The Niche

More induced pluirpotency; Yamanaka’s work reconfirmed

An independent laboratory has been able to reprogram differentiated human skin cells to an embryonic-like state using the method originally reported by Shinya Yamanaka of Kyoto University.1,2 Also late last year, two other laboratories announced reprogramming differentiated human cells via slightly different methods.3,4 The latest work, by Kathrin Plath of the University of California, Los Angeles, indicates that the technique is broadly feasible and introduces a potentially easier method to pluck reprogrammed cells from culture.5

Notably, all the groups that have reported success with this method have experience working with embryonic stem cells. At least two have derived human embryonic stem cells, and the genes that must be introduced to reprogram cells were identified through embryonic stem cells.

Meanwhile, the California Institute of Regenerative Medicine announced that it has received 50 applications for up to 16 grants totaling $25 million for creating new human pluripotent cell lines either from embryos from fertility clinics or from other sources of cells. In an executive order and his state of the union address, President Bush has directed the NIH to direct more funds to methods to create pluripotent stem cell lines without destroying embryos, but no dedicated funding programs have yet been announced.

The scientist leading the work just published in the Proceedings of the National Academy of Sciences has received a $1.5 million NIH grant set aside for “exceptionally innovative investigators.” Only 41 such grants were awarded, less than one of 50 applicants including biomedical researchers across the nation. She also received a $2.2 million grant from CIRM to study how reprogramming works in mice. Two other scientists who have successfully reprogrammed human cells (Yamanaka and James Thomson) have part-time appointments in California that will allow them to access some funds from the CIRM Medicine, but Plath is the first full-time faculty California to lead such an effort.

Plath’s team infected cultured skin cells originally collected from circumcisions and infected them with retroviruses carrying the four genes originally identified in reprogramming work. After about two weeks they began to see colonies proliferating. They looked clearly different from the cultured skin cells, but they did not go on to become induced pluripotent cells, and analysis showed that they had not taken in copies of all four genes. Colonies of cells destined to become reprogrammed lines showed up about 21 days after infection; they clustered together as human embryonic stem cells do, and they also displayed a variety of cell-surface markers characteristic of ES cells. A week later, Plath and researchers hand-selected colonies staining positive for one particular cell-surface marker. All cells analyzed from these colonies contained copies of all four genes.

Of the 30 colonies isolated, cells from seven were closely analyzed. As expected, all of these cells were expressing endogenous genes associated with pluripotency, and the viral genes were silenced. Though the cells express markers of the three main types of tissues forming the body, the team has not yet tested the functionality of the cells by differentiating them into teratomas or other cell types.

1. Takahashi K. et al. Induction of pluripotent stem cells from adult human fibroblasts by defined factors. Cell advance online publication 20 November 2007. doi: 10.1016/j.cell.2007.11.019 | Article |

2. Takahashi K. & Yamanaka S. Induction of pluripotent stem cells from mouse embryonic and adult fibroblast cultures by defined factors. Cell 126, 663–76 (2006) | Article | PubMed | ISI | ChemPort |

3. Yu, J. et al. Induced pluripotent stem cell lines derived from human somatic cells. Science advance online publication 20 November 2007. doi: 10.1126/science.1151526 | Article |

4. Park, I. H. et al. Reprogramming of human somatic cells to pluripotency with defined factors. Nature advance online publication, doi:doi: 10.1038/nature06534 (23 December 2007).

5. Lowry, W.E. et al. Generation of human-induced pluripotent cells from dermal fibroblasts. Advance online publication www.pnas.org_cgi_doi_10.1073_pnas.0711983105 11 February 2008

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