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Abstract
In asthmatics, the lung stroma undergoes significant remodeling in response to chronic airway inflammation. Nonetheless, there are limited studies characterizing the pulmonary lymphatic vasculature in asthma, nor whether these vessels expand or regress due to disease. In this thesis, I aim to address the role the pulmonary lymphatics undertake in allergic airway inflammation and provide a reconciliation between such behavior and current literature. Furthermore, how pulmonary lymphatics can exacerbate allergic airway disease, by facilitating induction of bronchus associated lymphoid tissue (BALT) and raising local and systemic IgE levels. Finally, we explore the use of a tolerogenic platform which leverages the natural tolerance inducing mechanisms of the body to drive allergen specific tolerance and ameliorate allergic airway disease.In these studies, we utilize human lung samples and the house dust mite (HDM) model of allergic airway inflammation to address the role of the pulmonary lymphatic endothelium in response to allergic airway disease. Here, we found that in asthmatic lungs, the pulmonary lymphatic density is increased, a phenomenon that was recapitulated in our HDM mouse model of allergic airway disease. These findings are not in conflict with recent literature that suggests that two cytokines central to the allergic response, IL-4 and IL-13, have potent anti-lymphangiogenic capacity, as lymphatic expansion was temporally decoupled from allergen inhalation. Furthermore, we found that by determining the levels of the pro-lymphangiogenic growth factors, VEGF-C and VEGF-D, and gauging these against the anti-lymphangiogenic cytokines, IL-4 and IL-13, we can get a measurement that coincides with the state of the pulmonary lymphatic vasculature and correlates with the numbers of pulmonary lymphatic endothelial cells. Based on lung protein analysis, we posit that when there is a balance between the pro-lymphangiogenic growth factors and the anti-lymphangiogenic cytokines, the pulmonary lymphatic vasculature will be constrained. However, an imbalance between these molecules can drive pulmonary lymphangiogenesis once the allergen-mediated inflammation subsides along with the anti-lymphangiogenic cytokines. Given this temporal decoupling between lymphangiogenesis and allergen-mediated inflammation, we pondered what roles could the pulmonary lymphatic vasculature mediate if it were to expand during allergic airway disease.
Our cohort of human asthmatic lung samples exhibited BALT, and specifically in asthmatic sera, the levels of VEGF-C correlated with the levels of IgE. Such correlation was recapitulated in mice undergoing chronic allergic inflammation, as well as the induction of BALT. Interestingly, we found that when we introduce VEGF-C into mice undergoing chronic HDM-mediated inflammation, their local and systemic IgE levels were raised, as well as the extent of BALT. In allergic mice, VEGFR3 signaling was both necessary and sufficient to mediate induction of BALT, and its expression by lymphatic endothelial cells was required for induction and for the VEGF-C mediated exacerbation. Mice with an expanded lymphatic vasculature where particularly susceptible to developing BALT and IgE upon allergen inhalation, suggesting that the pulmonary lymphatic vasculature could promote a lung microenvironment susceptible to allergic disease. Finally, we found that pulmonary lymphatics could upregulate CXCL13 in response to allergic airway inflammation, and expression of CXCL13 was required for the VEGF-C mediated BALT exacerbation. Following this train of thought, CXCL13 blocking antibodies showed promise as a potential therapeutic in allergic asthma, as their use ameliorated BALT in mice undergoing allergic airway disease. Altogether, our data suggests that in the context of allergic airway disease, the pulmonary lymphatic vasculature undertakes active roles in mediating BALT through CXCL13 dependent mechanisms.
Finally, we assessed the possibility of using a tolerogenic platform to generate allergen specific tolerance and ameliorate allergic airway disease. In collaboration with the Hubbell laboratory, we utilized a synthetically glycosylated form of a model allergen to leverage the tolerance inducing mechanisms of the liver microenvironment to ameliorate the phenotypes associated with allergic airway inflammation. Prophylactic treatment with a glycosylated model allergen reduced the effects of allergen inhalation, including airway and lung eosinophilia, systemic IgE and allergen specific IgG1 levels, as well as mucus hypersecretion. Prophylactic treatment with this tolerogenic platform induced allergen specific regulatory T cells, which inversely correlated with hallmarks of allergic inflammation, and restricted allergic T cell responses. Last, we found that therapeutic intervention with the glycosylated allergen in previously sensitized mice could still ameliorate some features of allergic airway disease, highlighting the potential for this tolerogenic platform in desensitizing former allergic individuals, including those suffering from allergic asthma.