Fig. 6

Cortical astrocytes become reactive following mild TBI. A) Gene set variation of custom-curated astrocyte gene sets (columns) across injury and time point groups (rows) reveals two distinct expression patterns: gradual increase by 3-5xCHI (left cluster) and rapid decrease after 1xCHI (right cluster). Gene sets are z-scored and clustered by Euclidean distance. B) Multiple linear regression models were calculated for each gene set using fixed effects of time point after most recent injury, binary injury status (injured, sham-injured), and number of injuries. The model shows that the effects of injury (both numeric and binary) outweigh time-dependent changes for most gene sets. Red boxes indicate statistical significance (p < 0.05). C) Astrocyte gene sets Gliotransmission and Plasticity are representative of right cluster, displaying a decrease after 1xCHI followed by return to sham-injured levels by 4-5xCHI (p = 0.013, p = 0.032, t-test with Bonferroni adjustment) (mean ± SEM). Data points are colored by the z-score of the log of astrocyte reactivity marker gene Gfap. D) Astrocyte gene sets Stress Response and General Metabolism are representative of left cluster, displaying a gradual increase after 3-5xCHI (Stress Response 3xCHI p = 0.038, 4xCHI p < 0.0001, 5xCHI p = 0.015; General Metabolism 4xCHI p = 0.00012, 5xCHI p = 0.020; t-test with Bonferroni adjustment) (mean ± SEM). Data points are colored by the z-score of the log of astrocyte reactivity marker gene Gfap, which is associated with high expression in either set. E) Glial fibrillary acidic protein (GFAP) is significantly upregulated 30 min and 4 h post-injury vs. pre-injury (p = 0.00061, p = 0.0082 respectively; multiple linear regression model, pre/post and number of injuries) (mean ± SEM). Note that sham-injured is equivalent to pre-1xCHI, 2xCHI at 24 h is equivalent to pre-3xCHI, and 4xCHI at 24 h is equivalent to pre-5xCHI in this representation. F) Heatmap of Pearson’s correlation coefficient of cortical GFAP versus each measured cortical protein (columns) within each injury group and time point (rows); red boxes indicate FDR-corrected p < 0.05. Panels A-D present transcriptional data from female somatomotor cortex. Panels E-F present protein data from female frontal cortex