Journal: iScience

Article Title: Disease phenotypic screening in neuron-glia cocultures identifies blockers of inflammatory neurodegeneration

doi: 10.1016/j.isci.2024.109454

Figure Lengend Snippet: High-content analysis allows interrogation of additional cell types for secondary outcome data (A and B) Average normalized microglia counts from neuron-glia cultures treated without (A) and with (B) LPS and each compound in the screen, and ordered by increasing microglia count. RM two-way ANOVA with Dunnett’s post-hoc test comparing all treatments to DMSO in the presence and absence of LPS. Darker regions of each graph indicate p < 0.05 significance. Data are represented as mean ± SD, N = 4. (C) Average normalized neuron counts plotted against average normalized microglia counts for all compounds ± LPS, with LPS-treated data from hit compounds highlighted and colored by target class. Data are represented as mean ± SD, N = 4. (D) Representative Hoechst-IB 4 labeled images (cropped) and corresponding segmented microglial masks from cultures treated with DMSO for 3 days ( DIV10 ) in the presence and absence of LPS, illustrating the microglial morphology analysis. Scale bars = 100μm. Mask colors are arbitrary. (E) Average microglial FormFactor (circularity) from neuron-glia cultures treated ± LPS and ± each compound in the screen. Data are represented as mean ± SD, N = 4. (F) Average microglial FormFactor plotted against average normalized microglia counts for all compounds in the presence of LPS, with data from hit compounds highlighted and colored by target class. Data are represented as mean ± SD, N = 4. See also Figures S6–S8 .

Article Snippet: Microglial FormFactor was highly variable in LPS-untreated wells, and microglia converged on a rounder and more ameboid shape with LPS activation ( D, 5E, and ).

Techniques: High Content Screening, Labeling

Journal: iScience

Article Title: Disease phenotypic screening in neuron-glia cocultures identifies blockers of inflammatory neurodegeneration

doi: 10.1016/j.isci.2024.109454

Figure Lengend Snippet: Clustered heatmap analysis of all screen data reveals 3 distinct mechanisms of neuroprotection (A) Clustered heatmap of data from neuroprotective hits and DMSO (counts of each cell type and microglial morphology in the absence and presence of LPS). E.g. “Neuron –“ and “Neuron +” columns display mean neuron counts in the absence and presence of LPS respectively. In the analysis of all compounds ( Figure S9 A), each data type was min-max normalized between 0 and 1 based on min and max values across both LPS-treated and LPS-untreated conditions to ensure comparability, and this data was then filtered for hits and DMSO. Legend and left-hand side color labels indicate the target pathway for each hit. Compound IDs and category numbers provided on the right-hand side. (B–G) Average data (variable: see Y axes) for hit compounds of each category. Each datapoint is the average value for one hit compound over all screen repeats. Categories 1, 2, and 3 included 13, 10, and 7 hit compounds respectively. One-way ANOVA with Tukey’s post hoc test. Blue and orange dashed lines indicate the average data value in LPS-untreated and LPS-treated DMSO wells respectively. Data are represented as mean ± SD. ∗∗p < 0.01, ∗∗∗p < 0.001, ∗∗∗∗p < 0.0001. See also Figure S9 .

Article Snippet: Microglial FormFactor was highly variable in LPS-untreated wells, and microglia converged on a rounder and more ameboid shape with LPS activation ( D, 5E, and ).

Techniques: