Assessment of the ultrastructural and proliferative properties of human embryonic stem cell-derived cardiomyocytes

作者：Mirit Snir, Izhak Kehat, Amira Gepstein, Raymond Coleman, Joseph Itskovitz‐Eldor, Erella Livne, Lior Gepstein · 发表于：American Journal of Physiology-Heart and Circulatory Physiology · 年份：2003 · DOI：10.1152/ajpheart.00020.2003 · 被引用次数：336 · 研究领域：Pluripotent Stem Cells Research、Tissue Engineering and Regenerative Medicine、Congenital heart defects research

Assessment of early ultrastructural development and cell-cycle regulation in human cardiac tissue is significantly hampered by the lack of a suitable in vitro model. Here we describe the possible utilization of human embryonic stem cell (ES) lines for investigation of these processes. With the use of the embryoid body (EB) differentiation system, human ES cell-derived cardiomyocytes at different developmental stages were isolated and their histomorphometric, ultrastructural, and proliferative properties were characterized. Histomorphometric analysis revealed an increase in cell length, area, and length-to-width ratio in late-stage EBs (>35 days) compared with early (10-21 days) and intermediate (21-35 days) stages. This was coupled with a progressive ultrastructural development from an irregular myofibrillar distribution to an organized sarcomeric pattern. Cardiomyocyte proliferation, assessed by double labeling with cardiac-specific antibodies and either [3H]thymidine incorporation or Ki-67 immunolabeling, demonstrated a gradual withdrawal from cell cycle. Hence, the percentage of positively stained nuclei in early-stage cardiomyocytes ([3H]thymidine: 60 +/- 10%, Ki-67: 54 +/- 23%) decreased to 36 +/- 7% and 9 +/- 16% in intermediate-stage EBs and to <1% in late-stage cardiomyocytes. In conclusion, a reproducible temporal pattern of early cardiomyocyte proliferation, cell-cycle withdrawal, and ultrastructural maturation was noted in this model. Establishment of this unique i...