Distinct Functional States of Neutrophils by Actin Disassembly and NF-kB/STAT3 Signaling

Polymorphonuclear neutrophils (PMNs) can differentiate into distinct states, which can either exacerbate or resolve inflammation. Our study shows that mice challenged with TLR agonists exhibited PMN differentiation along two major paths characterized by the expression of CCR5 or PD-L1. Similar differentiation was observed in the blood of severe COVID-19 patients and the synovial fluid of osteoarthritis patients. Prolonged in vitro priming of human PMNs modeled the differentiation paths. Actin disassembly favored CCR5 upregulation, while NF-kB activation stabilized the actin cytoskeleton and suppressed the development of CCR5+ PMNs. Additionally, PD-L1 upregulation was triggered by STAT3 signaling and NF-kB activation. Functionally, CCR5 expressing PMNs were pro-NETotic, while PD-L1+ PMNs showed immunosuppressive functions by inhibiting T cell proliferation via PD1. Together, PMN differentiation depended on the priming conditions, and the balance between actin disassembly and NF-kB/STAT3 activation translated the present micro-milieu into phenotypic and functional diversification of PMNs. SynopsisNeutrophils underwent phenotypical and functional diversification both in vivo and in vitro. Actin disassembly led to the generation of CCR5high neutrophils with increased spontaneous NETosis, whereas NF-kB and STAT3 induced PD-L1 expression with T-cell suppressive properties as a deviation from the default pathway. O_LIPMN of mice challenged with TLR agonists develop two distinct phenotypes, CCR5high and PD-L1high. C_LIO_LICCR5 and PD-L1-defined neutrophil phenotypes were found in blood of patients with severe COVID-19 and in the synovial fluid of osteoarthritis patients. C_LIO_LIIn vitro priming induced a similar bifurcation of PMN phenotypes marked by either CCR5 or PD-L1. C_LIO_LIActin disassembly preceded canonical development of CCR5+ PMN. C_LIO_LINF-kB halted actin disassembly by LPL regulation. C_LIO_LIDuring neutrophil priming, STAT3 aided NF-kB in the expression of PD-L1. C_LI Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=159 HEIGHT=200 SRC="FIGDIR/small/548975v1_ufig1.gif" ALT="Figure 1"> View larger version (29K): org.highwire.dtl.DTLVardef@10305f3org.highwire.dtl.DTLVardef@17bada8org.highwire.dtl.DTLVardef@c61021org.highwire.dtl.DTLVardef@14ce7f3_HPS_FORMAT_FIGEXP M_FIG C_FIG