Synergy between interstitial flow and VEGF directs capillary morphogenesisin vitrothrough a gradient amplification mechanism

Abstract

Cell organization is largely orchestrated by extracellular gradients of morphogenetic proteins. VEGF, an essential factor for capillary formation, is stored in the extracellular matrix, but the mechanisms by which it and other matrix-bound morphogens are mobilized to form spatial gradients are poorly understood. Here, we suggest an efficient mechanism for morphogen gradient generation by subtle biophysical forces in an in vitro model of capillary morphogenesis. Using a fibrin-bound VEGF variant that is released proteolytically to mimic the in vivo situation, we report that low levels of interstitial flow act synergistically with VEGF to drive endothelial organization, whereas each stimulus alone has very little effect. To help account for this synergy, we show how these slow flows can bias the distribution of cell-secreted proteases, which leads, interestingly, to the creation of an increasing VEGF gradient relative to the cell and skewed in the direction of flow. In contrast, diffusion alone can only account for symmetric, decreasing autocrine gradients. Indeed, branching of capillary structures was biased in the direction of flow only with the combination of VEGF and flow. This work thus demonstrates a general mechanism of morphogen gradient generation and amplification by small ubiquitous mechanical forces that are known to exist in vivo.fibrin ͉ angiogenesis ͉ lymphatic ͉ biotransport ͉ mechanobiology T issue morphogenesis results from the organization of cells into functional structures by means of the actions of biomolecular morphogens. The efficacy of a morphogen depends largely on its local gradient relative to the cell rather than its absolute amount (1). Such morphogens, particularly growth factors and some cytokines, commonly bind to the extracellular matrix (ECM) to be later released by cells via proteolytic enzymes. Although Alan Turing (2) first showed how patterns in morphogenesis could arise from diffusion and interaction kinetics, his arguments were derived for directly cell-secreted morphogens. The question of how cells create morphogen gradients by proteolytically mobilizing matrix-bound factors has not yet been addressed, and we wondered whether small biophysical forces that are ubiquitously present in vivo may help facilitate cell utilization of such factors.

Role of Lymphatic Vessels in Tumor Immunity: Passive Conduits or Active Participants?

“…In addition to regulating function, fluid flow is also an important regulator of lymphatic morphogenesis or lymphangiogenesis, and has been shown to drive lymphatic capillary organization in dermal wound healing models [45,46] as well as in vitro culture models [47][48][49]. Without flow, as in the case of lymphedema, lymphatic endothelium becomes hyperplastic (i.e.…”

A multichamber fluidic device for 3D cultures under interstitial flow with live imaging: Development, characterization, and applications

“…where K 0 is the hydraulic conductivity of the gel, m is the viscosity of the medium ($0.75 cp for 20% serumcontaining endothelial cell medium at 378C (Helm et al, 2005)), and 5P is the imposed pressure gradient (¼ dP/dr). We can integrate over the radius and estimate K 0 from the slope of DP versus Q:…”

“…We compared static versus flow conditions on LECs, as flow was previously shown to promote lymphatic capillarogenesis (Boardman and Swartz, 2003;Ng et al, 2004), as well as the effects of matrix-bound VEGF on LEC and BEC morphogenesis under flow (Helm et al, 2005) for 10 days. We found that interstitial flow of 1-2 mm/s induced from a 5 mm pressure drop (Fig.…”

Portal hypertension-related inflammatory phenotypes: From a vitelline and amniotic point of view

“…The interstitial fluid flow associated with edema, even though it can be extremely slow, can have important effects on tissue morphogenesis and function, cell migration and differentiation and matrix remodeling, among other processes [108]. Abnormally increased interstitial flow rates can occur during low-grade inflammation and can also trigger fibroblasts to differentiate or remodel the extracellular matrix, contributing thus to the development of tissue fibrosis [107,[109][110][111][112]. Interstitial flow may significantly alter the distribution of metalloproteinases and lymphatic growth factors, inducing lymphatic endothelial cell migration and capillary morphogenesis [107,110].…”