Lymphangiogenesis guidance by paracrine and pericellular factors

Abstract

Lymphatic vessels are important for tissue fluid homeostasis, lipid absorption, and immune cell trafficking and are involved in the pathogenesis of several human diseases. The mechanisms by which the lymphatic vasculature network is formed, remodeled, and adapted to physiological and pathological challenges are controlled by an intricate balance of growth factor and biomechanical cues. These transduce signals for the readjustment of gene expression and lymphatic endothelial migration, proliferation, and differentiation. In this review, we describe several of these cues and how they are integrated for the generation of functional lymphatic vessel networks.

Keywords: VEGF-C; VEGFR3; interstitial fluid pressure; lymphangiogenesis; lymphatic vessel basement membrane; lymphatic vessel sprouting; lymphedema.

Figure 1.

Pericellular cues that guide lymphatic vessel growth. (A,A′) Arterial endothelial cells and SMCs secrete lymphangiogenic guidance cues that contribute to the alignment of large lymphatic collectors with arteries. VEGF-C binds to pericellular matrix and LEC surface proteins, such as VEGFR3, neuropilin 2 (NRP2), and syndecan-4, and is processed upon its interaction with extracellular matrix (ECM) adapter, collagen- and calcium-binding EGF domain-containing protein 1 (CCBE1), and the ADAMTS3 protease as shown in A′. In zebrafish and mice, CXCL12 produced by blood vascular endothelial cells guides lymphatic growth via binding to its receptor, CXCR4, on LECs. Adrenomedullin (AM) binds to the RAMP2 and CALCRL receptors in mice. The chemokine sink CXCR7 regulates these interactions by sequestering both CXCL12 and adrenomedullin. (B) Upon growth factor-induced activation, both VEGFR3 and VEGFR2 can stimulate LEC proliferation, and VEGFR3 interaction with β1 integrins, such as α5β1, enhances the lymphangiogenic signals. (C) The sprouting and branching of lymphatic vessels is dependent on VEGF-C signaling via the VEGFR3–NRP2 receptor complex. Integrin α5β1 ligands fibronectin and collagen in the ECM increase VEGFR3 phosphorylation in the absence of a VEGFR3 ligand; they also potentiate VEGF-C-induced VEGFR3 activation and LEC migration. Macrophages provide a major source of VEGF-C in lymphangiogenesis associated with inflammation. The growth-promoting factors are counteracted by inhibitory signals, such as TGF-β and INF-γ, which act directly on LECs or affect VEGF-C production by, e.g., macrophages (see the overview figure). (D) The deflection of lymphatic vessel sprouts away from arteries has been suggested to be driven by arterial expression of semaphorin 3G (SEMA3G), which induces LEC repulsion via a plexin 1 (PLXN1)–NRP2–VEGFR3 receptor complex.

Figure 2.

Examples of ongoing lymphangiogenesis in mouse embryos and postnatal mice. (A) Embryonic skin dermis at embryonic day 14 (E14) stained for CD31 (blue), PROX1 (red), and NRP2 (green). (B) LEC clusters in the process of assembling to form mesenteric lymphatic vessels at E14, here stained for PROX1 (red) and NRP2 (green). (C) LYVE1 (green) staining of developing lymphatic vessels in the ventral part of the ear at postnatal day 16 (P16). The inset shows one of the growing lymphatic vessel tips, with the LEC nuclei indicated using PROX1 (red). (D,E) CD31 (blue), PROX1 (red), and VEGFR3 (green) whole-mount staining of the trachea (D) and tail dermis (E) at P5. (F) CD31 (blue) and LYVE1 (green) staining of the pleural side of a P5 diaphragm. (G,H) CD31 (blue), α-smooth muscle actin (αSMA; red), and LYVE1 (green) staining of (lacteal) lymphatic vessels in intestinal villi (G) and the intestinal wall (H) in adult mice. (I) CD31-stained (blue) and PROX1-stained (red) mesenteric lymphatic vessels at P7.