Physiologically Modeled Pulse Dynamics to Improve Function inIn Vitro-Endothelialized Small-Diameter Vascular Grafts

Abstract

The lack of a functional endothelium on small-diameter vascular grafts leads to intimal hyperplasia and thrombotic occlusion. Shear stress conditioning through controlled hydrodynamics within in vitro perfusion bioreactors has shown promise as a mechanism to drive endothelial cell (EC) phenotype from an activated, pro-inflammatory wound state toward a quiescent functional state that inhibits responses that lead to occlusive failure. As part of an overall design strategy to engineer functional vascular grafts, we present a novel two-phase shear conditioning approach to improve graft endothelialization. Axial rotation was first used to seed uniform EC monolayers onto the lumenal surface of decellularized scaffolds derived from the human umbilical vein. Using computer-controlled perfusion circuits, a flow-ramping paradigm was applied to adapt endothelia to arterial levels of fluid shear stress and pressure without graft denudation. The effects of constant pulse frequencies (CF) on EC quiescence were then compared with pulse frequencies modeled from temporal fluctuations in blood flow observed in vivo, termed physiologically modeled pulse dynamics (PMPD). Constructs exposed to PMPD for 72 h expressed a more functional transcriptional profile, lower metabolic activity (39.8% ± 8.4% vs. 62.5% ± 11.5% reduction, p = 0.012), and higher nitric oxide production (80.42 ± 23.93 vs. 48.75 ± 6.93 nmol/10(5) cells, p = 0.028) than those exposed to CF. By manipulating in vitro flow conditions to mimic natural physiology, endothelialized vascular grafts can be stimulated to express a nonactivated phenotype that would better inhibit peripheral cell adhesion and smooth muscle cell hyperplasia, conditions that typically lead to occlusive failure. Development of robust, functional endothelia on vascular grafts by modulation of environmental conditions within perfusion bioreactors may ultimately improve clinical outcomes in vascular bypass grafting.

Shear-resistant hydrogels to control permeability of porous tubular scaffolds in vascular tissue engineering

“…Vascular smooth muscle cells (VSMCs) and endothelial cells (ECs) of the tunica media and intima, respectively, have different requirements for growth and this represents a key morphological and functional characteristic of native vessels. Thus, both luminal and outer layers of a vascular scaffold should support the adhesion of the different cells, while the degradation of the different layers should be consistent with the different cell growth kinetics [8][9][10][11]. Engineering gradients, also in vascular tissue engineering, is of outmost importance [12].…”

“…The selection of suitable scaffolds is of outmost importance. The simplest scaffolds mostly used as vascular grafts include decellularized swine arteries [19] and umbilical arteries or veins [8,20], owing to their comparable mechanical properties to those of native tissue [21,22], neglectable or low porosity [23], and high EC-adhesive properties [21]. As drawbacks, these scaffolds impair VSMC infiltration, making the re-cellularization process hard and time-dispersive [19].…”

Sequential adaptation of perfusion and transport conditions significantly improves vascular construct recellularization and biomechanics

“…Additional development of an endothelial layer to avoid thrombosis of the vessel upon implantation is also crucial (Sánchez, Brey, & Briceño, 2018). It has previously been demonstrated that the re-endothelialization of the acellular human umbilical vein can similarly be performed using sequential conditioning steps (Uzarski, Cores, & McFetridge, 2015). In this case, an initial step consisted in 5 days of static cell adhesion to the lumenal surface and was followed by 24 hr of progressive flow rate increase up to reaching arterial shear stress and pressure.…”

Vascular Smooth Muscle Cells (VSMCs) in Blood Vessel Tissue Engineering: The Use of Differentiated Cells or Stem Cells as VSMC Precursors

“…For this purpose, chemically defined serum-free or serum-low media are used, for example, media supplemented with transforming growth factor-β (TGF-β) [26][27][28] or with heparin [29]. At the same time, heparin supports endothelialization of the prosthesis [30], which also contributes to the development of the contractile phenotype in VSMCs, for example, by producing sulfated heparinlike glycosaminoglycans [2,18,31], nitric oxide [32,33] and by developing contacts between VSMC and endothelial cells, that is, myoendothelial gap junctions [27].…”