Investigating miR-375-mediated regulation of intestinal helminth infection

研究 miR-375 介导的肠道蠕虫感染调节

基本信息

批准号：
10371515
负责人：
Praveen Sethupathy
金额：
$ 21.57万
依托单位：
CORNELL UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2021
资助国家：
美国
起止时间：
2021-09-24 至 2023-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10371515
关键词：
3&apos Untranslated Regions Address Age Architecture Binding Binding Sites Bioinformatics Biological Assay Bone Marrow CRISPR/Cas technology Cell Count Cell Differentiation process Cell Lineage Cell Maturation Cells Data Development Epithelial Epithelial Cells Exhibits Follow-Up Studies Foundations Future Gene Expression Genes Genetic Genetic study Goals Helminths Hookworm Infections Human Immune Immune response Individual Infection Infection prevention Interleukin-13 Intestines Knock-out Knockout Mice Knowledge LoxP-flanked allele Luciferases Measures Mediating MicroRNAs Modeling Molecular Morbidity - disease rate Mus Nematospiroides dubius Parasites Pathway interactions Public Health Regulation Resistance Role Secretory Cell Signal Transduction Small Intestines Soil Testing Therapeutic Tissues Transgenic Mice Villus Wild Type Mouse base cell type gastrointestinal epithelium helminth infection improved in vivo intestinal crypt intestinal epithelium novel protective effect response sex single-cell RNA sequencing stem stem cells tissue repair transcription factor transcriptomics villin

项目摘要

PROJECT SUMMARY: Soil-transmitted intestinal helminth parasites infect billions of people worldwide, causing substantial morbidity and posing a significant global public health problem. Recent hallmark studies have shown that following helminth infection, specialized cells in the gut epithelium called tuft cells expand and trigger a host- protective type 2 immune response that promotes parasite expulsion and tissue repair. Tuft cells are derived from stem cells at the base of the intestinal epithelial crypt, and transcription factors such as Pou2f3 that drive the maturation of tuft cells have been identified. However, despite these advances, the molecular mechanisms that control tuft cell abundance and anti-helminth function remain incompletely understood. Although it is established that microRNAs (miRNAs) as a class of regulatory molecules are critical for proper intestinal architecture and function, the roles of individual miRNAs in the intestinal epithelium are just now starting to emerge. We recently discovered that the whole-body genetic deletion of a single miRNA, miR-375, was associated with lower worm burdens after infection with Heligmosomoides polygyrus, a helminth parasite of mice used as a model for human hookworm infection. Moreover, the 375-/- mice exhibited increased abundance of tuft cells in the small intestine. Notably, we also determined through a bioinformatic screen that Pou2f3 is a predicted target of miR-375. This proposal is focused on bridging two critical knowledge gaps: (1) Is the effect on H. polygyrus worm burden mediated by loss of miR-375 function in the intestinal epithelial cell (IEC) lineage, immune cell lineage, or both? and (2) Does the effect of miR-375 loss depend on Pou2f3-mediated maturation of tuft cells? In the first Aim, we will define the impact of IEC-specific miR-375 loss on resistance to H. polygyrus. To accomplish this goal, we will apply two independent, complementary strategies. Specifically, we will: (i) generate bone marrow chimeric mice, in which miR-375 deficiency is restricted to non-immune cells, and also (ii) breed miR-375fl/fl;Vil1-Cre mice, in which miR-375 deficiency is specific to IECs, and compare H. polygyrus worm burden after infection with what we have observed previously in whole-body miR-375-/- mice. In the second Aim, we will establish whether tuft cells are required for the effects of miR-375 loss on H. polygyrus worm burden. We will first determine whether Pou2f3 is a direct target of miR-375 and also leverage single cell transcriptomics to identify additional candidate miR-375 target genes whose expression levels are significantly increased in intestinal epithelial stem and/or secretory progenitor cells of 375-/- mice during the response to helminth infection. We will then perform genetic studies to determine whether loss of Pou2f3 is sufficient to negate the positive effects of miR-375 loss on H. polygyrus worm burden. The completion of these studies will substantially advance the field by establishing that an IEC miRNA regulates tuft cell-mediated anti-helminth function during H. polygyrus infection. Improved understanding of the key epithelial regulators of the host response to helminth will be critical for developing new strategies to treat and prevent infection.

项目摘要：土壤传播的肠道蠕虫寄生虫感染了全球数十亿人大量的发病率并带来了重大的全球公共卫生问题。最近的Hallmark研究表明在蠕虫感染后，肠道上皮中的专门细胞称为簇细胞扩张并触发宿主 - 保护性2型免疫反应，可促进寄生虫驱动和组织修复。簇细胞得出从肠上皮隐窝底部的干细胞以及驱动诸如POU2F3之类的转录因子已经鉴定出簇细胞的成熟。但是，尽管有这些进步，分子机制控制簇细胞丰度和抗螺旋功能仍然不完全理解。虽然是确定microRNA（miRNA）作为一类调节分子对于适当的肠道至关重要建筑和功能，单个miRNA在肠上皮中的作用刚刚开始出现。我们最近发现，单个miRNA的全身遗传缺失，miR-375是与Heligmosomoides Polygyrus感染后，蠕虫寄生虫是小鼠寄生虫，与较低的蠕虫负担相关用作人钩虫感染的模型。此外，375 - / - 小鼠的丰度增加小肠中的簇细胞。值得注意的是，我们还通过生物信息学屏幕确定了POU2F3是一个预测miR-375的靶标。该建议的重点是弥合两个关键的知识差距：（1）效果是在肠道上皮细胞（IEC）谱系中miR-375功能的损失介导的多孔猪螺旋体负担上，免疫细胞谱系，还是两者兼而有之？（2）miR-375损失的影响取决于POU2F3介导的成熟簇细胞？在第一个目的中，我们将定义IEC特异性miR-375损失对抗多犬H. polygyrus的影响。为了实现这一目标，我们将采用两种独立的互补策略。具体来说，我们将：（i）产生骨髓嵌合小鼠，其中miR-375缺乏仅限于非免疫细胞，也是（ii）mir-375fl/fl; vil1-cre小鼠，其中miR-375缺乏特定于IEC，并比较polygyrus H. polygyrus 感染后我们先前在全身miR-375 - / - 小鼠中观察到的蠕虫负担。在第二个目的，我们将确定miR-375损失对h. polygyrus蠕虫负担的影响是否需要簇状细胞。我们将首先确定POU2F3是否是miR-375的直接靶标，并且还利用单细胞转录组学确定其他候选miR-375靶基因，其表达水平在在对蠕虫的反应期间，375 - / - 小鼠的肠上皮茎和/或分泌祖细胞感染。然后，我们将进行遗传研究，以确定POU2F3的丢失是否足以消除 miR-375损失对h. polygyrus蠕虫负担的积极影响。这些研究的完成将大大通过确定IEC miRNA调节簇细胞介导的抗螺旋功能，以推进该领域 H. Polygyrus感染。对主机对蠕虫的反应的关键上皮调节器的理解得以提高对于制定新的策略来治疗和防止感染至关重要。