Widespread apoptosis or loss of vasculature does not occur in the brains of mice with ECM.

<p><b>(A-C and K)</b> 10⁶ DsRed⁺CD8⁺ OT-I T cells were adoptively transferred into CFP⁺ P14 host mice, which were subsequently infected with 10⁶ SIINFEKL-expressing <i>Pb</i>-TG pRBCs (n = 4) or left uninfected (n = 5). Mice were intravenously injected with CellEvent Caspase-3/7 Detection Reagent when displaying signs of ECM (day 6 p.i). Brains were subsequently isolated and processed for histological examination. <b>(A)</b> Representative snapshot of an apoptotic cell (green) associated with the cortical vasculature (blue) of the brain taken from a mouse with ECM. <b>(B)</b> Quantification ± SD of apoptotic cells stained with CellEvent per mm³ in brains from infected and uninfected control mice. <b>(C)</b> Representative snapshot showing an apoptotic cell (green) in contact with a parasite-specific CD8⁺ OT-I T cell (red). Endothelial cells (blue) were visualized by CFP expression. <b>(D-J)</b> C57BL/6 mice were infected with 10⁴<i>Pb</i> ANKA pRBCs or left uninfected. Brains were removed and processed for histological examination when <i>Pb</i> ANKA mice developed signs of ECM (day 7 p.i.). Representative images demonstrating the <b>(D)</b> absence and <b>(E)</b> rarity of apoptotic cells (green) within the cortex of brains from uninfected and ECM affected mice respectively. Cell nuclei are shown in blue. <b>(F)</b> Quantification ± SD of apoptotic cells stained for activated caspase 3 per mm² in brains from infected and uninfected control mice. <b>(G-H)</b> Co-staining of activated caspase 3 (green) and CD31 (red) demonstrating an <b>(G)</b> apoptotic endothelial cell and an <b>(H)</b> apoptotic leukocyte within the cortical vasculature of brains taken from ECM affected mice. Cell nuclei are shown in blue. Quantification ± SD of total <b>(I)</b> number and <b>(J)</b> area per mm² of cortical blood vessels in the brain from uninfected and ECM affected mice. <b>(K)</b> Representative tile scan showing the localization of CellEvent⁺ apoptotic cells (green/asterisk) in relation to vascular leakage (red) within the brain of an ECM affected (day 6 p.i.) CFP⁺ P14 mouse. Endothelial cells (blue) were identified by CFP. Scale bars: (A,C) 25 μm, (D,E) 50 μm, (G,H) 25 μm, (L) 1 mm. Results are pooled from mice infected in one experiment. Significance determined by unpaired t test with Welch’s correction.