大黄附子汤改善小鼠重症急性胰腺炎肠动力障碍的机制研究

Mechanism of Dahuang Fuzi decoction in alleviating intestinal motility disorder in mice with severe acute pancreatitis

目的揭示大黄附子汤改善重症急性胰腺炎小鼠早期肠动力障碍的机制。方法将27只SPF级雄性C57BL/6小鼠随机分为对照组、重症急性胰腺炎组(SAP)和大黄附子汤治疗组(DHFZT),每组9只。SAP组和DHZFT组采用腹腔注射20%L-精氨酸溶液复制SAP小鼠模型,对照组腹腔注射等渗盐水。DHFZT组分别于造模后12、24、36 h以0.2 mL大黄附子汤灌肠,对照组和SAP组分别以0.2 mL等渗盐水灌肠。造模后48 h,每组取3只小鼠测定小肠推进率,其余小鼠麻醉后取血、取材。ELISA法检测小鼠血清淀粉酶、血钙、内毒素、TNF-α和IL-6含量。HE染色观察胰腺和回肠组织病理学变化。Western blot定量检测回肠nNOS、ChAT、VIP、SP蛋白表达。结果与对照组相比,SAP组的抑制性神经元nNOS和VIP数量显著增加,兴奋性神经元ChAT和SP数量显著下降,小肠推进率显著降低,对应血清淀粉酶、内毒素、TNF-α和IL-6水平显著升高,差异有统计学意义(P均<0.05);与SAP组相比,DHFZT组的抑制性神经元nNOS和VIP数量显著下降,而兴奋性神经元ChAT和SP数量显著增加,小肠推进率显著增加,对应血清淀粉酶、内毒素、TNF-α和IL-6水平显著降低,血钙含量显著上升,差异有统计学意义(P均<0.05)。结论SAP小鼠早期发生肠动力障碍的机制可能存在肠神经系统的重塑,表现在抑制性神经元数量增加、兴奋性神经元数量减少,肠动力受抑制;大黄附子汤能够逆转肠神经系统的重塑,从而促进SAP小鼠肠动力的恢复。

Objective To reveal the possible mechanism of intestinal motility disorder in mice with severe acute pancreatitis(SAP)and the effect of Dahuangfuzi decoction on it.Methods Twenty seven SPF male C57BL/6 mice were randomly divided into control group,severe acute pancreatitis group(SAP)and rhubarb and aconite decoction treatment group(DHFZT),with 9 mice in each group.SAP group and DHZFT group were intraperitoneally injected with 20%L-arginine solution to replicate SAP mouse model,and the control group was intraperitoneally injected with isotonic saline.In DHFZT group,0.2 mL rhubarb and aconite decoction was used for enema 12,24 and 36 hours after modeling,while 0.2 mL isotonic saline was used for enema in control group and SAP group.At 48h after modeling,3 mice in each group were taken to measure the small intestine propulsion rate,and the rest of the mice were taken blood and materials after anesthesia.Detection of serum amylase,blood calcium,endotoxin and TNF in mice by ELISA-αAnd IL-6 content.Histopathological changes of pancreas and ileum were observed with HE staining.Western blot was used to detect the expression of nNOS,ChAT,VIP and SP proteins in ileum.Results Compared with the control group,the expression of nNOS and VIP in the SAP group were increased significantly,and the ChAT and SP were decreased significantly;the propulsion rate of small intestine was decreased significantly,with the lower levels of serum amylase,endotoxin,TNF-αand IL-6,and the blood calcium content also significantly decreased.Compared with the SAP group,the expression of nNOS and VIP in the DHFZT group were decreased significantly,and the ChAT and SP were increased significantly;the propulsion rate of small intestine was increased significantly,with the lower levels of serum amylase,endotoxin,TNF-αand IL-6,and the blood calcium content also significantly increased.Conclusion The mechanism of intestinal motility disorder in the early course of SAP mice may be related to the ENS remodeling,the increased number of inhibitory neurons,the decreased number of excitatory neurons,and therefore the proportion was imbalanced,leading to intestinal motility inhibition.Dahuangfuzi decoction may reverse the remodeling of ENS neurons and promote the recovery of intestinal motility in SAP mice.