| Background and ObjectivesCerebral small vascular disease(CSVD)refers to a series of clinical,imaging and pathological syndromes,such as small arteries,arterioles,capillaries,venules and small veins,which have no collateral anastomosis.About 25% of ischemic stroke patients in the western countries are CSVD,and some studies show that 50% of them are in China.CSVD often lead to intracerebral space infarction,white matter degeneration,micro hemorrhage,subcortical infarct,brain atrophy,and other symptoms such as dizziness,vertigo,gait disorder,cognitive impairment and so on.At present,most of the animal models of cerebrovascular disease are acute cerebral ischemia models.It is great of significance to construct a model of chronic cerebral ischemia to simulate the clinical CSVD patients.It is great of significance for studing the pathogenesis and treatment of CSVD.Adenosine 5‘monophosphate(AMP)-activated protein kinase is a regulator of cellular energy metabolism.Most studies have shown that AMPK is activated during acute cerebral ischemia,and the consumption of brain ATP accelerates,which aggravates cerebral ischemia injury,especially in the ischemic penumbra area.In the process of chronic cerebral ischemia,the mechanism of the action of AMPK is not clear.Some people think that AMPK activation plays a protective role in chronic cerebral ischemia,and some people think it will aggravate the injury of chronic cerebral ischemia.In this study,we observed the role of AMPK in chronic cerebral ischemia and explored its action mechanism,so as to provide new ideas for finding more effective therapeutic targets of CSVD.Methods1.The bilateral common carotid artery of 3 months old male C57BL/6 mice was ligate with the inner diameter 0.18 mm micro spring.The bilateral common carotid artery stenosis(BCAS)was used to achieve chronic cerebral ischemia in mice,and the cerebral blood flow was monitored during the operation.The sham operation group only had separated bilateral common carotid artery and no micro spring.At 1 and 3 months after operation,Morris water maze and other methods were used to detect the cognitive function of mice.Magnetic resonance imaging(MRI)was performed before the execution.The brain tissues were paraffin fixed and stained for histological examination.2.After BCAS,mice were given intragastric administration of AMPK agonist(metformin)(250mg/kg)every day.Morris water maze and other behavioral tests were performed in 1 and 3 months after operation.The head MRI was performed before the execution.The brain tissues were paraffin fixed and stained for histological examination.3.AMPK alpha 2 gene knockout mice were constructed by Cre-loxp recombinant enzyme system,and PCR,Western blot and immunohistochemistry were used to verify the effect of gene knockout.The 3 months old male AMPK alpha 2 knockout mice were treated with BCAS,and brain damage was detected by behavioral and histological methods in mice.Results1.After BCAS,the mice’s learning and memory ability were decreased,and gradually increased with the prolongation of ischemia time.Demyelination,astrocyte proliferation and oligodendrocytes decrease appear in mice brain.The above results indicate that BCAS can induce chronic cerebral ischemia in mice,and successfully prepare chronic cerebral ischemia model.2.After BCAS,the mice were treated with AMPK activator metformin,and it was found that the learning and memory impairment of the mice was lighter than that the unfilled medicine mice.The demyelination of brain tissue and the number of glial cells were also lighter than those in the unfilled medicine mice.It indicates that activation of AMPK can improve chronic cerebral ischemia injury induced by BCAS in mice.3.Compared with the wild type(WT)C57BL/6 male mice after BCAS,the learning and memory ability of the AMPK alpha 2 knockout male mice after operation was markedly reduced,and the chronic cerebral ischemia injury in the brain tissue was obvious,especially the glial cell proliferation.It indicates that the absence of AMPK will aggravate chronic cerebral ischemia in mice.Conclusion1.Micro spring ligation of bilateral CCA in mice can successfully prepare chronic cerebral ischemia models.2.Activating AMPK activity in mice has the effect of resistance chronic cerebral ischemia injury.3.Knockout mouse AMPK alpha 2 gene function aggravates chronic cerebral ischemia injury. |
