Fig. 2

Transcriptomic changes in bulk tissue indicate ongoing microgliosis and astrocytosis associated with loss of synaptic function genes. Transcriptomic analysis using NanoString (Neuropathology panel) was performed on bulk cortical tissue from three independent experiments either freshly harvested from mouse brain or collected from brain slices after 1–14 days in organotypic culture. A Principal component analysis indicated a robust change in the bulk tissue transcriptome at DIV1. The transcriptome gradually shifted away from this early state and became stable by DIV6. 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 Hierarchical clustering of the full data set revealed a subset of genes whose differential expression shifted from robust increases early in culture (DIV1–4) to decreases later in culture (≥ DIV6). Manhattan hierarchical distances are shown to the right of full heat map and further highlight the differences in early vs. late transcriptomes. Scales for each heat map shown underneath and indicate Z-scores derived from Log₂[Fold Change]. D Gene panels for specific aspects of the overall neuroinflammatory signature. Disease associated microglial gene panel indicates strong DAM2 state. Astrocytes were broadly activated and existed in both the neurotoxic (A1) and neuroprotective (A2) state. Complement C3 was robustly increased across all time points, however NLRP3 was modestly reduced. Expression of neurotransmitter receptor genes across all transmitter classes was robustly decreased. Scales for each heat map shown to the side and indicate Z-scores derived from Log₂[Fold Change]