Background: The resolution of inflammation involves the efficient removal of apoptotic neutrophils (PMN). However, a subpopulation of PMN that are resistant to apoptosis may contribute to PMN persistence in tissues, an early hallmark of chronic inflammation. We previously made observations that neonatal PMN with prolonged survival had augmented expression of CD18/CD11b, an adhesion molecule critical to inflammation. Objectives: The objectives of this study were to test the hypothesis that surviving neonatal PMN retain the capacity to secrete key mediators associated with chronic inflammation. Methods: We profiled cytokine and chemokine secretion patterns of lipopolysaccharide (LPS)-stimulated neonatal and adult PMN using multicytokine array and ELISA. Results: We observed that surviving 24-hour neonatal PMN stimulated with LPS had enhanced secretion of interleukin (IL)-8, a chemokine involved in PMN activation and recruitment. In addition, 24-hour neonatal PMN secreted levels of monocyte inhibitory protein (MIP)-1β that were higher than those secreted by 0-hour PMN, but amounts of IL-1 receptor antagonist (IL-1Ra) were lower. Conclusions: The results of the present study extend previous observations of augmented function in surviving neonatal neutrophils, and further suggest their potential contribution to the pathogenesis of inflammatory disorders in neonates.

Neonatal PMN (PMN) exhibit delayed apoptosis both constitutively and under inflammatory conditions, and evidence has linked PMN longevity to the presence of anti-apoptotic proteins. Activation of the survival-associated transcription factor, nuclear factor kappa B (NF-κB), promotes the synthesis of several anti-apoptotic proteins including Flice inhibitory protein (FLIP). Neonatal and adult PMN were compared in the present studies to test the hypothesis that FLIP modulates age-related apoptosis. Expression of the short isoform, FLIP-S, was prominent at baseline and persisted during spontaneous apoptosis in neonatal PMN, while basal expression was lower and decreased under the same conditions in adult PMN. Stable FLIPS expression in neonatal PMN was associated with a relative resistance to apoptosis in response to the protein synthesis inhibitor, cycloheximide (CHX), or the NF-κB inhibitor, gliotoxin. In contrast, similar treatment of adult PMN promoted greater overall apoptosis accompanied by FLIP degradation. Nuclear levels of phosphorylated p65, a critical NF-κB dimer, were relatively robust in neonatal PMN under basal conditions or following stimulation with TNF-α, a cytokine that induces FLIP. In conclusion, persistent FLIP-S expression is involved in the longevity of neonatal PMN, and our data suggest a contribution of NF-κB signaling and related survival mechanisms.