iPSC-derived reactive astrocytes from patients with multiple sclerosis protect cocultured neurons in inflammatory conditions
Janis Kerkering, … , Marlen Alisch, Volker Siffrin
Janis Kerkering, … , Marlen Alisch, Volker Siffrin
Published May 23, 2023
Citation Information: J Clin Invest. 2023;133(13):e164637. https://doi.org/10.1172/JCI164637.
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Research Article Inflammation

iPSC-derived reactive astrocytes from patients with multiple sclerosis protect cocultured neurons in inflammatory conditions

Abstract

Multiple sclerosis (MS) is the most common chronic central nervous system inflammatory disease. Individual courses are highly variable, with complete remission in some patients and relentless progression in others. We generated induced pluripotent stem cells (iPSCs) to investigate possible mechanisms in benign MS (BMS), compared with progressive MS (PMS). We differentiated neurons and astrocytes that were then stressed with inflammatory cytokines typically associated with MS phenotypes. TNF-α/IL-17A treatment increased neurite damage in MS neurons from both clinical phenotypes. In contrast, TNF-α/IL-17A–reactive BMS astrocytes cultured with healthy control neurons exhibited less axonal damage compared with PMS astrocytes. Accordingly, single-cell transcriptomic BMS astrocyte analysis of cocultured neurons revealed upregulated neuronal resilience pathways; these astrocytes showed differential growth factor expression. Furthermore, supernatants from BMS astrocyte/neuronal cocultures rescued TNF-α/IL-17–induced neurite damage. This process was associated with a unique LIF and TGF-β1 growth factor expression, as induced by TNF-α/IL-17 and JAK-STAT activation. Our findings highlight a potential therapeutic role of modulation of astrocyte phenotypes, generating a neuroprotective milieu. Such effects could prevent permanent neuronal damage.

Authors

Janis Kerkering, Bakhrom Muinjonov, Kamil S. Rosiewicz, Sebastian Diecke, Charlotte Biese, Juliane Schiweck, Claudia Chien, Dario Zocholl, Thomas Conrad, Friedemann Paul, Marlen Alisch, Volker Siffrin

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Figure 3

MS patient astrocytes and cocultures with NGN2 neurons without and with exposure to inflammatory cytokines.

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MS patient astrocytes and cocultures with NGN2 neurons without and with ...
(A) Immunofluorescence staining of iPSC-derived astrocytes. Mature astrocytes differentiated for 6 weeks stain positive for GFAP, s100β, AQP4, IL-17R, and TNFR1. Scale bars: 50 μm. (B) iPSC-derived astrocytes cocultured with NGN2-neurons show close colocalization. Treatment with TNF-α/IL-17A for 24 hours (50 ng/mL) increased SMI32/SMI31 ratio in NGN2-neurons cultured with PMS astrocytes. Scale bars: 50 μm. Immunofluorescently stained SMI32/SMI31 neurons were analyzed with ImageJ and are presented as surface ratio of SMI32/SMI31 ± SD, normalized to the control. (C and D) Neurons in coculture with BMS astrocytes (C) were protected against TNF-α/IL-17A exposure, whereas PMS astrocytes (D) did not show protection against TNF-α and TNF-α/IL-17A. (E) Neurons in coculture with healthy control astrocytes (HC1, HC2, HC3) also showed increased SMI32/SMI31 ratios after TNF-α/IL-17A exposure. Each data point represents a microscopic field of view (641 × 479 μm) of 3 independent experiments depicted by different symbols; pooled data represent the mean from 3 individual patients (different colors) and 3 independent experiments (different symbols). Statistical significance was tested with a Kruskal-Wallis test; *P < 0.05, **P < 0.01, ***P < 0.001.