Comprehensive Evaluation of YJ-2 as a PAD4 Inhibitor in Alleviating Ischemic Brain Injury: From NETs-Induced Neurotoxicity to In Vivo Neuroprotection.
To evaluate the neuroprotective potential of YJ-2, a novel peptidylarginine deiminase 4 (PAD4) inhibitor, against ischemia/reperfusion brain injury by targeting neutrophil extracellular trap (NET) formation.
In vitro, a NETs-induced injury model was established using SH-SY5Y and bEnd.3 cells. YJ-2's effects on viability, apoptosis, oxidative stress, and barrier permeability were assessed via CCK-8, flow cytometry, and FITC-dextran assays. In vivo, a rat middle cerebral artery occlusion/reperfusion (MCAO/R) model received YJ-2 (10 μmol/kg) intravenously. Outcomes included infarct volume (TTC staining), neurological score, neuronal apoptosis (TUNEL), and oxidative markers (ELISA). PAD4 activity and histone H3 citrullination (H3cit) were examined by western blot and immunofluorescence.
YJ-2 reduced NET-mediated neuronal death and oxidative stress in vitro, and improved endothelial barrier integrity. In MCAO/R rats, YJ-2 significantly lowered infarct volume (44.2% → 30.6%), improved neurological function, and suppressed apoptosis. It also decreased PAD4 and H3cit expression in ischemic brain tissue, confirming target engagement.
YJ-2, by preserving blood-brain barrier (BBB) integrity and reducing neuronal apoptosis, highlights its therapeutic potential for ischemic stroke.
In vitro, a NETs-induced injury model was established using SH-SY5Y and bEnd.3 cells. YJ-2's effects on viability, apoptosis, oxidative stress, and barrier permeability were assessed via CCK-8, flow cytometry, and FITC-dextran assays. In vivo, a rat middle cerebral artery occlusion/reperfusion (MCAO/R) model received YJ-2 (10 μmol/kg) intravenously. Outcomes included infarct volume (TTC staining), neurological score, neuronal apoptosis (TUNEL), and oxidative markers (ELISA). PAD4 activity and histone H3 citrullination (H3cit) were examined by western blot and immunofluorescence.
YJ-2 reduced NET-mediated neuronal death and oxidative stress in vitro, and improved endothelial barrier integrity. In MCAO/R rats, YJ-2 significantly lowered infarct volume (44.2% → 30.6%), improved neurological function, and suppressed apoptosis. It also decreased PAD4 and H3cit expression in ischemic brain tissue, confirming target engagement.
YJ-2, by preserving blood-brain barrier (BBB) integrity and reducing neuronal apoptosis, highlights its therapeutic potential for ischemic stroke.
Authors
Jia Jia, Taledaohan Taledaohan, Wang Wang, Tuohan Tuohan, Chen Chen, Chan Chan, Shen Shen, Zhu Zhu, Wang Wang
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