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Previous research in our lab suggested that heart valve tissues cultured without mechanical stimulation do not retain their in vivo microstructure, i.e., cell density decreased within the deep tissue layers and increased at the periphery. In this study, a splashing rotating bioreactor was designed to apply mechanical stimulation to a mitral valve leaflet segment. Porcine valve segments (n = 9-10 per group) were cultured in the bioreactor for 2 weeks (dynamic culture), negative controls were cultured without mechanical stimulation (static culture), and baseline controls were fresh uncultured samples. Overall changes in cellularity and extracellular matrix (ECM) structure were assessed by H&amp;E and Movat pentachrome stains. Tissues were also immunostained for multiple ECM components and turnover mediators. After 2 weeks of culture, proliferating cells were distributed throughout the tissue in segments cultured in the bioreactor, in contrast to segments cultured without mechanical stimulation. Most ECM components, especially collagen types I and III, better maintained normal expression patterns and magnitudes (as found in baseline controls) over 2 weeks of dynamic organ culture compared to static culture. Lack of mechanical stimulation changed several aspects of the tissue microstructure, including the cell distribution and ECM locations. In conclusion, mechanical stimulation by the bioreactor maintained tissue integrity, which will enable future in vitro investigation of mitral valve remodeling.

Design and Validation of a Novel Splashing Bioreactor System for Use in Mitral Valve Organ Culture