Modeling pain in vitro using nociceptor neurons reprogrammed from fibroblasts

Abstract

Reprogramming somatic cells from one cell fate to another can generate specific neurons suitable for disease modeling. To maximize the utility of patient-derived neurons, they must model not only disease-relevant cell classes but also the diversity of neuronal subtypes found in vivo and the pathophysiological changes that underlie specific clinical diseases. Here, we identify five transcription factors that reprogram mouse and human fibroblasts into noxious stimulus-detecting (nociceptor) neurons that recapitulate the expression of quintessential nociceptor-specific functional receptors and channels found in adult mouse nociceptor neurons as well as native subtype diversity. Moreover, the derived nociceptor neurons exhibit TrpV1 sensitization to the inflammatory mediator prostaglandin E2 and the chemotherapeutic drug oxaliplatin, modeling the inherent mechanisms underlying inflammatory pain hypersensitivity and painful chemotherapy-induced neuropathy. Using fibroblasts from patients with familial dysautonomia (hereditary sensory and autonomic neuropathy type III), we show that the technique can reveal novel aspects of human disease phenotypes in vitro.

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.…”

“…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. Interestingly, the same pan-neurogenic factors employed to generate central nervous system neurons -namely Ascl1, Myt1l and Neurog1 -are sufficient to induce an A-δ nociceptor phenotype in 14% of transduced cells when combined with the terminal differentiation factors Isl1 and Klf7 (Wainger et al, 2015). These latter two factors, Isl1 and Klf7, are important in maintaining the expression of TrkA (Ntrk1) in peripheral sensory neurons (Marmigere and Ernfors, 2007).…”

“…These latter two factors, Isl1 and Klf7, are important in maintaining the expression of TrkA (Ntrk1) in peripheral sensory neurons (Marmigere and Ernfors, 2007). Of the resulting induced nociceptor cells, about half were functional, as evidenced by their response to capsaicin stimulation and sensitization by prostaglandin E2 (Wainger et al, 2015).…”

Direct somatic lineage conversion

“…Therefore, access to authentic human peripheral sensory neurons represents an important scientific goal with potentially large clinical implications. 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].…”