Figure 5.

Astrocyte activation can be controlled in the brain hydrogel. a) Human primary astrocytes were encapsulated in hydrogels for 24 h and subsequently dosed with cytokines IL-1α, TNF-α, and C1q for an additional 24 h. b) Representative images of astrocytes after 48 h in brain hydrogels and collagen and incubated with either standard culture medium (control), defined quiescent medium, dosed with cytokines (+) IL-1α (3 ng mL⁻¹), TNF-α (30 ng mL⁻¹), and C1q (400 ng mL⁻¹), or dosed with a 2× dose of cytokines (++) IL-1α (6 ng mL⁻¹), TNF-α (60 ng mL⁻¹), and C1q (800 ng mL⁻¹). Scale bar is 20 μm. c) Quantification of cells from (b) stained for GFAP, vimentin, and DAPI. N = 2, n = 250 cells per condition. d) Distribution of cell morphologies and e) representative cell morphologies in the different populations identified as round (gray), stellate (blue), or polarized (light blue). N = 3, n = 100 cells per condition. f) Quantification of astrocyte process length N = 6, n = 3. g) Single cell volume and h) single cell surface area for astrocytes encapsulated for 48 h. N = 3, n = 3. i) Comparison of migration speed astrocytes seeded on plastic (TCPS), or in the brain hydrogels with and without cytokines. N = 2. All plots show mean + SEM. Data in (c) were analyzed using a one-way analysis of variance (ANOVA). Data in (f)–(h) were analyzed using a two-way ANOVA followed by a Tukey’s multiple comparison test with 95% confidence interval. Data in (i) were analyzed using a one-way ANOVA followed by a Dunnett’s multiple comparison test with 95% confidence interval. *, **, and **** indicate P < 0.05, P < 0.01, and P < 0.0001, respectively; N.S., not significant.