ARTICLE IN BRIEF
Two different research groups induced rodent fibroblasts to become oligodendrocyte precursor cells by inserting transcription factors that caused the cells to change their behavior and start producing myelin.
Two new papers lay out similar methods of transforming fibroblasts directly into oligodendrocyte precursor cells (OPCs) capable of myelinating axons in shiverer mice, bred to produce virtually no myelin of their own.
The work, published online April 14 ahead of the print edition of Nature Biotechnology, demonstrates the feasibility of transforming such cells without first turning the fibroblasts into pluripotent stem cells, which can become any cell in the body. Instead, by inserting genes for three transcription factors into the fibroblasts, both groups managed to induce the cells to become OPCs.
“You can cut out the middle man and go straight from a skin cell to a neuron or some other cell,” said Marius Wernig, MD, PhD, assistant professor of pathology in the Institute for Stem Cell Biology and Regenerative Medicine at the Stanford University School of Medicine, lead author of one of the papers.
The author of the other paper, Paul Tesar, PhD, called the work a form of alchemy. “We have readily accessible cell types akin to the common base metals, and we're reprogramming them into highly prized cell types akin to the noble metals, like gold,” said Dr. Tesar, assistant professor in the department of genetics and genome sciences at Case Western Reserve University School of Medicine, and a New York Stem Cell Foundation Robertson Investigator.
Both groups induced rodent fibroblasts to become OPCs by inserting transcription factors that caused the cells to change their behavior and start producing myelin. Both groups used Sox10, a transcription factor that promotes the encoding of a protein involved in the regulation of embryonic development, and Olig2, involved in motor neuron and oligodendrocyte differentiation.
For the third transcription factor, the Wernig group used zinc finger protein 536 (Zfp536), involved in brain cell differentiation, while the Tesar group opted for Nkx6 homeobox 2 (Nkx6.2), involved in myelination. The two transcription factors appear to work equally well, although Zfp536 enables the cells to give rise to both astrocytes and oligodendrocytes.
While both sets of transcription factors achieved the desired effect, both proved to be less efficient at proliferation than induced pluripotent stem cells.
“This is really hard to measure, but we found about a one-third to one-quarter drop of efficiency in the ability of cells to develop mature oligodendrocytes,” Dr. Wernig said. “When you take primary OPCs and put them together with primary neurons, about 60 percent turn into oligodendrocytes in two to three weeks. In our case only about 20 percent or so could do that. They can do their job in vivo and myelinate, but not as well as primary OPCs. That's something we really have to work on now — to improve the efficiency and the quality of the cells.”
Dr. Tesar emphasized that although induced OPCs may not be as efficient, producing a usable number of them will be much faster than it would be with induced pluripotent stem (iPS) cells. “Making iPS cells can take months,” he said. “Then growing the cells and characterizing them can take another couple of months. Then redifferentiating them into oligodendrocytes can take three to five months. Now we're talking on the order of possibly a year.”
The type of direct reprogramming he and Dr. Wernig use will cut that time to about two weeks. Also, transforming a patient's own cells reduces the danger of rejection and the need for immunosuppression.
The findings of both groups represent years of work.
Dr. Tesar and his colleagues, in a 2011 Nature Methods paper, described a platform for quickly differentiating mouse pluripotent stem cells into myelinating oligodendrocytes in vitro and in vivo.
The next step, both researchers agree, will be to perform the same feat on human fibroblasts.
The results reported by Drs. Wernig and Tesar amount to a “huge step forward” in terms of developing cells for transplantation, according to Bruce Trapp, PhD, chairman of the department of neurosciences at the Cleveland Clinic's Lerner Research Institute.
“As far as I know, this is the first description of generating induced oligodendrocyte progenitor cells, and because of that, these are significant observations,” he said. “They've shown it's easy to get these cells from some individual for transplant. If the disease isn't mediated by an inherited gene defect, then if you took fibroblasts from the individual, turned them into OPCs, and transplanted them, you're not at risk for rejection. When you use pluripotent cells from a fetus, that's not your genetic background, so you'll have to deal with the risk of rejection, which is much more complicated.”
