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 4

Single-cell transcriptome analysis of MS patient astrocyte/NGN2 neuron cocultures.

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Single-cell transcriptome analysis of MS patient astrocyte/NGN2 neuron c...
Cells were treated for 24 hours with TNF-α/IL-17A or left untreated (control) and harvested with Accutase. Single-cell suspensions of 6 technical repeats per sample were filtered, and subsamples were labeled with CMOs to allow pooling for multiplexed libraries. (A) UMAP of all cells with projection of expression of MAP2 and TUBB3 (neuronal markers) and ALDH1L1 and AQP4 (astrocyte markers). (B) UMAP and pie chart to analyze the proportional distribution of cells in the color-coded subclusters, which were named according to the most prevalent defining genes (ranked by log2 fold change) and their prospective function. Neurons of PMS cultures were found to be higher in the inflammation cluster compared with BMS. (C) Enrichr analysis of KEGG 2021 and Reactome 2016 pathways comparing PMS and BMS. A black box represents the presence of a gene. (D) IPA analysis comparing samples according to potential upstream mediators (short list; for complete analysis see Supplemental Table 1). The z score represents the activation score of a predicted regulator based on the expression of the input genes; a positive score represents an activation and a negative score an inhibition. Each column of BMS or PMS represents the data set of 1 experiment (TNF-α/IL-17A treated vs. control); “all BMS” and “all PMS” are the mean of samples BMS1–BMS3 or PMS1–PMS3, respectively. The analysis predicted a JAK/STAT activation and a negative score for tofacitinib in all PMS samples.