Figure 1
Figure 1. Inhibition of erythroid cell growth and formation of HNE surface adducts in erythroid cells cocultivated with HZ-fed human monocytes, cocultivated with HZ, or treated with HNE. (A-C) Erythroid cells were cocultivated with unfed (Mo), HZ-fed (Mo + HZ), or RBC-fed (Mo + RBC) monocytes in liquid culture. Adherent monocytes were unfed, fed with either HZ (25μM HZ-heme, corresponding to 12 RBCs/monocyte in terms of heme content), or 50 opsonized RBCs/monocyte for 3 hours. After removal of nonphagocytosed HZ or RBCs, erythroid cells were added to the monocytes at a cell ratio of 1:1, corresponding to a 10:1 ratio at day 4 of coincubation. (A) Growth of erythroid cells was calculated from cell numbers counted in the expanding cultures (means ± SE of 3 independent experiments). (B) HNE-adducts on erythroid cell surface were quantified by flow cytometry after immune staining and expressed as MFI (means ± SE of 3 independent experiments). The significance of differences (P < .05) between erythroid cells cocultivated with HZ-fed monocytes and unfed (*) or RBC-fed (§) monocytes are indicated. (D-F) Erythroid cells were cocultivated with HZ (25μM HZ-heme), treated with HNE (7μM), or kept as untreated controls (CTRL) in liquid culture. (D) Growth of erythroid cells was calculated from cell numbers counted in the expanding cultures (means ± SE of 3 independent experiments). The significance of differences (P < .05) between HZ-treated (*) or HNE-treated (§) cells and untreated controls and (P < .1) between HZ-treated (**) cells and controls are indicated. (E) HNE adducts were quantified by flow cytometry after immune staining and given as relative MFI (means ± SE of 2-5 independent experiments). Relative MFI is the ratio of the MFIs of treated to untreated cells. The significances of differences (P < .05) between HZ-treated (*) or HNE-treated (§) cells and untreated controls are indicated. (C,F) Representative expression profiles of cell-surface HNE adducts measured by flow cytometry in erythroid cells cocultivated with HZ-fed (solid line filled space), RBC-fed (dotted line unfilled space) monocytes (C), HZ (solid line filled space), or treated with HNE (7μM; dotted line unfilled space) (F) for 1 day, and in control erythroid cells (solid line unfilled space). Background is plotted as dashed line.

Inhibition of erythroid cell growth and formation of HNE surface adducts in erythroid cells cocultivated with HZ-fed human monocytes, cocultivated with HZ, or treated with HNE. (A-C) Erythroid cells were cocultivated with unfed (Mo), HZ-fed (Mo + HZ), or RBC-fed (Mo + RBC) monocytes in liquid culture. Adherent monocytes were unfed, fed with either HZ (25μM HZ-heme, corresponding to 12 RBCs/monocyte in terms of heme content), or 50 opsonized RBCs/monocyte for 3 hours. After removal of nonphagocytosed HZ or RBCs, erythroid cells were added to the monocytes at a cell ratio of 1:1, corresponding to a 10:1 ratio at day 4 of coincubation. (A) Growth of erythroid cells was calculated from cell numbers counted in the expanding cultures (means ± SE of 3 independent experiments). (B) HNE-adducts on erythroid cell surface were quantified by flow cytometry after immune staining and expressed as MFI (means ± SE of 3 independent experiments). The significance of differences (P < .05) between erythroid cells cocultivated with HZ-fed monocytes and unfed (*) or RBC-fed (§) monocytes are indicated. (D-F) Erythroid cells were cocultivated with HZ (25μM HZ-heme), treated with HNE (7μM), or kept as untreated controls (CTRL) in liquid culture. (D) Growth of erythroid cells was calculated from cell numbers counted in the expanding cultures (means ± SE of 3 independent experiments). The significance of differences (P < .05) between HZ-treated (*) or HNE-treated (§) cells and untreated controls and (P < .1) between HZ-treated (**) cells and controls are indicated. (E) HNE adducts were quantified by flow cytometry after immune staining and given as relative MFI (means ± SE of 2-5 independent experiments). Relative MFI is the ratio of the MFIs of treated to untreated cells. The significances of differences (P < .05) between HZ-treated (*) or HNE-treated (§) cells and untreated controls are indicated. (C,F) Representative expression profiles of cell-surface HNE adducts measured by flow cytometry in erythroid cells cocultivated with HZ-fed (solid line filled space), RBC-fed (dotted line unfilled space) monocytes (C), HZ (solid line filled space), or treated with HNE (7μM; dotted line unfilled space) (F) for 1 day, and in control erythroid cells (solid line unfilled space). Background is plotted as dashed line.

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