Mechanisms and consequences of human milk oligosaccharide growth and bile stress across diverse strains of the potential therapeutic bacterium, Akkermansia muciniphila.

潜在治疗性细菌阿克曼氏菌 (Akkermansia muciniphila) 不同菌株的母乳寡糖生长和胆汁应激的机制和后果。

• 批准号: 10436303
• 负责人: Gilberto Flores
• 金额: $ 36.25万
• 依托单位: CALIFORNIA STATE UNIVERSITY NORTHRIDGE
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-07-09 至 2024-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10436303
• 关键词: Anti-Inflammatory Agents; Bacteria; Bile Acids; Bile fluid; Binding; Biological Assay; Blood Glucose; Cardiometabolic Disease; Cells; Cloning; Coculture Techniques; Collection; Complement; Consumption; Coupled; Data; Desire for food; Disease; Enzymes; Epithelial Cells; Fermentation; Gastrointestinal Diseases; Gene Expression; Glycoside Hydrolases; Goals; Growth; Health; Human; Human Activities; Human Milk; Immune response; In Vitro; Life; Measures; Metabolic Diseases; Mission; Molecular; Oligosaccharides; Outcome; Physiological; Polysaccharides; Probiotics; Production; Property; Proteins; Proteomics; Public Health; Research; Shapes; Stress; Testing; Therapeutic; Therapeutic Uses; United States National Institutes of Health; Vitamin B 12; Work; base; biological adaptation to stress; cardiometabolism; cofactor; design; disability; experimental study; extracellular; gut colonization; gut microbiome; immunogenic; innovation; insight; member; metabolomics; microbial; multiple omics; novel; obesity treatment; prebiotics; rational design; resistance mechanism; response; targeted treatment; therapy design; transcriptomics; treatment strategy

The current paradigm that Akkermansia muciniphila is a beneficial member of the human gut microbiome is based on an incomplete understanding of the physiological diversity and mechanistic activity across the lineage, as all previous work has focused on one described strain. The long-term goal is to help develop targeted, therapeutic uses of Akkermansia, either through stimulating endogenous strains with prebiotics or by administering specific strains as probiotics. The overall objectives for this application are to characterize the molecular mechanisms and immunogenic properties of genomically diverse strains of human-associated Akkermansia grown (i) on human milk oligosaccharides (HMO) and (ii) in the presence of bile. The central hypothesis of this work is that human-associated Akkermansia have evolved different growth efficiencies on HMO and bile stress responses that shape their immunogenic potential in a strain dependent manner. The rationale for this work is that if we are to use Akkermansia for the therapeutic treatment of metabolic disorders or other gastrointestinal diseases, then we need to design biologically informed treatment strategies that promote or introduce select strains for optimal health outcomes. The central hypothesis will be tested by pursuing two specific aims: 1) Identify the molecular mechanisms and immunogenic properties of Akkermansia grown on HMO; 2) Identify the bile resistance mechanisms and immunogenic properties of Akkermansia grown in the presence of bile. Under the first aim, targeted gene expression studies coupled with cloning and functional assays will be used to identify glycoside hydrolase enzymes involved in growth on HMO in three genomically diverse Akkermansia isolates. Concurrently, targeted metabolomics analyses will be used to quantify HMO consumption and fermentation end products. Lastly, co-culture experiments with HMO grown Akkermansia cells and human epithelial cells will be used to measure bacterial binding efficiency and the immune response of the epithelial cells. For the second aim, transcriptomic and proteomic profiling, coupled with targeted metabolomics (i.e. bile acid composition and microbial extracellular polysaccharides) will be used to characterize the bile stress response of the three Akkermansia isolates. Similar to aim 1, co- culture experiments with bile grown Akkermansia strains and human epithelial cells be used to measure bacterial binding efficiency and the host immune response. The research proposed in this application is innovative because it represents a substantive departure from the status quo by providing insights into the physiological diversity and molecular mechanisms across the Akkermansia lineage. The proposed research is significant because it will help define the physiological landscape across the lineage, thereby opening new horizons for biologically informed treatment strategies that promote or introduce select Akkermansia strains for optimal health outcomes.

当前的范式认为，Akkermansia muciniphila 是人类肠道的有益成员 微生物组基于对生理多样性和机械活动的不完全理解 跨越整个谱系，因为之前的所有工作都集中在一种所描述的菌株上。长期目标是帮助 通过刺激内源菌株来开发 Akkermansia 的有针对性的治疗用途 益生元或通过施用特定菌株作为益生菌。该应用程序的总体目标是 表征基因组多样化菌株的分子机制和免疫原性特性 人类相关的阿克曼氏菌 (i) 在人乳寡糖 (HMO) 上生长，以及 (ii) 在存在 胆汁。这项工作的中心假设是与人类相关的阿克曼氏菌进化出了不同的生长方式 HMO 和胆汁应激反应的效率决定了它们在菌株依赖性中的免疫原性潜力 方式。这项工作的基本原理是，如果我们要使用阿克曼氏菌来治疗 代谢紊乱或其他胃肠道疾病，那么我们需要设计基于生物学的治疗方法 促进或引入精选菌株以获得最佳健康结果的策略。中心假设将是 通过追求两个特定目标进行测试：1）确定分子机制和免疫原性特性 在 HMO 上生长的 Akkermansia； 2) 确定胆汁抵抗机制和免疫原性 阿克曼氏菌在胆汁存在下生长。在第一个目标下，结合了靶向基因表达研究 克隆和功能测定将用于鉴定参与生长的糖苷水解酶 三种基因组多样化的阿克曼氏菌分离株中的 HMO。同时，靶向代谢组学分析将 用于量化 HMO 消耗和发酵最终产品。最后，共培养实验 HMO 培养的 Akkermansia 细胞和人上皮细胞将用于测量细菌结合效率 以及上皮细胞的免疫反应。对于第二个目标，转录组和蛋白质组分析， 结合靶向代谢组学（即胆汁酸成分和微生物胞外多糖） 将用于表征三种阿克曼氏菌分离株的胆汁应激反应。与目标 1 类似，共同 使用胆汁生长的 Akkermansia 菌株和人上皮细胞进行培养实验，用于测量 细菌结合效率和宿主免疫反应。本申请中提出的研究是 创新，因为它通过提供对现状的深刻见解，代表了对现状的实质性偏离。 Akkermansia 谱系的生理多样性和分子机制。拟议的研究 意义重大，因为它将有助于定义整个谱系的生理景观，从而开辟新的领域 促进或引入特定阿克曼氏菌菌株的生物学治疗策略的前景 以获得最佳的健康结果。