An edited version will appear on the site on later this month.
Differentiated cells can be reset to an embryonic-stem-cell-like state, but doing so requires using retroviruses to insert a suite of genes into a culture of cells. Because the viruses insert their cargo at random into the genome, infected cell populations from the same individual are genetically different, and it’s hard to know whether differences between the resulting stem cell lines are due to this genetic variation, the epigenetic state of the original cells, or chance events. Worse, the current use of retroviruses renders the cells unusable for clinical applications.
Now researchers led by Rudolf Jaenisch at the Whitehead Institute in Cambridge Massachusetts show a convenient way to generate genetically identical cell populations that can be converted to induced pluripotent stem (iPS) cells by adding a drug . What’s more, they show that cells from multiple organs can be successfully reprogrammed.
In previous work, Jaenisch and other laboratories had reprogrammed cells using genes that turn on in the presence of a drug called doxycycline. This allowed them to study how long transgenes need to stay active for reprogramming to occur. To prove that the cells with drug-inducible reprogramming genes were pluripotent, researchers mixed the cells with mouse embryos to create chimeric mice.
In this paper, the researchers show that cells from multiple organs within these chimeric mice can be efficiently reprogrammed with the addition of doxycycline: reprogrammed cells include neural progenitors, mesenchymal stem cells, and keratinocytes as well as cells taken from muscle, intestinal epithelium, the adrenal gland, and the hematopoietic lineage.
iPS cells generated from different tissues taken from the same mouse are genetically identical, allowing researchers to examine the effects of cell types and retroviral insertion sites. For example, the researchers were able to reprogram intestinal epithelium derived from one iPS cell line, but not another, suggesting that reprogramming requirements vary between cell types. In particular, the expression of transgenes seemed to vary with both cell type and site of insertion in the genome.
The reprogramming rate for `secondary iPS cells’ was 4 to 8 fold higher than for the production of primary iPS cells, presumably because cells in the mice already had the favorable number of proviruses inserted at appropriate sites of the genomes. Still the overall reprogramming rate is low, only between 2% and 4%.
The researchers believe this could be because the drug-inducible transgenes may be more or less responsive to doxycycline even within genetically identical cells and also because reprogramming depends on stochastic events, several of which are required for complete reprogramming.
A source of genetically identical cells will help researchers home in on these and other variables and greatly simplify the search for methods to create clinically acceptable reprogrammed cells.
Wernig, M. et al. A drug-inducible transgenic system for direct reprogramming of multiple somatic cell types. Nat. Biotechnol. doi:10.1038/nbt1483 (Advance online publication July 1, 2008)