Traditional Chinese Medicine formula, Sanwujiao granule, attenuates ischemic stroke by promoting angiogenesis through early administration

Ischemic stroke (IS) is one of the most lethal diseases with the insufficient pharmacology therapeutic approach. Sanwujiao granule (SW) is widely used for IS in China with little known about its underlying mechanism. To investigate the characteristics of therapeutic effects and potential mechanisms of SW against IS. The fingerprint of SW was applied by high-performance liquid chromatography-mass spectrometry (HPLC-MS). Three different drug treatment strategies, including prophylactic administration, early administration and delayed administration, were applied in rats' permanent middle cerebral occlusion (pMCAO) model. The Garcia neurological deficit test, adhesive removal test, rotarod test, TTC and TUNEL staining were performed to evaluate the pathological changes. The transcriptomic analysis was used to predict the potential mechanism of SW. The vascular deficiency model of Tg(kdrl:eGFP) zebrafish larvae and oxygen-glucose deprivation model on bEnd.3 cells were used to verify SW's pharmacological effect. qRT-PCR, immunofluorescent staining and Western Blot were applied to detect the expression of genes and proteins. The network pharmacology approach was applied to discover the potential bioactive compounds in SW that contribute to its pharmacological effect. SW early and delayed administration attenuated cerebral infarction, neurological deficit and cell apoptosis. The transcriptomic analysis revealed that SW activated angiogenesis-associated biological processes specifically by early administration. CD31 immunofluorescent staining further confirmed the microvessel intensity in peri-infarct regions was significantly elevated after SW early treatment. Additionally, on the vascular deficiency model of zebrafish larvae, SW showed the angiogenesis effect. Next, the cell migration and tube formation were also observed in the bEnd.3 cells with the oxygen-glucose deprivation induced cell injury. It's worth noting that both mRNA and protein levels of angiogenesis factor, insulin-like growth factor 1, were significantly elevated in the pMCAO rats' brains treated with SW. The network pharmacology approach was applied and chasmanine, karacoline, talatisamine, etc. were probably the main active compounds of SW in IS treatment as they affected the angiogenesis-associated targets. These results demonstrate that SW plays a critical role in anti-IS via promoting angiogenesis through early administration, indicating that SW is a candidate herbal complex for further investigation in treating IS in the clinical.