Fig. 1

Transcriptomic changes in microglia indicate entry into a chronic neuroinflammatory state. RNA-seq was performed on microglia either freshly purified from mouse brain or isolated from organotypic brain slice cultures in three independent experiments after 1 or 6 days in organotypic culture. A Principal component analysis indicated good clustering of individual microglial preparations and robust separation of each time point. B Gene set enrichment analysis of driver genes for each principal component was performed to identify salient biology described by each axis. ‑Log₁₀[adj p-val] for enriched Kegg pathways is shown. C Transcription factor enrichment of the driver genes was assessed for each principal component. Note the high representation of transcription factors associated with inflammation in Principal Component 1 and immediate early genes and microglial development in Principal Component 2. D The canonical murine disease-associated microglial gene panel [23] and human AD microglia gene panel were used to assess the functional state of microglia with time in organotypic culture. Shown are two heat maps of Z-scores derived from Log₂[Fold Changes] (scale to the right of the heat map). Below each main heatmap are summary heatmaps indicating the area under the curve for each gene category. Within 1 day of culture, microglia adopted a strong DAM1 (Trem2-independent) state and shifted toward the HAM state. This pattern persisted into DIV6