(-)-黄皮酰胺对葡聚糖硫酸钠诱导的小鼠溃疡性结肠炎的保护作用及机制

Study on the therapeutic effect and mechanism of (-)-clausenamide on experimental colitis in mice

摘要

目的 研究传统药用植物黄皮的活性成分(-)-黄皮酰胺(clausenamide,Clau)对葡聚糖硫酸钠(dextran sulfate sodium,DSS)诱导的小鼠溃疡性结肠炎(ulcerative colitis,UC)的治疗作用及潜在机制。方法 将40只BALB/c小鼠随机分为正常对照组、模型组、5-ASA组及(-)-黄皮酰胺组。正常对照组小鼠饮用双蒸水,其他组连续7 d饮用4% DSS溶液诱导UC模型,5-ASA组(50 mg·kg^-1·d^-1 5-ASA)及(-)-黄皮酰胺组[40 mg·kg^-1·d^-1(-)-黄皮酰胺]在第1天UC造模的同时灌胃给药,连续给药10 d,正常对照组和模型组给予生理盐水灌胃。期间监测小鼠一般状况、体重、粪便情况并计算疾病活动指数(disease activity index,DAI)。第11天取血后处死小鼠,提取腹腔巨噬细胞(Mϕs),测量结肠长度,称重并计算各主要脏器系数,采用H&amp;E染色观察结肠组织病理变化、采用ELISA法测定血清和结肠组织中炎症因子的水平、Western blot检测结肠组织中p-STAT3表达。体外构建Mϕs与小鼠结肠上皮细胞(MCECs)共培养体系,划痕实验观察(-)-黄皮酰胺对MCECs迁移能力的影响。结果 与模型组相比,(-)-黄皮酰胺干预显著改善小鼠一般状况,抑制体重下降(P<0.01),降低DAI评分,逆转结肠缩短(P<0.01)及结肠组织病理学损伤,降低脾脏系数(P<0.05)。血清及结肠组织中IL-6、IL-1β、TNF-α含量显著下降(P<0.05或P<0.01)。Western blot显示(-)-黄皮酰胺明显下调结肠p-STAT3表达(P<0.05)。共培养实验和细胞划痕实验显示,(-)-黄皮酰胺处理的小鼠来源的Mϕs能显著增强MCECs的迁移能力(P<0.01)。结论 (-)-黄皮酰胺对DSS诱导的小鼠UC具有显著保护作用,其机制可能与抑制p-STAT3介导的炎症信号、促进肠上皮修复密切相关,提示其作为新型UC治疗候选小分子药物的潜在价值。

Abstract

Objective To investigate the therapeutic effects and underlying mechanisms of (-)-clausenamide (Clau),a bioactive component isolated from the traditional medicinal herb Clausena lansium,on dextran sulfate sodium (DSS)-induced ulcerative colitis (UC) in mice. Methods Forty BALB/c mice were randomly assigned into four groups:normal control group,model group,5-ASA (50 mg·kg^-1) group,and (-)-clausenamide (40 mg·kg^-1) group. Mice in the control group received double-distilled water,while those in the other groups received 4% DSS solution for 7 consecutive days to induce UC. The DSS+5-ASA group and DSS+(-)-clausenamide group received intragastric administration of the respective agents starting from day 1 of UC induction,continuing for 10 days. The normal control and model groups received intragastric normal saline. During the experimental period,general conditions,body weight,and stool characteristics were monitored,and the disease activity index (DAI) was calculated. On day 11,blood was collected,and the mice were euthanized. Peritoneal macrophages (MΦs) were isolated. Colon length was measured,major organ coefficients were calculated,and H&amp;E staining was performed to observe histopathological changes in colon tissue. Levels of inflammatory cytokines in serum and colon tissue were determined by ELISA. p-STAT3 expression in colon tissue was detected by Western blot. An in vitro co-culture system of MΦs and mouse colonic epithelial cells (MCECs) was established,and a scratch assay was performed to evaluate the effect of (-)-clausenamide on MCECs migration. Results Compared with the model group,(-)-clausenamide intervention significantly improved the general status of mice,inhibited body weight loss (P<0.01),reduced DAI scores,reversed colon shortening (P<0.01) and colonic histopathological damage,and decreased the spleen coefficient (P<0.05). The levels of IL-6,IL-1β,and TNF-α in serum and colon tissue were significantly reduced (P<0.05 or P<0.01). Western blot showed that (-)-clausenamide intervention significantly downregulated colonic p-STAT3 expression (P<0.01). Co-culture and scratch assays demonstrated that (-)-clausenamide-treated mouse-derived MΦs significantly enhanced the migration ability of MCECs (P<0.05). Conclusions These findings indicate that (-)-clausenamide exerts a significant protective effect against DSS-induced UC in mice. The mechanism may involve the inhibition of p-STAT3-mediated inflammatory signaling and promotion of intestinal epithelial repair,highlighting its potential as a novel small-molecule drug candidate for UC therapy.

