Improved migration and tube formation functions of BM ECs from young donors
On the 7th day of culture, BM ECs from young donors showed significantly higher abilities of tube formation (Fig. 2A, 9476 ± 746.1 vs. 5748 ± 950.1, P =0.02) and migration (Fig. 2B, 146.6 ± 5.3 vs. 110.8 ± 5.2, P =0.002) than those of cells from old donors. No significant difference was found in double-positive staining between the young and old groups (Fig. 2C, 62.2 ± 2.1 vs. 59.3 ± 1.3, P=0.48), which was in accordance with the flow cytometric EC frequency results.
ECs from young donors regulated T cells toproduce fewer proinflammatory cytokines
To investigate the effect of ECs from individuals with different ages on T cell cytokine production capacity, CD3+ T cells from middle-aged donors were cocultured with ECs from young and old donors for 3 days, respectively, and then T cell subsets were examined via flow cytometry.
Fig. 3A shows the schematic diagram of the study design for the coculture of ECs with CD3+ T cells. After coculture, significantly fewer type 1 T cells, including Th1 cells (Fig. 3B, 4.60% ± 1.21% vs. 17.46% ± 4.28%, P =0.001) and Tc1 cells (Fig. 3E, 5.09% ± 0.73% vs. 19.3% ± 2.86%, P =0.0006), were observed in young donors than in old donors. No statistic differences were found in the Th2 (Fig. 3C) and Tc2 cell frequencies (Fig. 3F) between the two groups. Therefore, the ratios of Th1/Th2 cells (Fig. 3D, 1.15 ±0.35 vs. 3.68 ± 0.94, P =0.01) and Tc1/Tc2 cells (Fig. 3G, 1.33 ± 0.36 vs. 3.43 ± 0.79, P =0.02) were notably less in young group than in old group. No significant differences were observed in CD4+/CD8+ T cell ratio (Fig. 3H), Th17 (Fig. 3I) or Tregs (Fig. 3J) between the two groups.