Immunomodulation by splenic megakaryocytes and platelets in sepsis

Project Summary Sepsis is a dysregulated host response to infection that culminates in organ failure leading to millions of deaths worldwide each year with an increasing incidence as the population ages. There is a fundamental lack of understanding of the complex host immune response in sepsis that has limited the development of targeted therapeutics for which there are none beyond antibiotics and supportive care measures. At its core, there is substantial immunopathology in sepsis with contributions from an overly exuberant immune response and ineffective pathogen clearance. We and others have studied the critical role of platelets in immune responses during acute infections, including the role of the lung in extramedullary platelet biogenesis. In this application, we will explore the role of the spleen, a central immune organ, in extramedullary megakaryopoiesis and platelet production in sepsis. Based on preliminary data, we hypothesize that the spleen co-opts a significant role in platelet biogenesis during sepsis and that the platelets produced from the spleen are immunomodulatory and important in host defense. In Aim 1, we will utilize a mouse model of peritonitis and polymicrobial sepsis resulting in thrombocytopenia to understand the mechanics of ‘stressed’ platelet biogenesis in this setting. We will study the role of adrenergic-dependent hematopoietic progenitor mobilization from the bone marrow during sepsis and the niche-promoting factors that regulate this process. In Aim 2, we will interrogate the engraftment of circulating hematopoietic progenitors in the spleen, their maturation into megakaryocytes, and the mediators (SCF, CXCL12, IL-3) regulating this process. Using state-of-the-art techniques such as intravital imaging and lineage tracing enabled by splenic transplantation, we will test the hypothesis that the spleen significantly contributes to platelet biogenesis during sepsis. In Aim 3, we will use novel methods of single-cell RNA sequencing of platelets to test for platelet heterogeneity during homeostasis and sepsis in mice and humans. Within this aim, we will test a novel cellular therapy for sepsis by transfusing immune-skewed platelets into septic mice and testing for therapeutic benefit. In summary, these studies will produce paradigm-shifting knowledge on the role of the spleen in extramedullary megakaryopoiesis and platelet production and the importance of platelet driven immunity, which will be foundational in the design of new therapeutic approaches to treat sepsis.