Background/Purpose: Neutrophil activation has been implicated to contribute to the systemic lupus erythematosus (SLE) pathogenesis. However, factors and mechanisms promoting neutrophil activation in SLE have not been fully explored. We previously demonstrated that ribonucleoprotein (RNP) immune complexes (ICs) promoted neutrophil activation in a TLR7/8 manner. Still, whether this process occurs in patients, and the pathways downstream of IC activation in human neutrophils have not been clearly defined. In this study, we aim to determine the role of RNA recognition by TLR7/8 in plasma-mediated neutrophil activation in SLE.

Methods: Plasma neutrophil activation markers (calprotectin, MPO-DNA complexes, and NE-DNA complexes) were detected in SLE patients (n=151) and healthy controls (HCs, n=31) by ELISAs.  Circulating IC levels were measured by flow cytometry. Neutrophils, isolated from healthy controls, were incubated with TLR agonists or plasma from SLE patients or HCs, and assessed for CD66b and CD11b expression by flow cytometry in presence or absence of TLR inhibitors, RNase, or Fc gamma receptor (FcγR) II inhibitors to define mechanisms of neutrophil activation by lupus plasma.

Results: SLE patients have increased plasma levels of calprotectin, MPO-DNA complexes, and NE-DNA complexes when compared to HCs (p< 0.05, p< 0.001, and p< 0.05, respectively). Plasma from SLE patients induced higher levels of cell surface CD66b (p=0.0002) and CD11b (p=0.01) expression than those from HCs. These data suggest presence of soluble components in SLE circulation able to induce de novo neutrophil activation. Targeting ICs by inhibiting FcgRIIA, and/or targeting RNA sensing by adding RNase, and/or blocking TLR7/8, TLR8 only, or IRAK4, markedly decreased plasma-mediated neutrophil activation (all p< 0.05). Consistent with the ability of selective TLR8 inhibitor to block plasma-mediated neutrophil activation, TLR8 agonists, but not TLR7 agonists induced neutrophil activation. Further, neutrophil mRNA expression of TLR8 was significantly higher than for TLR7. Finally, SLE patients that had plasma samples showing reduced capacity to promote neutrophil activation upon RNase treatment also had increased markers of inflammation, including higher levels of interferon alpha, IP-10 (p< 0.05), ICs (p< 0.05), and reduced complement C3 levels (p< 0.01), indicative of IC-driven disease.

Conclusion: These data support the concept of IC-driven activation of RNA-sensing TLR8 in neutrophils as one of the main mechanisms of neutrophil activation in human lupus. Patients with excess neutrophil activation and/or presence of RNA-containing ICs, may benefit from selective TLR8 inhibition and/or other strategies to target RNA removal.

« Back to ACR Convergence 2024

ACR Meeting Abstracts - https://acrabstracts.org/abstract/immune-complexes-mediated-activation-of-neutrophils-in-systemic-lupus-erythematosus-is-dependent-on-rna-recognition-by-tlr8/