CONSERVED ASTROCYTE REACTIVITY MODULES ACROSS SPECIES IN MULTIPLE SCLEROSIS

Astrocytes are central regulators of CNS homeostasis and neuroinflammation, yet in vitro modeling is confounded by serum exposure, species differences, and context-dependent reactivity. To systematically dissect distinct grades of astrocyte reactivity, we compared primary murine astrocytes cultured with or without serum to serum-free human iPSC-derived astrocytes generated from three independent lines, and benchmarked their inflammatory responses to TNFα with a final cross-comparison to Multiple Sclerosis patient data. Immunofluorescence and scanning electron microscopy provided initial insights into morphological and protein-level differences, while bulk RNA sequencing followed by cross-species transcriptomic integration enabled the identification of conserved and model-specific astrocyte states. This analysis clearly characterized the impact of serum on primary murine astrocyte cultures and revealed a robust Serum-Induced Astrocyte Signature (SIAS). Although species identity emerged as the dominant axis of transcriptional variance, TNFα stimulation of hiPSC-derived astrocytes uncovered a highly conserved TNFα-Induced Astrocyte Signature (TIAS) shared across human and mouse systems, highly relevant to MS (Multiple Sclerosis). The 48-gene cross-species TIAS were projected onto scRNA seq data from MS patients, confirming this relevance by identifying previously uncharacterized, consistently regulated genes with potential therapeutic relevance. Together, these results provide an integrated framework for interpreting astrocyte reactivity across experimental systems, define SIAS and TIAS as two principal reactive modules, and establish a scalable strategy for selecting and benchmarking astrocyte models in neuroinflammatory research.