Fluid forces control endothelial sprouting

Abstract

During angiogenesis, endothelial cells (ECs) from intact blood vessels quickly infiltrate avascular regions via vascular sprouting. This process is fundamental to many normal and pathological processes such as wound healing and tumor growth, but its initiation and control are poorly understood. Vascular endothelial cell growth factor (VEGF) can promote vessel dilation and angiogenic sprouting, but given the complex nature of vascular morphogenesis, additional signals are likely necessary to determine, for example, which vessel segments sprout, which dilate, and which remain quiescent. Fluid forces exerted by blood and plasma are prime candidates that might codirect these processes, but it is not known whether VEGF cooperates with mechanical fluid forces to mediate angiogenesis. Using a microfluidic tissue analog of angiogenic sprouting, we found that fluid shear stress, such as exerted by flowing blood, attenuates EC sprouting in a nitric oxide-dependent manner and that interstitial flow, such as produced by extravasating plasma, directs endothelial morphogenesis and sprout formation. Furthermore, positive VEGF gradients initiated sprouting but negative gradients inhibited sprouting, promoting instead sheet-like migration analogous to vessel dilation. These results suggest that ECs integrate signals from fluid forces and local VEGF gradients to achieve such varied goals as vessel dilation and sprouting.3D angiogenesis on a chip | collagen gel | structural remodeling | alternative to animal model | vessel analog A ngiogenesis, the expansion or extension of existing vasculature, is necessary to deliver oxygen and nutrients to ischemic or avascular regions in wounds and solid tumors (1), and a fundamental understanding of the determinants of angiogenesis would accelerate progress in the fields of regenerative medicine, tissue engineering, and oncology. Angiogenesis requires the coordinated growth and migration of endothelial cells (ECs): each EC residing in a vessel wall integrates local signals to determine whether it remains quiescent (2, 3), participates in dilation or contraction (1), undergoes morphogenesis to form an angiogenic sprout (4, 5), or intussusceptive involution (6). Implicit in these processes are endothelial proliferation during expansion of the vasculature and their loss during vessel contraction and pruning (7).Growth factors have pleiotropic effects on ECs and are undoubtedly key controllers of vascular morphogenesis. The best studied vascular morphogen, vascular endothelial growth factor (VEGF) (8), stimulates EC migration (9), proliferation (10), and matrix degradation (2). VEGF also controls vessel morphogenesis by inducing Delta-like ligand 4 (Dll4)-a membrane-bound ligand for the Notch family of receptors-at the advancing front of sprouts (11), thereby promoting the formation of specialized "tip cells," which extend protrusions or filopodia that sense growth factor concentrations to guide sprouts (4, 5). However, given the distinct phenotypes exhibited by ECs during sprouting, dilation, co...

Biomimetic model to reconstitute angiogenic sprouting morphogenesis in vitro

“…Finally, in addition to the simple coordination of tip and stalk cells to form linear vessels, our system also seems to support higher-order events such as branching, a key mechanism to the patterning of sprouts controlled by the dynamic interconversion of stalk cells and filopodia-containing tip cells (25,(43)(44)(45)(46), as well as loss of filopodial activity and regression upon eventual perfusion of the neovessel, a critical component of microvascular pruning and remodeling (47). The basis for this type of pruning could be explained by recent studies reporting that shear stress could suppress VEGF-induced invasion (37). Thus, the system introduced here faithfully recapitulates key features of in vivo angiogenesis and provides the ability to link specific stimuli to defined morphogenetic processes, further illustrating the power of such a model.…”

“…been presented for studying sprouting in the presence of flow (36)(37)(38). These use microfluidic channels with square rather than circular cross-sections, where three walls are silicone or glass and one sidewall is the edge of an ECM matrix compartment that contains PDMS posts for structural support.…”

In vitro microvessels for the study of angiogenesis and thrombosis

“…Three-dimensional system have been developed to study sprouting angiogenesis in vitro, for example, endothelial sprouting from a planar endothelium into bulk collagen gel (24), lateral endothelial sprouting into a collagen gel isolated between two microfluidic channels formed in PDMS silicone (25), and 3D endothelial sprouting from microbead supports in a bulk gel (26). None of these systems allows for the initiation of angiogenesis from native-like vessels with luminal flow and simultaneous control of the chemical and cellular microenvironment of the endothelium.…”

Optimizing design and fabrication of microfluidic devices for cell cultures: An effective approach to control cell microenvironment in three dimensions

“…[11][12][13][14][15] To verify that the device design with non-straight pillars allows the formation of a concentration gradient within the central channel, we performed preliminary numerical simulations in COMSOL. Generally, the generation and maintenance of a stable concentration gradient between the source and the sink depend on the diffusion coefficient of the diffusing molecule within the hydrogel, as well as on the geometrical features of the device.…”

“…Different devices based on this approach have been developed in order to study and unravel the intricate cell-signal interactions that occur in vivo. [13][14][15] Although these lFds may differ in terms of shape, dimensions, and culturing conditions, they all rely on the resistance effects of a cell laden biopolymeric gel to the flow of soluble molecules of interest. The presence of short cuts within the cell containing 3D matrix affects the spatial distribution of molecules and matrix collapse or the local densification impairs the diffusion mechanisms.…”