If you want to study human neurons, where can you get them? One promising approach is reprogramming, where skin cells are converted into induced pluripotent stem (iPS) cells, which are then differentiated to neuronal cells. Last year, Marius Wernig and his colleagues at Stanford University School of Medicine in Palo Alto, California streamlined that strategy by skipping the iPS step and converting fibroblasts directly into neurons. That, however, was in mice. Today, Wernig’s group reports in Nature that they have created functioning neuronal cells directly from human postnatal fibroblasts. Their technique may simplify the production of patient-specific neurons, which have potential uses in in vitro disease modeling, drug screening, and regenerative medicine.
Using mouse cells, the researchers had shown that lentivirus-based expression of three transcription factors converted fibroblasts into neuronal cells (Vierbuchen et al., 2010; also see news story at the Alzheimer Research Forum). They now report that just one additional transcription factor, NeuroD1, is needed to create human induced neuronal (iN) cells with mature neuronal morphology, gene expression, and action potential firing.
When plated with mouse cortical neurons, the cells showed post-synaptic currents, indicating the presence of functional synapses. On other measures, however, the neurons appeared relatively immature, and more work will be needed to create fully differentiated neurons of various types.
Made-to-order neurons like these could offer powerful models for neurological disease, including a wide range of pain conditions, and serve as much-needed platforms for drug discovery. In a review in Neuron, also published today, Kevin Eggan and colleagues at Harvard University, Cambridge, Massachusetts describe results so far using neuronal cells generated from iPS cells. They highlight the pressing need for human neurons, because results in animal models are failing again and again to translate into humans—a situation well known to pain researchers.
Comments
Clifford Woolf , Children's Hospital Boston
There are rare moments in science when the ground suddenly shifts, a new discovery changes everything, nothing will be the same again. This is I believe true for the work of Marius Wernig and his colleagues at Stanford, first with their discovery published last year in Nature (Vierbuchen et al., 2010) that it was possible to directly change cell fate of mouse embryonic fibroblasts into neurons with just three transcription factors without going back first to a pluripotential stem cell, and now even more so with the paper of Pang et al which shows that an additional transcription factor can convert postnatal non-neural human somatic cells into induced neurons.
Why is this so important? Because it offers a way of generating patient-specific human neurons suitable for modeling disease or screening for new therapies. At the moment Wernig has only made a “generic” neuron whose phenotype is definitely neuronal. They have the morphology, membrane excitability properties and synaptic properties of neurons, a major achievement, but have not yet been able to specify a particular neuron of a defined cell fate. This further refinement is just around the corner – watch this space. Imagine using this technology to make nociceptors from somatic cells from a patient with primary erythromelalgia due to a gain of function mutation in Nav1.7. channelopathy. Imagine making nociciceptors from a patient with painful peripheral diabetic neuropathy and finding a phenotypic characteristic reflecting disease susceptibility. We are truly on the brink of transformative opportunities for neurobiology in general and most certainly the pain field. From a cell biological perspective it is to me at least, truly amazing to think each of our somatic cells is just a small handful of transcription factors away from potentially becoming a completely different and fully differentiated cell type. Changing human postnatal foreskin fibroblasts into action potential firing neurons is neither magic nor alchemy, this simply is modern science at its very best and most exciting.
Megan Talkington, Pain Research Forum
A confirmation, and more progress, on the iN front
This week, a second group, led by Malin Parmar at Lund University, Sweden, reported findings similar to those of Pang et al.: direct conversion of human postnatal fibroblasts into neurons. Their paper was published online June 6 in PNAS (Pfisterer et al., 2011). Additionally, Parmar and coworkers showed that adding two more genes to the original set of three allowed them to convert human embryonic fibroblasts directly into dopaminergic neurons. The authors say their results offer proof of principle that other neuronal subtypes should be within reach.
PRF News Editor, Harvard NeuroDiscovery Center
More on reprogramming of fibroblasts to neurons: microRNAs join the mix, and a short cut to dopaminergic neurons
In a paper published online July 13 in Nature, Gerald Crabtree and colleagues from Stanford University, California, report that a pair of microRNAs (miRNAs) was able to transform human fibroblasts to neurons. Until now, reprogramming of somatic cells like fibroblasts has been accomplished by the forced expression of transcription factors, but this study shows that developmental programs directed by miRNAs can also be tapped to drive cells to new fates. The miRNAs were not very efficient at reprogramming the cells, but adding a trio of neurogenic transcription factors led to more, and more mature, neurons. MicroRNAs add another tool to the box for researchers trying to generate neurons from somatic cells. Combining miRNAs with different sets of transcription factors may be one strategy for generating varieties of neurons, the authors suggest.
In another recent report, Italian researchers tackled the problem of generating specific types of neurons. Vania Broccoli of the San Raffaele Scientific Institute, Milan, and Alexander Dityatev, Istituto Italiano di Technologia, Genoa, and colleagues converted mouse or human fibroblasts to dopaminergic (DA) neurons using three transcription factors previously implicated in normal DA cell development. The researchers found that they could generate cells that had the gene expression profile and electrical activity of mature DA neurons, and released dopamine. The conversion appeared to be direct—the investigators saw no sign of any intermediate progenitor-type cells. The mouse cells could be transplanted and generate functional DA neurons in vivo. Previously, a similar fibroblast to DA neuron conversion was reported in embryonic and postnatal fibroblasts, but using five factors (Pfisterer et al, 2011 and see previous comment on this post). Broccoli and coworkers thus simplified the procedure, and showed they could convert adult fibroblasts as well. Their paper appeared July 3 in Nature online.
References:
MicroRNA-mediated conversion of human fibroblasts to neurons.
Yoo AS, Sun AX, Li L, Shcheglovitov A, Portmann T, Li Y, Lee-Messer C, Dolmetsch RE, Tsien RW, Crabtree GR.
Nature. 2011 Jul 13. [Epub ahead of print]
Direct generation of functional dopaminergic neurons from mouse and human fibroblasts.
Caiazzo M, Dell'anno MT, Dvoretskova E, Lazarevic D, Taverna S, Leo D, Sotnikova TD, Menegon A, Roncaglia P, Colciago G, Russo G, Carninci P, Pezzoli G, Gainetdinov RR, Gustincich S, Dityatev A, Broccoli V.
Nature. 2011 Jul 3. [Epub ahead of print]