Syndecan-1介导双歧杆菌对新生大鼠坏死性小肠结肠炎保护作用的研究

Protective effect of syndecan-1 medicated Bifidobacterium on necrotizing enterocolitis in rats

目的探索Syndecan-1蛋白在新生大鼠坏死性小肠结肠炎(necrotizing enterocolitis,NEC)模型肠组织中的表达情况,明确双歧杆菌对Syndecan-1蛋白的修复作用。方法建立新生大鼠NEC模型。按随机数字表法将新生大鼠分为3组(n=10):健康对照组、NEC组和双歧杆菌组。HE染色观察回盲部肠组织病理学变化。qRT-PCR技术检测各组TNF-α、IL-1β、MMP-7和Syndecan-1 mRNA的表达水平。Western blot检测分析各组Syndecan-1蛋白的表达水平。免疫组化分析各组Syndecan-1蛋白的定位表达。结果 NEC组TNF-α、IL-1β和MMP-7 mRNA表达水平高于健康对照组,而双歧杆菌可以抑制TNF-α、IL-1β和MMP-7的表达。NEC组Syndecan-1 mRNA的表达水平略高于健康对照组,双歧杆菌组略低于NEC组,但差异均无统计学意义。Western blot结果显示,健康对照组Syndecan-1蛋白呈高水平表达,NEC组几乎未检测到Syndecan-1蛋白的表达,而双歧杆菌组可见Syndecan-1蛋白的表达。免疫组化结果显示,对照组绒毛上皮细胞膜呈深棕色,而NEC组绒毛上皮细胞膜棕色染色明显减弱,双歧杆菌组上皮细胞膜着浅棕色。结论 NEC炎症时,Syndecan-1蛋白被切割,双歧杆菌可保护Syndecan-1蛋白的表达。双歧杆菌对肠黏膜的保护作用可能是通过抑制MMP-7等因子的表达,从而保护了Syndecan-1蛋白。

Objectives To explore the expression of syndecan-1 in the rat model of necrotizing enterocolitis （NEC）, and confirm the recovery effect of Bifidobacterium on syndecan-1. Methods Rat model of NEC was established. A total of 30 Sprague-Dawley rats were randomly divided into 3 groups at the first day after birth, health control group, NEC group and Bdobacterium intervention group, with 10 rats in each group. HE staining was carried out to observe the histological changes in the intestinal tissue of ileoeecal segment. The mRNA expression levels of TNF-α、IL-1β, MMP-7 and syndecan-1 were measured by qRT- PCR. The protein expression level of syndecan-1 was detected by Western blotting, and its cellular localization on the intestine was observed through immunohistochemical staining. Results The mRNA levels of TNF-α、IL-1β and MMP-7 were significantly up-regulated in the NEC group than the normal control. Bifidobacterium intervention reduced the mRNA levels of above molecules. The mRNA level of syndecan-1 was mildly higherin the NEC group than the healthy control group, but lower than in the Bifidobacterium group, though no obvious differences. Syndecan-1 protein was highly expressed in the healthy rats, not expressed in the NEC rats, and expressed in the intervention rats. The results of immunohistochemical staining showed that syndecan-1 protein were mainly located on the membrane of intestinal epithelial cells in the healthy control group, but mildly in the NEC group and the intervention group, which consistent with the results of Western blotting. Conclusions Shedding of syndecan-1 happens in the process of NEC inflammation, and Bifidobacterium could protect its protein expression. The protective effect of Bifidobacterium on intestinal mucosal may be through inhibiting MMP-7 to repair the syndecan-1 protein.