Role of Mast cells in regulating inflammatory pathology of secondary lymphedema

PROJECT SUMMARY/ABSTRACT Secondary lymphedema is a chronic, debilitating condition that commonly occurs due to infections or to injury to the lymphatic system in patients who undergo surgical resection of the tumor, lymph node dissection, radiation, or chemotherapy for breast or other solid tumors. This condition is characterized by tissue swelling, recurrent infections, and diminished quality of life. Currently, there is no drug FDA-approved specifically for treating lymphedema, and existing therapies are primarily palliative, focused on symptom management rather than a cure. Thus, there is an urgent need to better understand the underlying pathology of lymphedema and develop novel treatments. Recent studies have highlighted the crucial role of Th2-differentiated CD4+ T cell adaptive immune responses in lymphedema pathology, which exacerbate lymphatic damage, slow down lymphatic regeneration, and contribute to fibroadipose tissue deposition, as well as the key role of keratinocytes in CD4+ T cell pathology. However, the cellular mechanisms that activate T cell adaptive immune responses following lymphatic injury remains unknown. Interestingly, our preliminary studies suggest that mast cells, an innate immune cell type, may play an important role in T cell activation and lymphedema pathology. Using patient biopsies and mouse models, we observed a significantly increased presence of mast cells in lymphedema tissues, along with elevated expression of mast cell activator proteins (IL-33, stem cell factor [SCF], and thymic stromal lymphopoietin [TSLP]) on keratinocytes following lymphatic injury. We also identified mast cells as a major source of Th2 cytokines IL- 4 / IL-13 in lymphedema biopsies; neutralizing these cytokines reduced mast cell numbers in our pilot clinical trial. Building on our preliminary work and considering the role of mast cells in the pathophysiology of other skin and fibrotic disorders, such as atopic dermatitis, we hypothesize that lymphatic injury induces the expression of mast cell activators and Th2-inducing proteins (IL-33, SCF, and TSLP) by keratinocytes. These activators facilitate the recruitment and activation of mast cells, thereby amplifying CD4+ Th2 inflammatory responses in lymphedema. Our proposed study is novel because the role of mast cells in lymphedema has not been studied. To address this important gap in our knowledge of lymphedema pathology, we will test our hypothesis by investigating how mast cells are recruited and activated in lymphedema following lymphatic injury using patient tissues and animal models. We will perform spatiotemporal and single-cell RNA sequencing analyses, and we will explore the roles of lymph fluid and keratinocytes on mast cell activation. We will also use knockout and transgenic mice models to analyze how mast cell-derived Th2 cytokines (IL-4 / 13) contribute to Th2 differentiation of CD4+ T cells and overall lymphedema pathology.