Figure 1.

PBEF is expressed in neutrophils and monocytes exposed to inflammatory stimuli. (A) Neutrophils expressed mRNA transcripts for PBEF in response to LPS, TNF-α, and IL-1β, stimuli that inhibit neutrophil apoptosis. (B) LPS (1 ∝g/ml) induced transcripts for caspase-1 (open circles), reaching maximal concentrations by 1 hour, IL-1β (filled circles) peaking at 3 hours, and PBEF (filled triangles) reaching maximal levels at 10 hours. The mRNA expression was quantified by real-time PCR; data are expressed as fold increase over basal levels of expression for each mRNA species; n = 4. (C) Western blot analysis showed low-level expression of PBEF in control cells, while LPS (1 ∝g/ml) increased PBEF protein, with maximum expression evident 10 hours after exposure. Blots were reprobed with β-actin to confirm equal loading; data are representative of three separate experiments. (D) PBEF mRNA transcripts were also expressed in peripheral blood monocytes, but not lymphocytes, in response to LPS, TNF, and IL-1. Blots are representative of three separate experiments; corresponding mRNA for GAPDH is shown to evaluate sample loading. (E) HL-60 cells induced to granulocytic differentiation by 1 ∝M all-trans retinoic acid showed increased message for PBEF, and transcript levels evaluated by real-time PCR were further increased by exposure of differentiated HL-60 cells to LPS (1 ∝g/ml) added to cultures with all-trans retinoic acid at day 0. Data are normalized to levels of PBEF transcripts in undifferentiated HL-60 cells; *P < 0.05 versus all-trans retinoic acid alone; n = 4. ATRA, all-trans retinoic acid.