Fig. 7
TGFβ1 decreases pericyte proliferation but does not affect their viability. Primary human brain pericytes were treated with vehicle (1 mM citric acid, pH 3 with 0.1 % BSA) or 10 ng/mL TGFβ1 for 0–72 h. Cells were fixed and immunostained for cleaved caspase 3 (CC3). Nuclei were counterstained with Hoechst. Cells treated for 24 h with 50 nM okadaic acid (OA) were used as a positive control. Representative images from the 24 h time point are shown (a). The percentage of CC3 positive cells was determined by automated image analysis from three independent experiments (b). Primary human brain pericytes were treated with vehicle (1 mM citric acid, pH 3 with 0.1 % BSA) or 10 ng/mL TGFβ1 for 0–72 h. For the final 24 h, cells were incubated with 10 μM EdU. Cells were fixed, EdU visualised and nuclei counterstained with Hoechst. Representative images from the 72-h time point are shown (c). The percentage of EdU positive cells (d), and the number of cells/field of view was determined by automated image analysis from three independent experiments (e). Primary human brain pericytes were treated with vehicle (1 mM citric acid, pH 3 with 0.1 % BSA) or 10 ng/mL TGFβ1 for 0–72 h. Cytotoxicity was determined from three independent experiments by an LDH release assay using cells lysed with DMEM/F12 containing 1 % Triton-X 100 as a positive control (f). Viability was determined from three independent experiments by a two hour treatment with Alamar Blue (g). *p < 0.05, **p < 0.01, ***p < 0.001 versus vehicle control for each time point (two-way ANOVA). Scale bar = 50 μm