Comprehensive evaluation of Dragon's Blood in combination with borneol in ameliorating ischemic/reperfusion brain injury using RNA sequencing, metabolomics, and 16S rRNA sequencing

Ischemia/reperfusion (IR) can induce deleterious responses such as apoptosis, inflammation, and oxidative stress; however, there are currently no efficient therapeutics to treat IR brain injury. Dragon's blood (DB) plays a significant role in treating ischemic stroke in China. Borneol (B) is an upper ushering drug that guides drugs to the target organs, including the brain. Therefore, we hypothesized that the combination of DB and B (DB + B) would provide cooperative therapeutic benefits for IR brain injury. To confirm this, we first investigated the protective effect of DB + B in an IR brain injury rat model using the modified neurological severity score (mNSS), infarction size measure, HE staining, and laser speckle contrast imaging. Then, we comprehensively evaluated the mechanism of DB + B in ameliorating IR brain injury based on RNA sequencing, serum untargeted metabolomics, and 16S rRNA sequencing. We have confirmed that DB + B enhanced the efficacy of the ischemic stroke treatment compared to DB or B alone for the first time. Our study provisionally confirms that the mechanism by which DB + B prevents IR brain injury is related to the maintenance of intestinal microecological balance and regulation of metabolic dysfunction, thereby suppressing inflammation and regulating immunity. DB + B may effectively regulate intestinal flora including o_Pseudomonadales, s_Bacteroides_caecimuris, o_unidentified_Bacilli, f-Pseudomonadaceae, and g-Pseudomonas, mainly regulate serum metabolites including improve the protective benefit of IR brain injury lysoPCs and lysoPEs, thus inhibiting TLR4/MyD88/NF-kappa B and IL-17 signing pathway to reduce inflammatory reactions. hat this mechanism is associated with the maintenance of intestinal flora balance and the regulation of metabolic dysfunction, thereby suppressing inflammation and regulating immunity. This provides scientific support for the clinical translation of DB + B in the prevention and treatment of ischemic stroke and establishes a basis for further investigation of its therapeutic mechanism.