Micropatterned cell sheets as structural building blocks for biomimetic vascular patches

Abstract

To successfully develop a functional tissue-engineered vascular patch, recapitulating the hierarchical structure of vessel is critical to mimic mechanical properties. Here, we use a cell sheet engineering strategy with micropatterning technique to control structural organization of bovine aortic vascular smooth muscle cell (VSMC) sheets. Actin filament staining and image analysis showed clear cellular alignment of VSMC sheets cultured on patterned substrates. Viability of harvested VSMC sheets was confirmed by Live/Dead cell viability assay after 24 and 48 h of transfer. VSMC sheets stacked to generate bilayer VSMC patches exhibited strong inter-layer bonding as shown by lap shear test. Uniaxial tensile testing of monolayer VSMC sheets and bilayer VSMC patches displayed nonlinear, anisotropic stress-stretch response similar to the biomechanical characteristic of a native arterial wall. Collagen content and structure were characterized to determine the effects of patterning and stacking on extracellular matrix of VSMC sheets. Using finite-element modeling to simulate uniaxial tensile testing of bilayer VSMC patches, we found the stress-stretch response of bilayer patterned VSMC patches under uniaxial tension to be predicted using an anisotropic hyperelastic constitutive model. Thus, our cell sheet harvesting system combined with biomechanical modeling is a promising approach to generate building blocks for tissue-engineered vascular patches with structure and mechanical behavior mimicking native tissue.

Generation of a Purified iPSC-Derived Smooth Muscle-like Population for Cell Sheet Engineering

“…Mouse iPSC-derived Acta2 hrGFP+ cell sheets had significantly higher elastic modulus compared with the MOVAS control cell sheets but were not significantly different from that of Acta2 hrGFP− cell sheets. The moduli of the samples were in the 50- to 250-kPa range, which is in the range of elastic moduli of bovine vascular SMC sheets previously reported by our group (Backman et al., 2017a, Rim et al., 2018); however, this range of moduli is on the lower end of reported stiffnesses for blood vessels and engineered vascular grafts (0.1–50 MPa) (Guo and Kassab, 2003, Lashkarinia et al., 2018, Wagenseil and Mecham, 2009). The Acta2 hrGFP+ cell sheets did, however, have significantly higher max stress at failure when compared with Acta2 hrGFP− cell sheets and MOVAS cell sheets.…”

“…To determine whether our iPSC derivatives could serve as building blocks for engineered smooth muscle tissue constructs, we next utilized a micropatterned enzyme-degradable hydrogel system (Figure 6A) (Rim et al., 2018) to generate releasable aligned cell sheets that maintain cell-cell junctions and extracellular matrix (ECM) proteins. After seeding, sorted human or mouse iPSC-SMCs aligned in the direction of the micropatterns, consistent with our previously published work using multiple micropatterned substrate systems (Backman et al., 2017a, Rim et al., 2018). Notably, the Acta2 hrGFP+ population retained expression of the GFP reporter following alignment on the alginate scaffold (Figure 6B).…”

Polydopamine and collagen coated micro-grated polydimethylsiloxane for human mesenchymal stem cell culture

“…Micro-topography controlled cellular behavior has been widely examined by researchers in the past few years. For example, Rim et al used aligned microgrooves with 5 μm height and 30 μm width to develop aligned vascular smooth muscle cell sheets, which can further be used as building blocks to fabricate tissue-engineered vascular patches [ 11 ]. In another study, aligned microgroove and micro-wavy surface with 5 μm groove height and 20 μm width showed enhanced EC alignment and adhesion [ 12 ].…”

Development of a bio‐MEMS device for electrical and mechanical conditioning and characterization of cell sheets for myocardial repair

“…The first technique fills in and coats the device with either alginate or cellulose hydrogel. This can later be selectively enzymatically degraded, keeping the cellular connections intact and releasing the cell sheet as has been previously achieved for planar and patterned substrates (Rim et al, ). The second method uses a custom synthetic oligopeptide, CCRRGDWLC, (Sigma‐Aldrich, TX) that forms a thiol bond with gold at either end, exposing a backbone with an RGD bonding site for cells (Enomoto et al, ).…”