Cooperative Effects of Vascular Angiogenesis and Lymphangiogenesis

In this study, we modeled lymphangiogenesis and vascular angiogenesis in a microdevice using a tissue-engineering approach. Lymphatic vessels (LV) and blood vessels (BV) were fabricated by sacrificial molding with seeding human lymphatic endothelial cells and human umbilical vein endothelial cells into molded microchannels (600-m diameter). During subsequent perfusion culture, lymphangiogenesis and vascular angiogenesis were induced by addition of phorbol 12-myristate 13-acetate (PMA) and VEGF-C or VEGF-A and characterized by podoplanin and Prox-1 expression. Our results showed that while blood capillaries consistently exhibited zipper-like junctions,lymphatic capillaries formed button-like junctionswhen treated with dexamethasone. To test the potential for screening anti-angiogenic (vascular and lymphatic) factors, antagonists of VEGF were introduced. We found that an inhibitor of VEGF-R3 did not completely suppress lymphatic angiogenesis with BVs present, although lymphatic angiogenesis was selectively prevented by addition of a VEGF-R3 inhibitor without BVs. To probe the mechanism of action, we focus on matrix metalloproteinase (MMP) secretion by vascular endothelial cells and lymphatic endothelial cells under monoculture or co-culture conditions. We found that vascular angiogenesis facilitated lymphangiogenesis via remodeling of the local microenvironment by the increased secretion of MMP, mainly by vascular endothelial cells. Applications of this model include a drug-screening assay for corneal disease and models for tumorigenesis including lymphatic angiogenesis and vascular angiogenesis.Lay SummaryLymphangiogenesis involves the formation of new lymphatic vessels from pre-existing ones during embryonic development, wound healing and in various pathological conditions. In this work, we simulate such physiological and pathological conditions in which there are important bi-directional effects between the lymphatic and vascular cells using an in vitro perfusion culture. Mechanical and chemical stimuli in vitro lead to maturation of the lymphatic structures as evidenced by lymphatic-specific cell-cell junctions and cell orientation. The combined vascular and lymphatic angiogenesis model could be useful to investigate the pathophysiology of tumorigenesis and corneal inflammation involving interactions between vascular angiogenesis and lymphatic angiogenesis.