Siglec-9 and SHP-1 Are Differentially Expressed in Neonatal and Adult Neutrophils

Abstract

Neonatal PMN (polymorphonuclear neutrophils) exhibit altered inflammatory responsiveness and greater longevity compared with adult PMN; however, the involved mechanisms are incompletely defined. Receptors containing immunoreceptor tyrosine-based inhibitory motif (ITIM) domains promote apoptosis by activating inhibitory phosphatases, such as Src homology domain 2-containing tyrosine phosphatase-1 (SHP-1), that block survival signals. Sialic acid-binding immunoglobulin-like lectin (Siglec)-9, an immune inhibitory receptor with an ITIM domain, has been shown to induce cell death in adult PMN in association with SHP-1. To test our hypothesis that neonatal PMN inflammatory function may be modulated by unique Siglec-9 and SHP-1 interactions, we compared expression of these proteins in adult and neonatal PMN. Neonatal PMN exhibited diminished cellular expression of Siglec-9, which was phosphorylated in the basal state. Granulocyte-macrophage colony-stimulating factor (GM-CSF) treatment decreased Siglec-9 phosphorylation levels in neonatal PMN but promoted its phosphorylation in adult PMN, observations associated with altered survival signaling. Although SHP-1 expression was also diminished in neonatal PMN, GM-CSF treatment had minimal effect on phosphorylation status. Further analysis revealed that Siglec-9 and SHP-1 physically interact, as has been observed in other immune cells. Our data suggest that age-specific interactions between Siglec-9 and SHP-1 may influence the altered inflammatory responsiveness and longevity of neonatal PMN. invasion and contribute to the clearance of pathogens. During resolution of the inflammatory process, neutrophils undergo apoptosis and are removed through ingestion by resident macrophages (1,2). However, a delay in the clearance of activated neutrophils promotes their persistence in tissues, a contributing factor in the pathogenesis of a variety of chronic inflammatory conditions (3). We reported a prolonged survival of neonatal neutrophils with enhanced inflammatory and cytotoxic activity (4,5). Altered inflammatory responses and delayed apoptosis may contribute to a variety of chronic inflammatory disorders in neonates, including bronchopulmonary dysplasia (6 -8). Understanding the mechanisms modulating neonatal neutrophil responses to inflammation is therefore paramount in designing targeted therapeutic approaches.Neonatal neutrophils have an intrinsic resistance to spontaneous and Fas-mediated apoptosis associated with decreased expression of proapoptotic proteins (4,9,10); however, the underlying mechanisms remain unclear. Compromise of this neutrophil function in neonates suggests a dysregulation of mechanisms important to inflammation and its resolution. Normally, the initiation and termination of inflammatory processes are tightly regulated through a combination of activating and inhibitory signals (11). An imbalance in this regulation could result in prolongation of processes that induce tissue damage and neutrophil reactivation, setting the stage for chronic infla...

Neonatal Neutrophils with Prolonged Survival Secrete Mediators Associated with Chronic Inflammation

“…Thus, the heightened expression of IL-8 in neonatal neutrophils may be related to alterations in regulatory signaling pathways. We recently reported prominent Akt-mediated survival signaling [42] and have observed a unique pattern of NF-B-regulated function in neonatal neutrophils (manuscript in preparation). In contrast to the pattern of IL-8 secretion, surviving neutrophils released negligible amounts of other inflammatory cytokines, including IL-1 ␣ , IL-1 ␤ , IL-6 and TNF-␣ .…”

Role of siglecs and related glycan-binding proteins in immune responses and immunoregulation

“…19, 22 Similarly, Siglec-9-induced neutrophil cell death is caspase and ROS-dependent in resting cells but only ROS-dependent in activated cells. 23 However, Siglec-9 and Siglec-E also have ITIM/SHP-1/inhibitory signaling related functions in that both are constitutively associated with SHP-1 24, 25 and Siglec-E-deficient mice exhibit enhanced neutrophil recruitment in endotoxin-induced acute pulmonary inflammation. 25 Intriguingly, Siglec-8 signaling and function in a different cell type, namely mast cells, is distinct from that in eosinophils and involves inhibition of mast cell FcεRI-mediated activation and function rather than cell death.…”

Neonatal Sepsis and Neutrophil Insufficiencies

“…They found that the prolonged survival of neonatal neutrophils was due to less abundant sialic acid-binding immunoglobin-like lectin-9 (Siglec-9) and Src homology domain-2 containing tyrosine phosphatase-1 (SHP-1), two pro-apoptotic proteins. Rashmi suggested that diminished SHP-1 function in neonatal neutrophils can explain the high basal phosphorylation of LYN previously reported by Yan et al [47]. Marodi et al investigated a third intracellular signaling pathway, the STAT family of proteins, using neonatal monocytes.…”

“…Pretreatment of cord blood neutrophils with anti-Fas antibodies induced significantly less apoptosis (24% v. 63%) than seen in adult neutrophils [110]. The limited responsiveness to apopototic stimuli is related to decreased surface receptor expression of the Fas receptor, decreased pro-apoptotic intracellular signaling molecule caspase-3, and decreased activity of the pro-apoptotic proteins Siglec-9 and SHP-1[47, 111, 112]. Adult and neonatal cells were thought to be equally responsive to anti-apoptotic factors including GM-CSF, IFN-γ, and LPS [111].…”