Desmodium caudatum对胃炎大鼠胃肠道激素和肠道菌群的影响
Summary
本方案描述了使用基于Illumina的16S rRNA基因测序分析肠道菌群的方法，并为评估草药汤剂的有效性提供了框架。
Abstract
为初步探究 尾鲷对 大鼠胃炎及肠道菌群的影响，采用经典水杨酸钠法建立慢性胃炎大鼠模型。将18只SPF大鼠分为对照组（C组）、模型组（M组）和治疗组（T组）3组。从每组大鼠中取出胃壁的病理切片。此外，通过ELISA测定各组大鼠血清中胃泌素和丙二醛的浓度。此外，通过对三组肠道细菌群落进行详细比较，采用基于Illumina的16S rRNA基因测序，探索了D . caudatum 对胃炎大鼠肠道菌群的影响。结果表明， 尾醛 汤可降低丙二醛含量，增加胃泌素含量。此外，还发现 尾苔 汤可增强肠道菌群的多样性和丰度，通过调节和恢复肠道菌群对胃炎的治疗产生积极影响。
Introduction
慢性胃炎（CG）是最常见的临床疾病之一，其特征是胃黏膜上皮的慢性和持续的炎症性变化，经常受到各种致病因素的反复攻击1。CG的发病率在所有胃病中排名第一，占胃肠科门诊服务率的40%-60%2。此外，发病率通常随着年龄的增长而增加，尤其是在中老年人中 3.毫无疑问，CG大大降低了人们的生活质量，强调了发现新治疗药物的迫切需要。
大量研究报告称，CG的发生和发展与胃肠道激素（如胃泌素（GAS））的分泌有关4,5。GAS是消化道中常见的胃肠道肽激素，通过促进嗜酸性粒细胞释放组胺来刺激细胞分泌胃酸。此外，它还能改善胃粘膜的营养和血液供应，促进胃粘膜和壁细胞的增殖6。因此，胃泌素可以作为评价CG发展水平的指标。此外，由活性氧化物触发的脂质过氧化产物可以激活炎症细胞，导致CG。丙二醛（MDA）是一种脂质过氧化标志物，是测量氧化应激程度的常用指标。它在一定程度上反映了胃粘膜中自由基的水平。MDA水平可以表明由自由基引起的局部胃粘膜中不饱和脂肪酸的攻击7,8。
肠道微生态学由数百万种微生物组成，它们存在于宿主肠道中，在维持宿主健康和调节宿主免疫力方面发挥着至关重要的作用。健康的肠道菌群具有高度丰富、多样性和稳定的微生物群功能，通过参与新陈代谢作为防止病原微生物入侵的保护屏障。肠道菌群的紊乱使个体更容易患上急性和慢性胃肠道疾病9,10。近年来，胃肠道疾病的微生物群治疗进展迅速，并显示出显著的疗效11。总之，肠道菌群的考虑对于理解和解决胃肠道疾病至关重要。
民间药材作为中医药宝库中不可或缺的组成部分，对临床实践和民族医药现代化具有重要意义。Desmodium caudatum （Thunb.） 的根部和整个部分在绵阳市北川羌地区长期被广泛用于缓解胃部不适。一些文章指出了用尾蛾治疗胃肠道疾病的科学依据，引用了其止血、抗氧化和胃肠道保护等作用12,13,14。然而，由于缺乏深入研究，尚未建立临床药效学机制。本文旨在基于胃肠道激素和肠道菌群研究尾曲霉治疗胃炎的效果，为其合理的临床应用提供依据。
Protocol
动物的护理和使用程序得到了绵阳师范大学伦理委员会的批准，并严格遵守了所有有关动物伦理使用的制度和政府规定。在本研究中，使用了SPF大鼠（昆明物种，雄性和雌性，体重180-220 g）。这些动物是从商业来源获得的（见 材料表）。所有动物都被饲养在无病原体的环境中，并可以 随意 获取食物。 1.试剂的制备 用电子秤称取2.5g水?…
Representative Results
胃壁病理切片的结果如 图1所示。与C组相比，M组表现出轻度胃壁萎缩和轻度炎症。然而，与M组相比，T组没有明显的炎症、肠化生或萎缩。这表明 D.caudatum 汤可以有效改善胃炎。 血清胃肠道激素测定结果如 图2所示。M组丙二醛（MDA）含量显著高于C组。用 D.caudatum 汤处理后，它显着减少。胃泌素含量（GAS）明显低于C组…
Discussion
羌族12人常用的民间药材，在治疗胃肠道疾病方面显示出显著的疗效。随着现代药理学研究的进步，胃肠道微生态失衡导致的菌群失衡被确定为急性和慢性胃肠道疾病的关键因素22,23。肠道中的某些微生物在维持体内的动态平衡方面起着至关重要的作用，血清中的激素反映了身体的生理和病理状态。因此，该研究的重点是肠道菌?…
Disclosures
The authors have nothing to disclose.
Acknowledgements
本工作由四川省科技厅重点研发项目（2020YFS0539）资助。
Materials
| Alpha diversity analysis | Mothur 1.30.2 | ||
| AxyPrep deoxyribonucleic acid (DNA) gel extraction kit | Axygen Biosciences | AP-GX-50 | |
| Beta diversity analysis | Qiime 1.9.1 | ||
| Cryogenic refrigerator | Forma-86C ULT freezer | ||
| Desmodium caudatum | The Traditional Chinese Medicine Hospital of Beichuan Qiang Autonomous County | ||
| E.Z.N.A. soil kit | Omega Bio-tek | D5625-01 | |
| Illumina MiSeq Platform | Illumina Miseq PE300/NovaSeq PE250 | ||
| Multiskan Spectrum | spectraMax i3 | ||
| OTU clustering | Uparse 7.0.1090 | ||
| OTU statistics | Usearch 7.0 | ||
| PCR instrument | TransGen AP221-02 | ||
| PCR instrument | ABI GeneAmp 9700 | ||
| QuantiFluor-ST double-stranded DNA (dsDNA) system | Promega Corp. | ||
| Sequence classification annotation | RDP Classifier 2.11 | ||
| Sodium salicylate | Sichuan Xilong Chemical Co., Ltd | 54-21-7 | |
| SPF rats | Chengdu Dashuo Experimental Animal Co., Ltd | ||
| SPSS 18.0 | IBM |
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Cite This Article
Bu, L., Tan, C., Zhang, B., Hu, J., Zhang, X., Han, X., Tian, H., Ma, X. Effects of Desmodium caudatum on Gastrointestinal Hormones and Intestinal Flora in Rats with Gastritis. J. Vis. Exp. (205), e65744, doi:10.3791/65744 (2024).