Fig. 1: High throughput screening of pericytes identifies inflammatory-modulating compounds.

a, b Pericytes were treated with IL-1β for 24 h and immunostained for CCL2 and ICAM-1 protein expression. Integrated intensity per cell was calculated using total cells (Hoechst), n = 3, significance determined using one-way ANOVA with Dunnett’s correction for multiple comparisons, mean ±  SEM.****p < 0.0001. Case used H239. c, d Pericytes were screened for compounds that can modify IL-1β-induced CCL2 or ICAM-1 expression using the Prestwick chemical library. Mean integrated intensity per cell was calculated using total cell counts and normalised to IL-1β + vehicle condition (arrow) (blue arrow-digoxin, red arrow-lanatoside C, black arrows-hit compounds tested in (g)). Internal controls were present on each drug plate (TGFβ₁, 1 or 10 ng mL⁻¹). Case used: H239. e, f Hits identified using cut-off criteria from primary screen that modified CCL2 or ICAM-1 expression with known therapeutic uses. g Conditioned media from pericytes pre-treated with hit compounds for 24 h, then stimulated with IL-1β (0.05 ng mL⁻¹) for 24 h was analysed by CBA, data was normalised to cell number and presented as the logged value of cytokine secretion in pg/mL/10,000 cells. Control (0.01% BSA in PBS) + dimethylsulfoxide (DMSO), Vehicle (IL-1β (0.05 ng mL⁻¹ + (DMSO)), remainder are IL-1β + drug (n = 2, media pooled from triplicate wells for analysis). Cases used E206, E213. h Drug screening pipeline in primary human brain pericytes. Initial screen in pericytes completed in duplicate wells and secondary screen in 83 compounds that met cut-off criteria was completed in triplicate wells. Effects that were reproduced were examined using a three-point concentration test, then tested for effects on cell secretion (data in Fig. S1).