Generation of Human Nociceptor-Enriched Sensory Neurons for the Study of Pain-Related Dysfunctions

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2022
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Wohlfarth, Harald
Loser, Dominik
Dirks, Wilhelm G.
Pardo González, Emilio
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European Union (EU): 825759
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TOX-Free Toxicity assessment on neurons and cardiomyocytes by means of FluoRescence Emitting Electrodes
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Stem Cells Translational Medicine. Oxford University Press (OUP). 2022, 11(7), pp. 727-741. ISSN 2157-6564. eISSN 2157-6580. Available under: doi: 10.1093/stcltm/szac031
Zusammenfassung

In vitro models of the peripheral nervous system would benefit from further refinements to better support studies on neuropathies. In particular, the assessment of pain-related signals is still difficult in human cell cultures. Here, we harnessed induced pluripotent stem cells (iPSCs) to generate peripheral sensory neurons enriched in nociceptors. The objective was to generate a culture system with signaling endpoints suitable for pharmacological and toxicological studies. Neurons generated by conventional differentiation protocols expressed moderate levels of P2X3 purinergic receptors and only low levels of TRPV1 capsaicin receptors, when maturation time was kept to the upper practically useful limit of 6 weeks. As alternative approach, we generated cells with an inducible NGN1 transgene. Ectopic expression of this transcription factor during a defined time window of differentiation resulted in highly enriched nociceptor cultures, as determined by functional (P2X3 and TRPV1 receptors) and immunocytochemical phenotyping, complemented by extensive transcriptome profiling. Single cell recordings of Ca2+-indicator fluorescence from >9000 cells were used to establish the "fraction of reactive cells" in a stimulated population as experimental endpoint, that appeared robust, transparent and quantifiable. To provide an example of application to biomedical studies, functional consequences of prolonged exposure to the chemotherapeutic drug oxaliplatin were examined at non-cytotoxic concentrations. We found (i) neuronal (allodynia-like) hypersensitivity to otherwise non-activating mechanical stimulation that could be blocked by modulators of voltage-gated sodium channels; (ii) hyper-responsiveness to TRPV1 receptor stimulation. These findings and several other measured functional alterations indicate that the model is suitable for pharmacological and toxicological studies related to peripheral neuropathies.

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570 Biowissenschaften, Biologie
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nociceptors; peripheral nervous system diseases; allodynia; oxaliplatin; TRPV cation channels; receptors, purinergic P2X3
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ISO 690HOLZER, Anna-Katharina, Christiaan KARREMAN, Ilinca SUCIU, Lara-Seline FURMANOWSKY, Harald WOHLFARTH, Dominik LOSER, Wilhelm G. DIRKS, Emilio PARDO GONZÁLEZ, Marcel LEIST, 2022. Generation of Human Nociceptor-Enriched Sensory Neurons for the Study of Pain-Related Dysfunctions. In: Stem Cells Translational Medicine. Oxford University Press (OUP). 2022, 11(7), pp. 727-741. ISSN 2157-6564. eISSN 2157-6580. Available under: doi: 10.1093/stcltm/szac031
BibTex
@article{Holzer2022-07-20Gener-57821,
  year={2022},
  doi={10.1093/stcltm/szac031},
  title={Generation of Human Nociceptor-Enriched Sensory Neurons for the Study of Pain-Related Dysfunctions},
  number={7},
  volume={11},
  issn={2157-6564},
  journal={Stem Cells Translational Medicine},
  pages={727--741},
  author={Holzer, Anna-Katharina and Karreman, Christiaan and Suciu, Ilinca and Furmanowsky, Lara-Seline and Wohlfarth, Harald and Loser, Dominik and Dirks, Wilhelm G. and Pardo González, Emilio and Leist, Marcel}
}
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