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. 2022 Jul 21;25(8):104813. doi: 10.1016/j.isci.2022.104813

Figure 4.

Figure 4

Response of the Brain-Chip to neuroinflammation

(A) Schematic illustration of the induction of neuroinflammation by perfusion of TNF-α through the brain channel.

(B–D) Secreted levels of IL-1β, IL-6, and IFNγ in control or TNF-α-treated Brain-Chips including, or not, microglia and/or astrocytes. n = four to five independent chips; data are represented as mean ± SEM NS: Not Significant, ∗p < 0.05, ∗∗p < 0.01, ∗∗∗∗p < 0.0001; statistical analysis with Student’s t-test.

(E and F) Representative immunofluorescent staining for microglia (CD68, red), neurons (MAP2, green), astrocytes (GFAP, magenta), and nuclei (DAPI, blue) in TNF-α-treated or control chips (bar, 100 nm).

(G) Quantification of the CD68-positive events/field of view in four randomly selected different areas/chip, n = 3 Brain-Chips; data are represented as mean ± SEM, ∗∗p < 0.01 compared to the untreated control group, statistical analysis by Student’s t-test.

(H) Quantification of the number of GFAP-positive and MAP2 events/field of view in n = 4 randomly selected different areas/chip, n = 3 Brain-Chips; data are represented as mean ± SEM, ∗∗p < 0.01, compared to the untreated control group; statistical analysis with Student’s t-test.

(I) Immunofluorescence images show an example of the two types of astrocyte morphology in cultures (polygonal shape toward more elongated shape), after immunostaining with an antibody against GFAP (bar, 100 nm).

(J) Representative immunofluorescent staining for pericytes (NG2, red) in TNF-α-treated or control chips (bar, 100 nm).

(K) Quantification of NG2 fluorescent intensity in n = 5 randomly selected different areas/chip, n = 3 Brain-Chips; data are represented as mean ± SEM, ∗∗∗∗p < 0.0001 compared to the untreated control group; statistical analysis with Student’s t-test.

(L) Quantification of the number of MAP2 events/field of view in n = 4 randomly selected different areas/chip, n = 3 Brain-Chips; data are represented as mean ± SEM, ∗∗∗∗p < 0.0001, compared to the untreated control group; statistical analysis with Student’s t-test.

(M) Levels of secreted glutamate in the brain channel on culture day 7 (n = six to seven independent chips. ∗∗∗∗p < 0.0001, compared to the untreated group. Data are represented as mean ± SEM, statistical analysis by Student’s t-test.