The next step, he believes, will involve fine-tuning the transcription factors used.
“They may need factors that suppress the differentiation of the fibroblast genes or of the oligodendrocyte precursor cell,” Dr. Trapp said. “They'll probably need an enhancement of certain genes and the repression of others. They've focused here on enhancers only, and that's appropriate at this stage, but may have to fine-tune it.”
Steven Goldman, MD, PhD, Rykenboer professor of neurology and co-director of the Center for Translational Neuromedicine at the University of Rochester Medical Center, said that the major advantage to the technique developed by Drs. Wernig and Tesar is speed.
“These guys skipped the stem cell stage altogether, and have come up with a means of generating the OPCs directly from fibroblasts.”
Also, the transcription factor code that defines oligodendrocytes and allows reprogramming directly from somatic to oligodendroglial cells is an important insight, he added.
“It tells me that the different stages of development in the oligodendroglial lineage that we knew existed can be defined by different transcription factor codes,” he said. “That's important. The comparison of the two studies is every bit as revealing as the individual studies themselves.”
The downside to their procedure, according to Dr. Goldman, is that it doesn't produce large numbers of cells. “The oligodendrocyte progenitor doesn't have the same mitotic competence as the stem cell from which it's derived,” he said.
Translating the work to humans may also pose a challenge because the oligodendrocyte progenitor cell biology of rodents and humans is quite different. “The transcription factor combinations that work in rodents may not work in humans,” Dr. Goldman said. “So the direct reprogramming approach describes fascinating and important cell biology, but I don't think that it will go clinical any time soon, whereas the tissue-derived and iPS and ES cell-based strategies are already in the clinical pipeline.”
However, say Drs. Wernig and Tesar, their technique allows for much faster creation of cells, and improvements in technique could improve scalability. “It is likely that the optimization of the reprogramming efficiency could provide sufficient numbers of cells needed for clinical purposes,” Dr. Tesar said.
“Human embryonic stem cell and iPS cell-based and tissue-derived trials for remyelination are already in the pipeline, and will serve as a framework for potential direct reprogramming-based trials in the future,” Dr. Tesar said.
Dr. Trapp, of Cleveland Clinic, concluded: “Five or six years ago, when this started, it sounded like science fiction. They're blazing the trail for fibroblasts to be as efficient as stem cells. I would think within next five years they could be designing a clinical study with cells they generate.”
LINK UP FOR MORE INFORMATION:
•. Yang N, Zuchero JB, Wernig M, et al. Generation of oligodendroglial cells by direct lineage conversion. Nat Biotechnol
2013; E-pub 2013 Apr 14.
•. Najm FJ, Lager AM, Tesar PJ. Transcription factor–mediated reprogramming of fibroblasts to expandable, myelinogenic oligodendrocyte progenitor cells. Nat Biotechnol
2013;E-pub 2013 Apr 14.
•. Vierbuchen T, Ostermeier A, Wernig M, et al. Direct conversion of fibroblasts to functional neurons by defined factors. Nature
•. Marro S, Pang ZP, Wernig M, et al. Direct lineage conversion of terminally differentiated hepatocytes to functional neurons. Cell Stem Cell
•. Lujan E, Chanda S, Wernig M, et al. Direct conversion of mouse fibroblasts to self-renewing, tripotent neural precursor cells. Proc Nat Acad Sci
•. Najm FJ, Zaremba A, Tesar PJ, et al. Rapid and robust generation of functional oligodendrocyte progenitor cells from epiblast stem cells. Nat Methods
2011;8(11):957-1062Wang S, Bates J, Li X, Schanz S, Chandler-Militello D, Goldman SA, et al. Human iPSC-derived oligodendrocyte progenitor cells can myelinate and rescue a mouse model of congenital hypomyelination. Cell Stem Cell