Neonatal neutrophils with prolonged survival secrete mediators associated with chronic inflammation

Abstract

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.

Fig. 1

IL-8 secretion by freshly-isolated and surviving PMN. In paired studies, AD and CB PMN that were freshly isolated (0-hour), or enriched for the annexin V-negative (nonapoptotic, surviving) fraction (24-hour), were cultured for 4 h in the presence of LPS (10 ng/ml final concentration) or media alone, and the cellfree supernatants were tested for IL-8 content. The graph represents individual data points derived from 5–9 separate studies of paired cultures of AD and CB PMN. Labels in the x-axis represent cultures of 0-hour and 24-hour PMN in media alone or in the presence of LPS (+). The horizontal bars represent the mean values for each data set.

Fig. 2

MIP-1β secretion by freshly-isolated and surviving PMN stimulated with LPS. In paired studies, AD and CB PMN that were freshly isolated (0-hour), or enriched for the annexin V-negative (nonapoptotic, surviving) fraction (24-hour), were cultured for 4 h in the presence of LPS (10 ng/ml final concentration) and the cell-free supernatants tested for MIP-1β content. The graph represents individual data points derived from 5 separate studies of paired cultures of AD and CB PMN. Labels in the x-axis represent cultures of 0-hour and 24-hour PMN in the presence of LPS (+). The horizontal bars represent the mean values for each data set.