Heyu Yang,1– 3,* Bingqian Luo,1– 3,* Yifan Du,3,4,* Jiafu Guo,1– 3 Shiqi Zhang,1– 3 Ping Wang,1– 3 Yuan Dai,3,4 Yun Lu,3,5 Shijun Xu1– 3 

1State Key Laboratory of Southwestern Chinese Medicine Resources, Chengdu University of Traditional Chinese Medicine, Chengdu, Sichuan, 611137, People’s Republic of China; 2School of Pharmacy, Chengdu University of Traditional Chinese Medicine, Chengdu, Sichuan, 611137, People’s Republic of China; 3Institute of Material Medica Integration and Transformation for Brain Disorders, Chengdu University of Traditional Chinese Medicine, Chengdu, Sichuan, 611137, People’s Republic of China; 4School of Health Preservation and Rehabilitation, Chengdu University of Traditional Chinese Medicine, Chengdu, Sichuan, 611137, People’s Republic of China; 5Hospital of Chengdu University of Traditional Chinese Medicine, Chengdu, Sichuan, 610072, People’s Republic of China

*These authors contributed equally to this work

Correspondence: Yun Lu, Email luyun999@126.com Shijun Xu, Email xushijun@cdutcm.edu.cn

Background: Perihematomal neuroinflammation serves as a pivotal pathogenic driver of secondary brain injury during the acute stage of intracerebral hemorrhage (ICH). The traditional Chinese medicine Zhongfeng Xingnao Liquid exhibits anti-neuroinflammatory effects. This study aims to elucidate the optimal timing for ZFXN administration within hours of symptom onset and its underlying mechanisms, focusing on NF-κB/NLRP3-mediated neuroinflammation.
Methods: ICH was induced by injection of autologous arterial blood into the left caudal nucleus. Neurological deficits scores, hematoma volume, cerebral blood flow (CBF), H&E and Nissl staining were conducted at 24 hours following ICH. The levels of neuroinflammation response and NF-κB/NLRP3 axis surrounding the hematoma were measured using immunofluorescent staining and Western blot. The inhibition of ZFXN on NF-κB/NLRP3 axis was further confirmed in Lipopolysaccharide (LPS)-induced BV-2 cells.
Results: Post-ICH pathology was characterized by progressive hematoma expansion, elevated neurological deficit scores, neuronal damage, and reduced CBF, accompanied by neuroinflammatory. Early ZFXN intervention within 6 hours post-ICH significantly reduced hematoma volume and improved neurological scores (mNSS, Bederson, Zea Longa) at 24 hours, while markedly alleviating perihematomal neuronal damage and enhancing CBF, with optimal efficacy observed following one-hour administration. The treatment also effectively suppressed IL-1β/TNF-α release and microglial activation through NF-κB/NLRP3 pathway inhibition. Consistently, ZFXN diminished NF-κB-p65 nuclear translocation and downregulated NLRP3 inflammasome components (ASC, Cleaved Caspase-1) in LPS-stimulated BV-2 cells.
Conclusion: ZFXN emerges as a promising neuroprotective agent for ICH through targeted inhibition of the NF-κB/NLRP3 inflammatory axis, demonstrating optimal efficacy within the critical 6-hour hyperacute phase by mitigating secondary neuroinflammation and addressing current therapeutic gaps in ICH management.

Keywords: Zhongfeng Xingnao liquid, intracerebral hemorrhage, neuroinflammation, microglia, NF-κB/NLRP3 axis