Fig. 6. CD4⁺ T cells induce apoptosis of endothelial cells through death receptor signaling.

CD4⁺ T cells were isolated from splenocytes or brain of mice 24 hours after autologous blood injection induced ICH or sham control. FACS-sorted CD4⁺ T cells were used for RT2 Profiler PCR Array analysis of 87 apoptosis related genes. (A) Heatmap shows relative changes of expression of cell apoptotic-related genes in CD4⁺ T cells from the brain or spleen of ICH mice versus sham mice. (B) Quantification of top 15 up-regulated apoptosis associated genes. (C) Bar graph shows genes of death receptor ligands in CD4⁺ T cells of ICH or sham mice. In (B) and (C), n = 3 duplicates from 15 mice. One-way ANOVA followed by Tukey post hoc test. (D) Immunostaining of death receptor ligand TRAIL in brain CD4⁺ T cells of ICH or sham mice. Quantification was averaged as percentage of death receptor ligand–positive cells in CD4⁺ T cells. Scale bars, 40 μm and (inset) 20 μm. n = 6 mice per group. Mann-Whitney test. (E) Flow cytometry plots and bar graph show expression of TRAIL receptor (DR5) in different brain cell types of sham and ICH mice, including microglia (CD45^intCD11b⁺), astrocytes [glial fibrillary acidic protein positive (GFAP⁺)], neuron (NeuN⁺), and endothelial cells (CD31⁺). FMO, fluorescence minus one. n = 6 mice per group. One-way ANOVA followed by Tukey post hoc test. (F) CD31⁺ endothelial cell apoptosis was analyzed by TUNEL staining in sham mice and ICH mice with anti-DR5 monoclonal antibody (mAb) or IgG group. Scale bars, 40 μm and (inset) 20 μm. Quantification was averaged as positive cells per mm² in the perihematomal region. n = 6 mice per group. Mann-Whitney test. Means ± SD. *P < 0.05 and **P < 0.01.