Low density culture of dissociated human ES cells in the absence (a) and presence (b) of 10 μM Y-27632.

Unlike mouse ES cells, human ES cells are virtually impossible to grow as dissociated cells in suspension. That makes it difficult to clonally isolate cells, which researchers want to do in order to assess how cells are differentiating or to probe the effects of gene transfer or homologous recombination. Watanabe et al set out to find inhibitors of cellular signaling molecules that would help dissociated cells survive. This June they report in Nature Biotechnology that a Rho-associated kinase (ROCK) inhibitor Y-27632 had the most potent effect. Treatment with 10 micromolar ROCK inhibitors allowed growth of completely dissociated hES cells, as well as individual hES cells seeded into 96-well plates. Dissociated cells treated with the ROCK inhibitor retained their pluripotency, and could be induced to differentiate into different cell types and form teratomas in mice. The data suggests that ROCK inhibitors promote cell survival by acting on the apoptosis pathway. However, understanding just how they do so awaits further study. ROCK inhibitors such as Y-27632 and Fasudil are already in clinical use for cardiovascular disease, so should be compatible with protocols to use hES cells for human therapies.