Extended bioreactor conditioning of mononuclear cell-seeded heart valve scaffolds

The tissue-engineered heart valve may be the ideal valve replacement option but still must overcome challenges in leaflet recellularization. This study sought to investigate the potential for leaflet matrix restoration and repopulation following mononuclear cell seeding and extended periods of bioreactor conditioning. Human aortic heart valves were seeded with mononuclear cells and conditioned in a pulsatile bioreactor for 3 days, 3 weeks, or 6 weeks. The results of this study determined that a mononuclear cell population can be readily localized within the leaflet tissue in as little as 3 days. Furthermore, as extended bioreactor condition continued to the 3- and 6-week time points, the mesenchymal stem cell subfraction proliferated and appeared to become the predominant cell phenotype. This was evident through positive expression of mesenchymal stem cell markers and no expression of mononuclear cell markers observed by immunohistochemistry in the 3- and 6-week groups. In addition, cells in the 3- and 6-week groups exhibited an up-regulation of mesenchymal stem cell-associated genes (THY1, NT5E, and ITGB1) and a down-regulation of mononuclear cell-associated genes (CD14, ICAM1, and PECAM1) compared to the initial seeded cell population. However, repopulation of the leaflet interstitium was less extensive than anticipated. Valves in the 6-week time point also exhibited retracted leaflets. Thus, while the 3-week bioreactor-conditioning period used in this study may hold some promise, a bioreactor-conditioning period of 6 weeks is not a viable option for clinical translation due to the negative impact on valve performance.