Objectives: To investigate the effect of Peitu Yimu(strengthening spleen and soothing liver) acupuncture on intestinal mucosal barrier function and corticotropin-releasing factor (CRF)/CRF receptor 1 (CRFR1) pathway in rats with diarrhea-predominant irritable bowel syndrome (IBS-D), so as to explore its underlying mechanism in alleviating IBS-D.
Methods: Forty female SD rats were randomly divided into blank, model, electroacupuncture (EA), and agonist groups, with 10 rats in each group. Except for the blank group, rats in the other groups were given folium sennae infusion by gavage combined with chronic unpredictable mild stress to establish IBS-D model. Rats in the EA group received acupuncture at "Tianshu"(ST25) and EA at "Zusanli"(ST36) and "Taichong"(LR3) (2 Hz/15 Hz) on one side for 20 min, with the side chosen alternately every other day, for 14 days after modeling. Rats in the agonist group received acupuncture 30 min after intravenous injection of CRFR1 agonist urocortin, with the same manipulation method and time as the EA group. Before and after intervention, visceral pain threshold and stool Bristol scores were measured. Elevated plus maze test and open field test were used to detect anxiety and depression like behavior of rats. ELISA was used to detect the contents of CRF and CRFR1 in rats serum. Immunohistochemistry was used to detect the positive expressions of CRF, CRFR1, zonula occludens protein 1(ZO-1), occlusal protein(Occludin), and closure protein 1 (Claudin-1) in colon tissue.
Results: Compared with the blank group, the visceral pain threshold, open arm time percentage (OT%), total distance of movement in the open field test, and positive expression of ZO-1, Occludin, and Claudin-1 in colon were decreased (P<0.01, P<0.05), while Bristol stool scores, serum CRF and CRFR1 contents, and positive expressions of CRF and CRFR1 in colon were increased (P<0.01) in the model group. After intervention and compared with the model group, the visceral pain threshold, OT%, total distance of movement in the open field test, and positive expressions of ZO-1, Occludin, and Claudin-1 in colon were increased (P<0.05, P<0.01), while Bristol stool scores, serum CRF and CRFR1 contents, and positive expressions of CRF and CRFR1 in colon were decreased (P<0.01) in the EA group；the Bristol stool scores, serum CRF content, and CRF positive expression in colon were significantly decreased in the agonist group (P<0.01).
Conclusions: Peitu Yimu acupuncture can significantly improve visceral hypersensitivity and anxiety-depression state in IBS-D rats. Its mechanism may be related to the inhibition of CRF/CRFR1 pathway and restoration of intestinal tight junction protein expressions.
目的: 基于促肾上腺皮质激素释放因子（CRF）/CRF受体1（CRFR1）通路探讨培土抑木针法对腹泻型肠易激综合征（IBS-D）大鼠肠黏膜屏障功能的影响及机制。方法: 将40只雌性SD大鼠随机分为空白组、模型组、电针组、激动剂组，每组10只。除空白组外，其余各组大鼠采用番泻叶浸液灌胃联合慢性不可预知性温和刺激构建IBS-D大鼠模型。针刺组大鼠于造模后针刺一侧“天枢”，电针“足三里”“太冲”（2 Hz/15 Hz），每次20 min，隔日左右交替，干预14 d；激动剂组尾静脉注射CRFR1激动剂尿促皮素后30 min进行针刺，针刺方法及时间均同电针组。干预后检测各组大鼠内脏痛阈值，进行粪便Bristol评分，采用高架十字迷宫实验和旷场实验评价大鼠焦虑抑郁行为，采用ELISA法检测大鼠血清CRF、CRFR1的含量，采用免疫组织化学法检测结肠组织中CRF、CRFR1，紧密连接蛋白闭锁连接蛋白1（ZO-1）、咬合蛋白（Occludin）、闭合蛋白1（Claudin-1）阳性表达。结果: 与空白组相比，模型组大鼠内脏痛阈值，高架十字迷宫开臂时间比（OT%）、旷场实验运动总距离，结肠ZO-1、Occludin、Claudin-1阳性表达下降（P<0.01，P<0.05），粪便Bristol评分，血清CRF、CRFR1含量，结肠CRF、CRFR1阳性表达上升（P<0.01）。与模型组相比，干预后电针组内脏痛阈值，OT%、旷场实验运动总距离，结肠ZO-1、Occludin、Claudin-1阳性表达均上升（P<0.05，P<0.01），粪便Bristol评分，血清CRF、CRFR1含量，结肠CRF、CRFR1阳性表达下降（P<0.01）；激动剂组粪便Bristol评分，血清CRF含量和结肠CRF阳性表达显著下降（P<0.01）。结论: 培土抑木针法可以显著改善IBS-D大鼠的内脏高敏、焦虑抑郁状态，其机制可能与抑制CRF/CRFR1通路，恢复肠道紧密连接蛋白表达有关。.
Keywords: Corticotropin-releasing factor; Diarrhea-predominant irritable bowel syndrome; Electroacupuncture; Intestinal mucosal barrier; Peitu Yimu (soothing the liver and strengthening the spleen) acupuncture; Tight junction protein.