Browsing by Author "David, Dency"

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    Effect of substrate topography on human vascular smooth muscle cell proliferation and phenotype change
    (University of Waterloo, 2025-01-22) David, Dency
    Cardiovascular diseases (CVDs) remain the leading cause of mortality worldwide, with vascular occlusion being a primary contributor. Bypass grafting is a common surgical intervention to restore blood flow, traditionally using autologous grafts such as saphenous veins and internal thoracic arteries. However, the limited availability and invasive harvesting process of autologous grafts have prompted the development of synthetic small-diameter vascular grafts (sSDVGs) as alternatives. Despite advancements, the clinical efficacy of sSDVGs remains unsatisfactory due to high rates of thrombotic occlusion, intimal hyperplasia (IH), and restenosis, primarily caused by dysregulated vascular smooth muscle cell (VSMC) behavior. VSMCs play a critical role in the progression of IH through their proliferation, migration, and phenotypic plasticity following vascular injury. While extensive studies have explored the influence of substrate topography on endothelial cell (EC) response, the effects on VSMCs remain underexplored. This study investigates the hypothesis that substrate topographies with varying geometries, isotropy, and sizes can differentially regulate VSMC behavior, potentially mitigating IH and improving the functionality of sSDVGs. To test this hypothesis, a 16-pattern multiarchitecture (MARC) chip was employed to screen various surface patterns for their ability to modulate VSMC phenotype. Five promising patterns were selected and individually fabricated on polydimethylsiloxane (PDMS) substrates for further evaluation. The influence of these topographies on VSMC behavior was assessed under normal and platelet-derived growth factor (PDGF)-stimulated conditions by analyzing protein markers associated with VSMC phenotypic states, including α-smooth muscle actin (α-SMA), phosphorylated myosin light chain kinase (pMLCK), F-actin, desmin, vimentin, phosphorylated focal adhesion kinase (pFAK), and yes associated protein (YAP). Among the tested patterns, the 2μm grating emerged as the most effective in inducing a contractile VSMC phenotype. VSMCs cultured on this pattern exhibited reduced proliferation, an elongated spindle-like morphology, and increased expression of muscle-specific proteins, irrespective of PDGF presence. Conversely, VSMCs on the 1.8μm convex microlens and unpatterned substrates showed higher proliferation rates and a diminished contractile phenotype. Remarkably, the beneficial effects of the 2 μm grating pattern were retained when incorporated into a fucoidan-modified polyvinyl alcohol (PVA) hydrogel, a biomaterial known to support EC adhesion and exhibit low thrombogenicity. The 2μm grating suppressed PDGF-induced proliferation while promoting a contractile phenotype and enhancing directional motility. Mechanistic studies revealed elevated pMLCK expression, increased cytoplasmic localization of YAP, and enhanced focal adhesion maturation on 2μm gratings, supporting contractility and reducing proliferation. In contrast, unpatterned and 1.8μm convex lens substrates induced nuclear YAP localization and reduced pMLCK expression, favoring a proliferative phenotype. This study introduces a promising strategy for regulating VSMC behavior through substrate topography, leveraging biophysical cues to promote a contractile phenotype while suppressing proliferation. By incorporating these insights into the design of biomimetic graft surfaces, this approach holds significant potential to address the limitations of sSDVGs, reduce complications such as IH, and improve long-term graft patency. Furthermore, the integration of topographical and biochemical modifications into PVA-based hydrogels represents an innovative avenue for the development of next-generation vascular grafts that combine mechanical strength with enhanced biological functionality. This work paves the way for advancing sSDVGs toward better clinical outcomes, reduced graft failure, and improved patient prognosis.