Fig. 5

Activation of microglia treated with ISG15. Primary microglial cultures were prepared from p1 mouse pups. Primary astrocyte cultures were prepared from p3-4 mouse pups. Human IPSC-derived neurons were differentiated from NSC in media containing cAMP, BDNF, IGF1, GDNF, and StemPro neurosupplement for > 2 weeks. Astrocytes (DIV29) and microglia (DIV7) were treated with ISG15 at the indicated dose. Chemokine and cytokine mRNA (A) and protein (B) expression 24 h following treatment of murine microglia with ISG15 as assessed by RT-PCR and cytometric bead array, respectively. Relative expression of mRNA for cytokines, chemokines, and transcription factors in murine astrocytes (C) or IPSC-derived human neurons (D) 24 h following treatment with ISG15 as assessed by RT-PCR. E Relative expression of activation markers in microglia treated as in panels A and B following co-treatment with neutralizing antibodies against ISG15 or CD11b, co-treatment with the TLR4 inhibitor TAK242, or treatment with LPS. P values are indicated for one-way ANOVA. *P < 0.01, **P < 0.001, ***P < 0.0001. Mean ± SEM are shown