Hypoxia and Extracellular Matrix Proteins Influence Angiogenesis and Lymphangiogenesis in Mouse Embryoid Bodies

Abstract

Regulatory mechanisms for angiogenesis are relatively well established compared to lymphangiogenesis. Few studies have shown that a combination of vascular endothelial growth factor VEGF-A/C with hypoxia or collagen matrix promotes lymphatic structures along with blood vessel development in mouse embryoid bodies (EB). In this study we tested the hypothesis that while hypoxia combined with prolonged VEGF-A/C treatment would induce early lymphangiogenesis in addition to angiogenesis in mouse EBs, under similar conditions specific extracellular matrix (ECM) proteins would promote lymphatic vessel-like structures over angiogenesis. EBs were subjected to four conditions and were maintained under normoxia and hypoxia (21% and 2.6% O2, respectively) with or without VEGF-A/C. Microarray analyses of normoxic and hypoxic EBs, and immunofluorescence data showed very low expression of early lymphatic endothelial cell (LEC) markers, lymphatic vessel endothelial hyaluronan receptor 1 (LYVE1), and prospero-related homeobox 1 (Prox1) at early time points. Double immunofluorescence using MECA-32 and Prox1/LYVE1 demonstrated that combined hypoxia and VEGF-A/C treatment promoted formation of blood vessel-like structures, whereas only Prox1+/LYVE1+ LECs were detected in EBs at E22.5. Furthermore, EBs were grown on laminin or collagen-I coated plates and were subjected to the four treatments as described above. Results revealed that LECs in EBs at E36.5 attached better to collagen-I, resulting in an organized network of lymphatic vessel-like structures as compared to EBs grown on laminin. However, blood vessel-like structures were less favored under these same conditions. Collectively, our data demonstrate that hypoxia combined with growth factors promotes angiogenesis, whereas combination of these conditions with specific ECM proteins favors lymphangiogenesis processes in mouse EBs.

Insulin-Regulated Protein Palmitoylation Impacts Endothelial Cell Function

“…Membrane-associated proteins that utilize palmitoylation to increase hydrophobicity for impacting cell motility or membrane trafficking could be suitable targets for novel therapeutics in metabolic syndrome and type 2 diabetes. We identified several appropriate candidates including FBLN3 (fibulin-3), which associates with the extracellular matrix and promotes cell migration 21 , 22 , LYVE1 (Lymphatic vessel endothelial hyaluronic acid receptor 1 23 , BST2 (Bone marrow stromal antigen 2) 24 , and PAFAH1b3 25, 26 (pursued in detail below).…”

VEGF‐C Induces Lymphangiogenesis and Angiogenesis in the Rat Mesentery Culture Model

“…. Induction of either angiogenesis or lymphangiogenesis from embryoid bodies depending on control of the local chemical environment also highlights the common lineage between lymphatic and blood endothelial cells. These examples of lymphatic/blood endothelial cell plasticity and our observation of lymphatic/blood vessel connections in cultured rat mesenteric tissues motivate future studies using the rat mesentery culture model to determine whether cell plasticity in an intact microvascular network scenario leads to conversion of vessel types.…”

Studies of postpartum mammary gland involution reveal novel pro-metastatic mechanisms

“…In breast cancer, increased LVD, lymph node involvement, and lymph vessel invasion are predictive of higher risk of metastasis; further, increased LVD and lymph node metastasis are commonly observed in PPBC [16] . While the role of collagen in lymphangiogenesis has not been extensively characterized, one study has shown that collagen I increases lymphangiogenesis and angiogenesis in mouse embryoid bodies under hypoxic conditions [160] . Therefore, it is plausible that other fibrillar collagens may contribute to lymphangiogenesis during mammary tumorigenesis.…”