Figure 2
Figure 2. CD34+ BMDECs produce NO in response to rhRLX. Human CD34+ BMDECs were isolated in the absence (A-B) and presence (C-D) of 100μM L-NAME and then incubated with DAF-FM before imaging by confocal microscopy. Panels A and C are brightfield images of panels B and D, respectively, the same fields imaged with 495 nm excitation and 515 nm emission. Arrows indicate some of the CD34+ BMDECs. Original magnification ×200. (E) CD34+ BMDECs were isolated from a healthy volunteer and labeled with DAF-FM for 30 minutes before removing probe and waiting 10 minutes for de-esterification. The cells were monitored for 30 minutes, to confirm a stable baseline of bioavailable NO before vehicle (○; arrow) or 50 ng/mL of rhRLX (●; arrow) was added to the CD34+ cells. Shown is the mean fluorescence (± SD) of at least 8 cells in which fluorescence was continuously monitored. *P < .01 versus last baseline value. (F) BMDEC-CFU were isolated from a healthy volunteer and labeled with DAF-FM. After 30 minutes to stabilize NO baseline, the indicated concentration of rhRLX was added and after 30 minutes fluorescence was determined. Data are mean ± SD. *P < .001 versus 0 ng/mL of rhRLX. (G) Cells were incubated with 10μM L-NAME before being placed in the Boyden chamber with 50 ng/mL of rhRLX. The mean percentage (± SD) of fluorescence of cells migrating relative to control is shown. *P < .001 versus all other treatments.

CD34+ BMDECs produce NO in response to rhRLX. Human CD34+ BMDECs were isolated in the absence (A-B) and presence (C-D) of 100μM L-NAME and then incubated with DAF-FM before imaging by confocal microscopy. Panels A and C are brightfield images of panels B and D, respectively, the same fields imaged with 495 nm excitation and 515 nm emission. Arrows indicate some of the CD34+ BMDECs. Original magnification ×200. (E) CD34+ BMDECs were isolated from a healthy volunteer and labeled with DAF-FM for 30 minutes before removing probe and waiting 10 minutes for de-esterification. The cells were monitored for 30 minutes, to confirm a stable baseline of bioavailable NO before vehicle (○; arrow) or 50 ng/mL of rhRLX (●; arrow) was added to the CD34+ cells. Shown is the mean fluorescence (± SD) of at least 8 cells in which fluorescence was continuously monitored. *P < .01 versus last baseline value. (F) BMDEC-CFU were isolated from a healthy volunteer and labeled with DAF-FM. After 30 minutes to stabilize NO baseline, the indicated concentration of rhRLX was added and after 30 minutes fluorescence was determined. Data are mean ± SD. *P < .001 versus 0 ng/mL of rhRLX. (G) Cells were incubated with 10μM L-NAME before being placed in the Boyden chamber with 50 ng/mL of rhRLX. The mean percentage (± SD) of fluorescence of cells migrating relative to control is shown. *P < .001 versus all other treatments.

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