葛根素对腹泻型肠易激综合征肠道平滑肌神经递质调节机制的影响研究

目的　观察葛根素对腹泻型肠易激综合征（IBS-D）大鼠的治疗作用并探讨其对肠道平滑肌的神经调节机制。方法　2021年11月至2022年5月将24只SPF级4周龄雄性SD大鼠随机分为对照组，模型组，葛根素低、高剂量组。采用慢性束缚应激诱导建立IBS-D模型。造模完成后葛根素低、高剂量灌胃给药，剂量分别为22、44 mg/（kg·d），每日1次，持续14 d；对照组与模型组以等量生理盐水灌胃，持续14 d。定期测定各组大鼠内脏敏感性、排便情况，采用苏木精-伊红（HE）染色法观察大鼠结肠组织病理改变，末次给药后测定结肠匀浆乙酰胆碱（Ach）、一氧化氮（NO）、五羟色胺（5-HT）的含量。统计学方法采用独立样本t检验。结果　光学显微镜观察示模型组大鼠无明显病理改变，符合IBS-D定义与组织学表现。与对照组比较，模型组大鼠腹壁回缩反射评分显著升高，第21、28天排便面积均显著增加，结肠组织Ach、5-HT表达均显著升高，NO表达显著降低，差异均有统计学意义（均P<0.05）。各项数据变化均符合IBS-D模型相关改变。与模型组比较，葛根素低、高剂量组的腹壁回缩反射评分均显著降低［（0.48±0.06）分、（0.54±0.06）分比（0.84±0.03）分］，差异均有统计学意义（均P<0.05）；与模型组比较，葛根素低、高剂量均使第21、28天排便面积显著减少（均P<0.05）；模型组的Ach、5-HT表达为（3.67±0.87）μg/mgprot、（131.97±16.64）μg/L，葛根素低剂量组的Ach、5-HT表达为（2.65±0.87）μg/mgprot、（115.01±12.07）μg/L、葛根素高剂量组的Ach、5-HT表达为（2.26±1.05）μg/mgprot、（109.01±6.59）μg/L，葛根素低、高剂量组的Ach、5-HT表达均显著降低（均P<0.05）；模型组NO表达为（9.61±1.50）μg/mgprot、葛根素低剂量组NO表达为（16.32±4.15）μg/mgprot、葛根素高剂量组NO表达为（9.34±5.80）μg/mgprot（P<0.05）。结论　葛根素对慢性束缚应激诱导的IBS-D大鼠模型具有一定的治疗作用，其作用机制或与上调结肠NO表达水平，下调结肠Ach、5-HT表达水平有关。

Study on the neuroregulatory mechanism of puerarin on IBS-D gastrointestinal smooth muscle

Objective　To observe the therapeutic effect of puerarin on irritable bowel syndrome with predominant diarrhea (IBS-D) in rats and to explore its neuroregulatory mechanism on gastrointestinal smooth muscle. Methods　From November 2021 to May 2022, 24 SPF 4-week-old male SD rats were randomly divided into a control group, a model group, and low and high dose puerarin groups. The IBS-D model was established by induction of chronic restraint stress. After the completion of modeling, low and high dose puerarin were intragastrically administered at doses of 22 and 44 mg/(kg·d), respectively, once daily for 14 days. The control group and the model group were intragastrically administered with the same amount of saline for 14 days. The visceral sensitivity and defecation were measured regularly in each group, the pathological changes of colon tissues were observed by hematoxylin-eosin (HE) staining, the contents of serotonin (5-HT), nitric oxide (NO), and acetylcholine (Ach) in colon homogenates were measured after the last administration. Independent sample t test was used for statistical analysis. Results　Light microscopy showed no obvious pathological changes in rats of the model group, in accordance with the definition of IBS-D and histological findings. Compared with those in the control group, the abdominal wall retraction reflex score of rats in the model group was significantly increased, the defecation area was significantly increased on the 21st and 28th day, the expressions of Ach and 5-HT in the colon tissue were significantly increased, and the expression of NO was significantly decreased, with statistically significant differences (all P<0.05). Compared with that in the model group (0.84±0.03) points], low and high dose puerarin significantly decreased the abdominal wall retraction reflex score of IBS-D rats (0.48±0.06) points and (0.54±0.06) points], with statistically significant differences (both P<0.05). Compared with that in the model group, low and high dose puerarin significantly decreased the defecation areas on the 21st and 28th day (all P<0.05). The expressions of Ach and 5-HT in the model group were (3.67±0.87) μg/mgprot and (131.97±16.64) μg/L, those in the low-dose puerarin group were (2.65±0.87) μg/mgprot and (115.01±12.07) μg/L, those in the high-dose puerarin group were (2.26±1.05) μg/mgprot and (109.01±6.59) μg/L, and the expressions of Ach and 5-HT in the low and high dose puerarin group were significantly decreased (all P<0.05). The expression of NO was (9.61±1.50) μg/mgprot in the model group, (16.32±4.15) μg/mgprot in the low-dose puerarin group, and (9.34±5.80) μg/mgprot in the high-dose puerarin group (P<0.05). Conclusion　Puerarin has a certain therapeutic effect on IBS-D rat models induced by chronic restraint stress, and its mechanism may be related to the up-regulation of NO expression level and the down-regulation of Ach and 5-HT expression levels in the colon tissue.