通过体内电子自旋共振光谱法评估实验性结肠炎小鼠的黏膜巯基化合物

Mucosal sulfhydryl compounds evaluation by in vivo electron spin resonance spectroscopy in mice with experimental colitis.

作者信息

Togashi H, Oikawa K, Adachi T, Sugahara K, Ito J, Takeda T, Watanabe H, Saito K, Saito T, Fukui T, Takeda H, Ohya H, Kawata S

机构信息

Second Department of Internal Medicine, Yamagata University School of Medicine, 2-2-2 Iida-Nishi, Yamagata 990-9585, Japan.

出版信息

Gut. 2003 Sep;52(9):1291-6. doi: 10.1136/gut.52.9.1291.

DOI:10.1136/gut.52.9.1291

PMID:12912860

原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC1773775/

Abstract

BACKGROUND

Sulfhydryl (SH) compounds are essential in maintaining mucosal integrity in the gastrointestinal tract. A decrease in colonic mucosal SH compounds affects the redox status of the mucosa, resulting in vulnerability to further attacks. Therefore, there is a strong need for in vivo evaluation of SH compounds in the colonic mucosa.

AIMS

The aim of the current study was to establish a method of evaluating levels of SH compounds in the colonic mucosa of live animals before and after induction of colitis.

METHODS

Murine experimental colitis was induced by instillation of trinitrobenzene sulphonic acid (TNBS) dissolved in 50% ethanol into the colon via the anus. For evaluation of mucosal SH compounds in the colon, 3-carbamoyl-2,2,5,5-tetramethylpyrrolidine-1-oxyl (carbamoyl-PROXYL), a stable nitroxide radical, was instilled into the colonic lumen of live mice and the spin clearance rate was measured by L-band electron spin resonance (ESR) spectroscopy.

RESULTS

Morphological study showed that mucosal damage was severe one or two days after TNBS instillation. The colonic mucosa started to regenerate at four days, and looked normal at seven days, after induction of colitis. The spin clearance rate of carbamoyl-PROXYL decreased significantly at 0.5, 1, 2, and 4 days after induction of colitis compared with mice before TNBS instillation. Surprisingly, although the colonic mucosa looked normal seven days after TNBS administration, the spin clearance rate still remained significantly slow. The spin clearance rate returned to normal 14 days after induction of colitis. The change in in vivo spin clearance rate was consistent with the time dependent change in mucosal reduced glutathione, a major component of SH compounds.

CONCLUSION

The spin clearance rate obtained by L-band ESR spectroscopy in combination with carbamoyl-PROXYL can give an estimate of the level of colonic mucosal SH compounds in live animals and is useful for evaluating the mucosal defence system against oxidative stress.

摘要

背景

巯基（SH）化合物对于维持胃肠道黏膜完整性至关重要。结肠黏膜SH化合物减少会影响黏膜的氧化还原状态，导致其更易受到进一步攻击。因此，迫切需要对结肠黏膜中的SH化合物进行体内评估。

目的

本研究的目的是建立一种评估活体动物在诱导结肠炎前后结肠黏膜中SH化合物水平的方法。

方法

通过经肛门向结肠内注入溶解于50%乙醇中的三硝基苯磺酸（TNBS）诱导小鼠实验性结肠炎。为评估结肠黏膜中的SH化合物，将一种稳定的氮氧自由基3-氨甲酰基-2,2,5,5-四甲基吡咯烷-1-氧基（氨甲酰基-PROXYL）注入活体小鼠的结肠腔，并通过L波段电子自旋共振（ESR）光谱法测量自旋清除率。

结果

形态学研究表明，TNBS注入后1或2天黏膜损伤严重。结肠炎诱导后，结肠黏膜在第4天开始再生，并在第7天看起来正常。与TNBS注入前的小鼠相比，在结肠炎诱导后0.5、1、2和4天，氨甲酰基-PROXYL的自旋清除率显著降低。令人惊讶的是，尽管TNBS给药后7天结肠黏膜看起来正常，但自旋清除率仍然显著缓慢。结肠炎诱导后14天，自旋清除率恢复正常。体内自旋清除率的变化与黏膜还原型谷胱甘肽（SH化合物的主要成分）随时间的变化一致。