IL-17A Promotes NETs Formation via the PKCζ-ERK-ROS-PAD4 Pathway in a Mouse Model of Ischemic Stroke.

AIMS: Interleukin-17A (IL-17A) aggravates poststroke neurological damage and enhances neutrophil extracellular traps (NETs) formation, yet the underlying mechanism remains unclear. This study aimed to elucidate how IL-17A regulates NETs generation after ischemic stroke. METHODS: Using a mouse middle cerebral artery occlusion (MCAO) model, we administered the Peptidylarginine deiminase 4 (PAD4) inhibitor GSK484 or an IL-17A-neutralizing antibody (IL-17mAb). Infarct volume and neurological function were assessed, and protein expression of PAD4, myeloperoxidase (MPO), citrullinated histones H3 (CitH3), protein kinase Cζ (PKCζ), and phosphorylated extracellular signal-regulated kinase (p-ERK) was evaluated. IL-17A(-)/(-) mice and primary neutrophils were used to further validate the signaling pathway. RESULTS: Inhibiting PAD4 with GSK484 can significantly reduce infarct size, improve neurological outcomes, and decrease PAD4, MPO, and CitH3 protein levels. Subsequently, we found that IL-17mAb treatment can reduce the expression of PAD4, MPO, and CitH3 in the peri-infarct area, as well as the expression of PKC-ζ and p-ERK. Similarly, the expression of PAD4 and CitH3 decreased in the peri-infarct area of IL-17A(-)/(-) MCAO mice. Finally, we verified IL-17A induced PAD4 upregulation via the PKCζ-ERK-ROS axis in vitro. CONCLUSION: IL-17A promotes NETs formation by upregulating PAD4 through the PKCζ-ERK-ROS pathway, exacerbating ischemic brain injury. Targeting this axis may offer a novel therapeutic strategy for stroke.