Clonally derived human embryonic stem cell lines maintain pluripotency andproliferative potential for prolonged periods of culture

Embryonic stem (ES) cell lines derived from human blastocysts have the deve lopmental potential to form derivatives of all three embryonic germ layers even after prolonged culture. Here we describe the clonal derivation of two human ES cell lines, H9.1 and H9.2. At the time of the clonal derivation o f the H9.1 and H9.2 ES cell lines, the parental ES cell line, Ho, had alrea dy been continuously cultured for 6 months. After an additional 8 months of culture, H9.1 and H9.2 ES cell lines continued to: (1) actively proliferat e, (2) express high levels of telomerase, and (3) retain normal karyotypes. Telomere lengths, while somewhat variable, were maintained between 8 and 1 2 kb in high-passage H9.1 and H9.2 cells. High-passage H9.1 and H9.2 cells both formed teratomas in SCUD-beige mice that included differentiated deriv atives of all three embryonic germ layers. These results demonstrate the pl uripotency of single human ES cells, the maintenance of pluripotency during an extended period of culture, and the long-term self-renewing properties of cultured human ES cells. The remarkable developmental potential, prolife rative capacity, and karyotypic stability of human ES cells distinguish the m from adult cells. (C) 2000 Academic Press.