Mice with Lymphatic Dysfunction Develop Pathogenic Lung Tertiary Lymphoid Organs that Model an Autoimmune Emphysema Phenotype of COPD

We have previously shown that mice with loss of C-type lectin-like type II (CLEC2), which have lymphatic dysfunction due to the role of CLEC2 in platelets for maintaining separation between the venous and lymphatic system, develop lung tertiary lymphoid organ (TLO) formation and lung injury that resembles an emphysema phenotype of chronic obstructive pulmonary disease (COPD). We now sought to investigate whether and how TLOs in these mice may play a pathogenic role in lung injury that is relevant to human disease. We found that inhibiting TLO formation using an anti-CD20 antibody in CLEC2-deficient mice partially blocked the development of emphysema. TLOs in CLEC2-deficient mice were rich in plasma cells and were a source of a broad array of autoantibodies. Chronic cigarette smoke exposure increased the size and number of lung TLOs in CLEC2-deficient mice, and was associated with increased markers of antigen presentation and maturation, leading to increased autoantibody deposition. Using lung tissue from COPD patients, we found an increase in lymphatic markers in patients with an emphysema phenotype and autoreactive TLOs compared to COPD patients without emphysema that lack prominent TLOs. Taken together, these results demonstrate that emphysema in mice with lymphatic dysfunction can be partially rescued by blocking TLO formation, and that these TLOs are source of autoantibodies that are exacerbated by cigarette smoke. Our work suggests that lymphatic dysfunction in mice may recapitulate some aspects an autoimmune emphysema phenotype that is seen in a subset of patients with COPD. Overall design: Mice carrying the Prox1-EGFP reporter (in which all LECs are labeled with GFP) were injected with AlexaFluor647-conjugated isolectin (Invitrogen) for intravital labeling of endothelial cells just prior to sacrifice. The lungs were then digested using dispase/collagenase in HBSS to generate a single cell suspension for FACS, with additional staining using antibodies for CD31 (Biolegend 102418), lineage cocktail (Bioloegend 133311), and Epcam (Biolegend 118225). LECs were sorted using positive selection for isolectin, CD31, GFP, and negative gating for Epcam and Lineage markers.