目的 基于 PI3K/Akt 信号通路探讨四神丸对溃疡性结肠炎（UC）小鼠记忆性 B 细胞的调控作用。方法 将 48 只 C57BL/6 小鼠随机分为正常组、正常+四神丸组、模型组及模型+四神丸组，每组 12 只。采用葡聚糖硫酸 钠（DSS）诱导慢性复发性 UC 小鼠模型。模型组、模型+四神丸组小鼠自由饮用 2.0%DSS 溶液以诱导肠道炎症， 连续饮用 7 d 后更换为纯水，第 14～21 天继续饮用 DSS 溶液。第 7～21 天，正常+四神丸组、模型+四神丸组 小鼠给予四神丸混悬液（2.5 g·kg-1 ·d-1 ）灌胃，每日 1 次，连续 15 d。观察小鼠体征，包括行为活动、精神状 态、体质量变化、粪便黏稠度及隐血情况，进行疾病活动指数（DAI）评分。测量小鼠结肠质量、长度，计算结 肠质量指数、结肠质量/结肠长度比值。采用 HE 染色法观察结肠组织病理形态变化；流式细胞术测定肠系膜 淋巴结及派尔集合淋巴结中 B 细胞亚群的表达水平；Western Blot 法测定结肠组织中 PI3K、Akt、p-Akt、 NF-κB p65、PTEN 蛋白表达水平。结果 与正常组比较，模型组小鼠精神萎靡，毛发枯槁无光泽，喜蜷缩聚 团，摄食量显著减少，粪便稀溏，部分可见肉眼血便；第 5～21 天的体质量显著下降（P＜0.01），第 1～21 天 的 DAI 评分显著升高（P＜0.01）；结肠长度显著缩短（P＜0.01），结肠质量、结肠质量指数及结肠质量/结肠长 度比值均显著升高（P＜0.01）；结肠组织病理损伤明显，肠壁完整性破坏，黏膜表面可见明显溃疡，腺体结构 紊乱，隐窝结构部分杂乱，同时伴炎性细胞浸润，病理损伤评分显著升高（P＜0.01）；肠系膜淋巴结及派尔集 合淋巴结中 CD19+ CD27+细胞（记忆性 B 细胞）表达水平显著升高（P＜0.01），而 CD19+ CD27+ CD5+ CD25+细胞 （CD25+ mBreg 细胞）、CD19+ CD27+ CD5+ Foxp3+ 细胞（Foxp3+ mBreg 细胞）表达水平显著降低（P＜0.05，P＜0.01）； 结肠组织中 PI3K、Akt、p-Akt、NF-κB p65 蛋白表达水平均显著升高（P＜0.01），PTEN 蛋白表达水平显著降 低（P＜0.01）。与模型组比较，模型+四神丸组小鼠精神状态得到明显改善，毛发渐趋润泽，粪便形态趋于正 常，便血症状明显缓解；第 17～21 天的体质量明显升高（P＜0.05，P＜0.01），第 11～21 天的 DAI 评分明显降 低（P＜0.05，P＜0.01）；结肠长度显著延长（P＜0.01），结肠质量、结肠质量指数及结肠质量/结肠长度比值均 显著降低（P＜0.01）；结肠组织病理损伤有明显改善，肠上皮细胞有不同程度修复，部分腺体增生，仅伴少量 炎性细胞浸润及溃疡，病理损伤评分显著降低（P＜0.01）；肠系膜淋巴结及派尔集合淋巴结中记忆性 B 细胞表 达水平显著下降（P＜0.01），而 CD25+ mBreg、Foxp3+ mBreg 细胞表达水平显著升高（P＜0.05，P＜0.01）；结肠 组织中 PI3K、Akt、p-Akt、NF-κB p65 蛋白表达水平均显著降低（P＜0.01），PTEN 蛋白表达水平显著升高 （P＜0.01）。结论 四神丸可有效缓解 DSS 诱导的 UC 小鼠结肠炎症，可能是通过抑制 PI3K/Akt 信号通路的活 化，重塑记忆性 B 细胞亚群稳态来实现的。

