Lactogenic immunity/probiotics: Effect on neonatal gut immunity

描述（由申请人提供）：母乳含有促进婴儿健康和中度腹泻的母体抗体（MatAb），以及可溶性免疫调节剂（TGF 2、IL 4、sCD 14等），其作用在很大程度上尚未探索。轮状病毒（RV）是全球婴儿腹泻的主要原因。新批准的口服RV疫苗在发展中国家的效力未经证实或较低，高成本仍然是普遍采用的障碍。虽然乳酸菌（LAB）减少RV腹泻的婴儿，机制是不确定的，并提供并行MatAb/介体与LAB的影响是未经探讨的。我们的创新研究将首先评估初乳（col）/牛奶和关键的母体免疫调节剂（TGF 2，IL 4）对益生菌定植的影响。然后，我们确定了单独的和合并的影响，col/牛奶和LAB对免疫成熟，缓和RV腹泻和稳态在新生儿肠道。母乳喂养的婴儿与配方奶粉喂养的婴儿的双歧杆菌和双歧杆菌菌株不同。我们假设，col/milk促进了这2种益生菌LAB的定植，需要更少或更低的LAB剂量，这种协同作用增强了肠道免疫成熟和体内平衡，导致轮状病毒腹泻的缓和。初步研究表明，鼠李糖乳杆菌GG（LGG）长期定居在col/牛奶喂养的猪后，单次口服剂量支持我们的新想法。我们独特的新生无菌（Gn）猪模型是初乳剥夺的（无MatAb预乳/乳），允许MatAb/介体操作并且不含微生物（剖腹产衍生的，无菌饲养），允许评价限定的益生菌。重要的是，它们的胃肠道生理学、粘膜免疫应答和对人RV腹泻的易感性与婴儿相似。在目的1中，我们确定含有MatAb/介质（或TGF 2 + IL 4）的col/乳是否影响a）乳双歧杆菌和LGG在肠道中的定殖、持久性和分布，以及B）新生儿肠道免疫应答的成熟。在目的2中，我们阐明了a）具有低滴度RV抗体的col/milk（自然暴露的母猪）与LAB是否调节HRV发病机制和免疫，从而缓和HRV腹泻和B）肠道致病性病毒感染是否影响肠道稳态和LAB微生物群落。我们将使用体内生物发光成像来研究LGG与/不与B的相互作用。乳杆菌在新生儿肠道中的相互作用，以及col/牛奶或HRV感染对这些相互作用的影响。这种体内细菌生态学的新方法将阐明宿主内益生菌LAB的相互作用。待评估的肠道和全身免疫学参数包括：1）先天性应答; 2）病毒特异性、LAB特异性和总同种型B细胞应答，重点关注肠道伊加; 3）T细胞应答（Th 1、Th 2、Th 3、Tr 1细胞因子）。我们的创新研究将提供对新生儿免疫成熟的更好理解，以及对新型col/milk MatAb/mediators与益生菌在RV保护性免疫中的有益作用的新见解。此外，我们的研究结果将建议通过对母乳喂养的婴儿或孕妇短期联合施用益生菌和col/mlk介体来限制发展中国家婴儿的RV腹泻的新策略。这种方法将促进婴儿中稳定的LAB定殖和肠道免疫成熟，从而导致疾病缓和。免疫调节和新生儿免疫应答的增强也对食物抗原或抗生素的耐受性具有根本意义，以控制食物过敏或炎性肠综合征以及过敏性肠炎。

DESCRIPTION (provided by applicant): Breast milk contains maternal antibodies (MatAbs) that promote infant health and moderate diarrhea, and soluble immunomediators (TGF2, IL4, sCD14, etc) whose role is largely unexplored. Rotavirus (RV) is a leading cause of diarrhea in infants worldwide. Newly licensed oral RV vaccines have unproven or lower efficacies in developing countries and high costs remain obstacles to universal adoption. Although lactic acid bacteria (LAB) reduce RV diarrhea in infants, mechanisms are undefined and the effect of providing concurrent MatAb/mediators with LAB is unexplored. Our innovative studies will first assess the impact of colostrum (col)/milk and key maternal immunomediators (TGF2, IL4) on colonization by probiotics. We then determine the individual and combined effects of col/milk and LAB on immunologic maturation, moderation of RV diarrhea and homeostasis in the neonatal gut. Bifidobacteria and lactobacillus strains differ in breastfed vs formula fed infants. We hypothesize that col/milk promotes colonization by these 2 probiotic LAB, requiring fewer or lower LAB doses, and this synergism enhances gut immune maturation and homeostasis leading to moderation of rotavirus diarrhea. Preliminary studies showing that L rhamnosus GG (LGG) colonized col/milk-fed pigs long-term after a single oral dose support our novel idea. Our unique neonatal gnotobiotic (Gn) pig model is colostrum- deprived (no MatAb pre-col/milk), allowing MatAb/mediator manipulation and free of microbes (caesarean-derived, sterile housing), permitting evaluation of defined probiotics. Importantly their gastrointestinal physiology, mucosal immune responses and susceptibility to human RV diarrhea mimic that of infants. In Aim 1, we determine if col/milk containing MatAb/mediators (or TGF2+IL4) influence a) Bifidobacterium lactis and LGG colonization, persistence and distribution in the gut and b) maturation of neonatal gut immune responses. In Aim 2 we elucidate a) if col/milk with low titer RV antibody (naturally exposed sows) in concert with LAB modulates HRV pathogenesis and immunity, thereby moderating HRV diarrhea and b) if an enteropathogenic viral infection affects gut homeostasis and the LAB microflora. We will use in vivo bioluminescent imaging to study LGG interactions with/without B. lactis in the neonatal gut, and the influence of col/milk or HRV infection on these interactions. This novel approach to in vivo bacterial ecology will elucidate probiotic LAB interactions within the host. Intestinal and systemic immunologic parameters to be assessed include: 1) innate responses; 2) virus- specific, LAB-specific and total isotype B cell responses focusing on gut IgA; 3) T cell responses (Th1, Th2, Th3, Tr1 cytokines). Our innovative studies will provide improved understanding of maturation of neonatal immunity and new insights into the beneficial effects of novel col/milk MatAb/mediators in concert with probiotics in protective immunity to RV. Further, our findings will suggest new strategies to limit RV diarrhea in infants in developing countries through short-term co- administration of probiotics and col/mlk mediators to breastfed infants or pregnant mothers. This approach will promote stable LAB colonization and gut immune maturation in infants leading to disease moderation. Immunomodulation and enhancement of neonatal immune responses also have fundamental implications for tolerance to food antigens or commensals to control food allergies or inflammatory bowel syndromes as well as antibiotic-induced diarrheas.