研究目的：

    本研究旨在通过痤疮丙酸杆菌生物膜构建寻常痤疮（Acne Vulgaris，AV）炎症小鼠模型，探讨中药有效成分槐属二氢黄酮G（Sophoraflavanone G，SFG）对寻常痤疮炎症的抑制效应及其初步作用机制。

研究方法：

1.寻常痤疮炎症小鼠模型的建立：选取健康的6-8周雌性野生型（Wild-type，WT）SPF级ICR小鼠，建立痤疮丙酸杆菌生物膜诱导的寻常痤疮炎症小鼠模型。通过测量耳厚度、耳重量、皮肤组织病理（HE染色）和免疫组化法评估寻常痤疮炎症小鼠模型。

2. 研究中药成分槐属二氢黄酮G对寻常痤疮炎症小鼠模型的炎症抑制作用：选取健康的6-8周龄、雌性野生型SPF级ICR小鼠43只，除正常对照组（3只）外，随机分成模型组、溶解基质对照组、槐属二氢黄酮G组、阳性药物对照组。除对照组外，其余4组造模方式同前，该天指定为0天。24h后，以注射点为中心在双侧鼠耳相同位置局部外用20μL溶解基质、体积分数2.5%槐属二氢黄酮G及氯霉素洗剂。继续在SPF环境中饲养24h，检测前述指标评估槐属二氢黄酮G对寻常痤疮炎症小鼠模型的抑制作用。

3．初步探索中药成分槐属二氢黄酮G对寻常痤疮炎症小鼠模型的抑制作用机制：按课题组既往方法建立痤疮丙酸杆菌体外生物膜模型，使用临床常用抗生素米诺环素及中药苦参成分槐属二氢黄酮G共孵育，对比其体外抗生物膜能力；取前述经SFG处理的寻常痤疮炎症小鼠，行蛋白免疫印迹法检测鼠耳组织中NLRP3、Caspase-1蛋白表达量，检测评估SFG对寻常痤疮相关炎症通路的抑制作用。

研究结果：

1．寻常痤疮炎症小鼠模型的建立：与对照组相比，造模组耳部皮肤出现炎性红斑基础上的丘疹脓疱，周边皮肤毛细血管明显扩张；鼠耳厚度较对照组明显升高，差异具有统计学意义；皮损部位HE染色可见经痤疮丙酸杆菌生物膜感染48h后表皮角质层增厚、毛囊皮脂腺开口部位角化过度，皮脂腺体积增大，真皮内炎性细胞浸润，并可见中性粒细胞聚集形成脓腔结构的急性炎症表现；免疫组化染色可见鼠耳皮肤组织的IL-1β表达量明显增加，与对照组相比差异具有统计学意义。

2．中药成分槐属二氢黄酮G对寻常痤疮炎症小鼠模型的炎症抑制作用：与模型组相比，外用2.5%槐属二氢黄酮G后，小鼠耳部红斑明显减轻，周边皮肤毛细血管扩张程度减轻，未见明显脓疱皮损产生，耳厚度及耳重量较对照组明显降低，差异具有统计学意义；皮损部位HE染色未见明显炎症细胞浸润，免疫组化染色可见鼠耳皮肤组织的IL-1β表达量较对照组明显降低，差异具有统计学意义。

3．初步探索中药成分槐属二氢黄酮G对寻常痤疮炎症小鼠模型的抑制作用机制：与空白对照组相比，激光共聚焦显微镜示中、高浓度的槐属二氢黄酮G，类似米诺环素，能通过破坏成熟生物膜结构起到抗生物膜作用，蛋白质免疫印迹示经槐属二氢黄酮G处理后鼠耳组织中NLRP3、Caspase-1蛋白表达量降低。

研究结论：

    鼠耳皮内注射痤疮丙酸杆菌生物膜能够成功构建寻常痤疮炎症小鼠模型；外用传统中药苦参的有效成分槐属二氢黄酮G能够有效抑制痤疮丙酸杆菌生物膜所诱导的寻常痤疮炎症小鼠模型的炎症反应；该药物能够通过破坏痤疮丙酸杆菌生物膜结构起到抗生物膜作用及抑制NLRP3炎症小体通路来实现抗炎作用。

Research Objectives:

This study aims to establish an Acne Vulgaris Inflammation Mouse Model and investigates the anti-inflammatory effect of traditional Chinese medicine active ingredient Sophoraflavanone G (SFG) on acne vulgaris, as well as explores its preliminary mechanism.

Research Methods:

1.Establishment of Acne Vulgaris Inflammation Mouse Model: Healthy 6-8 weeks-old female wild-type (WT) SPF-grade ICR mice were selected to construct an Acne Vulgaris Inflammation Mouse Model. The success of this model was evaluated by measuring ear thickness, ear weight, histopathological examination (HE staining), and immunohistochemical analysis of skin tissue.

2.Evaluation of the anti-inflammatory effects of SFG on Acne Vulgaris Inflammation Mouse Model: Forty-three healthy 6-8 weeks-old female ICR mice were randomly divided into control group (n=3), model group（n＝10）, SFG-treated group（n=10,Vehicle control group (n=3)and Standard treatment control (chloramphenicol lotion) group（n=10）. Except for the control group, other groups were subjected to the same modeling procedure (designated as Day 0). After 24 h, 20 μL of solvent, 2.5% SFG, or chloramphenicol lotion was topically applied to the injection site. Following treatment, mouses were housed under SPF conditions for a further 24-hour period prior to assessment of the specified parameters to determine the anti-inflammatory activity of Sophoraflavanone G on Acne Vulgaris Inflammation Mouse Model.

3.Preliminary exploration of the mechanism of SFG’s anti-inflammatory effects: We firstly established an in-vitro biofilm model of Cutibacterium acnes according to our previous methods. The clinically used antibiotic minocycline and the Chinese herbal component Sophoraflavanone G were co-incubated to compare their in vitro anti-biofilm activities. Additionally, Western blotting was performed to detect the expression levels of NLRP3 and Caspase-1 proteins in the ear tissues of treated mice.

Research Results:

Successful establishment of Acne Vulgaris Inflammation Mouse Model: Compared with the control group, the model group exhibited inflammatory erythema, papμLes, and pustules on the ears, with significant dilation of surrounding capillaries. The mouse ear thickness was significantly increased compared to the control group, and the difference was statistically significant(p < 0.05). HE staining revealed hyperkeratosis, sebaceous gland hyperplasia, and dense inflammatory cell infiltration, including neutrophil aggregation forming micro-abscesses. Immunohistochemistry confirmed elevated IL-1β expression (p < 0.05).

2.SFG alleviates acne inflammation in the mouse model: Compared with the model group, topical 2.5% SFG significantly alleviates erythema, capillary dilation, and pustule formation. Ear thickness and weight were markedly decreased (p < 0.05).HE staining showed minimal inflammatory infiltration, and immunohistochemistry demonstrated reduced IL-1β expression (p < 0.05).

3.Mechanistic insights into SFG’s anti-inflammatory effects: Confocal microscopy revealed that mid-concentration and high-concentration of  SFG, similar to minocycline, disrupted mature C.acnes biofilms. Western blotting indicated that SFG downregulated NLRP3 and Caspase-1 protein expression.

Conclusion:

The traditional Chinese medicine active ingredient Sophoraflavanone G, derived from Sophora flavescens, alleviates C.acnes biofilm-induced  inflammation by disrupting bacterial biofilms and suppressing the NLRP3 inflammasome pathway.