Selective conversion of fibroblasts into peripheral sensory neurons

Abstract

Humans and mice detect pain, itch, temperature, pressure, stretch and limb position via signaling from peripheral sensory neurons. These neurons are divided into three functional classes (nociceptors/pruritoceptors, mechanoreceptors and proprioceptors) that are distinguished by their selective expression of TrkA, TrkB or TrkC receptors, respectively. We found that transiently coexpressing Brn3a with either Ngn1 or Ngn2 selectively reprogrammed human and mouse fibroblasts to acquire key properties of these three classes of sensory neurons. These induced sensory neurons (iSNs) were electrically active, exhibited distinct sensory neuron morphologies and matched the characteristic gene expression patterns of endogenous sensory neurons, including selective expression of Trk receptors. In addition, we found that calcium-imaging assays could identify subsets of iSNs that selectively responded to diverse ligands known to activate itch- and pain-sensing neurons. These results offer a simple and rapid means for producing genetically diverse human sensory neurons suitable for drug screening and mechanistic studies.

More than one way to induce a neuron

“…Previous work by the group that performed the current study showed that a pair of transcription factors from the basic helix-loophelix (bHLH) and Pit-Oct-Unc (POU) families can induce the expression of neuronal markers through direct reprogramming 6 . Tsunemoto et al were inspired by this finding in their current work.…”

Direct neuronal reprogramming: learning from and for development

“…Indeed, this approach has been used for inducing many other neuronal subtypes lost in traumatic injury or neurodegenerative diseases, and generation of which in vitro could be potentially useful for cellbased therapies as well as for disease modelling (Blanchard et al, 2015;Caiazzo et al, 2011;Gascón et al, 2016;Rouaux and Arlotta, 2013;Son et al, 2011;Victor et al, 2014;Wainger et al, 2015). Especially for the latter, direct reprogramming has the advantage of maintaining the age of the starter cell (Mertens et al, 2015), in contrast to the re-setting that occurs when generating induced pluripotent stem cells (Lapasset et al, 2011).…”

“…From a developmental perspective, these neurons differentiate from neural crest cells (Pavan and Raible, 2012), and hence a promising approach for generating them has been to reprogramme human fibroblasts into neural crest cells, which can then be further differentiated into peripheral neurons . However, direct neuronal reprogramming has proven to be more successful in obtaining specific subtypes of peripheral neurons than the neural crest cell route (Blanchard et al, 2015;Wainger et al, 2015). As peripheral sensory neurons are different to those of the central nervous system, an important question is how similar or different the molecular requirements are for inducing these cells.…”

“…A second pool of factors, Neurog1/2 and Pou4f1 (also known as Brn3), was successful in converting fibroblasts into sensory neurons, albeit with a low efficiency (4-6%; Blanchard et al, 2015). Single-cell expression analysis revealed non-overlapping expression of TrkA, TrkB (Ntrk2) and TrkC (Ntrk3), suggesting that the reprogramming protocol generated different sensory subtypes.…”

Direct somatic lineage conversion

“…Two groups recently reported exciting findings that suggest that mouse and human fibroblasts can be converted to iN cells with even such a high degree of subspecialization as the three main classes of peripheral sensory neurons, including nociceptive neurons that would be highly relevant for pain research [59,60]. The Baldwin group found that just two transcription factors, either Ngn1 or Ngn2, in combination with Brn3a, a critical lineage determination factor for peripheral neurons, was sufficient to convert both mouse primary and human iPS cell-derived fibroblasts to iN cells with peripheral identity representing the three major classes of sensory neurons: nociceptive, mechanosensitive and proprioceptive, which express the key specific markers TrkA, B and C, respectively [59]. The neurons exhibited a typical pseudounipolar morphology, expressed several additional markers of sensory neurons and responded to appropriate stimuli like capsaicin, menthol and mustard oil.…”