嗜黏蛋白阿克曼菌改善肠道屏障损伤的作用机制研究

韦博艺; 李冰; 赵浩如; 纪正业; 陈如意; 裴斐; 杨文建; 苏安祥

doi:10.13386/j.issn1002-0306.2025090083

嗜黏蛋白阿克曼菌改善肠道屏障损伤的作用机制研究

Role of Akkermansia muciniphila in Ameliorating Intestinal Barrier Damage

摘要

摘要: 本课题组前期研究发现，猴头菇多肽（Lys-Ser-Pro-Leu-Tyr，KSPLY）能够改善肠道屏障损伤，同时能增加肠道菌嗜黏蛋白阿克曼菌（Akkermansia muciniphila）的丰度，为了探讨A.muciniphila在改善肠道屏障损伤中的作用，本研究采用脂多糖（LPS）诱导Caco-2细胞建立屏障损伤细胞模型，分别采用不同浓度A.muciniphila、菌体组分和菌代谢产物处理细胞，结合非靶向代谢组学、CCK-8法及荧光定量PCR技术，评估细胞活力、黏液相关因子表达、炎症因子、凋亡相关因子及MAPK信号通路关键蛋白的mRNA表达变化。结果显示与LPS组相比，当A.muciniphila浓度为10⁷和10⁵CFU/mL时，Caco-2细胞Muc2 mRNA的表达量显著升高，分别增加16.75%和11.57%，TFF3 mRNA的表达量分别增加12.38%和9.71%；A.muciniphila细胞壁处理后，与LPS组相比，Caco-2细胞Muc2表达量增加18.71%，TFF3表达量显著增加13.28%；A.muciniphila主要代谢产物L-天冬氨酸（L-aspartic acid）、L-组氨酸（L-histidine）、柠檬酸（Citric acid）和犬尿酸（Kynurenic acid）诱导Caco-2细胞Muc2 mRNA的表达量增加18.09%、20.29%、23.67%、18.61%；细胞TFF3 mRNA的表达量分别上升10.22%、10.21%、10.29%、10.98%。与LPS组相比，A.muciniphila及其细胞壁和主要代谢产物处理后，Caco-2细胞促炎因子TNF-α、IL-1β、IL-6和IL-8 mRNA的表达量显著下降；细胞促凋亡因子Bax、Caspase-3、Caspase-8 以及促凋亡蛋白p21和p27 mRNA的表达量显著降低；MAPK信号通路中三个关键蛋白p38、JNK、ERK mRNA的表达量显著下降。综上所述，A.muciniphila可通过细胞壁及主要代谢产物，抑制MAPK信号通路激活，修复受损的肠道屏障。本研究为完善A.muciniphila在改善肠道屏障损伤中的作用物质及作用机制提供了理论基础。

Abstract: Previous research found that Hericium erinaceus-derived peptide (Lys-Ser-Pro-Leu-Tyr, KSPLY) ameliorates intestinal barrier injury and increases the abundance of the gut bacterium Akkermansia muciniphila. To investigate the role of A.muciniphila in improving intestinal barrier damage, a barrier injury model was established using lipopolysaccharide (LPS)-induced Caco-2 cells. The cells were treated with different concentrations of A.muciniphila, its cellular components, or its metabolic products. Untargeted metabolomics was employed to analyze metabolite profiles, cell viability was assessed by the CCK-8 assay, and mRNA expression levels of mucin-related factors, inflammatory cytokines, apoptosis-related factors, and key MAPK signaling pathway proteins were measured using quantitative real-time PCR. Compared to the LPS group, significant upregulation of Muc2 mRNA expression was observed by 16.75% and 11.57% after treatment with A.muciniphila at 10⁷ and 10⁵ CFU/mL, respectively, while TFF3 mRNA expression was increased by 12.38% and 9.71%. Treatment with the A.muciniphila cell wall resulted in an 18.71% increase in Muc2 and a 13.28% increase in TFF3 expression. The major metabolites of A.muciniphila—L-aspartic acid, L-histidine, citric acid, and kynurenic acid—were found to enhance Muc2 mRNA expression by 18.09%, 20.29%, 23.67%, and 18.61%, respectively, and TFF3 mRNA expression by 10.22%, 10.21%, 10.29%, and 10.98%. Furthermore, the mRNA expression of pro-inflammatory cytokines (TNF-α, IL-1β, IL-6, IL-8), pro-apoptotic factors (Bax, Caspase-3, Caspase-8), pro-apoptotic proteins (p21, p27), and key MAPK pathway components (p38, JNK, ERK) was significantly downregulated by A. muciniphila, its cell wall, and its key metabolites. In conclusion, A.muciniphila and its cell wall components and primary metabolites can alleviate intestinal barrier injury, likely by inhibiting the activation of the MAPK signaling pathway. This study provides a theoretical basis for understanding the active substances and mechanisms by which A.muciniphila improves intestinal barrier damage.