四磨汤口服液改善功能性消化不良模型大鼠的作用机制

Mechanism of Simotang oral liquid in improving functional dyspepsia model rats

目的 探讨四磨汤口服液调控PTEN诱导假定激酶1(Pink1)/E_(3)泛素连接酶(Parkin)轴对功能性消化不良(FD)模型大鼠的改善作用及机制。方法 取SD雄性大鼠,利用不可预知的慢性应激法建立FD模型,然后随机分为模型组、阳性对照组(多潘立酮片,3.5 mg/kg)、四磨汤组(四磨汤口服液,5.4 mL/kg),另设不造模的空白组,每组20只。各给药组大鼠灌胃相应药物,空白组和模型组大鼠灌胃等体积生理盐水,每天1次,持续14 d。末次给药结束后,检测大鼠胃排空率和小肠推进率;检测大鼠血清中胃肠激素[胃动素(MTL)、胃泌素(GAS)、胆囊收缩素(CCK)]含量;观察大鼠胃窦组织中卡哈尔间质细胞(ICC)线粒体结构;观察大鼠胃窦组织中细胞色素C氧化酶Ⅳ(COXⅣ)和Parkin在线粒体中的共定位表达;检测大鼠胃窦组织中微管相关蛋白轻链3(LC3)、p62、Pink1、Parkin蛋白的表达。结果 与空白组比较,模型组大鼠胃排空率、小肠推进率和血清中MTL、GAS含量以及p62蛋白表达水平均显著降低(P<0.05),血清中CCK含量和胃窦组织线粒体中COXⅣ、Parkin的共定位表达以及LC3、Pink1、Parkin蛋白表达水平均显著升高(P<0.05);胃窦组织中ICC线粒体数量减少,线粒体空泡化,线粒体嵴模糊,自噬溶酶体较多。与模型组比较,阳性对照组和四磨汤组大鼠上述指标水平均显著逆转(P<0.05);ICC线粒体结构逐渐恢复,线粒体嵴清晰,自噬溶酶体减少。结论 四磨汤口服液可改善FD模型大鼠的胃肠动力和胃肠激素水平,其作用机制可能与抑制Pink1/Parkin轴、阻断自噬有关。

OBJECTIVE To explore the improvement effect and mechanism of Simotang oral liquid on functional dyspepsia(FD) model rats by regulating PTEN-induced putative kinase 1(Pink1)/E_(3) ubiquitin ligase(Parkin) axis.METHODS SD male rats were subjected to unpredictable chronic stress to establish an FD model,and were randomly grouped into the model group,positive control group(Domperidone tablets,3.5 mg/kg),Simo decoction group(Simotang oral liquid,5.4 mL/kg),with 20 rats in each group.The blank group without modeling was set up.Administration groups were given relevant medicine intragastrically,blank group and model group were given constant volume of normal saline intragastrically,once a day,for consecutive 14 d.After the last administration,the gastric emptying rate and small intestine propulsion rate of rats were detected.The contents of gastrointestinal hormones [motilin(MTL),gastrin(GAS) and cholecystokinin(CCK)] in rats were determined;the mitochondrial structure of ICC in gastric antrum tissue of rats was observed;the immunofluorescence co-localization method was applied to observe the expression of cytochrome C oxidase Ⅳ (COX Ⅳ) and Parkin in gastric antrum tissue of rats;the protein expressions of LC3,p62,Pink1 and Parkin in gastric antrum tissue of rats were all detected.RESULTS Compared with the blank group,the gastric emptying rate,small intestinal propulsion rate,the serum contents of MTL and GAS,and protein expression of p62 in model group were all reduced significantly(P<0.05);the serum content of CCK,the co-localization expression of COX Ⅳ and Parkin in mitochondria,the protein expressions of LC3,Pink1 and Parkin were all increased significantly(P<0.05).The number of ICC mitochondria in gastric antrum tissue decreased,mitochondria became vacuolated,mitochondrial cristae were blurry,and there were more autophagic lysosomes.Compared with the model group,the above indexes of the positive control group and the Simo decoction group were reversed significantly(P<0.05);the mitochondrial structure of ICC gradually recovered,with clear mitochondrial cristae and reduced autophagic lysosomes.CONCLUSIONS Simotang oral liquid can improve gastrointestinal motility and gastrointestinal hormone levels in FD model rats,the mechanism of which may be related to inhibiting Pink1/Parkin axis and blocking autophagy.