Date of Award

2014

Degree Type

Thesis

Department

Chemical and Biomedical Engineering

First Advisor

Kothapalli, Chandra

Subject Headings

Nitric oxide -- Physiological effect, Vascular smooth muscle, Aorta -- Cytology, Muscle cells, Nitric Oxide NO Human Aortic Smooth Muscle Cell HA-SMC Coculture S-Nitrosoglutathione GSNO Human Aortic Endothelial Cell HA-EC Microfluidic Aneurysm Extracellular Matrix ECM Vascular Tissue

Abstract

Vascular diseases such as atherosclerosis and aneurysms are characterized by the over-proliferation and migration of aortic smooth muscle cells (SMCs), and degradation of extracellular matrix (ECM) within the vessel wall, leading to compromise in cell-cell and cell-matrix signaling pathways. Recent tissue engineering approaches to regulate SMC over-proliferation and enhance healthy ECM synthesis showed promise, but resulted in low crosslinking efficiency and matrix deposition yields. In this study, the benefits of exogenous nitric oxide cues, delivered from S-Nitrosoglutathione (GSNO), to cell proliferation and matrix deposition by adult human aortic SMCs (HA-SMCs) within 3D biomimetic cultures have been explored. The first experiment utilized a microfluidic platform with two adjacent, permeable 3D culture chambers, to enable paracrine signaling between vascular cell cocultures. Healthy HA-SMCs were cultured in these devices within 3D collagen hydrogels, either alone or in the presence of human aortic endothelial cell (HA-ECs) cocultures, and exogenously supplemented with varying GSNO dosages (0-100 nM) for 21 days. Results showed that EC cocultures stimulated SMC proliferation within GSNO-free cultures. However, with increasing GSNO concentration, HA-SMC proliferation decreased in the presence or absence of EC cocultures, while HA-EC proliferation increased. GSNO (100 nM) significantly enhanced the total protein amount synthesized by HA-SMCs, in the presence or absence of EC cocultures, while lower dosages (1-10 nM) offered marginal benefits. On a per cell basis, multi-fold increases in the synthesis and deposition of elastin, glycosaminoglycans, hyaluronic acid and lysyl oxidase crosslinking enzyme (LOX) were noted at higher GSNO dosages, and coculturing with ECs significantly furthered these trends. The matrix yields of these proteins reached almost 40 - 51 within selective cocultures receiving GSNO. Similar increases in TIMP-1 and MMP-9 levels were noted within cocultures with increasing GSNO dos

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