Stacking of aligned cell sheets for layer-by-layer control of complex tissue structure.

Children suffering from congenital heart defects (CHD) often require vascular reconstruction. Pediatric patients would greatly benefit from a cell-based tissue engineered vascular patch (TEVP) that has potential for growth. As artery structure and function are intimately linked, mimicking native tissue organization is an important design consideration. In this study, we cultured human mesenchymal stem cell on patterned thermo-responsive substrates. Cell alignment improved over time up to 2 wk in culture when sheets were ready for harvest. We then used cell sheets as "functional units" to build complex tissue structures that mimic native vascular smooth muscle cell organization in the medial layer of the artery. Cell sheets could be stacked using a gelatin stamp such that individual sheets in the construct were well aligned with each other (mimic of circumferential orientation) or at angles with respect to each other (mimic of herringbone structure). Controlling tissue organization layer-by-layer will be a powerful approach to building tissues with well defined and complex structure.