Reduction in neutrophil cell surface expression of tumor necrosis factor receptors but not Fas after transmigration - Implications for the regulationof neutrophil apoptosis

Objectives: To test the hypothesis that loss of polymorphonuclear neutrophi l tumor necrosis factor alpha (TNF-alpha) receptors during transmigration r enders the exudate neutrophil refractory to TNF-alpha-mediated stimulation of apoptosis; and to investigate the surface expression of Fas on both circ ulating and exudate neutrophils. Design: A prospective cohort study. Setting: Surgical laboratory of a tertiary care hospital. Participants: Twenty-one healthy human volunteers. Interventions: All subjects had circulating neutrophils and exudate neutrop hils collected by venipuncture and skin window methods, respectively. Main Outcome Measures: Circulating and exudate neutrophils were incubated i n culture medium (1.0 x 10(6) neutrophils per milliliter) alone or with TNF -alpha (100 ng/mL). Apoptosis was evaluated by flow cytometry (annexin V-fl uorescein isothiocyanate and propidium iodide). Tumor necrosis factor alpha -phycoerythrin and anti-human Fas-fluorescein isothiocyanate were used to e valuate neutrophil TNF-alpha receptors and surface expression of Fas. Results: Exudate neutrophils had a significant delay in apoptosis races whe n compared with circulating neutrophils. The percentage of neutrophils expr essing TNF-alpha receptors was significantly diminished after exudation (80 % +/- 15% vs 33% +/- 9%; P<.001), as was the median channel number of TNF-a lpha phycoerythrin fluorescence (8.1 +/- 1.6 vs 5.2 +/- 0.5; P = .001). How ever, the expression of Fas was unchanged after transmigration (percentage positive for Fas: 98.7% +/- 0.7% vs 92.8% +/- 3.4%, P = .89; Fas antibody-f luorescein isothiocyanate median channel fluorescence: 12.2 +/- 1.1 vs 13.1 +/- 1.2; P = .80). Exposure of exudate neutrophils to TNF-alpha failed to increase their rate of apoptosis. Conclusions: Exudate polymorphonuclear neutrophils are confirmed to have de layed apoptosis. Loss of TNF-alpha receptors during transmigration is neces sary for neutrophil survival in the extravascular inflammatory milieu.