Selection of viable cardiomyocytes for cell transplantation using three-dimensional tissue culture

Background We developed a three-dimensional cardiac tissue culture system t o select viable and contractile cells for the purpose of cardiomyocyte tran splantation. In this study we will show that reaggregation of cardiomyocyte s in culture is an active process indicative of cardiomyocyte viability and functionality. Methods. Myocardial tissue from newborn mice has been enzymatically digeste d, incubated in culture inserts, and studied by phase contrast microscopy, conventional histology, immunohistochemistry, electron microscopy, and TUNE L assay. Results. Cells that are plated on nonadhesive surfaces, reaggregate to spon taneously contracting cell aggregates. The capacity to reaggregate was sign ificantly dependent from the age of the tissue donor (P < 0.0001) and on th e method of enzymatic dissociation (P < 0,0001), The majority of cells with in the aggregates consisted of cardiomyocytes, After 24 hr incubation, sign ificant amounts of laminin and fibronectin had been deposited between the c ells. Ultrastructural analysis revealed viable cardiomyocytes attached to e ach other by tight junctions. The apoptotic rate within the aggregates was 11.4 +/- 4.6 vs. 44.5 +/- 10.5% immediately after dissociation (P < 0.05). Conclusions. The capacity to form spontaneously contracting aggregates is a n inherent characteristic of viable cardiomyocytes in 3-dimensional culture s, which could be successfully exploited for cellular cardiomyocyte transpl antation.