Isolation and Immunomodulatory Evaluation of Glycyrrhiza Polysaccharide-Induced Probiotics from Murine Gut Microbiota

While low-molecular-weight Glycyrrhiza polysaccharide (LGP) demonstrates immunomodulatory effects through gut microbiota modulation, the specific LGP-responsive bacterial taxa and their therapeutic potential as next-generation probiotics (NGPs) remain poorly characterized. In this study, we isolated dominant LGP-induced bacterial strains (Bacteroides vulgatus and Parabacteroides sp.) from murine intestinal contents and systematically evaluated their probiotic potential using comprehensive NGPs selection criteria: 1) it has excellent gastrointestinal tolerance, and the survival rate in gastric juice with pH 2.5 and 0.5% cholate is over 90%; 2) broad-spectrum antibiotic susceptibility; 3) potent pathogen inhibition (Escherichia coli, Enterobacter cloacae, and Proteus vulgaris), and 4) absence of cytotoxicity or phagocytic interference in RAW264.7 macrophages (confirmed for heat-killed cells, supernatants, and lysates). In vivo experiments, we found that NGPs exhibited significant immunomodulatory abilities, B. vulgatus fresh culture (BX) stimulated 2.49-fold leukocyte proliferation, and parabacteroides sp. lysate (PL) elevated IL-10 production (1.33-fold). NGPs slao ameliorated hepatic injury markers, restored intestinal barrier integrity, enriched beneficial taxa (Akkermansia, Rikenella, UCG-010), enhanced SCFA production (isobutyrate, isovalerate, butyrate, valerate), and modulated critical metabolic pathways (LPS biosynthesis, TCA cycle, gluconeogenesis). These findings not only elucidate the microbial mechanisms underlying LGP's immunomodulation but also provide two well-characterized, gut-derived NGP candidates with validated multi-functional efficacy, meeting stringent safety and functionality requirements for therapeutic development.

低分子量甘草多糖（low-molecular-weight Glycyrrhiza polysaccharide, LGP）可通过调控肠道菌群发挥免疫调节作用，但其响应LGP的特异性细菌类群以及其作为下一代益生菌（next-generation probiotics, NGPs）的治疗潜力仍未得到充分阐明。本研究从小鼠肠道内容物中分离得到优势LGP诱导菌株——普通拟杆菌（Bacteroides vulgatus）和副杆菌属（Parabacteroides sp.），并采用严格的下一代益生菌筛选标准对其益生潜力进行了系统评估：1）具备优异的胃肠道耐受能力，在pH 2.5的胃液及0.5%胆盐中的存活率均超过90%；2）具有广谱抗生素敏感性；3）可有效抑制致病菌（大肠埃希菌（Escherichia coli）、阴沟肠杆菌（Enterobacter cloacae）及普通变形杆菌（Proteus vulgaris））；4）对RAW264.7巨噬细胞无细胞毒性或吞噬干扰（针对热灭活菌体、培养上清及菌体裂解液均得到验证）。体内实验结果显示，所筛选的下一代益生菌展现出显著的免疫调节能力：普通拟杆菌新鲜培养液（BX）可使白细胞增殖水平提升2.49倍，副杆菌属菌体裂解液（PL）可使白细胞介素-10（interleukin-10, IL-10）分泌量升高1.33倍。这类下一代益生菌还可改善肝损伤标志物水平、恢复肠道屏障完整性，富集有益菌属（嗜黏蛋白阿克曼菌属、理研菌属、UCG-010），促进短链脂肪酸（short-chain fatty acids, SCFA）生成（包括异丁酸、异戊酸、丁酸及戊酸），并调控关键代谢通路（脂多糖（lipopolysaccharide, LPS）生物合成、三羧酸循环、糖异生）。本研究结果不仅阐明了LGP免疫调节作用背后的微生物学机制，同时还提供了两株经过充分表征的肠道来源下一代益生菌候选菌株，其多功能功效已得到验证，满足治疗开发所需的严苛安全性与功能学要求。