4
membrane, a key ECM remodeling process which occurs as a result of EC-pericyte
interactions during capillary tube assembly [94]. Vascular basement membrane
assembly generally is thought to be a tube maturation stimulus, and hence, decreased
morphogenesis is coincident with its appearance around the abluminal surface of
EC-lined tubes. This point needs to be investigated in more detail, but early
Fig. 1.1 Temporal analysis of endothelial tube formation events in 3D collagen matrices during
vasculogenesis and angiogenic sprouting. (a) A time-lapse series is shown whereby two ECs are
shown to form intracellular vacuoles that eventually coalescence inside each cell and then following cell-cell contact with the neighboring cells form a lumenal space in between the two cells.
Vacuole fusion events are observed while they are contacting each other. L indicates EC lumen;
arrowheads indicate vacuolating ECs. Bar equals 25 μm. (b) Confocal images of ECs expressing
GFP-Rac1 fusion proteins that label intracellular vacuoles, the developing lumenal membrane, as
well as the plasma membrane during lumen formation events at 24 h of culture. Upper panels- L
indicates EC lumenal space, and arrowheads indicate intracellular vacuoles. Bar equals 20 μm.
Over time ECs form interconnecting networks of tubes which are illustrated using fixed and
toluidine- blue stained cultures. Arrows indicate tube structures; L indicates EC lumen; Bar equals
200 μm. (c) EC sprouting was stimulated by the combination of sphingosine-1-phosphate (1 μm)
and SDF-1α (200 ng/ml) which were mixed in the collagen matrix. A time-lapse series (over a 46 h
period) was taken beneath the surface of the monolayer to examine sprouting events and lumen
development in assays mimicking angiogenesis. White arrow and arrowheads indicate EC tip cells
that are sprouting through the 3D collagen matrices; black arrowheads indicate intracellular vacuoles that are observed particularly in ECs directly trailing tip cells; black arrows indicate the cell
border of EC tubes; L indicates EC lumen. Bar equals 50 μm
G. E. Davis
membrane, a key ECM remodeling process which occurs as a result of EC-pericyte
interactions during capillary tube assembly [94]. Vascular basement membrane
assembly generally is thought to be a tube maturation stimulus, and hence, decreased
morphogenesis is coincident with its appearance around the abluminal surface of
EC-lined tubes. This point needs to be investigated in more detail, but early
Fig. 1.1 Temporal analysis of endothelial tube formation events in 3D collagen matrices during
vasculogenesis and angiogenic sprouting. (a) A time-lapse series is shown whereby two ECs are
shown to form intracellular vacuoles that eventually coalescence inside each cell and then following cell-cell contact with the neighboring cells form a lumenal space in between the two cells.
Vacuole fusion events are observed while they are contacting each other. L indicates EC lumen;
arrowheads indicate vacuolating ECs. Bar equals 25 μm. (b) Confocal images of ECs expressing
GFP-Rac1 fusion proteins that label intracellular vacuoles, the developing lumenal membrane, as
well as the plasma membrane during lumen formation events at 24 h of culture. Upper panels- L
indicates EC lumenal space, and arrowheads indicate intracellular vacuoles. Bar equals 20 μm.
Over time ECs form interconnecting networks of tubes which are illustrated using fixed and
toluidine- blue stained cultures. Arrows indicate tube structures; L indicates EC lumen; Bar equals
200 μm. (c) EC sprouting was stimulated by the combination of sphingosine-1-phosphate (1 μm)
and SDF-1α (200 ng/ml) which were mixed in the collagen matrix. A time-lapse series (over a 46 h
period) was taken beneath the surface of the monolayer to examine sprouting events and lumen
development in assays mimicking angiogenesis. White arrow and arrowheads indicate EC tip cells
that are sprouting through the 3D collagen matrices; black arrowheads indicate intracellular vacuoles that are observed particularly in ECs directly trailing tip cells; black arrows indicate the cell
border of EC tubes; L indicates EC lumen. Bar equals 50 μm
G. E. Davis