吲哚-3-甲醛对志贺菌感染的保护作用及其机制研究

Indole-3-carboxaldehyde protects mouse from Shigella infection

摘要:

【目的】探究吲哚-3-甲醛(indole-3-carboxaldehyde，I3A)对志贺菌感染性结肠炎的缓解作用及其机制研究。【方法】体外实验，通过微量肉汤稀释法测定I3A对志贺菌的最低抑菌浓度(minimum inhibitory concentration，MIC)和I3A的抗菌活性，运用酶标仪测定600 nm处不同浓度I3A作用的菌液吸光度(A₆₀₀)，并通过平板菌落计数检测志贺菌量。体内实验，将18只小鼠随机分为对照(NC)组、SF301组和SF301+I3A组。然后SF301+I3A组进行连续8 d的200 µL I3A灌胃处理，同时NC组和SF301组用200 µL无菌水灌胃处理，SF301组和SF301+I3A组于灌药第4天制备志贺菌感染性结肠炎小鼠模型，体重和疾病活动指数(disease activity index，DAI)用于评价小鼠状况。上述实验结束后将所有小鼠处死，取中段结肠进行组织病理学检测观察炎症情况，下段结肠检测白细胞介素-6 (interleukin-6，IL-6)和肿瘤坏死因子-α (tumor necrosis factor-α，TNF-α)的表达水平，粪便、盲肠内容物检测载菌量。同时进行14 d的I3A处理小鼠监测I3A的体内应用安全性。【结果】体外实验：I3A的MIC为128 µg/mL，不同浓度的I3A涂板计数和A₆₀₀结果与对照组比较，均有统计学差异(均P<0.05)。体内实验：与SF301组相比，SF301+I3A组小鼠体重和DAI更接近NC组，肉眼观结肠组织无明显损伤，组织病理学检查未见明显炎症，载菌量和炎性因子均有统计学差异(均P<0.05)；I3A对小鼠体重变化无显著影响(P>0.05)。【结论】本研究证明I3A在志贺菌感染性结肠炎中对肠道具有保护作用，I3A可通过抑制志贺菌载菌量、降低结肠损伤和炎症水平来缓解志贺菌感染性结肠炎，并且I3A在体内没有显示毒性。

Abstract:

[Objective]To investigate the alleviating effect of indole-3-carboxaldehyde (I3A) on the colitis caused by Shigella infection and decipher the underlying mechanism.[Methods]We conducted the in vitro experiment to measure the minimum inhibitory concentration (MIC) and activity of I3A against Shigella by micro broth dilution method.Further,we used a microplate reader to measure the absorbance (A₆₀₀) of the bacterial suspensions with different concentrations of I3A at 600 nm and counted the number of Shigella via the plate colony counting method.In the in vivo experiment,we randomly assigned 18 mice into a control (NC) group,a SF301 group,and a SF301+I3A group.The SF301+I3A group was administrated with 200µL I3A by gavage for 8 consecutive days,and the NC group and SF301 group with 200µL sterile water.The colitis model of Shigella infection was established on the 4th day in the SF301 group and SF301+I3A group.Body weight and disease activity index (DAI) were used to evaluate the conditions of mice.After the above experiments,we euthanized all the mice,collected the middle colon for histopathological detection to observe the inflammation,determined the expression levels of interleukin-6(IL-6) and tumor necrosis factor-α(TNF-α) in the lower colon,and measured the bacterial load in feces and cecum.The safety of I3A application in vivo was monitored in mice for 14 days.[Results] In vitro experiment:the MIC of I3A was 128µg/mL.The bacterial counts and A₆₀₀in the groups treated with different concentrations of I3A were different from those in the control group (P<0.05).In vivo experiment:compared with the SF301 group,the SF301+I3A group showed the body weight and DAI close to those in the NC group and significantly relieved colonic damage.No obvious inflammation was observed in SF301+I3A group,and the bacterial load and the expression levels of inflammatory cytokines were different between SF301+I3A group and SF301 group (P<0.05).I3A had no significant effect on the body weight of mice (P>0.05).[Conclusion] We demonstrate that I3A has a protective effect on the colitis caused by Shigella infection.Specifically,I3A alleviates the colitis by inhibiting Shigella load and reducing colonic damage and inflammation levels.Moreover,I3A shows no toxicity in vivo.