Fundamentals Of Human Lymphoid Cell Culture

Author : Ansel P. Nalin
Publisher :
ISBN 13 :
Total Pages : 0 pages
Book Rating : 4.:/5 (137 download)

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Download or read book Mechanisms of Human Innate Lymphoid Cell Development written by Ansel P. Nalin and published by . This book was released on 2021 with total page 0 pages. Available in PDF, EPUB and Kindle. Book excerpt: Human innate lymphoid cells (ILCs) comprise a diverse population of lymphocytes with various roles in tissue homeostasis, inflammation, host defense, and malignancy. ILCs collectively include the cytotoxic natural killer (NK) cells, which participate in direct lysis of infected or malignant cells, as well as distinct helper ILC populations (ILC1s, ILC2s, and ILC3s), which mediate their immunomodulatory roles via production of cytokines. Many physiologic pathways rely on the effector functions of NK cells and other ILC subsets; yet, the mechanisms that regulate their differentiation and maturation are not fully known. The overall goal of this research is to further characterize the pathways of human ILC development and to identify the mechanisms that regulate the differentiation and function of mature ILC populations. The results of these studies elucidate the roles of signal pathways and cellular interactions in the regulation of ILC biology. Activation of the Notch signaling pathway promoted the differentiation of non-NK helper ILC subsets at specific stages of development. While direct co-culture of ILC precursor cells with stromal feeder cells (and exogenous IL-7) was sufficient for the acquisition of NK cell markers (CD94 and NKG2A), the simultaneous presence of Notch ligand was required for the differentiation of functional ILC2s and ILC3s under these conditions. Although Notch was not required for NK cell differentiation, it did have an impact on the phenotype of in vitro-derived NK cells. Activation of Notch in the presence of stromal cells promoted the expression of the activating receptor NKp80, suggesting a role for Notch in the later stages of NK cell maturation. Based on the expression patterns of individual Notch receptors, it was hypothesized that there are stage-specific differences in the signaling mechanism. In support of this, it was shown that both NOTCH1 and NOTCH2 were involved in ILC2 and ILC3 differentiation from ILC precursor cells. Alternatively, NOTCH1 was the primary receptor involved in NKp80 acquisition from immature NK cells. It was also determined that the developmental potential for ILC2s, followed by ILC3s, was lost at successive stages of NK cell development. Compared to functional NK cells (those that express NKp80), earlier stages of immature NK cells and ILC precursor cells display greater developmental plasticity and heterogeneity at the bulk population level. It was observed that intermediate ILC populations were highly enriched for expression of CD200R1. Interestingly, the expression (or lack) of CD200R1 in specific populations correlated with significant differences in NK cell phenotypic and functional maturity. An intermediate NK cell subset lacking CD200R1 expression was found to have greater expression of markers associated with mature NK cells despite lacking other typical markers of terminal NK cell maturation, NKp80 and CD16. The phenotypic and functional properties associated with this population suggest that this minor subset of CD200R1- cells is out of sequence with the typical stages of NK cell maturation, further illustrating the heterogeneity of NK/ILC developmental intermediate populations. It was further hypothesized that the biology of mature ILC populations is regulated by tissue-specific interactions in the microenvironment. The NF-kB signaling pathway was identified as a regulator of ILC3 biology. Inhibition of NF-kB signaling resulted in decreased IL-22 production from ILC3s in the presence of IL-18. Likewise, mutation of specific NF-kB binding sites within the IL22 promoter led to decreased transcription. Because IL-18 can activate NF-kB, a tissue-specific mechanism was proposed based on the proximity of lymphocytes with ILC3-associated markers to IL-18-producing dendritic cells within the interfollicular regions of the tonsil. In additional studies of the tissue-specific mechanisms that regulate ILC biology, a novel ILC1 population was identified in the liver using data from single cell RNA Sequencing. These cells demonstrated ILC1-associated phenotypic and functional features and were distinct from other previously described human ILC1 populations, as well as from liver-resident NK cells. These liver-type ILC1s (ltILC1s) produced IFN-gamma; and expressed T-BET; yet, they did not degranulate, nor express the NK-associated markers NKp80 and EOMES. These results further elucidate the mechanisms regulating human ILC development and function. By characterizing the normal processes of ILC biology, these studies establish a foundation for understanding how these processes can become dysregulated in the setting of disease. It is further hoped that the discoveries related to this work can be applied in translational research efforts that seek to optimize the role of the immune system in clinical therapies.