Tissue engineering of functional trileaflet heart valves from human marrow stromal cells.
BACKGROUND
We previously demonstrated the successful tissue engineering and implantation of functioning autologous heart valves based on vascular-derived cells. Human marrow stromal cells (MSC) exhibit the potential to differentiate into multiple cell-lineages and can be easily obtained clinically. The feasibility of creating tissue engineered heart valves (TEHV) from MSC as an alternative cell source, and the impact of a biomimetic in vitro environment on tissue differentiation was investigated.


RESULTS
Human MSC were isolated, expanded in culture, and characterized by flow-cytometry and immunohistochemistry. Trileaflet heart valves fabricated from rapidly bioabsorbable polymers were seeded with MSC and grown in vitro in a pulsatile-flow-bioreactor. Morphological characterization included histology and electron microscopy (EM). Extracellular matrix (ECM)-formation was analyzed by immunohistochemistry, ECM protein content (collagen, glycosaminoglycan) and cell proliferation (DNA) were biochemically quantified. Biomechanical evaluation was performed using Instron(TM). In all valves synchronous opening and closing was observed in the bioreactor. Flow-cytometry of MSC pre-seeding was positive for ASMA, vimentin, negative for CD 31, LDL, CD 14. Histology of the TEHV-leaflets demonstrated viable tissue and ECM formation. EM demonstrated cell elements typical of viable, secretionally active myofibroblasts (actin/myosin filaments, collagen fibrils, elastin) and confluent, homogenous tissue surfaces. Collagen types I, III, ASMA, and vimentin were detected in the TEHV-leaflets. Mechanical properties of the TEHV-leaflets were comparable to native tissue.


CONCLUSIONS
Generation of functional TEHV from human MSC was feasible utilizing a biomimetic in vitro environment. The neo-tissue showed morphological features and mechanical properties of human native-heart-valve tissue. The human MSC demonstrated characteristics of myofibroblast differentiation.
D020341:Absorbable Implants; D000328:Adult; D019149:Bioreactors; D001854:Bone Marrow Cells / Q000166:cytology*; D002454:Cell Differentiation; D002478:Cells, Cultured; D005109:Extracellular Matrix / Q000737:chemistry; D005240:Feasibility Studies; D005434:Flow Cytometry; D006350:Heart Valve Prosthesis; D006351:Heart Valves / Q000033:anatomy & histology / Q000166:cytology / Q000502:physiology / Q000648:ultrastructure; D006801:Humans; D007150:Immunohistochemistry; D008855:Microscopy, Electron, Scanning; D017154:Stromal Cells / Q000737:chemistry / Q000166:cytology*; D013718:Tensile Strength; D023822:Tissue Engineering / Q000379:methods*