EC-derived PDGF-BB and FGF2 expand PαS-MSCs. (A) Immunohistologic staining for PDGFRα (green) and VE-Cadherin (red) in BM sections of tPA-treated mice. Scale bar, 50 µm. (B) Absolute numbers of VE-Cadherin⁺ ECs per femur from WT B6 mice after 2 days of injection with recombinant tPA or vehicle was determined by FACS analysis (n = 3/group). (C-D) Representative images of (C) FGF2 and (D) PDGF-BB in BM sections of tPA-treated mice. Nuclei were counterstained using DAPI. VE-Cadherin (red) and FGF2/PDGF-BB (green). Arrows indicate FGF2 or PDGF-BB coexpressing BM-ECs after tPA treatment. Scale bar, 50 µm. Staining was repeated 4 times. (E) Relative mRNA expression of PDGF-BB and FGF2 in CD45⁻CD31⁺ ECs isolated from tPA and vehicle-treated mice. Data are normalized to GAPDH (n = 3/group). (F-G) PDGF-BB serum level was measured by ELISA on day 2 of (F) tPA treatment of WT B6 mice in the presence or absence of c-Kit neutralizing antibody or of (G) KitL treatment of WT B6, Plg^−/−, and MMP-9^−/− mice. Same control from Figure 2E was used for reference (n = 3/group). (H-I) Relative mRNA expression of (H) PDGF-BB and (I) FGF2 as determined by reverse transcriptase (RT)-PCR in CD45⁻CD31⁺ ECs treated with vehicle, tPA, or KitL for 2 days. Data are normalized to GAPDH (n = 6/group). (J) Representative western blot of PDGF-BB and FGF2 in supernatant of HUVECs treated in vitro with vehicle, tPA, and KitL with or without c-Kit neutralizing antibody, ACK2 (n = 3/group). Mean ± SEM. *P < .05; **P < .01; ***P < .001 by Student t test or ANOVA with Tukey HSD tests for multigroup analysis.. Data are representative of 2 independent experiments.