Figure Legends
Fig 1. Comparable frequencies but lower ROS levels in BM ECs from young donors than from old donors. Representative gating strategy for BM ECs (CD34+CD309+CD133+) from healthy donors (A) . Frequencies of BM ECs in BMMNCs(B) and intercellular ROS levels in BM ECs (C) among young, middle-aged and old donors. The data are showed as the mean ± standard error of the mean (SEM).
Fig 2. Better migration and tube formation functions of BM ECs from young donors than from old donors. Representative tube formation images and tube length of 7-day primary cultured BM ECs (magnification ×4) in the young and old donor groups (A) . Typical figures of the transwell migration assay and counts of the migrated BM ECs (magnification ×10) from young and old donors(B) . Representative images of double-positive-stained ECs (yellow) with DiI–Ac-LDL uptake (red) and FITC–UEA-I binding (green) and the double-positive-stained cell numbers (magnification ×10) in the young and old donor groups (C) .
Fig 3. ECs from young donors regulated T cells to produce fewer proinflammatory cytokines. Schematic diagram of the study design of the coculture of ECs and CD3+ T cells and the measurement of effector T subsets after 3 days of coculture (A) . The frequencies of BM Th1 (CD3+CD8IFN-γ+)(B) , Th2 (CD3+CD8IL-4+)(C) , Tc1 (CD3+CD8+IFN-γ+)(E) , Tc2 (CD3+CD8+IL-4+)(F) , Th17 (CD3+CD8IL-17+)(I) and Tregs (CD3+CD8CD25+Foxp3+)(J) in young and old donors. The ratios of BM Th1/Th2(D) , Tc1/Tc2 (G) , CD4+/CD8+ T cells (H) in young and old donors.
Fig 4. Upregulated genes related to the cytokine-mediated signalling pathway and the regulation of T cell activation in BM ECs from old donors. Schematic diagram of the study design for RNA-seq analysis of ECs from donors (A) . Volcano plots of the downregulated genes (red) and upregulated genes (green) in the old group and comparable genes (blue) between the two groups (B) . GO enrichment analysis shows the top 10 biological processes enriched by upregulated genes in ECs from young and old donors (C) . Heatmaps show genes expression involved in the regulation of T cell activation and cytokine−mediated signalling pathways, including TLR1, TLR2, TLR4, CD80, CD86, CXCL10, CCR2, CCR5, PRF1, and NLRP3, in young and old ECs (scaled by row) (D) . The relative mRNA expression levels of aforementioned genes in ECs from young and old donors tested by qRT-PCR (E) . *P <0.05, ** P <0.005.
Fig 5. Lower expression of TLR4, CCR5 and CD80 in ECs from young donors than from old donors. The representative gating strategy and frequencies of TLR4+ ECs (A) , CCR5+ ECs(B) , CD80+ ECs (C) , and CD86+ ECs (D) from the young and old donors with LPS stimulation.
Fig 6. Graphical abstract of the current study. BM ECs from young donors had lower ROS levels, better functions, and more balanced T cell modulating effects than ECs from old donors. Lower TLR signaling and costimulatory molecules, and less inflammatory and chemotaxis cytokine secretion in ECs from young donors might cause young ECs to induce more balanced T cell subset differentiation than old ECs.