鼠皮穿刺实验，结合穿刺后鼠皮的苏木精-伊红（HE）染色组织病理学检查，综合评估微针的机械穿刺性能；同时考察微针的体外溶胀性能及在体皮肤屏障修复能力。建立脂多糖（LPS）诱导的大鼠急性乳腺炎模型，通过乳腺组织HE染色，观察病理损伤改善情况；免疫组化法检测乳腺组织中巨噬细胞标志物CD68⁺的表达水平；酶联免疫吸附测定法（ELISA）检测乳腺组织中肿瘤坏死因子（TNF）-α、白细胞介素（IL）-6、IL-1β 3种促炎因子的表达量。结果 确定最优制备工艺：真空干燥箱程序升温干燥，交联温度80℃；微针基质最优处方为S-97 0.97 g、PEG100000.43 g、Na₂CO₃ 0.11 g。该工艺下制备的微针铝箔刺孔率达（97.11±1.27）%，溶胀度为（578.55±50.45）%，且成型性良好。微针可稳定负载3%的木芙蓉叶最佳配伍组分（每克基质含45 mg配伍组分），每片微针含药量1.5 mg； HPLC检测显示，微针中芦丁、异槲皮苷、山柰酚的质量分数分别为（1.101 0±0.013 2）、（0.165 3±0.002 4）、（0.035 4±0.000 4） mg·g^-1。形态表征结果显示，微针为圆形贴片，表面光滑、阵列完整，共含385颗圆锥状针头，针体长度约550 μm、底部直径约300 μm、针尖间距约700 μm。透皮实验表明，48 h内微针中芦丁的累积渗透率达90.7%，异槲皮苷与山柰酚的累积渗透率均约为77.0%，透皮性能优异。药效学结果显示，与模型组相比，木芙蓉叶有效组分微针高剂量组可显著降低大鼠乳腺组织中CD68⁺的活性（P＜0.01），同时显著下调TNF-α、IL-1β、IL-6 3种促炎因子的表达水平（P＜0.01），且乳腺组织病理损伤明显减轻。结论 成功制备的木芙蓉叶有效组分水凝胶微针具有良好的机械穿刺性能、药物负载能力与透皮释药特性，可有效抑制LPS诱导的大鼠急性乳腺炎。其作用机制可能与降低乳腺组织中CD68⁺巨噬细胞活性，抑制TNF-α、IL-1β、IL-6促炎因子表达，从而减轻炎症反应相关。

Objective To prepare water gelatinous microneedles of effective components from Hibiscus mutabilis leaves, systematically investigate their physicochemical properties, and evaluate their therapeutic effects and mechanisms on a rat model of acute mastitis. Methods The water gelatinous microneedles were prepared by a chemical cross-linking method combined with a mold method. The formation, swelling degree, and puncture rate of the microneedles were used as evaluation indicators. The key influencing factors were screened by single-factor experiments, and the process parameters such as cross-linking temperature, drying method and matrix (S-97, PEG10000, and Na₂CO₃) dosage were optimized by Box-Behnken design-response surface methodology (BBD-RSM). The appearance morphology and structural characteristics of the microneedles were characterized by scanning electron microscopy (SEM) and stereomicroscopy. The drug loading and content of rutin, isorhamnetin, and kaempferol in the microneedles were determined by high-performance liquid chromatography (HPLC). The mechanical puncture performance of the microneedles was comprehensively evaluated by aluminum foil puncture experiments, ex vivo rat skin puncture experiments, and histopathological examination of the punctured rat skin by HE staining. Meanwhile, the in vitro swelling performance and skin barrier repair ability of the microneedles were investigated. A rat model of acute mastitis induced by lipopolysaccharide (LPS) was established. The improvement of pathological damage in the mammary gland tissue was observed by HE staining. The expression level of the macrophage marker CD68⁺ in the mammary gland tissue was detected by immunohistochemistry. The expression levels of three proinflammatory factors, TNF-α, IL-6, and IL-1β, in the mammary gland tissue were determined by enzyme-linked immunosorbent assay (ELISA). Results The optimal preparation process was determined as follows: drying in a vacuum drying oven with a cross-linking temperature of 80 ℃. The optimal prescription for the microneedle matrix was 0.97 g of S-97, 0.43 g of PEG10000, and 0.11 g of Na₂CO₃. Under this process, the aluminum foil puncture rate of the prepared microneedles was (97.11 ±1.27)%, the swelling degree was (578.55 ±50.45)%, and the formation was good. The microneedles could stably load 3% of the best combination of effective components from H. mutabilis leaves (45 mg of the combination per gram of matrix), with a drug content of 1.5 mg per microneedle. HPLC detection showed that the mass fractions of rutin, isorhamnetin, and kaempferol in the microneedles were (1.101 0 ±0.013 2), (0.165 3 ±0.002 4), and (0.035 4 ±0.000 4) mg·g^-1, respectively. Morphological characterization results showed that the microneedles were circular patches with smooth surfaces and complete arrays, containing 385 conical needle tips, with a needle length of approximately 550 μm, a bottom diameter of approximately 300 μm, and a tip spacing of approximately 700 μm. Transdermal experiments indicated that the cumulative penetration rate of rutin from the microneedles reached 90.7% within 48 h, and the cumulative penetration rates of isorhamnetin and kaempferol were approximately 77.0%, demonstrating excellent transdermal performance. Pharmacological results showed that compared with the model group, the high-dose group of H. manihot leaves effective component microneedles significantly reduced the activity of CD68⁺ macrophages in rat mammary gland tissue (P < 0.01), and significantly downregulated the expression levels of TNF-α, IL-1β, and IL-6 pro-inflammatory factors (P < 0.01), with significant alleviation of pathological damage in the mammary gland tissue. Conclusion The successfully prepared water gelatinous microneedles of effective components from H. mutabilis leaves have good mechanical puncture performance, drug loading capacity, and transdermal drug release characteristics, and can effectively inhibit LPS-induced acute mastitis in rats. The therapeutic mechanism may be related to the reduction of CD68⁺ macrophage activity in mammary gland tissue and the inhibition of TNF-α, IL-1β, and IL-6 pro-inflammatory factor expression, thereby alleviating the inflammatory response.

R944.4;R285.5

贵州省高层次创新型人才项目（黔科合平台人才-GCC[2023]037）；国家苗药工程技术研究中心能力提升项目（黔科合中引地[2023]006）；毕节医学高等专科学校科研项目（BJYZXK202415）