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Benefits of Nitric Oxide Cues to Matrix Synthesis by Healthy and Aneurysmal Human Smooth Muscle Cells within 3D Cocultures

Simmers, Phillip Charles
http://rave.ohiolink.edu/etdc/view?acc_num=csu1399977973

Abstract Details

2014, Master of Science in Biomedical Engineering, Cleveland State University, Washkewicz College of Engineering.
Vascular diseases such as atherosclerosis and aneurysms are characterized by the over-proliferation and migration of aortic SMCs, and degradation of 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 GSNO, to cell proliferation and matrix deposition by adult human aortic 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 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, SMC proliferation decreased in the presence or absence of EC cocultures, while EC proliferation increased. GSNO (100 nM) significantly enhanced the total protein amount synthesized by SMCs, in the presence or absence of EC cocultures, while lower dosages offered marginal benefits. On a per cell basis, multi-fold increases in the synthesis and deposition of elastin, GAGs, hyaluronic acid and 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 dosages, although MMP-2 levels remained attenuated. These quantitative assay data were strongly supported by immunofluorescence images under respective conditions. Based on the optimized dosage (100 nM) deduced from first experiment, we tested the utility of NO cues within aneurysmal HA-SMC cultures, under both 2D and 3D culture conditions. Results showed that cell proliferation was higher on 2D substrates compared to that within 3D cultures. In accordance to the trends noted within healthy SMC cultures, there was a decrease in cell proliferation in both 2D and 3D, with increasing GSNO dose. Similarly, the presence of GSNO stimulated elastin, GAGs, hyaluronic acid, LOX, MMP-2, 9 and TIMP-1 synthesis and release by aneurysmal SMCs, although the matrix deposition levels were lower compared to their healthy counterparts. Both experiments results attest to the tremendous benefits of delivering NO cues to suppress SMC proliferation and promote robust ECM synthesis and deposition by HA-SMCs, with significant applications in tissue engineering, biomaterial scaffold development, and drug delivery.
Chandra Kothapalli, PhD (Committee Chair)
Nolan Holland, PhD (Committee Member)
Moo-Yeal Lee, PhD (Committee Member)
99 p.
Anatomy and Physiology; Biochemistry; Biomedical Engineering; Biomedical Research; Cellular Biology; Engineering; Health; Health Care; Medicine; Pharmaceuticals
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

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Citations

  • Simmers, P. C. (2014). Benefits of Nitric Oxide Cues to Matrix Synthesis by Healthy and Aneurysmal Human Smooth Muscle Cells within 3D Cocultures [Master's thesis, Cleveland State University]. OhioLINK Electronic Theses and Dissertations Center. http://rave.ohiolink.edu/etdc/view?acc_num=csu1399977973

    APA Style (7th edition)

  • Simmers, Phillip. Benefits of Nitric Oxide Cues to Matrix Synthesis by Healthy and Aneurysmal Human Smooth Muscle Cells within 3D Cocultures. 2014. Cleveland State University, Master's thesis. OhioLINK Electronic Theses and Dissertations Center, http://rave.ohiolink.edu/etdc/view?acc_num=csu1399977973.

    MLA Style (8th edition)

  • Simmers, Phillip. "Benefits of Nitric Oxide Cues to Matrix Synthesis by Healthy and Aneurysmal Human Smooth Muscle Cells within 3D Cocultures." Master's thesis, Cleveland State University, 2014. http://rave.ohiolink.edu/etdc/view?acc_num=csu1399977973

    Chicago Manual of Style (17th edition)

csu1399977973
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© 2014, all rights reserved.
This open access ETD is published by Cleveland State University and OhioLINK.