Defining the structural and functional properties of select agricultural fibers underlying their immunomodulatory actions

该计划的长期目标是定义来自不同常见农产品的纤维的生理相关微生物介导的发酵途径，并揭示各种环境下纤维相关免疫调节途径的生化机制。纤维是不可消化的碳水化合物，依靠肠道微生物（细菌和真菌）的发酵，分解成可被人体细胞利用的副产物（如短链脂肪酸）。我们之前的工作显示了特定的益生菌群落（改变的微生物丰度/多样性），更重要的是，改变的微生物功能（酶）与减少的ß-果聚糖纤维发酵有关。这种改变的发酵与SCFA的产生减少和未发酵的ß-果聚糖与白细胞（巨噬细胞、T细胞和中性粒细胞）的免疫调节相互作用增加有关，特别是通过Th17细胞途径。我们的初步研究结果表明，这些纤维的结构在其可发酵性和白细胞反应过程中起着重要作用，因为聚合程度（DP，纤维长度）与炎性小体（NLRP3:IL-1ß）激活有关；较高的DP纤维增加了细胞因子的分泌和炎性小体基因的表达（例如，IL-1ß， IL-23）。拟议研究的短期目标是：(1)描述膳食纤维（如ß-葡聚糖，半乳糖寡糖，菊粉）的结构特征（如酚含量，聚合度）；纤维要么购买，要么从农业食品中提取。结构特征将与白细胞对每种膳食纤维化合物反应的细胞途径相关，使用中尺度发现和ELISA进行检查；(2)确定肠道微环境因素（如pH值、代谢物）参与调节微生物群培养的功能（酶），介导其纤维发酵活性；(3)揭示特定微生物酶在纤维发酵过程中的作用；(4)使用靶向显微镜和途径分析（qPCR, western blotting， ELISA）提供纤维-微生物相互作用如何影响白细胞反应（炎症小体途径的调节）的机制见解。该项目将促进我们对膳食纤维物理特性在调节与白细胞和发酵微生物相互作用中的作用的基本理解，同时为HQP在食品化学、微生物学和免疫学方面提供多学科培训机会。我们的工作将有助于提高对环境因素（微环境和农产品）与驻留在肠粘膜（纤维发酵的生理部位）的白细胞群之间发生的相互作用的理解。

The long-term goal of this program is to define the physiologically relevant microbe-mediated fermentation pathways for select fibers of interest, sourced from different common agricultural products and to uncover the biochemical mechanisms underlying fiber-associated immunomodulatory pathways in various environmental settings. Fibers are nondigestible carbohydrates that rely on fermentation by intestinal microorganisms (bacteria and fungi) for their decomposition into byproducts that can be utilized by human cells (e.g., short chain fatty acids). Our previous work showed specific dysbiotic microbial communities (altered microbe abundance/diversity), and more significantly, altered microbe functions (enzymes) were associated with reduced ß-fructan fibre fermentation. This altered fermentation was associated with reduced production of SCFA and increased immunomodulatory interactions of unfermented ß-fructans with leukocytes (macrophages, T cells, and neutrophils), particularly through Th17 cellular pathways. Our preliminary findings suggest that the structure of these fibers, which differs between fiber subtypes and the agricultural source from which they are isolated, plays a role in their fermentability and in leukocyte response processes, as the degree of polymerization (DP; length of the fiber) was associated with inflammasome (NLRP3:IL-1ß) activation; higher DP fibres increased cytokine secretion and expression of inflammasome genes (e.g., IL-1ß, IL-23). The short-term objectives of the proposed research are: (1) profile the structural features (e.g., phenolic content, degree polymerization) of dietary fibres (e.g., ß-glucans, galatooligosaccharides, inulins); fibres will be either purchased or extracted from agricultural food sources. Structural features will be correlated with the cellular pathways of leukocyte responses to each of the dietary fibre compounds, examined using mesoscale discovery and ELISA; (2) identify gut microenvironment factors (e.g., pH, metabolites) involved in regulating the functions (enzymes) of microbiota cultures that mediate their fibre fermentation activities; (3) reveal the role of specific microbial enzymes in mediating fibre fermentation processes; and (4) provide mechanistic insight into how fibre-microbe interactions impact leukocyte response (modulation of inflammasome pathways) using targeted microscopy and pathway analysis (qPCR, western blotting, ELISA). This program will advance our fundamental understanding of the role of dietary fibre physical properties in regulating interactions with leukocytes and fermentative microbes while providing multidisciplinary training opportunities for HQP in food chemistry, microbiology and immunology. Our work will aid to improve understanding of interactions that occur, between environmental factors (microenvironment and agricultural products) and leukocyte cell populations that reside in the intestinal mucosa, the physiological site of fibre fermentation.