Abstract
Endothelial progenitor cell (EPC) transplantation induces the formation of new blood-vessel networks to supply nutrients and oxygen, and is feasible for the treatment of ischemia and cardiovascular diseases. However, the role of EPCs as a source of proangiogenic cytokines and consequent generators of an extracellular growth factor microenvironment in three-dimensional (3D) microvessel formation is not fully understood. We focused on the contribution of EPCs as a source of proangiogenic cytokines on 3D microvessel formation using an in vitro 3D network model. To create a 3D network model, EPCs isolated from rat bone marrow were sandwiched with double layers of collagen gel. Endothelial cells (ECs) were then cultured on top of the upper collagen gel layer. Quantitative analyses of EC network formation revealed that the length, number, and depth of the EC networks were significantly enhanced in a 3D model with ECs and EPCs compared to an EC monoculture. In addition, conditioned medium (CM) from the 3D model with ECs and EPCs promoted network formation compared to CM from an EC monoculture. We also confirmed that EPCs secreted vascular endothelial growth factor (VEGF). However, networks cultured with the CM were shallow and did not penetrate the collagen gel in great depth. Therefore, we conclude that EPCs contribute to 3D network formation at least through indirect incorporation by generating a local VEGF gradient. These results suggest that the location of EPCs is important for controlling directional 3D network formation in the field of tissue engineering.
Highlights
Neovascularization is a crucial step in the development of reconstructed tissue applied to clinical practice
The number of networks in the Endothelial cells (ECs)+Endothelial progenitor cell (EPC) model reached 8 ± 3 networks/mm2, whereas that of the EC model was 3 ± 1 networks/mm2 at day 5 (Figure 3H). These results indicate that the ECs in the EC+EPC model frequently invade the underlying collagen gel to construct 3D endothelial networks
This study demonstrated the contribution of EPCs as a source of proangiogenic cytokines on 3D endothelial network formation using an in vitro 3D network model
Summary
Neovascularization is a crucial step in the development of reconstructed tissue applied to clinical practice. Some two-dimensional (2D) tissue-engineered products such as skin and cornea substitutes have already been in practical use [1,2], reconstruction of three-dimensional (3D) tissues, such as the liver and heart, remains difficult. This is due to the need for 3D tissues to have an extensive vascular system to support tissues by providing oxygen and nutrients, as diffusion of the oxygen and nutrients cannot penetrate more than a few hundred microns. Recent findings recognize that the concentration profile of extracellular growth factors, such as vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF), plays an important role in regulating the formation of 3D microvascular networks. Gerhardt et al [4]
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