| Object: Build middle cerebral artery occlusion model to get cerebral ischemia-reperfusion(CIR)injured rats,and when reperfusion occurs,rats inhale isoflurane(ISO)at once last for 1 hour to study the role of ISO postconditioning to neuronal pyroptosis in CIR injured rats.The use of MEF2 D shRNA is for specifically knocking down expression level of MEF2 D in rats’ bilateral hippocampus,and aimed to confirm that the myocyte enhancer factor 2D(MEF2D)participates in the process of brain protection during ISO postconditioning against CIR injury to predict selection of narcotics.Methods: Adult male Sprague-Dawley rats were subjected to middle cerebral artery occlusion(MCAO)for90 minutes and reperfusion for 24 h.Inhalation of ISO lasted for 1 h after reperfusion.Modified Neurological Severity Scores,2,3,5-triphenyl tetrazolium chloride staining,the step-through passive test,and the scanning electron microscope observation were used to evaluate the impairments in neurological function.Expression levels of caspase-1,GSDMD,IL-18 and MEF2 D protein were determined by immunofluorescence(IF)staining and western blotting and relative m RNA were determined by quantitative real-time polymerase chain reaction(q RT-PCR).To specifically knock down MEF2 D expression,MEF2 D short hairpin RNA(shRNA)was injected into the bilateral hippocampus 14 days before building MCAO model.Results:(1)In MI group,neurobehavioral deficit scores and the infarct volumes were lower compared with the MCAO group(P < 0.05),meanwhile,the expression levels of GSDMD and IL-18 were less too(P <0.05),but caspase-1 protein and m RNA expression levels had no significant differences(P > 0.05).The formation of cystic vesicles in the neuronal membrane was less,and the cytoplasm was clearer in MI group compared to M group(P < 0.05).(2)After MEF2 D was specifically knocked down in the hippocampus of rats,there were no significant differences in neurobehavioral scores and infarct volumes of rats between RMI and RM group(P > 0.05),and the protein and m RNA expressions of IL-18 were not significantly different either(P > 0.05),while protein and m RNA expressions of caspase-1 and GSDMD were lower(P < 0.05).The morphology of neurons did not change significantly.(3)After the injection of MEF2 D shRNA before ISO postcondition(RMI group)resulted in larger infarct volumes and higher neurologic deficit scores,higher numbers of vesicle of pyroptosis neuron,and higher levels of IL-18 protein and m RNA against ISO postconditioning CIR injury in rats(P < 0.05),while caspase-1 and GSDMD protein and m RNA expression levels were same as MI group(P < 0.05).The cytoplasm of neurons was concentrated and the continuity of cell membrane was interrupted.Conclusions: ISO postconditioning plays a key role in anti-pyroptosis in model of CIR injury and the MEF2 D participates in this protective process through regulation of downstream regulators IL-18. |
