oa Reproducing human heart valves anisotropy with a nanotechnology approach

Topology and organization of cell environment is an important regulator of differentiation and function. In view of heart valve tissue engineering, we devised a biomimetic scaffold that supports proper cell growth and cell-matrix interactions by reproducing the specific anisotropic fibrillar structure of valves extracellular matrix (ECM). We hypotheseized that high porosity levels of novel nanofibrillar matrices allow extensive cell colonization while nanofiber anisotropy influences cell phenotype. Iso- and anisotropic nanofibrillar structures were obtained with an innovative jet-spraying method that allows the formation polymer nanofibers by the air-stream diffraction of a poly (ε-caprolactone) solution in chloroform. Human adipose derived stem cells (hADSCs) and human valve interstitial cells (hVICs) were rotary seeded (600000 and 500000 cells respectively) on nanofibrillar discs and cultured for 20 days in 10 ml of complete medium under rotation (10 rpm). Isotropic and well aligned nanofibers (anisotropic, 600 nm diameter) were obtained from sprayed polymer (Fig. 1-A). Valve mechanical anisotropy was mimicked by scaffold elasticity, which was increased longitudinally to fiber alignment (from 0.3 to 0.7 Mpa) and decreased orthogonally (from 0.3 to 0.01 Mpa). Further fine-tuning of scaffolds mechanical anisotropy could be obtained by orthogonally associating anisotropic layers of different thickness. Iso- and anisotropic matrices allowed an extensive cellular proliferation (6-fold DNA increase) regardless of cell type. Although cells were more homogeneously distributed within anisotropic scaffolds after seeding, they populated the entire scaffold thickness within 10 days (Fig.1-B) and produced their own ECM (collagen I, III and elastin) regardless of scaffold architecture. Anisotropy induced elongated hADSCs morphology that followed nanofibers' main orientation (Fig. 1-C). hVICS followed the same nanofibers-induced orientation pattern when cultured in anisotropic scaffolds. In conclusion, highly porous anisotropic matrices reproducing valve ECM structure are excellent substrate for cell population while providing an efficient tool to study cells/environment interaction for producing engineered cardiovascular tissues.