Assistant Professor University of Missouri - Columbia Columbia, Missouri, United States
Introduction: Arterial grafts are used to replace small-diameter vessels in vascular disease treatments, but they often suffer from thrombosis due to inadequate endothelialization. Endothelial cells grown under laminar flow conditions mimic native vascular environments, promoting beneficial protein expression but suffering from high detachment and loss rates. This study investigated whether substrates with aligned fibrous orientations could similarly promote healthy endothelial behaviors without the drawbacks of direct laminar flow. The investigation focused on human umbilical vein endothelial cells (HUVECs) cultured on substrates with either random or aligned fiber orientations. We aimed to determine if aligned fibers encouraged HUVECs to undergo significant morphological changes, resembling the effects of laminar flow, enhanced migration, especially gene expression related to vascular health, and thrombosis prevention.
Materials and
Methods: Random and aligned substrates were produced from SG-80 polymer through electrospinning. The surface roughness, hydrophobicity, structure, tensile, and nanomechanical properties of the random substrates were evaluated and compared to aligned substrates. The random and aligned substrates were seeded with HUVECs and cultured in vitro for one month. Cell proliferation, migration, elongation, viability, ECM production, and gene expression related to healthy endothelialization were assessed on the substrates.
Results, Conclusions, and Discussions: The random and aligned substrates exhibited random and aligned fiber orientations, respectively (Figure: a-b). HUVECs were cultured on the surfaces of the substrates. The cells on the aligned substrates exhibited an elongated morphology in the direction parallel to the orientation of the fibers (Figure: c). Conversely, the cells cultured on the random substrates adopted a cobblestone morphology (Figure: d). Moreover, the cells on the substrates were stained for vWF and CD31, which is a crucial glycoprotein and adhesion molecule expressed in endothelial cells, respectively. The cells on the aligned substrates exhibited more vWF and CD31 expression than the cells on the random substrate (Figure: e-h). Moreover, the aligned substrate had cells oriented in the direction of the fibers, while the cells on the random substrate had a random orientation. The gene expression of VEGF, laminin, collagen IV, vWF, CD31, and VE-cadherin of the endothelial cells on the substrates was also measured through RT-PCR to identify the effect of the substrates on the biological behavior of the cells. The cells on the aligned substrates had higher gene expressed of VEGF, laminin, collagen IV, vWF, CD31, and VE-cadherin than those on the random substrates, which suggests that substrates with aligned fiber orientation improve the biological behavior of cultured endothelial cells (Figure: i). Collectively, these results indicate that the physical structure of the substrate plays a significant role in influencing HUVEC behavior. Specifically, aligned substrates appear to mimic the cellular effects typically induced by laminar flow conditions, underscoring the importance of substrate structure in healthy endothelial cell growth and the formation of an anti-thrombotic endothelium in vascular devices.
