Microfluidic Devices for Angiogenesis - Mechanical and Chemical Signaling in Angiogenesis - page 104

Assays for angiogenesis have varied in their design, ranging from the more con-
ventional gel systems, where endothelial cells are plated onto the surface of a gel
[ 82 ] (Fig. 3 a) or sandwiched between two layers of hydrogel [ 83 ] (Fig. 3 b) with
growth factors added to stimulate vascular network formation or sprouting, to
systems in which the cells are plated as a monolayer on a gel surface (Fig. 3 C), or
vessel rings [ 84 ] (Fig. 3 d) or endothelial-coated microbeads are suspended in
matrix [ 85 ] (Fig. 3 e).
Two different types of approach have been used, one to study vasculogenesis
and another for angiogenesis. While we focus most of the discussion here on
angiogenesis, a short summary of some of the important microfluidic work on
vasculogenesis is first presented.
Vasculogenesis occurs by the transformation of angioblasts into endothelial
cells that assemble to form a microvascular network [ 86 ]. This process usually
occurs during development (although there is some evidence that it can also occur
in adults), whereas angiogenesis can occur both in the embryo and the adult
organism [ 87 ]. Vasculogenesis, or at least one model for it, is accomplished by
sandwiching endothelial precursor cells or endothelial cells themselves, between
two layers of gel [ 83 ]. In this case, vascular networks form as a consequence of the
seeded cells stretching out toward their neighbors, making contact, forming a 3D
network and in many cases, ultimately creating an interconnected array of tubes
with diameters comparable to those of a capillary. Generally, these networks have
been non-functional in the sense that it is not possible to perfuse them due to the
manner in which they are connected to the channels of the device, but systems
having the capability to link with these formed networks now exist and it is just a
matter of time before these systems can be perfused and presumably used to
provide nutrients and gas exchange to other resident cells.
Several microfluidic assays for angiogenesis have been developed consisting of
medial channels that run parallel to a gel region. One such system consists of a
hydrogel (e.g., collagen) seeded with embryoid bodies or embryonic kidneys and
flanked on both sides by medium-filled channels [ 88 ] (Fig. 4 C). Molded from
PDMS and sealed by vacuum against a petri dish following the placement of
tissue-seeded gels, gradients were produced in this system by the flow of growth
factor-containing medium (VEGFA, VEGFC, FGF2) on one side and control
medium on the other. Vascular sprouts were observed to form preferentially in the
direction of higher growth factor concentrations. The authors noted the potential
for real-time imaging and the retrieval of tissue specimens at the end of the
experiment for subsequent biochemical analysis.
A major challenge in using this system was the containment of gel solution,
which these authors accomplished by means of PDMS plugs inserted into the
channels during gel polymerization. Another approach is to use posts cast in the
PDMS replica to define the gel region, and to rely on surface tension along these
posts to contain the gel and also to stabilize it after gelation (Fig. 4 A, B) [ 35 , 89 ].
Initial systems accomplished this by direct microinjection of the gel solution,
followed immediately (before polymerization) by attachment of a coverslip to the
plasma-treated PDMS surface. In these studies, medium and endothelial cells were