21
high VEGF environment of tumors is one reason why pericyte coverage is decreased
compared to normal vascular beds [48, 61]. Treatment with VEGF antagonists has
led to the finding that pericyte coverage increases, which results in improved microcirculatory function (i.e., vascular normalization) [61]. This approach represented a
new strategy to improve drug delivery into the tumor microenvironment, since poor
perfusion exists due to the abnormal microcirculatory network that is present.
1.3.13 Molecular Mechanisms Underlying Why Pericytes Are
Able to Stabilize EC-Lined Tube Networks
A major question that has not been sufficiently addressed is why pericyte coverage
stabilizes vessels and what their functions are when they arrive at the EC abluminal
surface. To address this question, we established novel EC-pericyte coculture models in 3D collagen matrices (Fig. 1.5). We developed systems using either bovine
retinal pericytes or human brain pericytes. In each case, the pericyte populations
express the pericyte markers NG2 proteoglycan, 3G5 ganglioside, smooth muscle
actin, and desmin. Perhaps the most important function of pericytes is to recruit to
microvascular capillary beds. Using our model of EC vasculogenic tube assembly,
we developed a system whereby we randomly mix together ECs and pericytes at a
5:1 or 5:1.25 ratio (i.e., 20–25% pericytes compared to 100% of ECs). Remarkably,
the ECs form tube networks, and then, pericytes are recruited to these tubes [94]
(Fig. 1.5). This ratio of ECs to pericytes is particularly optimal, and the reasons for
this are currently not clear. It may be that too many pericytes (through their production of TIMP-3) [87] interfere with morphogenesis, by inhibiting MT1-MMPdependent signaling, or that they are physically in the way and counteract the ability
of ECs to find neighbors to properly form multicellular tubes. It is clear that too
many pericytes can disrupt EC-pericyte tube co-assembly.
We further made the observation that EC tubes, from EC-only cultures, eventually became much wider than EC tubes from EC-pericyte cocultures. We examined
this issue over time and observed that vascular diameters reached a range of
20–25 μm in EC-pericyte cocultures, which are vessel diameters observed in vivo
during vasculogenesis, while EC-only cultures’ diameters can reach 80–100 μm
over a 5-day period [94]. Thus, pericytes have a marked ability to negatively regulate vascular tube diameters, which may have to do with the induction of TIMP-2
and TIMP-3. As discussed earlier, they are induced in EC-pericyte cocultures [87]
and can inhibit and restrict EC lumen diameters. A number of studies indicate that
vascular diameters are greater when pericyte recruitment is reduced or when ECM
components such as fibronectin are knocked out of ECs during vasculogenesis
in vivo [6, 42].
The mechanisms whereby pericytes are recruited to EC-lined tubes are still
being investigated although past data supports the concept that PDGF-BB plays a
role [5, 16, 54]. In past and ongoing studies from our laboratory, we have shown
that pericyte recruitment is dependent on signals derived from the combined action
1 Molecular Control of Capillary Tube Morphogenesis and Maturation Through…
high VEGF environment of tumors is one reason why pericyte coverage is decreased
compared to normal vascular beds [48, 61]. Treatment with VEGF antagonists has
led to the finding that pericyte coverage increases, which results in improved microcirculatory function (i.e., vascular normalization) [61]. This approach represented a
new strategy to improve drug delivery into the tumor microenvironment, since poor
perfusion exists due to the abnormal microcirculatory network that is present.
1.3.13 Molecular Mechanisms Underlying Why Pericytes Are
Able to Stabilize EC-Lined Tube Networks
A major question that has not been sufficiently addressed is why pericyte coverage
stabilizes vessels and what their functions are when they arrive at the EC abluminal
surface. To address this question, we established novel EC-pericyte coculture models in 3D collagen matrices (Fig. 1.5). We developed systems using either bovine
retinal pericytes or human brain pericytes. In each case, the pericyte populations
express the pericyte markers NG2 proteoglycan, 3G5 ganglioside, smooth muscle
actin, and desmin. Perhaps the most important function of pericytes is to recruit to
microvascular capillary beds. Using our model of EC vasculogenic tube assembly,
we developed a system whereby we randomly mix together ECs and pericytes at a
5:1 or 5:1.25 ratio (i.e., 20–25% pericytes compared to 100% of ECs). Remarkably,
the ECs form tube networks, and then, pericytes are recruited to these tubes [94]
(Fig. 1.5). This ratio of ECs to pericytes is particularly optimal, and the reasons for
this are currently not clear. It may be that too many pericytes (through their production of TIMP-3) [87] interfere with morphogenesis, by inhibiting MT1-MMPdependent signaling, or that they are physically in the way and counteract the ability
of ECs to find neighbors to properly form multicellular tubes. It is clear that too
many pericytes can disrupt EC-pericyte tube co-assembly.
We further made the observation that EC tubes, from EC-only cultures, eventually became much wider than EC tubes from EC-pericyte cocultures. We examined
this issue over time and observed that vascular diameters reached a range of
20–25 μm in EC-pericyte cocultures, which are vessel diameters observed in vivo
during vasculogenesis, while EC-only cultures’ diameters can reach 80–100 μm
over a 5-day period [94]. Thus, pericytes have a marked ability to negatively regulate vascular tube diameters, which may have to do with the induction of TIMP-2
and TIMP-3. As discussed earlier, they are induced in EC-pericyte cocultures [87]
and can inhibit and restrict EC lumen diameters. A number of studies indicate that
vascular diameters are greater when pericyte recruitment is reduced or when ECM
components such as fibronectin are knocked out of ECs during vasculogenesis
in vivo [6, 42].
The mechanisms whereby pericytes are recruited to EC-lined tubes are still
being investigated although past data supports the concept that PDGF-BB plays a
role [5, 16, 54]. In past and ongoing studies from our laboratory, we have shown
that pericyte recruitment is dependent on signals derived from the combined action
1 Molecular Control of Capillary Tube Morphogenesis and Maturation Through…