Morphogenesis of Lymphatic Vascular Networks: Insights from Connexin and Foxc2 Knockout Mice

Abstract

To maintain human health, the lymphatic system requires a structurally and functionally sound network of lymph vessels to absorb lipid-based nutrients, preserve extracellular fluid homeostasis, and mediate immune responses. Aside from lymphedema, investigations in the past few decades have found that impairment of the lymphatic vasculature is also involved in processes such as inflammation, tumor metastasis, fat metabolism, and obesity. However, despite a long history of study and rekindled vigor in the field of lymphatic vascular research, our knowledge of lymph vessel development and physiology is still quite limited. Recently, mutations in a protein family known as connexins (Cxs) were identified as the cause of lymphatic dysfunction in some cases of inherited lymphedema. This dissertation explores the role of primarily two specific connexins, Cx37 and Cx43, and the transcription factor Foxc2 in the morphogenesis and function of the lymphatic vasculature in mice. To accomplish this, phenotypic characterization of mice with genetic deficiencies (knockout mice) in Cx37, Cx43, and/or Foxc2 was performed principally via necropsy, histological techniques (immuno-fluorescence microscopy and H&E staining), and Evans blue dye (EBD) injections. Developmental abnormalities were found in lymphatic vascular growth, patterning, and remodeling in mice lacking Cx37, Cx43, Foxc2 or a combined deficiency of these proteins. Reductions or complete loss of lymphatic valves were a common finding in mice lacking one or more of these proteins. These valve deficits underlay lymphatic insufficiencies that resulted in lymphedema and chylothorax in some genotypes. Foxc2 was found to be a regulator of Cx37 expression. Moreover, Foxc2 was also dependent on Cx37 function for proper morphogenesis of lymph vessels. These findings pertaining to the expression of connexins in the lymphatic vasculature, their role in lymphatic valvulogenesis, and the interdependence of Cx37 and Foxc2 during lymph-vascular development represent my original contributions to human knowledge.