Source and homeostatic functions of anti-adduct IgM in humans

点击翻译按钮获取中文摘要

Project Summary Serum IgM have crucial “housekeeping” functions such as the clearance of apoptotic cells and cellular debris, preventing their accumulation in tissue. However, despite their important role, IgM have been largely understudied in comparison to other immunoglobulin isotypes, especially IgG. A central characteristic of serum IgM linked to their biological function is their reactivity to simple chemical moieties exposed on cell membranes and macromolecules. For instance, IgM bind to phosphorylcholine on apoptotic cells to promote their elimination. These moieties form adducts when attached to proteins or other macromolecules. Remarkably, only a few chemical adducts recognized by serum IgM have been identified thus far. Whether these are examples of a much larger group of chemical radicals targeted by serum IgM is unknown. To address this knowledge gap, we developed a high-dimensional platform to assess monoclonal IgM and serum IgM reactivity to 87 ubiquitous adducts. Results showed that: 1) monoclonal “polyreactive” IgM cloned from memory blood B cells and binding to apoptotic cells, hence displaying a typical “natural antibody” profile, react in fact to specific adducts; 2) the anti-adduct IgM repertoire is highly restricted in newborns and only includes reactivity to a limited number of adducts; 3) IgM reactivity diversifies abruptly around 6 months of age, marking a transition from a restricted neonatal repertoire to a broad adult repertoire and 4) this transition appears to coincide with exposure to environmental antigens. Overall, these studies uncovered a much more complex anti-adduct IgM immunity than was initially anticipated. Yet, several important questions remain unanswered: What are the functional niches of anti-adduct B cells/PC? Do anti-adduct B cells constitute a distinct subset with specific phenotypic characteristics? What is the role of anti-adduct IgM in efferocytosis and regulation of inflammation? Are these functions dependent on the IgM specificity to individual adducts? Our proposed studies will address these salient questions through the characterization of anti-adduct B cell and PC at the single-cell level. Specific aim-1. To characterize anti-adduct memory B cells and plasma cells and map their niches We will first detect anti-adduct memory IgM+ B cells and IgM-secreting cells in the bone marrow, spleen, thymus and gut-associated lymphoid tissue in order to identify their main niches. We will then characterize the phenotype of anti-adduct memory IgM+ B cells using flow cytometry and single-cell-RNA-seq combined with BCR-seq to profile their transcriptome and evaluate their clonal composition in vivo. Specific aim-2. To determine the role of anti-adduct IgM in regulation of efferocytosis and inflammation In specific aim 2 we will examine the capacity of IgM mab specific to individual adducts to opsonize apoptotic cells and enhance their clearance by different types of phagocytes. We will also investigate the role of complement in this function. In a second part, we will examine whether apoptotic cells opsonized by IgM reactive to different adducts trigger distinct cytokine secretion upon their uptake by phagocytes.