Figure 5
Figure 5. CXCR7 participates in the CXCL12 priming effect on PB CD34+ cell survival and colony formation. (A) PB CD34+ cells were stimulated for 48 hours in a serum- and cytokine-free StemαA medium in the presence or absence of CXCL12 (0.5 ng/mL), blocking anti-CXCR4 and/or -CXCR7 (11G8) antibodies or isotype control, or CCX704 or CCX733 compounds. After trypan blue labeling, the number of live cells were determined by counting on a Malassez cell. Histograms show live cell number modulations vs control as a mean of fold changes ± SD (n = 3). Asterisk indicates significance vs control (*P ≤ .05). Deltas indicate significance between conditions (ΔP ≤ .05, ΔΔP ≤ .01). (B) PB CD34+ cells were stimulated (2.105 cells/mL) for 24 hours in serum- and cytokine-free StemαA medium in the presence or absence of CXCL12 (0.5 ng/mL), blocking anti-CXCR4 and/or -CXCR7 (11G8) antibodies or isotype control, or CCX704 or CCX733 compound. Cells were then placed at a density of 2000 cells/mL in methylcellulose media supplemented with cytokines in 35-mm Petri dishes. Colonies were scored using an inverted microscope after 14 days of culture on the basis of morphological criteria. Histograms show modulations of the total number of colonies including BFU-E, CFU-G, CFU-GM, and CFU-MIX vs untreated cells. Results are expressed as the mean of fold changes ± SD (n = 3). Asterisks indicate significance vs untreated cells (*P ≤ .05, **P ≤ .01). Deltas indicate significance between conditions (ΔP ≤ .05, ΔΔP ≤ .01).

CXCR7 participates in the CXCL12 priming effect on PB CD34+cell survival and colony formation. (A) PB CD34+ cells were stimulated for 48 hours in a serum- and cytokine-free StemαA medium in the presence or absence of CXCL12 (0.5 ng/mL), blocking anti-CXCR4 and/or -CXCR7 (11G8) antibodies or isotype control, or CCX704 or CCX733 compounds. After trypan blue labeling, the number of live cells were determined by counting on a Malassez cell. Histograms show live cell number modulations vs control as a mean of fold changes ± SD (n = 3). Asterisk indicates significance vs control (*P ≤ .05). Deltas indicate significance between conditions (ΔP ≤ .05, ΔΔP ≤ .01). (B) PB CD34+ cells were stimulated (2.10 cells/mL) for 24 hours in serum- and cytokine-free StemαA medium in the presence or absence of CXCL12 (0.5 ng/mL), blocking anti-CXCR4 and/or -CXCR7 (11G8) antibodies or isotype control, or CCX704 or CCX733 compound. Cells were then placed at a density of 2000 cells/mL in methylcellulose media supplemented with cytokines in 35-mm Petri dishes. Colonies were scored using an inverted microscope after 14 days of culture on the basis of morphological criteria. Histograms show modulations of the total number of colonies including BFU-E, CFU-G, CFU-GM, and CFU-MIX vs untreated cells. Results are expressed as the mean of fold changes ± SD (n = 3). Asterisks indicate significance vs untreated cells (*P ≤ .05, **P ≤ .01). Deltas indicate significance between conditions (ΔP ≤ .05, ΔΔP ≤ .01).

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