<p>Endothelial dysfunction is the predominant pathophysiological state prior to the onset of atherosclerosis. Currently, treatments for endothelial dysfunction are evaluated in vitro using two-dimensional (2D) cell culture assays or in vivo animal models. Microphysiological systems are small-scale three-dimensional (3D) tissue models that recapitulate the native tissue structure and function. An ideal microphysiological system is comprised of human cells embedded within a 3D matrix introduced to physiological fluid perfusion. Immune challenge in the form of cytokines or immune cells further recapitulates the native microenvironment.</p><p>A vascular microphysiological system was developed from a small-diameter tissue engineered blood vessel (TEBV) in a perfusion culture circuit. TEBVs were created from collagen gels embedded with human neonatal dermal fibroblasts and plastically compressed to yield collagen constructs with high fiber densities. TEBVs are rapidly producible and can be directly introduced into perfusion culture immediately after fabrication. Endothelium-independent vasoconstriction in response to phenylephrine and endothelium-dependent vasodilation in response to acetylcholine were used to analyze the health and function of the endothelium non-destructively over time.</p><p>Endothelial dysfunction was induced through introduction of the pro-inflammatory cytokine tumor necrosis factor – α (TNF-α). Late-outgrowth endothelial progenitor cells derived from the peripheral blood of coronary artery disease patients (CAD EPCs) were evaluated as a potential endothelial source for autologous implantation in both a two-dimensional (2D) direct co-culture model as well as a 3D model as an endothelial source for a tissue engineered blood vessel. CAD EPCs demonstrated similar adhesive properties to a confluent, quiescent layer of smooth muscle compared to human aortic endothelial cells. Within the TEBV system, CAD EPCs demonstrated the capacity to elicit endothelium-dependent vasodilation. CAD EPCs were compared to adult EPCs from young, healthy volunteers. Both CAD EPCs and healthy volunteer EPCs demonstrated similar endothelium-dependent vasoactivity in response to acetylcholine; however, in response to TNF-α, CAD EPCs demonstrated a reduced response to phenylephrine at high doses.</p><p>The treatment of TEBVs with statins was explored to model the drug response within the system. TEBVs were treated with lovastatin, atorvastatin, and rosuvastatin for three days prior to exposure to TNF-α. In all three cases, statins prevented TNF-α induced vasoconstriction in response to acetylcholine within the TEBVs, compared to TEBVs not treated with statins. Overall, this work characterizes and validates a novel vascular microphysiological system that can be tested in situ in order to determine the effects of various patient populations and drugs on endothelial health and function under healthy and inflammatory conditions.</p> / Dissertation

<p>  </p> <p>Diabetes, a chronic condition characterized by elevated blood glucose levels, arises when pancreatic β-cells lose capacity to produce a robust, dynamic glucose-stimulated insulin secretion (GSIS) response. Accurate measurement of β-cell health and function <em>ex vivo</em> is thus fundamental to diabetes research, including studies evaluating disease mechanisms, novel drug candidates, and replacement β-cell populations. However, present-day dynamic GSIS assays typically represent end-point measurements, involve expensive commercial perifusion machines, and require time-consuming enzyme-linked immunosorbent assays (ELISA) for insulin detection. Microfluidic devices developed as accessible, low-cost alternatives still rely on secondary ELISAs and suspend islets in liquid medium, limiting their survival <em>in vitro</em>. Here, we present a novel, 3D-printed microphysiological system (MPS) designed to recreate components of <em>in-vivo</em> microenvironments through encapsulation in fibrillar type I collagen and restoration of favorable molecular transport conditions. Following computational-informed design and rapid prototyping, the MPS platform sustained collagen-encapsulated mouse islet viability and cytoarchitecture for 5 days and supported <em>in-situ</em> measurements of dynamic β-cell function. To rapidly detect insulin secretion from β-cells in the MPS, we then developed a highly sensitive electrochemical sensor for zinc (Zn2+), co-released with insulin, based on glassy carbon electrodes modified with bismuth and indium and coated with Nafion. Finally, we validated sensor detection of Zn2+ released from glucose-stimulated INS-1 β-cells and primary mouse islets, finding high correlation with insulin as measured by standard ELISA. Together, the 3D MPS and Zn2+ sensor developed in this dissertation represent novel platforms for evaluating β-cell health and function in a low-cost, user-friendly, and physiologically-relevant manner. </p>