CCBE1 Is Essential for Mammalian Lymphatic Vascular Development and Enhances the Lymphangiogenic Effect of Vascular Endothelial Growth Factor-C In Vivo

Abstract

Rationale: Collagen-and calcium-binding EGF domains 1 (CCBE1) has been associated with Hennekam syndrome, in which patients have lymphedema, lymphangiectasias, and other cardiovascular anomalies. Insight into the molecular role of CCBE1 is completely lacking, and mouse models for the disease do not exist.Objective: CCBE1 deficient mice were generated to understand the function of CCBE1 in cardiovascular development, and CCBE1 recombinant protein was used in both in vivo and in vitro settings to gain insight into the molecular function of CCBE1. Methods and Results: Phenotypic analysis of murine Ccbe1 mutant embryos showed a complete lack of definitive lymphatic structures, even though Prox1؉ lymphatic endothelial cells get specified within the cardinal vein. Mutant mice die prenatally. Proximity ligation assays indicate that vascular endothelial growth factor receptor 3 activation appears unaltered in mutants. Human CCBE1 protein binds to components of the extracellular matrix in vitro, and CCBE1 protein strongly enhances vascular endothelial growth factor-Cmediated lymphangiogenesis in a corneal micropocket assay. Conclusions:

Interaction of tumor cells and lymphatic vessels in cancer progression

“…4 This budding and migration of lymphatic endothelial progenitor cells is mediated by VEGF-C signals and is critically modulated by collagen-and calcium-binding EGF domain 1 protein. 6,7 COUP-TFII and Prox-1 are essential for the maintenance of the identity of the lymphatic endothelial cells following their differentiation. 8 The primary lymphatic networks then enlarge by sprouting lymphangiogenesis induced by the VEGFR-3 ligand VEGF-C. By upregulation of VEGFR-3 expression independently of Prox-1, T-box transcription factor 1 supports the growth and maintenance of the gastrointestinal lymphatic vessel network.…”

Ccbe1 expression marks the cardiac and lymphatic progenitor lineages during early stages of mouse development

“…Recently, mouse (m)Ccbe1 has been shown to be essential for budding and/or migration of lymphatic endothelial cells (LECs) from the anterior cardinal veins to form the lymph sacs (E10.5 to E13.5) and give rise to the lymphatic vasculature (Bos et al, 2011). Indeed, homozygous mCcbe1 knockout mice showed a reduction of the number of Prox-1 + and Lyve-1 + LECs at the level of the cardinal vein and lymph sacs which lead to defective lymphatic vasculature, severe edema and prenatal death (Bos et al, 2011).…”

“…Indeed, homozygous mCcbe1 knockout mice showed a reduction of the number of Prox-1 + and Lyve-1 + LECs at the level of the cardinal vein and lymph sacs which lead to defective lymphatic vasculature, severe edema and prenatal death (Bos et al, 2011). In zebrafish, zccbe1 transcripts were also found to be located along the migratory routes of lymphangioblast that bud from the cardinal veins to seed the horizontal myoseptum region and subsequently generate the primordial lymphatic vessels.…”

A Tale of Two Models: Mouse and Zebrafish as Complementary Models for Lymphatic Studies

“…Similar to many secreted ligands, VEGF-C is also produced in a preproform, and secreted as a pro-VEGF-C, which needs to be processed to be fully active (Lee= et al, 2005;McColl= et al, 2003). A disintegrin and metalloprotease with thrombospondin motifs-3 (ADAMTS3) has been recently shown to mediate proteolytic cleavage of pro-VEGF-C (Jeltsch=et al, 2014), and the process is facilitated by Collagen and calcium-binding EGF domain-containing protein 1 (CCBE1) (Bos=et al, 2011;Jeltsch= et al, 2014). In humans, mutations in VEGF-C, VEGFR3, and CCBE1 have been shown to cause hereditary lymphedemas (Brouillard= et al, 2014): For instance, mutations in VEGFR3 cause Milroy's disease (Irrthum et al, 2000;Karkkainen et al, 2000), and mutations in CCBE1 lead to Hennekam lymphan- .…”