VEGF binding sites were identified on the cell surface of vascular endothelial cells in
vitro and in vivo. Subsequently, it became apparent that receptors for VEGF also
occur on bone marrow-derived cells [50]. VEGF binds two highly related receptor
tyrosine kinases (RTK), VEGFR-1 and VEGFR-2. Both VEGFR-1 and VEGFR-2
have seven immunoglobulin-like (Ig-like) domains in the extracellualar domain, a
single transmembrane region, and a consensus tyrosine kinase sequence [51, 52].
VEGFR-3 (fms-like-tyrosine kinase (Flt)-4) is a member of the same family of RTKs,
but is not a receptor for VEGF, binding instead to VEGF-C and VEGF-D [53]. In
addition to these RTKs, VEGF interacts with a family of coreceptors, the neuropilins
(NRP).
VEGFRs share similar regulatory mechanisms with well-characterized receptor
tyrosine kinases, by which include receptor dimerization and activation of the tyrosine
kinase. Moreover, VEGFRs perform cellular processes that are common to many
growth factor receptors such as cell survival and proliferation. The summary of
VEGFRs shows in Figure 1, VEGF-A binds to VEGFR-1 and VEGFR-2, which are
expressed in the cell surface of most blood ECs. In contrast, PLGF and VEGFB
interact only with VEGFR-1. VEGFC and VEGFD bind VEGFR-2 and VEGFR-3,
which is largely restricted to lymphatic EC. There is much evidence that VEGFR-2 is
the major mediator of EC mitogenesis and survival, as well as angiogenesis and
micro-vascular permeability. VEGFR-1 has an established signaling role in mediating
monocyte chemotaxis. Also, in hematopoietic stem cells (HSC) or leukemic cells,
both VEGFR-1 and VEGFR-2 may mediate a chemotactic and a survival signal [43].
1.3.1 VEGFR-1
VEGFR-1 or Flt-1 (fms-like-tyrosine kinase-1) was the first discovery of VEGF
receptor [54]. The VEGFR-1 tyrosine kinase exhibits all the conserved motifs that are
required for kinase activity. The crystal structure of part of the VEGFR-1 extracellular
domain shows that the Ig domain-2 is the major ligand binding site on the receptor in
physiology and pathology. VEGFR-1 expression is unregulated by hypoxia by a
HIF-1 dependent mechanism [55].VEGFR-1 binds not only VEGF-A but also PLGF
and VEGF-B [40, 56]. The crucial role of VEGFR-1, as mentioned above, was
determined by Fong, 1995, which revealed that when disruption of VEGFR-1 gene in
mice resulted in embryonic lethality.
In addition to the full length of VEGFR-1, there is an alternatively spliced
soluble form of VEGFR-1 (sFlt-1), which has been shown to be an inhibitor of VEGF
activity [57]. Hence, not only the full length membrane bound form of VEGFR-1 but
sFlt-1 as well could perform a decoy function, which sequesters VEGF and prevent its
interaction with VEGFR-2 [58].
In some cases, VEGFR-1 is expressed by tumor cells and probably mediates a
chemotatic signal, thus potentially extending the role of this receptor in cancer growth.
For instance, Wu and collaborates indicated that VEGF-A autocrine growth activity is
acquired by certain human breast tumor cell lines defined by expression of VEGFR-1
[59].
Anchoring of the extracellular domain of VEGFR-1 to the cell membrane is
important, as 50% of the mice that lack both of the tyrosine kinase domains and the
transmembrane domain died at embryonic stage, owing to vascular malformation [60].
This study indicates that endothelial cells develop but fails to organize in vascular
channels. Excessive proliferation of angioblasts has been reported to be responsible
for such disorganization and leathality, indicating that, at least during early
development, VEGFR-1 is a negative regulator of VEGF action [61]. By contrarily,
many evidence also indicated that VEGFR-1 is a positive regulator of monocyte and
macrophage migration [62, 63].
1.3.2 VEGFR-2
At the same time VEGFR-1 was discovered, Shalaby et al. found VEGFR-2, another
receptor for VEGF, and proposed that VEGFR-2 had similar characteristics and
continuously shows today. Biochemical analyses showed that the second and third
Ig-like domains in VEGFR-2, also known as kinase domain region (KDR), is
important for the determination of ligand binding specificity for VEGF [64].
VEGFR-2 undergoes RTK dimerization and strong ligand-dependent tyrosine
phosphorylation in intact cells.
There are many studies indicating that VEGFR-2 is the major mediator of the
mitogenesis, survival, and permeability enhancing effects of VEGF-A in endothelial
cells [32]. VEGF binds to VEGFR-2 and stimulates activation of Ras in HUVECs.
Besides, Ras activation has been coupled to an angiogenic phenotype of endothelial
cells [65]. The early finding that the binding of VEGF to VEGFR-2 is enhanced by
heparin has been confirmed by recent studies, which shows that heparin amplifies
signalling by VEGF. Byzova et al. have reported that VEGFR-2 activation by VEGF
results in PI3 kinase/Akt-dependent activation of several integrins, which indicates
that VEGF enhanced cell adhesion, migration, soluble ligand binding [66]. Moreover,
VEGFR-2 activation has been shown to be required for the antiapoptotic effects of
VEGF for human umbilical vein endothelial cells (HUVECs) [29].
1.3.3 VEGFR-3 and Neuropilin
VEGFR-1 and VEGFR-2 are expressed on the surface of blood endothelial cells. By
contrast, VEGFR-3 (Flt-4) is largely restricted to lymphatic endothelial cells. In recent
studies, VEGFR-3 is important for lymphatic endothelial cell development and
function [53].
Neuropilin-1 (NRP1), as its name suggests, is a molecular that is implicated in
neuronal guidelince and had been previously shown to bind the collapsin/semaphorin
family. NRP1 is a cell surface glycoprotein that lacks intrinsic catalytic activity, a
receptor for the heparin-binding isoforms of VEGF, and seems to present VEGF165 to
VEGFR-2 in a manner that potentiates VEGFR-2 signaling [67]. This result shows
that neuropilin stabilizes the VEGF/VEGFR-2 signaling complex when expressed on
adjacent cells.