基于TRAF1/ASK1/JNK通路研究黄精水提物对过食“辛辣醇酒”致胃黏膜损伤小鼠的保护作用

Protective effect of Polygonati Rhizoma aqueous extract on gastric mucosal injury in mice induced by overtaking pungent substances and alcohol via TRAF1/ASK1/JNK pathway

为探讨黄精水提物对胃阴亏虚型胃黏膜损伤小鼠的改善作用和潜在机制,采用ICR小鼠每天灌胃1次辛辣物酒混溶液连续6周,建立胃阴亏虚型胃黏膜损伤小鼠模型。造模成功后,将模型小鼠按体质量随机分为模型组、黄精水提物0.5 g·kg^(-1)组、黄精水提物1.0 g·kg^(-1)组。模型组和各给药组每天继续灌胃1次辛辣物酒混溶液,同时给药组按照剂量灌胃相应药物,连续7周。给药期间,检测各组小鼠胃阴亏虚证相关中医证候指标;末次给药后解剖取胃组织观察其组织形态学变化,检测胃组织匀浆中抗氧化酶活性,酶联免疫吸附测定法、免疫组化法检测胃组织中炎症因子的表达,Western blot法检测肿瘤坏死因子受体相关因子1(TNF receptor-associated factor 1,TRAF1)/凋亡信号调节激酶1(apoptosis signal regulating kinase 1,ASK1)/c-Jun氨基末端激酶(c-jun N-terminal kinase, JNK)通路相关分子的蛋白表达,免疫荧光法检测凋亡因子的表达。结果显示,黄精水提物能有效改善小鼠的胃阴亏虚证;改善胃黏膜组织病变;提高胃组织中抗氧化酶活性(P<0.05,P<0.01);降低胃组织中促炎因子的水平(P<0.05,P<0.01);降低胃组织TRAF1/ASK1/JNK通路相关分子的蛋白表达(P<0.01);降低胃组织促细胞凋亡因子表达,提高B淋巴细胞瘤-2基因(B-cell lymphoma-2,Bcl-2)表达。以上提示,黄精水提物能缓解饥而不欲食、口干渴、大便干结等胃阴亏虚证证候,并能改善过食“辛辣醇酒”引起的胃黏膜损伤,其作用机制可能与抑制TRAF1/ASK1/JNK通路减少胃组织细胞凋亡和炎症有关。

This study aims to investigate the effect and potential mechanism of Polygonati Rhizoma aqueous extract in protecting mice from gastric mucosal injury(stomach Yin deficiency).ICR mice were administrated with the mixture of pungent substance extract and alcohol by gavage once a day for 6 weeks to establish the mouse model of gastric mucosal injury with gastric Yin deficiency.The modeled mice were randomized into three groups of model and Polygonati Rhizoma aqueous extract administrated at 0.5 g·kg^(-1) and 1.0 g·kg^(-1) according to the body weight.The mice were then continuously administrated with the molding agent once a day,and those in the Polygonati Rhizoma aqueous extract groups were additionally administrated the extract for 7 weeks.During this period,traditional Chinese medicine syndrome indicators related to gastric Yin deficiency were tested.After the last administration,the gastric tissue was collected to observe the histomorphological changes.The activities of antioxidant enzymes in the gastric homogenate were measured,and the levels and expression of proinflammatory cytokines in the gastric tissue were measured by enzyme-linked immunosorbent assay and immunohistochemistry,respectively.The expression levels of proteins in the tumor necrosis factor receptor(TNFR)-associated factor 1(TRAF1)/apoptosis signal regulating kinase 1(ASK1)/c-Jun N-terminal kinase(JNK)pathway were determined by Western blot,and those of apoptosis factors were determined by immunofluorescence.The results showed that the Polygonati Rhizoma aqueous extract alleviated the stomach Yin deficiency in mice,ameliorated gastric mucosal lesions,increased the activities of antioxidant enzymes in the gastric tissue(P<0.05,P<0.01),lowered the levels of pro-inflammatory cytokines in the gastric tissue(P<0.05,P<0.01),down-regulated the expression levels of proteins in the TRAF1/ASK1/JNK pathway(P<0.01)and pro-apoptotic factors in the gastric tissue,and up-regulated the expression of B-cell lymphoma-2(Bcl-2).The results suggested that the Polygonati Rhizoma aqueous extract could alleviate the symptoms of stomach Yin deficiency,such as hunger without desire to eat,thirst,and dry stool,and mitigate the gastric mucosal injury caused by overtaking pungent substances and alcohol by inhibiting the TRAF1/ASK1/JNK pathway to reduce apoptosis and inflammation in the gastric tissue.