Abstract: BACKGROUND: Previous studies by the research team have shown that the neuroprotective effect of ginsenoside Rb1 on improving cerebral ischemia-reperfusion injury may be related to the Wnt/β-catenin signaling pathway, but the specific mechanism of action remains unclear.
OBJECTIVE: To explore the molecular mechanism of ginsenoside Rb1 in alleviating cerebral ischemia-reperfusion injury in mice.
METHODS: 100 C57BL/6 mice were randomly divided into four groups. The sham operation group (n=25) did not undergo model establishment. In the cerebral ischemia-reperfusion injury group (n=25), the middle cerebral artery occlusion model was established by thread embolism. In the ginsenoside Rb1 group (n=25), the middle cerebral artery occlusion model was established by thread embolism and ginsenoside Rb1 was intraperitoneally injected every day for 3 days before the establishment. In ginsenoside Rb1+inhibitor group (n=25), the middle cerebral artery occlusion model was established by thread embolism and ginsenoside Rb1 and Wnt/β-catenin signaling pathway inhibitor XAV939 were intraperitoneally injected every day for 3 days before the establishment. Three days after modeling, Zea Longa score and balance beam test were used to evaluate the neurological deficits of mice. TTC staining was used to observe the volume of cerebral infarction. The dry-wet mass method was used to detect the degree of brain edema in mice. The activities of superoxide dismutase and glutathione peroxidase and the concentration of malondialdehyde in the parietal lobe of the ischemic side were detected by colorimetry. The co-expression of microglial marker Iba1 and inducible nitric oxide synthase (or arginase 1) was detected by immunofluorescence. The levels of aquaporin AQP4, inflammatory-related factors, and phosphorylation of Wnt/β-catenin pathway proteins glycogen synthase kinase 3β and β-catenin were detected by western blot assay. The mRNA expression of inflammatory factors was detected by q-PCR.
RESULTS AND CONCLUSION: (1) Compared with cerebral ischemia-reperfusion injury group, the neurological deficit symptoms, cerebral infarction foci and brain tissue edema, oxidative stress and inflammatory response of the mice were alleviated; the number of M1 microglia (Iba1 and inducible nitric oxide synthase co-expressed) decreased; the number of M2 microglia (Iba1 and arginase 1 co-expressed) increased; the expression of phosphorylated glycogen synthase kinase 3β and phosphorylated β-catenin protein decreased in ginsenoside Rb1 group. Compared with ginsenoside Rb1 group, the neurological deficit symptoms, cerebral infarction foci and brain tissue edema, oxidative stress and inflammatory response of the mice were aggravated; the number of M1 microglia increased; the number of M2 microglia decreased; the expression of phosphorylated glycogen synthase kinase 3β and phosphorylated β-catenin protein increased in ginsenoside Rb1+inhibitor group. (2) The results indicate that ginsenoside Rb1 can regulate the polarization of microglia to M2 type and alleviate oxidative stress damage and inflammatory response after cerebral ischemia-reperfusion injury. Its mechanism may be related to the nuclear translocation of β-catenin mediated by the Wnt/β-catenin signaling pathway.

Key words: cerebral ischemia-reperfusion injury, ginsenoside Rb1, microglia, Wnt/β-catenin signaling pathway, neuroinflammation, oxidative stress, neuroprotection, engineered tissue construction