Mechanisms of maternal high fat diet induced susceptibility to gut inflammation in offspring

坏死性小肠结肠炎（NEC）是一种毁灭性的疾病，每年花费美国医疗保健系统50亿美元。它主要影响早产儿，没有有效的预防或治疗策略。发病机制仍然知之甚少，但早产是最重要的风险因素，改变的微生物群和过早的免疫反应在发病机制中很重要。肥胖/超重母亲（美国50%的育龄妇女）所生婴儿的早产风险增加，我们的母体高脂肪饮食模型（mHFD）导致对NEC的易感性增加，这是由改变的细菌定植和先天免疫介导的。我们的目标是利用这个模型来阐明mHFD如何改变新生儿的定植和免疫力，以增加对NEC的易感性。该建议利用了一种新的小鼠mHFD模型，现在在我们的实验室建立。我们最近用这个模型表明，mHFD暴露导致后代中独特的出生后定植模式（微生物群）。这种改变的微生物群随后增加了炎性3型先天淋巴细胞（ILC 3），导致对NEC样损伤的易感性增加。我们有初步的数据，强烈表明，暴露于子宫内的细菌依赖性配体，通过Toll样受体（TLR）的信号是负责改变mHFD后代的微生物群。此外，引入产生芳烃受体（Ahr）配体的细菌（L. murinus）在新生后代中扩增这些ILC 3。L.在mHFD后代中，murinus特异性增加。因此，我们假设暴露于mHFD导致子宫内TLR信号传导的改变和随后的出生后Ahr配体产生细菌的定植，这些细菌扩增ILC 3，导致后代对肠道炎症的易感性增加。为了测试这一点，我们有两个具体的目标：（1）确定子宫内TLR介导的信号传导如何调节后代中出生后定植的mHFD改变，以及（2）确定mHFD微生物群的配体如何调节Ahr信号传导以在后代中培养ILC 3。我们将通过使用无菌小鼠、条件性MyD 88敲除小鼠（没有通过MyD 88的TLR信号传导）和置于mHFD上的特异性TLR缺陷小鼠来测试特异性目标1。具体目标2将利用已知Ahr配体和拮抗剂的体内和体外（分选纯化的ILC 3和肠外植体）引入来检查。还将在暴露于我们的NEC模型的新生小鼠中用预处理的ILC 3进行连续转移实验。在这些研究完成后，我们将大大提高我们对母体营养如何影响后代出生后定植的认识，并进一步确定可推广的和新颖的宫内和出生后方法，以改变新生儿的微生物群并调节ILC 3功能以预防NEC。

Necrotizing enterocolitis (NEC) is a devastating disease that costs the US healthcare system US$5 billion annually. It primarily affects preterm infants and there are no effective targeted strategies for prevention or treatment. The pathogenesis remains poorly understood but prematurity is the most significant risk factor and an altered microbiota and premature immune response are important in the pathogenesis. The risk for prematurity increases in babies born to mothers whom are obese/overweight (50% of childbearing age women in the US), and our maternal high fat diet model (mHFD) results in increased susceptibility to NEC mediated by altered bacterial colonization and innate immunity. Our objective is to utilize this model to unravel how mHFD alters neonatal colonization and immunity to increase susceptibility to NEC. This proposal utilizes a novel murine mHFD model, now established in our lab. We have recently shown with this model that mHFD exposure results a unique post-natal colonization pattern (microbiota) in offspring. This altered microbiota subsequently increases inflammatory type 3 innate lymphoid cells (ILC3), resulting in increased susceptibility to NEC-like injury. We have preliminary data that strongly suggests that exposure to bacteria-dependent ligands in-utero that signal via toll-like receptors (TLR) are responsible for the altered microbiota in mHFD offspring. In addition, introduction of an aryl hydrocarbon receptor (Ahr) ligand-producing bacteria (L. murinus) expands these ILC3 in neonatal offspring. L. murinus is specifically increased in mHFD offspring. We therefore hypothesize that exposure to mHFD results in altered in utero TLR signaling and subsequent post-natal colonization with Ahr-ligand producing bacteria that expand ILC3, resulting in increased susceptibility to intestinal inflammation in offspring. To test this, we have 2 specific aims: (1) Determine how in utero TLR-mediated signaling regulates mHFD alteration of post-natal colonization in offspring and (2) Determine how ligands of mHFD microbiota modulate Ahr signaling to educate ILC3 in offspring. We will test specific aim 1 by utilizing germ-free mice, conditional MyD88 knock-out mice (no TLR signaling via MyD88) and specific TLR deficient mice placed on mHFD. Specific aim 2 will be examined utilizing in vivo and in vitro (sort-purified ILC3 and intestinal explants) introduction of known Ahr ligands and antagonists. Adoptive transfer experiments will also be performed with pre-treated ILC3s in neonatal mice exposed to our NEC model. At the completion of these studies, we will dramatically advance our knowledge of how maternal nutrition affects post-natal colonization in offspring and further identify generalizable and novel in-utero and post-natal approaches to altering the microbiota in neonates and modulating ILC3 function to prevent NEC.