Inflammation-Mediated Lymphatic Coagulation: A New Role for Neutrophil Extracellular Traps

This work explores how endothelial cells respond to and regulate immune responses, specificallyfocusing on lymphatic endothelial cells and how their interactions with neutrophils can promote intralymphatic coagulation. In this work, I introduce lymphatic coagulation in the lungs and lung-draining lymph nodes as a clinical manifestation of COVID-19 and identify a correlation between lymphatic coagulation and the presence of neutrophils extracellular traps (NETs) within lymphatic vessels. Both intralymphatic NETosis and lymphatic coagulation also correlate with dysregulated germinal centers in these patients, and in a separate cohort of hospitalized COVID-19 patients, serum NET levels inversely correlated with antiviral antibody titers, suggesting that lymphatic clotting may impair the formation or maintenance of germinal centers necessary for robust antiviral antibody responses. Additionally, in mice degrading NETs with DNase 1 prevented TNFα-induced coagulation in lymphatic vessels, indicating that the correlation observed in COVID-19 decedents was indicative of a mechanistic role. After establishing that NETs do induce clotting in lymphatic vessels, I next investigated the role that lymphatic endothelial cells themselves may play in regulating both lymphatic coagulation and NETosis and compared this to that of blood endothelial cells. Using platelet-free plasma as a model of lymph, I observed that lymphatic endothelial cells exhibited a higher clotting threshold compared to blood endothelial cells, which is at least partially regulated by higher secretion of tissue plasminogen activator, and that this threshold is reduced by TNFα pre-treatment. TNFα also stimulated lymphatic endothelial cells to promote neutrophil recruitment and NETosis, which are regulated by CXCL8 and CCR7 in this context. These findings provide mechanistic insights into lymphatic clotting, offering potential avenues for therapeutic interventions in associated conditions. Lastly, this work investigates the role of a mechanosensitive LIM-domain protein, FHL2, in regulating endothelial cell inflammatory response.