Figure 5.

Endothelial inflammation after repetitive CHI, and improved cognitive outcome in adolescent IL-1R1 and caspase-1 KO mice. (a) Representative image of Lectin/DAPI staining of isolated CD31+ microvessels. (b) Western blot of isolated CD31+ microvessels or mouse cortex homogenate using NeuN, GFAP and Iba1 antibodies. (c) At 11 days after the last CHI, expression of pro-IL-1β was decreased in endothelial cells isolated from 3HD and 3HW but not 1HD mice, whereas cleaved IL-1β was increased in endothelium only after 3HD but not 3HW or 1 HD. (d) Densitometry data for panel A: 1HD (n = 5/group), 3HD (n = 5/group, *p < 0.05 for pro- IL 1β and **p < 0.001 for cleaved- IL 1β vs. sham), 3HW (n = 3/group, *p < 0.05 for pro- IL 1β). Note that cleaved IL-1β was not detected in the CD31- fraction. (e) Western blot and (f) densitometry of IL-18 in endothelial cells in sham vs. 1HD (p = ns), 3HD (*p < 0.05) and 3HW (p = ns). IL-18 was not readily detected in the CD31-fraction. (g) Western blot results in three-dimensional cultures of human brain endothelial cells subjected to concussive trauma ×3 in one day. Expression of cleaved IL-1β (but not pro- IL-1β) at 24 h was significantly increased in injured cultures. (h) Densitometry data for panel G (n = 4/group, *p < 0.05 for cleaved IL-1β vs. sham). (i) Naïve IL-1R1 KO mice had similar Morris water maze performance as naïve WT in hidden platform trials but slightly worse performance in visible platform trials (p = 0.02 for group, n = 10–11/group). However, after 3HD, IL-1R1 KO mice performed better in hidden trials compared to injured WT mice (p < 0.05 for group, n = 10/group) with no difference in visible platform trials (p = 0.05 vs. injured WT). (j) Naïve Cas1 KO and WT mice did not differ in MWM performance in the hidden or visible platform (p = ns, n = 11–12/group). After 3HD, Cas1 KO mice performed better in hidden trials compared to WT mice (p < 0.05 for group, n = 9–10/group).