Date of Award
Doctor of Philosophy (PhD)
Dan T. Simionescu
Background: Each year in the U.S., an estimated minimum of 40,000 infants are expected to be affected with some form of congenital heart defect (CHD), while a quarter of infants affected with CHD will require invasive treatment within the first year. The current treatments for CHD that involve valve intervention are not tailored to growing children. Mechanical and biological substitute of valves deteriorate and accumulate calcification when implanted in very young patients. In addition, children with current valve substitutes are faced with a lifetime of drug regimens, such as anti-coagulants, to prolong the longevity of the replacements and multiple surgeries for valve implantation. Long-term goal: Our overarching aim is to create a tissue engineered pulmonary valve (TEPV) as a living replacement, which addresses the short comings of current treatments. A TEPV approach for children with CHD would create a living scaffold with the potential of integrating into its environment, regenerating, adapting, and growing in parallel with the somatic growth of the child. This will alleviate the need for any additional surgeries throughout their life. Approach: To achieve the goal of creating a living TEPV, we proposed combining non-immunogenic valve scaffolds and autologous endothelial cells obtained from human adipose derived stem cells (hADSCs) by in vitro differentiation. This is only a first step in our larger project to seed the valve interstitium with autologous fibroblasts and the external surfaces with endothelial cells (ECs). The acellular scaffolds were obtained via decellularization of porcine pulmonary valves using a pressurized perfusion system. Biochemical assays and histology tests validated decellularization completeness and bioreactor studies confirmed valve hemodynamics. hADSCs were differentiated in vitro towards ECs using added growth factors and dynamic shear conditioning. After propagation, to optimize cell seeding on both surfaces of the valve scaffolds (atrial and ventricular), we used fibrin gel as a delivery vehicle and compared manual seeding with 3D bioprinting as a means of EC seeding and evaluated cell retention and viability using Live/Dead assays. After seeding, we exposed the seeded valves to physiological environments of pulmonary flow and pressures within a valve bioreactor for conditioning and maturation of the endothelium. Results: Our data show complete pulmonary valve decellularization in our system, including the sinus and the ascending pulmonary artery, efficient differentiation of hADSCs into ECs using growth factors and shear, improved seeding using the 3D bioprinter and adequate coverage of ECs on both surfaces of the valves after bioreactor conditioning. This initial success opens possibilities for complete cell repopulation in vitro with autologous cells before implantation.
Correction. Circulation 133, e599 (2016). Schoen, F.J. & Levy, R.J. Calcification of tissue heart valve substitutes: progress toward understanding and prevention. Ann Thorac Surg 79, 1072-1080 (2005). Cebotari, S., et al. Use of fresh decellularized allografts for pulmonary valve replacement may reduce the reoperation rate in children and young adults: early report. Circulation 124, S115-123 (2011).
Bruce, Maria Margarita Rodriguez, "Development of a Tissue Engineered Pulmonary Heart Valve for Pediatric Patients" (2021). All Dissertations. 2808.