DLL4-Notch1 inhibitory peptide (NDI) prevents sepsis-induced endothelial barrier disruption and detrimental neutrophil accumulation.

Sepsis remains a leading cause of death, driven by systemic inflammation and organ injury. A key contributor is the accumulation of hyperactive ICAM1hiCXCR1lo "reverse migrated" neutrophils. Recently, a distinct DLL4+ neutrophil subset was identified in the lungs, exacerbating lung injury, but its precise mechanism of action was unclear. Understanding how these specific neutrophils contribute to endothelial barrier disruption and promote further detrimental neutrophil migration is crucial for developing targeted sepsis therapies.

Researchers investigated how DLL4+ neutrophils activate alveolar macrophages (AMs) to disrupt endothelial barriers and promote neutrophil reverse migration in a cecal ligation and puncture (CLP) induced sepsis model. AMs were treated with DLL4+ neutrophils or recombinant mouse DLL4 (rmDLL4). ADAM17 mRNA and protein levels in AMs were assessed. Conditioned medium from treated AMs was applied to pulmonary vascular endothelial cells (PVECs) to measure junctional adhesion molecule-C (JAM-C) protein via Western blot. ICAM1hiCXCR1lo neutrophils were detected by flow cytometry. A small-molecule inhibitor of ADAM17 and a novel DLL4-Notch1 inhibitory peptide (NDI) were also tested.

During CLP-induced sepsis, DLL4+ neutrophils were found to interact with AMs via the Notch1 pathway, significantly increasing ADAM17 expression. This elevated ADAM17 subsequently decreased JAM-C on PVECs, leading to endothelial barrier disruption and the generation of detrimental ICAM1hiCXCR1lo neutrophils. The researchers demonstrated that a small-molecule inhibitor of ADAM17 effectively preserved pulmonary endothelial barrier integrity and reduced ICAM1hiCXCR1lo neutrophil accumulation, confirming the central role of ADAM17 in this pathway.

Importantly, the novel DLL4-Notch1 inhibitory peptide (NDI) effectively suppressed ADAM17 expression, restored JAM-C levels, and reduced the accumulation of ICAM1hiCXCR1lo neutrophils in the sepsis model. These findings highlight a critical inflammatory cascade initiated by DLL4+ neutrophils that exacerbates systemic inflammation and worsens sepsis outcomes.