Biodegradable Poly-ε-Caprolactone Scaffolds with ECFCs and iMSCs for Tissue-Engineered Heart Valves

Abstract

Clinically used heart valve prostheses, despite their progress, are still associated with limitations. Biodegradable poly-ε-caprolactone (PCL) nanofiber scaffolds, as a matrix, were seeded with human endothelial colony-forming cells (ECFCs) and human induced-pluripotent stem cells-derived MSCs (iMSCs) for the generation of tissue-engineered heart valves. Cell adhesion, proliferation, and distribution, as well as the effects of coating PCL nanofibers, were analyzed by fluorescence microscopy and SEM. Mechanical properties of seeded PCL scaffolds were investigated under uniaxial loading. iPSCs were used to differentiate into iMSCs via mesoderm. The obtained iMSCs exhibited a comparable phenotype and surface marker expression to adult human MSCs and were capable of multilineage differentiation. EFCFs and MSCs showed good adhesion and distribution on PCL fibers, forming a closed cell cover. Coating of the fibers resulted in an increased cell number only at an early time point; from day 7 of colonization, there was no difference between cell numbers on coated and uncoated PCL fibers. The mechanical properties of PCL scaffolds under uniaxial loading were compared with native porcine pulmonary valve leaflets. The Young’s modulus and mean elongation at Fmax of unseeded PCL scaffolds were comparable to those of native leaflets (p = ns.). Colonization of PCL scaffolds with human ECFCs or iMSCs did not alter these properties (p = ns.). However, the native heart valves exhibited a maximum tensile stress at a force of 1.2 ± 0.5 N, whereas it was lower in the unseeded PCL scaffolds (0.6 ± 0.0 N, p < 0.05). A closed cell layer on PCL tissues did not change the values of Fmax (ECFCs: 0.6 ± 0.1 N; iMSCs: 0.7 ± 0.1 N). Here, a successful two-phase protocol, based on the timed use of differentiation factors for efficient differentiation of human iPSCs into iMSCs, was developed. Furthermore, we demonstrated the successful colonization of a biodegradable PCL nanofiber matrix with human ECFCs and iMSCs suitable for the generation of tissue-engineered heart valves. A closed cell cover was already evident after 14 days for ECFCs and 21 days for MSCs. The PCL tissue did not show major mechanical differences compared to native heart valves, which was not altered by short-term surface colonization with human cells in the absence of an extracellular matrix.

Optimization of Enzymatic and Chemical Decellularization of Native Porcine Heart Valves for the Generation of Decellularized Xenografts

“…Although a high SDS concentration has been reported to have a critical role in decellularization and in lowering DNA residues [32], the absence of SDS in our protocol suggests that SDS can easily be replaced by DNase and RNase for its effect on removing DNA. In this way, the SDS interactions with the ECM [19,21], its collagen integrity disruption, and ECM structure damages [32,33] have been prevented (Figures 4 and 5).…”

“…The nuclear components were entirely missing and there was no reduction in thickness visible in comparison to the native valves. For recell purposes, human induced-pluripotent stem cells-derived MSCs (hMSCs, described before by [19]) were used. The recellularized compound mostly showed fusiform cell structures with an extracellular matrix in-between, but also confluent cell layers on top.…”

“…The immunohistochemistry was performed for human-induced pluripotent stem cellsderived MSCs (hMSCs, described by Lutter et al in [19]). Antigen retrieval was attained through boiling the slides in citrate buffer (10 mM citric acid monohydrate (Merck, USA), 0.05% Tween (AppliChem, Darmstadt, Germany) in ddH 2 O), pH 6.…”

“…The samples were stretched in both the radial and circumferential directions at 5.0 mm/min until complete rupture. Young's modulus, F max , and elongation at break were recorded for these measurements as described before by [19].…”

In Vivo Evaluation of PCL Vascular Grafts Implanted in Rat Abdominal Aorta

“…Furthermore, there are other significant requirements that reduce the use of well-known polymers and the demand for tailored synthesis of new compounds. Mechanical behavior, durability, extensibility, imitation of a native vessel in the replacement area, a lack of cytotoxicity in the polymer and the products of its degradation, a balance between the resorption rate and vascular wall formation de novo, and many other features are crucial for a good clinical outcome [4][5][6][7].…”