ECM Remodeling in Angiogenesis - Mechanical and Chemical Signaling in Angiogenesis - page 190

Fig. 3 Three-dimensional co-cultures of ECs and stromal cells generate stable, pericyte-invested
capillary networks in vitro. In these experiments, bone marrow-dereived mesenchymal stem cells
(MSCs) MSCs were distributed throughout a fibrin-based 3-D ECM in the presence of
microcarrier beads coated with ECs. a After 7 days, cultures were fixed and IF stained at day 7
for F-actin (green) and collagen IV (red) to visualize basement membrane deposition (white
arrows). Scale = 50 lm. b MSCs expressing GFP were interspersed with mCherry-transduced
ECs. Physical association of the ECs and MSCs were observed (white arrows). Scale = 25 lm.
c Cultures containing mCherry-transduced ECs and MSCs were fixed and IF stained for pericyte
markers aSMA (aqua) and NG2 (white). DAPI-stained nuclei are visible in the blue channel.
Scale = 50 lm. (Figure reproduced from [ 74 ] with permission from Elsevier.)
Once nascent capillaries have formed, a basement membrane, a hallmark of a
more mature vessel network, is deposited. Pericytes play a key role in this process.
These cells are recruited from the host stroma in part via the secretion of PDGF-b by
ECs. PDGF receptor-b signaling then initiates a cascade of events that control
pericyte-EC binding, migration to the site, and proliferation. Interestingly, all of
these processes are in some manner controlled by MT1-MMP [ 71 - 73 ]. Co-cultures
of ECs and stromal cells of various origins in 3-D gels yield stable, pericyte-invested
networks of capillaries characterized by the presence of basement membrane
subjacent to the ECs along with the periendothelial location of the stromal cells
(Fig. 3 )[ 74 ]. In the absence of stromal cells, ECs express MT1-MMP, with very
little basement membrane production, even at later time points. If stromal cells are
included with ECs in culture, MT1-MMP expression is undetectable in the capillary
stalks. The ECs in the stalk instead produce basement membrane proteins, and the
expression of MT1-MMP is restricted to the tip cells. Similar expression profiles
have also been see in vivo [ 45 ].
Further examination of the newly deposited basement membrane production
yields some other interesting observations. The EC TIE-2 receptor is preferentially
expressed in the stalk portion of a maturing capillary sprout, but are notably absent
in the tip ECs. This expression is thought to be controlled by the pericytes, which
produce Ang-1 to signal to the ECs via the TIE-2 receptor [ 45 , 75 ]. This Ang-1/
TIE-2 interaction is one mechanism by which pericytes and SMCs can regulate
MMP activity of ECs. An alternative, and much more direct, means to achieve this
control is via TIMP-1 secretion. TIMP-1 inhibits soluble MMPs, but also appears
to stabilize vessels by inducing basement membrane protein production [ 76 ]. Other
TIMPs also work in conjunction with both cell types to inhibit angiogenic
sprouting and stabilize nascent vessels. TIMP-3 is secreted by perivascular cells,