Additional file 1 of A primary neural cell culture model to study neuron, astrocyte, and microglia interactions in neuroinflammation
Creators: name: Goshi, Noah
name: Morgan, Rhianna K.
name: Lein, Pamela J.
name: Seker, Erkin
Publisher: name: figshare
Year: 2020
Subjects: subject: Medicine
subject: Cell Biology
subject: Neuroscience
subject: Physiology
subject: FOS: Biological sciences
subject: FOS: Biological sciences
subject: Science Policy
subject: Immunology
subject: FOS: Clinical medicine
subject: FOS: Clinical medicine
subject: Developmental Biology
subject: 111714 Mental Health
subject: FOS: Health sciences
subject: FOS: Health sciences

description: Additional file 1: Supplementary Data Table 1. Analysis of the simple main effects (Tukey test) from Figure 4B. The p-values from each pairwise result are shown on the table, with p-values less than 0.05 highlighted in green. Supplementary Data Table 2. Analysis of the simple main effects (Tukey test) from Figure 4C. The p-values from each pairwise result are shown on the table, with p-values less than 0.05 highlighted in green. Supplementary Data Table 3. Analysis of the simple main effects (Tukey test) from Figure 4E. The p-values from each pairwise result are shown on the table, with p-values less than 0.05 highlighted in green. Supplementary Data Table 4. Statistical analysis of Figure 5B-J. The p-values from the 2-way ANOVA and simple main effects analysis (Tukey Test) are shown. p-values &lt; 0.05 are highlighted in green. Supplementary Figure 1. Immunostained images of the co- and tri-cultures at DIV 14 showing the co-localization of f-actin (cyan) and β-III tubulin (red) indicative of dendritic spines. Scale bar = 20 μm. Supplementary Figure 2. Tri-culture media supplement requirements for microglia survival at DIV 7. The results indicate that IL-34 is required for microglial survival in the tri-culture. The figure shows the mean ± SD of the technical replicates (n = 4) of a single biological replicate. Supplementary Figure 3. The tri-culture shows reduced caspase 3/7 activity at DIV 9 (n = 6). The letters above the bars indicate statistically distinct groups (p &lt; 0.05), while the points indicate the values of the technical replicates. Supplementary Figure 4. Representative images of the co- and tri-cultures 48 h following a 1 h treatment with different concentrations of glutamate or vehicle control. The cultures were immunostained for the three cell types of interest: neurons – anti-βIII-tubulin (red), astrocytes – anti-GFAP (green), microglia – anti-Iba1 (orange) and the general nuclear stain DAPI (blue). Scale bar = 100 μM. Supplementary Figure 5 Complete proteomic profile from Figure 5A. Supplementary Figure 6. There are approximately 2% of the total cell population that was not clearly identifiable as neurons, astrocytes or microglia. (A) Mean ± SD of cells from each culture type not reactive for antibodies selective for neurons, astrocytes or microglia (n = 3). (B) Representative images from DIV 7 co- and tri-cultures immunostained for NG2 (red), a biomarker of oligodendrocyte precursor cells (OPCs), and reacted with DAPI (blue), scale bar = 100 μm. Supplementary Figure 7. Change in number of cells following incubation with LPS or 25 μM glutamate. (A) Percent change in cell number following incubation with LPS. (B) Percent change in cell number following incubation with 25 μM glutamate. All graphs display mean ± SD (n = 3).