Interleukin-10 does not directly affect the constitutive rate of human neutrophil or eosinophil apoptosis.

Abstract

IL-10 is a cytokine which is produced by most inflammatory cells and has been implicated as a potent anti-inflammatory regulator of the immune response [ l , 21. In addition, IL-10 suppresses proinflammatory cytokine biosynthesis and release (e.g., TNF-a) from inflammatory cells including neutrophils and eosinophils and reduces LPSinduced eosinophil survival [3,4]. The rate of eosinophil and neutrophil apoptosis can be modulated by cytokines such as GMCSF and TNF-a [5-7l. Thus IL-10 may be involved in the control of apoptosis, by directly acting on these cells via inhibition of cytokine synthesis or indirectly by influencing cytokine secretion from other inflammatory cells; alternatively it may act in concert with other modulators of apoptosis. This study investigated the direct effect of IL-10 on neutrophil and eosinophil apoptosis. Human neutrophils or eosinophils were isolated from the peripheral blood of health donors. Harvested cells were suspended in Iscove's modified Dulbecco's medium supplemented with 10% autologous serum as previously described [8]. Granulocytes were incubated at a density of 2 x 106/ml in 96-well flat-bottomed plates at 37 OC in a 5% CO2 atmosphere. Cells were cultured with IL-10 at concentrations of 1,5 or 10 ng/ml or in medium alone. Cytocentrifuge preparations (in triplicate) were made of harvested cells and assessed for apoptosis (YO mean f SEM). Cells with one or more darkly stained pyknotic nuclei were considered apoptotic. At least 5 fields of view (over 500 cells) were counted per cytospin. Cell viability was assessed by hypan blue exclusion. To demonstrate that the IL-10 used in this study was biologically active, we investigated the effects of this cytokme on the production of TNF-a by human blood-derived monocytes and macrophages.