A Transwell-Based Vascularized Model to Investigate the Effect of Interstitial Flow on Vasculogenesis

Abstract

Interstitial flow plays a significant role in vascular system development, mainly including angiogenesis and vasculogenesis. However, compared to angiogenesis, the effect of interstitial flow on vasculogenesis is less explored. Current in vitro models for investigating the effect of interstitial flow on vasculogenesis heavily rely on microfluidic chips, which require microfluidic expertise and facilities, and may not be accessible to biological labs. Here, we proposed a facile approach to building perfusable vascular networks through the self-assembly of endothelial cells in a modified transwell format and investigated the effect of interstitial flow on vasculogenesis. We found that the effect of interstitial flow on vasculogenesis was closely related to the existence of VEGF and fibroblasts in the developed model: (1) In the presence of fibroblasts, interstitial flow (within the range of 0.1–0.6 μm/s) facilitated the perfusability of the engineered vasculatures. Additional VEGF in the culture medium further worked synergically with interstitial flow to develop longer, wider, denser, and more perfusable vasculatures than static counterparts; (2) In the absence of fibroblasts, vasculatures underwent severe regression within 7 days under static conditions. However, interstitial flow greatly inhibited vessel regression and enhanced vascular perfusability and morphogenesis without the need for additional VEGF. These results revealed that the effect of interstitial flow might vary depending on the existence of VEGF and fibroblasts, and would provide some guidelines for constructing in vitro self-assembled vasculatures. The established transwell-based vascularized model provides a simple method to build perfusable vasculatures and could also be utilized for creating functional tissues in regenerative medicine.

The Edifice of Vasculature-On-Chips: A Focused Review on the Key Elements and Assembly of Angiogenesis Models

“…It also provides a specific mechanical environment necessary for the physiological activity of interstitial cells . In angiogenesis chips, researchers have predominantly used such devices to introduce and regulate interstitial flow into their culture system with the aim of better understanding endothelial cell behavior. , Deng et al demonstrated that the coculture of HUVECs and NHLFs under interstitial flow promoted lumenization and perfusability of engineered vasculature, enhanced vascular permeability, and stimulated morphogenesis without the need for additional VEGF . Interstitial flow has been demonstrated to upregulate matrix metalloproteinase-2 (MMP-2) and promote neovessel formation, demonstrating enhanced vessel formation, connectivity, and function in the brain microvasculature network .…”

“… 90 , 143 Deng et al demonstrated that the coculture of HUVECs and NHLFs under interstitial flow promoted lumenization and perfusability of engineered vasculature, enhanced vascular permeability, and stimulated morphogenesis without the need for additional VEGF. 144 Interstitial flow has been demonstrated to upregulate matrix metalloproteinase-2 (MMP-2) and promote neovessel formation, demonstrating enhanced vessel formation, connectivity, and function in the brain microvasculature network. 145 In brain microvasculature models, Winkleman et al described the role of interstitial pressure by devising a simple 3-channel microfluidic system comprising primary brain ECs, pericytes, and astrocytes within a 3D fibrin matrix.…”