【摘要】 目的对糖尿病大鼠肠黏膜屏障功能检测方法进行分析评价,并以此为靶点探讨蜂胶是否通过修复肠黏膜屏障发挥降血糖作用。方法建立糖尿病大鼠模型,以80及160 mg/kg蜂胶进行干预,持续4周。罗氏血糖仪检测尾血血糖(FBG);高效液相色谱法检测红细胞糖化血红蛋白(HbA1c)含量;ELISA实验检测血浆D-乳酸(D-LA)及连蛋白(zonulin)含量;透射电镜观察肠组织上皮细胞间连接装置超微结构变化,Western Blotting检测其紧密连接相关蛋白表达水平。结果成功建立糖尿病大鼠模型,80、160 mg/kg蜂胶干预4周后,大鼠血糖值分别达到17.57 mmol/L和18.24 mmol/L,HbA1c含量分别达到8.94%及8.22%,均较模型组明显降低(P<0.05)。补充蜂胶后,糖尿病大鼠回肠及结肠组织上皮细胞间紧密连接、黏着连接间隙异常增宽等现象得到有效缓解。补充蜂胶使回肠和结肠组织中claudin-1、occludin及ZO-1蛋白表达水平显著升高,且具有剂量依赖性(P<0.05)。80及160 mg/kg蜂胶干预后,血清D-LA含量分别为160.03 ng/mL及151.18 ng/mL,zonulin含量分别为650.14μg/mL及647.60μg/mL,均较模型组降低,但各组间比较,均未见显著性差异(P>0.05)。结论肠黏膜屏障超微结构观察及肠组织紧密连接相关蛋白表达水平检测较肠黏膜屏障功能血清学检测可能具有更高的灵敏性,可作为可靠的实验手段应用于蜂胶降糖的肠黏膜屏障修复机制研究。更多还原

【Abstract】 Objective To evaluate the method of detecting intestinal mucosal barrier damage, and investigate whether propolis can lower glucose levels through repairing the intestinal mucosal barrier in diabetic rats. Methods A rat model of diabetes mellitus was established by administering a high-sugar and high-fat diet with a single intraperitoneal injection of 1% streptozotocin(STZ) at a dose of 40 mg/kg. Then, the rats in the propolis treatment groups were orally supplemented with 80 or 160 mg/(kg·d) propolis by gavage. The rats of control and model groups were orally supplemented with soybean oil. After 4 weeks, a Roche glucometer was used to detect blood glucose concentration, high-performance liquid chromatography was used to detect whole-blood hemoglobin A1 c(HbA1 c), ELISA was used to detect the levels of plasma D-lactic acid(D-LA) and zonulin, western blotting was used to detect the expression levels of claudin-1, occludin, and ZO-1 in ileum and colon tissues, and the ultrastructure of ileum and colon tissues was examined by transmission electron microscopy. Results The diabetic rat model was established successfully. After 4 weeks of intervention, the blood glucose level in the 80 and 160 mg/kg propolis treatment groups reached 17.57 and 18.24 mmol/L, and their HbA1 c levels were 8.94% and 8.22%, respectively, which were significantly lower than that in the model group(P< 0.05). Transmission electron microscopy showed that after propolis intervention, the tight junctions and adherens junctions of intestinal epithelium were improved. Western blotting showed that the expression levels of claudin-1, occludin, and ZO-1 in the ileum and the colon of the propolis intervention groups were significantly higher than in the model group, in a dose-dependent manner(P< 0.05). ELISA data showed that the levels of plasma D-LA were 160.03 and 151.18 ng/mL in the 80 and 160 mg/kg propolis intervention groups, respectively, and their plasma zonulin levels were 650.14 and 647.60 μg/mL, with no significant differences between them(P> 0.05). Conclusions Ultrastructural observation of the intestinal mucosal barrier and detection of the expression levels of proteins closely associated with the intestinal tissue have higher sensitivity than the serological determination of intestinal mucosal barrier function, so they can be used as a reliable experimental method for studying the mechanism of the intestinal barrier repair associated with the hypoglycemic effect of propolis.更多还原