关键词

黄皮酰胺 / 溃疡性结肠炎 / 葡聚糖硫酸钠 / p-STAT3

Key words

Clausenamide / Ulcerative colitis / Dextran sulfate sodium (DSS) / p-STAT3

参考文献

[1] Segal JP, LeBlanc JF, Hart AL . Ulcerative colitis:an update[J]. Clin Med (Lond), 2021, 21(2): 135-139. DOI: 10.7861/clinmed.2021-0080.

[2] Le Berre C, Honap S, Peyrin-Biroulet L . Ulcerative colitis[J]. Lancet, 2023, 402(10401): 571-584. DOI: 10.1016/S0140-6736(23)00966-2.

[3] Wang L, Zhang H, Tang F, et al. Therapeutic effects of valeriana jatamansi on ulcerative colitis:insights into mechanisms of action through metabolomics and microbiome analysis[J]. J Proteome Res, 2023, 22(8): 2669-2682. DOI: 10.1021/acs.jproteome.3c00237.

[4] Liu Y, Li BG, Su YH, et al. Potential activity of traditional Chinese medicine against ulcerative colitis:a review[J]. J Ethnopharmacol, 2022, 289: 115084. DOI: 10.1016/j.jep.2022.115084.

[5] Chen X, Xu T, Lv X, et al. Ginsenoside Rh2 alleviates ulcerative colitis by regulating the STAT3/miR-214 signaling pathway[J]. J Ethnopharmacol, 2021, 274: 113997. DOI: 10.1016/j.jep.2021.113997.

[6] Zhang L, Guo W, Lu N, et al. Advances in research on unphosphorylated STAT3:a review[J]. Medicine (Baltimore), 2025, 104(30): e43476. DOI: 10.1097/MD.0000000000043476.

[7] Yu H, Pardoll D, Jove R . STATs in cancer inflammation and immunity:a leading role for STAT3[J]. Nat Rev Cancer, 2009, 9(11): 798-809. DOI: 10.1038/nrc2734.

[8] Yun JH, Park SW, Kim KJ, et al. Endothelial STAT3 activation increases vascular leakage through downregulating tight junction proteins:implications for diabetic retinopathy[J]. J Cell Physiol, 2017, 232(5): 1123-1134. DOI: 10.1002/jcp.25575.

[9] Robinson P, Montoya K, Magness E, et al. Therapeutic potential of a small-molecule STAT3 inhibitor in a mouse model of colitis[J]. Cancers (Basel), 2023, 15(11): 2977. DOI: 10.3390/cancers15112977.

[10] Fan Y, Yi W, Huang H, et al. Efficacy of herbal medicine (Gegen Qinlian Decoction) on ulcerative colitis:a systematic review of randomized controlled trials[J]. Medicine (Baltimore), 2019, 98(52): e18512. DOI: 10.1097/MD.0000000000018512.

[11] Chu SF, Zhang JT . Recent advances in the study of (-)-clausenamide:chemistry,biological activities and mechanism of action[J]. Acta Pharm Sin B, 2014, 4(6): 417-423. DOI: 10.1016/j.apsb.2014.10.004.

[12] Liu D, Yu XM, Huang L . Novel concise synthesis of (-)-clausenamide[J]. Chinese J Chem, 2013, 31(3): 344-348. DOI: 10.1002/cjoc.201201187.

[13] 周雪沁, 苏畅, 李晓桐, 等 . N-对氯苯磺酰基-4-氨基水杨酸抑制DSS诱导的溃疡性结肠炎的作用[J]. 药学学报, 2018, 53(10): 1652-1659. DOI: 10.16438/j.0513-4870.2018-0567.

[14] Wirtz S, Popp V, Kindermann M, et al. Chemically induced mouse models of acute and chronic intestinal inflammation[J]. Nat Protoc, 2017, 12(7): 1295-1309. DOI: 10.1038/nprot.2017.044.

[15] Zhu A, Qiu Q, Xu Z, et al. Investigating the role and underlying mechanisms of 18β-glycyrrhetinic acid in the therapy of ulcerative colitis through modulation of the PPAR-γ/NF-κB signaling pathway[J]. J Inflamm Res, 2025, 18: 7529-7543. DOI: 10.2147/JIR.S510949.

[16] Xu ZG, Zhang QL, Ding C, et al. Beneficial effects of hordenine on a model of ulcerative colitis[J]. Molecules, 2023, 28(6): 2834. DOI: 10.3390/molecules28062834.

[17] Samad MA, Ahmad I, Hasan A, et al. STAT3 signaling pathway in health and disease[J]. MedComm, 2025, 6(4): e70152. DOI: 10.1002/mco2.70152.

基金资助

山东省中医药科技项目(Q2023072)

山东省自然科学基金-青年基金项目(ZR2024QH068)