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. 2011 May;6(3):238–250. doi: 10.1016/j.scr.2011.02.001

Figure 1.

Figure 1

HBMSCs plated and cultured at high density differentiated toward functional ECs. To determine whether HBMSCs plated and cultured at high density differentiated toward EC over time, their morphology, EC functional properties, and expression of EC markers were determined. (A) Representative phase contrast images of (I) HBMSCs under standard culture conditions (MSC-St) for 48 h, (II) HBMSCs cultured at low density (HBMSC-L) for 14 days, or (III) HBMSCs cultured at high density (HBMSC-H) for 14 days. (IV) HUVECs under standard culture conditions were used as a positive cellular control. For culture conditions, see Materials and methods. Images were obtained using an Olympus (CK X41) microscope (4x objective). Inserts represent an enlarged region of each image. Scale bars = 200 μm. (B) (I–III) Immunofluorescence analysis showing DiI-Ac-LDL uptake in (I) HUVECs, (II) HBMSCs under standard culture conditions (HBMSC-St) for 48 h, and (III) HBMSCs cultured at high density (HBMSC-H) for 28 days. DiI-Ac-LDL uptake (red), DAPI (blue). Images obtained using an Olympus upright widefield fluorescence (BX51) microscope (20x objective). Scale bars = 20 μm. (IV–VI) Immunofluorescence analysis of VCAM-1 in cells stimulated with 10 μg/ml TNFα for 24 h. (IV) HUVECs under standard culture conditions for 48 h, (V) HBMSCs under standard culture conditions (HBMSC-St) for 48 h, and (VI) HBMSCs at high density (HBMSC-H) for 28 days. VCAM-1 (green), DAPI (blue). Images were obtained using a Nikon C1 upright confocal microscope (60x objective). Scale bars = 20 μm. (C,D) Immunofluorescence analysis of VE-cadherin, PECAM-1, vWF, and VEGFR1 expression in (I, IV) HUVECs under standard culture conditions for 48 h, (II, V) HBMSCs under standard culture conditions for 48 h, or (III, VI) HBMSCs cultured at high density (HBMSC-H) for (C) 28 days or (D) 14 days. VE-cadherin, PECAM-1, vWF, and VEGFR1 (green), phosphorylated VEGFR1 (red), DAPI (blue). Images were obtained using a Nikon C1 upright confocal microscope (60x objective) (C, I–VI; and D, IV–VI) or using an Olympus IX71 Deltavision microscope (40x objective) (D, I–III). Scale bars = 10 μm. White arrows highlight specific regions of localised immunoreactivity. Representative images from two independent experiments.