Objective To investigate the regulatory effect of Sishen Pills on memory B cells in ulcerative colitis （UC） mice based on the PI3K/Akt signaling pathway. Methods Forty-eight C57BL/6 mice were randomly divided into a normal group，normal + Sishen Pills group，model group，and model + Sishen Pills group，with 12 mice in each group. A dextran sulfate sodium （DSS）-induced chronic recurrent UC mouse model was established. Mice in the model group and model + Sishen Pills group were allowed to freely drink 2.0% DSS solution to induce intestinal inflammation. After 7 days，the DSS solution was replaced with pure water，and DSS administration was resumed from days 14 to 21. From days 7 to 21，mice in the normal + Sishen Pills group and model + Sishen Pills group were orally administered Sishen Pills suspension （2.5 g·kg-1 ·d-1 ） once daily for 15 consecutive days. Mouse signs， including behavioral activity， mental state，body mass changes，fecal consistency，and occult blood，were observed，and the disease activity index （DAI） was scored. Colon weight and length were measured，and the colon weight index and colon weight/length ratio were calculated. Histopathological changes in colon tissue were observed using HE staining. Flow cytometry was used to determine the expression levels of B cell subsets in mesenteric lymph nodes and Peyer’s patches. Western Blot was performed to measure the expression levels of PI3K，Akt，p-Akt，NF-κB p65，and PTEN in colon tissue. Results Compared with the normal group，mice in the model group exhibited lethargy，dull and dry fur，a tendency to huddle， significantly reduced food intake， loose stools， and visible bloody stools in some cases. Body mass from days 5 to 21 was significantly decreased （P＜0.01），while DAI scores from days 1 to 21 were significantly increased （P＜0.01）. Colon length was markedly shortened （P＜0.01），while colon weight，colon weight index，and colon weight/length ratio were significantly elevated （P＜0.01）. Colon tissue showed significant pathological damage，including disruption of intestinal wall integrity， visible ulcers on the mucosal surface， disorganized glandular structure， irregular crypt architecture， and inflammatory cell infiltration. The pathological injury score was significantly increased （P＜0.01）. The expression levels of CD19+ CD27+ cells （memory B cells） in mesenteric lymph nodes and Peyer’s patches were significantly elevated （P＜0.01），while the expression levels of CD19+ CD27+ CD5+ CD25+ cells （CD25+ mBreg cells） and CD19+ CD27+ CD5+ Foxp3+ cells （Foxp3+ mBreg cells） were significantly decreased （P＜0.05， P＜0.01）. The protein levels of PI3K，Akt，p-Akt，and NF-κB p65 in colon tissue were significantly increased （P＜0.01），while PTEN protein expression was significantly decreased （P＜0.01）. Compared with the model group，mice in the model + Sishen Pills group showed significant improvement in mental state， fur glossiness， and stool morphology， with markedly alleviated bloody stools. Body mass from days 17 to 21 was significantly increased （P＜0.05， P＜0.01）， while DAI scores from days 11 to 21 were significantly decreased （P＜0.05，P＜0.01）. Colon length was significantly extended （P＜0.01），while colon weight，colon weight index，and colon weight/length ratio were significantly reduced （P＜0.01）. Colon tissue exhibited significant improvement in pathological damage，with varying degrees of intestinal epithelial cell repair， partial glandular hyperplasia， only mild inflammatory cell infiltration and ulcers， and a significantly decreased pathological injury score （P＜0.01）. The expression levels of memory B cells in mesenteric lymph nodes and Peyer’s patches were significantly decreased （P＜0.01），while the expression levels of CD25+ mBreg and Foxp3+ mBreg cells were significantly increased （P＜0.05，P＜0.01）. The protein levels of PI3K，Akt，p-Akt， and NF-κB p65 in colon tissue were significantly decreased （P＜0.01），while PTEN protein expression was significantly increased （P＜0.01）. Conclusion Sishen Pills effectively alleviates DSS-induced colon inflammation in UC mice， possibly by inhibiting the activation of the PI3K/Akt signaling pathway and restoring the homeostasis of memory B cell subsets.

R285.5

国家自然科学基金项目（82360871）；江西省自然科学基金项目（20224BAB216110）；江西省卫生健康委员会科技计划项目（202211331）； 江西省教育厅科学技术研究项目（GJJ2403643，GJJ2200965）。