miR-379-5p改善大鼠脑缺血再灌注损伤的机制研究OA

Objective To investigate the effect of miR-379-5p on cerebral ischemia-reperfusion injury(CIRI)in rats and its underlying mechanism.Methods The CIRI rat model was established by transiently blocking the middle cerebral artery.The rats were randomly divided into the model group,the miR-379-5p agonist group(injection of miR-379-5p agonist into the lateral ventricle),the miR-379-5p antagonist group(injection of miR-379-5p antagonist into the lateral ventricle),the gasdermin D(GSDMD)inhibitor group(injection of 80 nmol necrosulfonamide into the lateral ventricle).In addition,rats with only the internal and external carotid arteries exposed were designated as the sham-operated group.The modified neurological severity score(mNSS)was used to evaluate the degree of neurological deficits of rats in each group.2,3,5-triphenyltetrazolium chloride(TTC)staining and hematoxylin-eosin(HE)staining were used to observe the cerebral infarction and morphological changes in the brain tissues of rats in each group.TUNEL staining was used to detect the neuronal apoptosis in the brain tissues of rats in each group.RT-qPCR was used to detect the expression level of miR-379-5p in the brain tissues of rats in each group.Immunofluorescence staining and Western blot were used to detect the fluorescence expression of GSDMD and the protein expression levels of NOD-like receptor protein 3(NLRP3),apoptosis-associated speck-like protein containing a CARD(ASC),pro-Caspase-1,GSDMD and N-terminal active fragment of GSDMD(GSDMD-N)in the brain tissues of rats in each group.Results Compared with the model group,the mNSS scores of rats in the miR-379-5p agonist group and the GSDMD inhibitor group were decreased,the infarct area of brain tissue was reduced,the disordered arrangement of neurons,cell swelling,and vacuolation in the cortical brain tissue were significantly improved,the neuronal apoptosis index was decreased,the miR-379-5p expression level was upregulated,the mean fluorescence density of GSDMD and the protein expression levels of NLRP3,ASC,pro-Caspase-1,GSDMD,and GSDMD-N were downregulated,and all these differences were statistically significant(P<0.05).Compared with the model group,the mNSS score of rats in the miR-379-5p antagonist group was increased,the infarct area of brain tissue was increased,the disordered arrangement of neurons,cell swelling and vacuolation were aggravated,the neuronal apoptosis index was increased,the miR-379-5p expression level was downregulated,the mean fluorescence density of GSDMD and the protein expression levels of NLRP3,ASC,pro-Caspase-1,GSDMD,GSDMD-N were further upregulated,and all these differences were statistically significant(P<0.05).Conclusion Upregulation of miR-379-5p expression has a protective effect on CIRI rats,and its mechanism of action may be related to the inhibition of GSDMD-mediated pyroptosis.