Some of the most challenging obstacles limiting the reprogramming of mature human cells into stem cells may not seem quite as daunting in the near future. Two independent research groups, one lead by Harvard Stem Cell Institute Principal Faculty member Konrad Hochedlinger, and another at the Whitehead Institute, are describing new tools that provide invaluable platforms for elucidating the molecular, genetic and biochemical mechanisms associated with reprogramming. The work of both groups is being published in the September 11th issue of the journal Cell Stem Cell.
Although scientists have successfully reprogrammed mature human skin cells into induced pluripotent stem cells (iPS) by expressing a few key transcription factors, the conversion has been extremely inefficient. “Little is known about the mechanisms by which reprogramming occurs, in part because of the low efficiency,” says Hochedlinger. In addition, the iPS cells created thus far have been generated with retroviruses and non-inducible lentiviruses, both of which have limitations that may limit clinical applications.
The Hochedlinger group created a drug-inducible lentiviral system to generate human iPS cells that were molecularly and functionally similar to human embryonic stem cells. The researchers found that once the “primary” iPS cells differentiated to mature cells, addition of the drug doxycycline reactivated the genes required for reprogramming and induced generation of “secondary” iPS cells at a frequency that was far greater that the initial “primary” conversion.
“The secondary system will enable chemical and genetic screening efforts to identify key molecular constituents of reprogramming, as well as important obstacles in this process, and will ultimately lend itself as a powerful tool in the development and optimization methods to produce human iPS cells,” explains Hochedlinger, an assistant professor in Harvard’s Department of Stem Cell and Regenerative Biology.
A separate team of scientists, led by Rudolf Jaenisch of the Whitehead Institute and MIT, had parallel success in deriving human secondary iPS cells using doxycycline-inducible transgenes. “The drug-inducible system we describe represents a novel, predictable, and highly reproducible platform to study the kinetics of iPS cell generation,” says Dr. Jaenisch.
Both research teams found that generation of secondary human iPS cells required less time than the initial reprogramming. Interestingly, the time required to generate iPS cells varied among the types of skin cells that were used. For instance, human fibroblasts required several weeks while keratinocytes required only about ten days. “The fast kinetics of reprogramming observed for keratinocytes suggests that these cells would be useful for development and optimization of methods to reprogram cells by transient delivery of factors,” suggests Hochedlinger.
The combined results from both research groups represent a major advance towards more efficient strategies for reprogramming differentiated human cells into iPS cells. In addition, the methods described here will provide critical insight into the reprogramming process and are likely to lead to the generation of cells that will be amenable for future clinical applications.
HSCI’s Chad Cowan, a member of the Principal Faculty, an Assistant Professor in Harvard’s Department of Stem Cell and Regenerative Biology, and a Stowers Medical Institute Investigator, was a member of the Hochedlinger team.