Anisotropicity and flexibility in trilayered microfibrous substrates promote heart valve leaflet tissue engineering

Heart valve leaflet substrates with native trilayer and anisotropic structures are crucial for successful heart valve tissue engineering. In this study, we used the electrospinning technique to produce trilayer microfibrous leaflet substrates using two biocompatible and biodegradable polymers-poly (L-lactic acid) (PLLA) and polycaprolactone (PCL), separately. Different polymer concentrations for each layer were applied to bring a high degree of mechanical and structural anisotropy to the substrates. PCL leaflet substrates exhibited lower unidirectional tensile properties than PLLA leaflet substrates. However, the PLLA substrates exhibited a lower flexural modulus than the PCL substrates. These substrates were seeded with porcine valvular interstitial cells (PVICs) and cultured for one month in static conditions. Both substrates exhibited cellular adhesion and proliferation, resulting in the production of tissue-engineered constructs. The PLLA tissue-engineered constructs had more cellular growth than the PCL tissue-engineered constructs. The PLLA substrates showed higher hydrophilicity, lower crystallinity, and more significant anisotropy than PCL substrates, which may have enhanced their interactions with PVICs. Analysis of gene expression showed higherα-smooth muscle actin and collagen type 1 expression in PLLA tissue-engineered constructs than in PCL tissue-engineered constructs. The differences in anisotropic and flexural properties may have accounted for the different cellular behaviors in these two individual polymer substrates.