大鼠脑缺血再灌注损伤后肠道不同节段内褪黑素受体的变化
Changes of melatonin receptors in various intestinal segments after cerebral ischemia reperfusion injury in rats
王洁 1肖培伦 1陈雨婷 1范佳琦 1张月霞 1王晓莉 2于树娜1
作者信息
- 1. 山东第二医科大学基础医学院人体解剖学教研室,潍坊 261053
- 2. 山东第二医科大学医学影像学院,潍坊 261053
- 折叠
摘要
目的:脑缺血再灌注损伤(CIRI)可通过脑-肠轴对胃肠道等远隔器官造成损害.已知褪黑素可通过激活特定的受体发挥神经保护作用.然而,脑组织缺血后胃肠道中褪黑素受体的变化及其与肠道损伤的关系仍不清楚.方法:将24只成年雄性SD大鼠随机分为假手术(Sham)组和CIRI组,CIRI组采用Zea-Longa线栓法制备脑缺血再灌注模型,缺血2 h,再灌注24 h后收集大鼠空肠、回肠及结肠组织.采用2,3,5-三苯四唑氯(TTC)染色和HE染色观察CIRI后脑组织和肠组织的损伤程度,免疫荧光染色观察肠道封闭蛋白1(ZO-1)和Claudin5的表达,实时荧光定量PCR(RT-qPCR)、免疫荧光染色和Western Blot检测褪黑素受体1(MT1)和褪黑素受体2(MT2)的表达变化.通过一般线性回归分析CIRI后褪黑素受体与肠道紧密连接蛋白的相关性.结果:TTC染色显示,CIRI组大鼠脑梗死面积明显高于Sham组;HE染色显示,CIRI组大鼠肠绒毛断裂、缩短,杯状细胞减少.免疫荧光染色结果显示CIRI组大鼠肠组织内ZO-1+和Claudin5+的表达明显低于Sham组(P＜0.05);RT-qPCR显示,CIRI组MT1和MT2 mRNA的表达明显低于Sham组(P＜0.05),其中,结肠组织的降低最为明显.免疫荧光染色和Western Blot结果也显示,与Sham组相比,CIRI组MT1和MT2蛋白的表达显著降低(P＜0.05);一般线性回归分析显示,Sham组与CIRI组的MT1+和M12+平均荧光强度的差值与两组间Clau-din5+和ZO-1+平均荧光强度的差值均呈正相关.结论:CIRI后空肠、回肠和结肠内MT1和MT2的表达均降低,且结肠组织降低的最为显著,提示脑卒中的愈后不良可能与褪黑素受体的降低有一定相关性.
Abstract
Objective:Cerebral ischemia reperfusion injury(CIRI)can cause damage to distant organs,such as the gastrointestinal tract,through the gut-brain axis.Melatonin is known to play a neuroprotective role by activating specific receptor pathways.However,the changes of melatonin receptors in the gastrointestinal tract after brain injury and their relationship with intestinal injury are still unclear.Methods:Twenty-four male Sprague-Dawley rats were randomly di-vided into the Sham group and CIRI group.The CIRI model was prepared by Zea-Longa method.The jejunum,ileum,and colon tissues of the rats were collected 2 h after ischemia and 24 h after reperfusion.The damage of intestinal and brain tissues was observed by using 2,3,5-triphenyl tetrazolium chloride(TTC)and HE staining.The positive expres-sion of zonula occludens-1(ZO-1)and Claudin5 was observed by immunofluorescence staining.The melatonin receptor 1(MT1)and melatonin receptor 2(MT2)expression was detected by reverse transcription-quantitative polymerase chain reaction(RT-qPCR),immunofluorescence staining and Western Blot.The correlation between the melatonin receptors and intestinal tight junction proteins was analyzed by general linear regression.Results:TTC staining showed that the infarct size of rats in the CIRI group was significantly higher than that in the Sham group.HE staining showed that intestinal villi was broken and shortened,and goblet cells were reduced in the CIRI group.The results of immuno-fluorescence staining revealed that the positive expression of ZO-1 and Claudin5 in the intestinal tissues of rats in the CIRI group was significantly lower than that in the Sham group(P＜0.05).Compared with the Sham group,the RT-qPCR revealed a significantly lower expression of MT1 and MT2 mRNA in the CIRI group(P＜0.05),and the decrease in colon tissue was the most obvious.The results of immunofluorescence staining and WB also showed that the expression of MTl and MT2 in the CIRI group was significantly lower than that in the Sham group(P＜0.05).A gen-eral linear regression analysis revealed the difference in mean fluorescence intensities of MT1+and MT2+between the Sham group and the CIRI group were positively correlated with the difference of ZO-1+and Claudin5+between the two groups.Conclusion:After CIRI,the expression of both MT1 and MT2 receptors in various intestinal segments was de-creased,and the decrease in colon tissue was the most significant.It is suggested that the poor recovery of stroke may be related to the decrease of melatonin receptor.
关键词
脑缺血再灌注损伤/脑-肠轴/肠/褪黑素受体/大鼠Key words
cerebral ischemia reperfusion injury(CIRI)/brain-gut axis/intestines/melatonin receptor(MT)/rat引用本文复制引用
基金项目
山东省医药卫生科技发展计划项目(202001020642)
山东省中医药科技发展计划(Q-2023071)
出版年
2024
NETL
NSTL
万方数据