Naochuxue formula attenuates early brain injury following subarachnoid hemorrhage by inhibiting neuronal apoptosis via network pharmacology and in vivo experiments

Objectives: Investigate Naochuxue formula's mechanism in the treatment of subarachnoid hemorrhage.

Methods: Analyzed Naochuxue formula's active components via nontargeted metabolomics, and predicted the core targets using network pharmacology. Sprague‒Dawley rats were randomly divided into six groups: sham, model, Naochuxue formula low, medium and high dose groups, and edaravone. Neurological deficits were assessed using the modified Garcia score and tissue damage was assessed by measuring the brain water content. Blood‒brain barrier permeability was assessed using the Evans blue procedure and pathological changes in the lesion site were observed through HE staining and Nissl staining. TUNEL staining and Caspase-3 immunofluorescence were used to observe the Apoptosis of neurons in the hippocampus. The distribution and expression of p-PI3K and p-AKT were determined using immunohistochemistry. The expression of the apoptosis-related genes Caspase-3, Bcl-2, and Bax was determined using RT‒PCR.

Key findings: Compared with the model group, rats in the high-dose Naochuxue formula group exhibited significant improvements in neurological defects, brain histopathology, blood‒brain barrier permeability and brain edema on Day 3 posttreatment, downregulated Bax and Caspase-3 expression, and significantly upregulated p-PI3K, p-AKT, and Bcl-2 expression (all P < 0.05).

Conclusions: High-dose formula for 3 days activated PI3K/Akt signaling pathway, inhibitd neuronal Apoptosis, and exerted neuroprotective effects.

PI3K/AKT signaling pathway; apoptosis; early brain injury; naochuxue formula; subarachnoid hemorrhage.