Highly neurogenic glia from human and mouse myenteric ganglia generate functional neurons following culture and transplantation into the gut

• Published in: Cell reports (Cambridge) 2024-11, Vol.43 (11), p.114919, Article 114919
• Main Authors: Mueller, Jessica L., Leavitt, Abigail R., Rahman, Ahmed A., Han, Christopher Y., Ott, Leah C., Mahdavian, Narges S., Carbone, Simona E., King, Sebastian K., Burns, Alan J., Poole, Daniel P., Hotta, Ryo, Goldstein, Allan M., Stavely, Rhian
• Format: Article
• Language: English
• Subjects: Animals; cell therapy; Cells, Cultured; enteric glial cell; enteric nervous system; enteric neural stem cell; enteric neuron; Humans; Mice; myenteric ganglia; Myenteric Plexus - cytology; Myenteric Plexus - metabolism; Neural Stem Cells - cytology; Neural Stem Cells - metabolism; Neural Stem Cells - transplantation; Neurogenesis; Neuroglia - metabolism; Neurons - cytology; Neurons - metabolism; neuroregeneration; postnatal; regeneration
• ISSN: 2211-1247; 2211-1247
• DOI: 10.1016/j.celrep.2024.114919

Enteric neural stem cell (ENSC) therapy offers great promise for neurointestinal diseases; however, current isolation methods yield insufficient neurons for regenerative applications. Multiomic profiling of enteric glial cells (EGCs) suggests that subpopulations within myenteric ganglia (MyGa) are a reservoir of highly neurogenic ENSCs. Here, we describe protocols to enrich for intraganglionic EGCs by isolating intact fragments of MyGa, generating cultures with higher neuronal purity than traditional methodologies isolating intramuscular single cells (IM-SCs). MyGa-derived EGCs transdifferentiate into more neurons than IM-SC-derived EGCs do, confirming their neurogenic predisposition. Following transplantation to the mouse intestine, MyGa-derived neurons generate calcium transients and activate smooth muscle in response to optogenetic stimulation. In the human intestine, MyGa-derived cells are similarly highly neurogenic, are enriched for a distinct progenitor population identified by single-cell RNA sequencing (scRNA-seq), and exhibit neuromuscular connectivity following xenogeneic transplantation into mice. Highly neurogenic ENSCs are preferentially located within the MyGa, and their selective isolation offers considerable potential for therapy. [Display omitted] •Glia in the MyGa have higher neurogenic potential than intramuscular glia do•MyGa can be isolated by their enzymatic resistance and counter filtration•Enteric neural stem cells (ENSCs) cultured from MyGa improve neuronal differentiation•Human MyGa-derived cells engraft and exert neuromuscular connectivity in mice Mueller et al. show that selectively isolated ENSCs from MyGa have a high neuronal differentiation potential. These findings are leveraged to develop protocols to generate ENSC cultures from mouse and human specimens that show functional neuromuscular connectivity post transplantation, holding potential for advancing regenerative therapies for neurointestinal diseases.