Natural Killer Cell Immunity Induced by ß-1,3-Glucan

免疫系统的发展是为了维持体内平衡，同时保护宿主免受感染微生物的侵袭。现在很明显，环境因素激活和塑造了免疫力。-1，3-葡聚糖是一种在饮食谷物和微生物中被吸入和消费的分子。有证据表明，饮食中的-1，3-葡聚糖能激活包括自然杀伤(NK)细胞在内的免疫功能；然而，其机制及其在宿主防御发育中的作用尚不清楚。这项建议的目的是了解-1，3-葡聚糖如何调节和导致NK细胞免疫分化。 自然杀伤细胞(Natural Killer，NK)是一种先天的淋巴细胞，可以在没有预先刺激的情况下直接杀死致病微生物，如细菌、寄生虫、真菌以及肿瘤细胞和病毒感染的细胞。然而，最近的研究表明，它们可以形成一种类似于适应性免疫细胞(如T细胞)的记忆特性。适应性免疫细胞对以前遇到的病原体表现出增强的反应，这增强了它们清除入侵微生物病原体的能力。 我们之前已经证明，NK细胞直接杀死隐球菌和念珠菌，这是一种能够破坏动态平衡并导致病理的环境真菌。最近，我们确定NKp30是识别和介导NK细胞对这些真菌的细胞毒作用的NK细胞模式识别受体(PRR)。此外，我们还确定真菌细胞壁中的病原体相关分子模式(PAMP)-1，3-葡聚糖是NKp30的配体，它介导NK细胞的识别和杀伤。有趣的是，我们发现-1，3-葡聚糖不仅在PRR-PAMP介导的NK细胞杀伤中起关键作用，而且能够增强NK细胞对新生念珠菌和白色念珠菌的杀伤作用。进一步的实验表明，-1，3-葡聚糖增强的杀伤作用在最初模拟后的几周内逐渐减弱，然后在第四周表现出更强的NK细胞杀伤作用，这与适应性免疫回忆反应类似。目前尚不清楚1，3-葡聚糖如何增强NK细胞杀伤，以及它是否导致激活事件、分化事件或调节NK细胞免疫的记忆获得。 在这个方案中，我们计划使用转基因和/或人源化的小鼠来确定-1，3-葡聚糖是否增强了NK细胞的杀伤作用，并了解了NKp30与-1，3-葡聚糖在增强NK细胞杀伤作用中的相互作用。我们将使用RNA-seq和质量细胞术来分析NK细胞在-1，3-葡聚糖增强的细胞毒作用中的差异基因表达。本项目的最终目的是了解-1，3-葡聚糖增强杀伤的机制，以及-1，3-葡聚糖是否诱导抗原特异性的NK细胞克隆扩增和记忆发育，或者是导致NK细胞杀伤活性增强的表观遗传学改变，并确定增强的NK细胞杀伤活性是真菌还是更广泛的病原微生物所特有的。

The immune system has developed to maintain homeostasis while protecting the host from infectious microorganisms. It is now clear that environmental factors activate and shape immunity. -1,3-glucan is one such molecule that is both inhaled and consumed in dietary grains and microbes. Evidence shows that dietary -1,3-glucan activates immunity including natural killer (NK) cells; however, the mechanisms and their role in host defense development is unknown. The goal of this proposal is to understand how -1,3-glucan modulates and causes differentiation of NK cell immunity. Natural Killer (NK) cells are innate lymphocytes that directly kill pathogenic microorganisms, e.g., bacteria, parasites and fungi as well as tumor cells and virus-infected cells without prior stimulation. However, recent studies demonstrate that they can develop a memory-like property similar to adaptive immune cells such as T cells. Adaptive immune cells demonstrate enhanced responses to previously encountered pathogens that enhances their ability to clear invading microbial pathogens. We previously showed that NK cells directly kill Cryptococcus and Candida, environmental fungi that have the ability to disrupt homeostasis and cause pathology. Recently we determined that NKp30 is the NK cell pattern recognition receptor (PRR) that recognizes and mediates NK cell cytotoxicity against these fungi. Furthermore, we identified that -1,3-glucan, a pathogen-associated molecular pattern (PAMP) in the fungal cell wall is the ligand for NKp30, which mediates NK cell recognition and killing. Interestingly, we found that -1,3-glucan is not only critical for PRR-PAMP-mediated NK cell killing, but also able to enhance NK cell cytotoxicity against C. neoformans and C. albicans. Further experiments suggest that the -1,3-glucan-enhanced killing tapers off over few weeks after initial simulation and then showed greater NK cell killing at the fourth week, a similar phenomenon to adaptive immune recall responses. It remains unknown how -1,3-glucan enhances NK cell killing and whether it leads to an activation event, a differentiation event, or acquisition of memory that modulates NK cell immunity. In this proposal, we plan to use transgenic and/or humanized mice to determine whether -1,3-glucan enhances NK cell cytotoxicity and understand the NKp30 interaction with -1,3-glucan in the enhanced NK cell cytotoxicity. We will profile differential gene expression of NK cells in the -1,3-glucan-enhanced cytotoxicity using RNA-seq and mass cytometry. The ultimate goal of this program is to understand the mechanisms that -1,3-glucan-enhanced killing, and whether -1,3-glucan induces antigen specific NK cell clonal expansion and memory development or epigenetic change leading to enhanced NK cell cytotoxicity, and determine whether the enhanced NK cell cytotoxicity is specific to fungi or to a wider range of pathogenic microorganisms.