Objectives: To observe the effect of moxibustion intervention on the hypothalamus-spinal cord-colon axis of rats with irritable bowel syndrome with diarrhea (IBS-D) and explore the mechanism of moxibustion in improving visceral hypersensitivity in rats with IBS-D.
Methods: A total of 36 SD rats were randomly divided into normal, model, and moxibustion groups, with 12 rats in each group. The IBS-D model was established by maternal separation + acetic acid stimulation + chronic restraint. Rats of the moxibustion group received bilateral moxibustion on "Tianshu" (ST25) and "Shangjuxu" (ST37) for 15 min, once a day for 7 consecutive days. The body weight, loose stool rate, and minimum threshold volume of abdominal withdrawal reflex (AWR) were measured before and after moxibustion intervention, respectively. The histopathological changes in the colon tissue were observed after HE staining. The number of colonic mucosal mast cells (MCs) was measured by toluidine blue staining. The activation of MCs was determined by tryptase positive expression level and examined by immunohistochemical staining. The content, protein and mRNA expression levels and positive expression levels of corticotropin releasing factor (CRF), substance P (SP), and calcitonin gene-related peptide (CGRP) in the hypothalamus, spinal cord and colon tissues were measured by ELISA, Western blot, real-time fluorescent quantitative PCR and immunofluorescence staining, respectively.
Results: Compared with the normal group, the loose stool rate was increased (P<0.01)；the body weight and minimum threshold volume of AWR were decreased (P<0.01)；the inflammatory infiltration of colon tissues was obvious；the number of MCs and positive expression level of tryptase in the colon tissue were increased (P<0.01)；the contents, positive expression le-vels, protein and mRNA expression levels of CRF, SP and CGRP in the hypothalamus, spinal cord and colon tissues were increased (P<0.01, P<0.05) in the model group. After the intervention, compared with the model group, all these indicators showed opposite trends (P<0.01, P<0.05) in the moxibustion group.
Conclusions: Moxibustion can improve visceral hypersensitivity in rats with IBS-D, and its mechanism may be related to regulating the hypothalamic-spinal-colon axis to reduce the release of CRF, SP and CGRP, and thus to inhibite MC in colon tissue.
目的: 观察艾灸干预对腹泻型肠易激综合征（IBS-D）大鼠下丘脑-脊髓-结肠轴的影响，探讨艾灸改善IBS-D大鼠内脏高敏感的作用机制。方法: 将36只SD大鼠随机分为正常组、模型组、艾灸组，每组12只。采用母子分离＋醋酸刺激＋慢性束缚法复制IBS-D模型。艾灸组大鼠给予艾灸双侧“天枢”“上巨虚”15 min，每日1次，连续7 d。分别于实验第34、45、53天检测各组大鼠体质量、稀便率和腹部回撤反射（AWR）的最小容量阈值；HE染色法检测结肠组织病理学变化；甲苯胺蓝染色法检测结肠黏膜肥大细胞（MC）数量；免疫组织化学染色法检测类胰蛋白酶阳性表达水平；ELISA、Western blot、实时荧光定量PCR和免疫荧光染色法分别检测大鼠下丘脑、脊髓、结肠组织中促肾上腺皮质激素释放因子（CRF）、P物质（SP）、降钙素基因相关肽（CGRP）的含量、蛋白和mRNA相对表达量、阳性表达。结果: 干预结束后与正常组比较，模型组大鼠稀便率升高（P<0.01），体质量和AWR的最小容量阈值降低（P<0.01），结肠组织局部炎性细胞浸润明显，结肠组织中MC数量及类胰蛋白酶阳性表达水平升高（P<0.01），下丘脑、脊髓和结肠组织中CRF、SP、CGRP含量、阳性表达、蛋白和mRNA表达水平升高（P<0.01，P<0.05）；与模型组比较，艾灸组大鼠稀便率降低（P<0.01），体质量和AWR的最小容量阈值升高（P<0.01），结肠黏膜下炎性细胞减少，结肠组织中MC数量及类胰蛋白酶阳性表达水平降低（P<0.01），下丘脑、脊髓和结肠组织中CRF、SP、CGRP 含量、阳性表达、蛋白及mRNA表达水平降低（P<0.01，P<0.05）。结论: 艾灸可改善IBS-D大鼠内脏高敏感，其作用机制可能与艾灸调节下丘脑-脊髓-结肠轴，减少CRF、SP、CGRP释放，降低结肠组织中MC数量有关。.
Keywords: Calcitonin gene-related peptide; Corticotropin releasing factor; Irritable bowel syndrome with diarrhea; Mast cell; Moxibustion; Substance P.