ENDOTHELIAL PROGENITOR CELL RECRUITMENT IN A WOUND HEALING MICROFLUIDIC VASCULAR MODEL
Lewis, Daniel Matthew
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During wound healing, endothelial progenitor cells (EPCs) are recruited from the bone marrow and directed to the site of injury. At the site of injury, hypoxic conditions promote TNF-α, which up regulates intercellular adhesion molecule-1 (ICAM-1). EPCs use ICAM-1 to attach to endothelial cells (ECs) lining blood vessels. Here, we design, develop and test a three-dimensional microbioreactor system (3-D MBR) with precise control and monitoring of oxygen and media flow rate. We first analyze the transport of oxygen in the proposed device. Following fabrication of the 3-D MBR, we next utilized a step-wise seeding technique, which resulted in confluency of human umbilical vein endothelial cells (HUVECs) on all four sides of the device. We next examine endothelial colony forming cell (ECFC) attachment and retention onto HUVECs using conventional 2-D cultures. HUVECs are pre-stimulated with one of four conditions: 21% oxygen (atmospheric), atmospheric with TNF-α-supplemented media, 1% oxygen (hypoxia), and lastly hypoxia with TNF-α-supplemented media. We show the highest attachment and retention of ECFCs on HUVECs pre-treated with TNF-α and 1% oxygen, which correlated with the highest expression levels of ICAM-1. Using the new 3-D MBR system we next demonstrate that TNF-α and hypoxia, when used in conjunction, significantly increase EPC attachment on ECs under pathologically relevant flow conditions. The 3-D MBR system allows us to mimic the oxygen and shear stress environment in the vasculature, thus providing a step between traditional in-vitro and in-vivo experimentation to model a variety of vascular-related disorders, especially wound healing.