| Objective Stroke is one of the most prevalent diseases with high incidence,mortality and disability in the world.In the previous study,there was evidence that IGRS could inhibit the protein expression of endoplasmic reticulum stress with neuroprotective effect.In summary,it can be inferred that IGRS could exert protective effects on CIRI and attenuate CIRI by inhibiting endoplasmic reticulum stress-mediated neuronal apoptosis.（1）To study the effects of IGRS on cerebral ischemia reperfusion injury in rats.（2）To study the effects of IGRS on neuronal apoptosis induced by cerebral ischemia reperfusion in rats.（3）To study the effects of IGRS on endoplasmic reticulum stress induced by cerebral ischemia reperfusion in rats.（4）To provide the experimental and theoretical basis for the development of IGRS becoming into the clinical applied new drugs.Methods（1）The sprague-dawley rats were randomly divided into sham-operation（Sham）,cerebral ischemia reperfusion group with saline treatment（Vehicle）,vehicle with IGRS-treated group at dose of 30 mg·kg-1,vehicle with IGRS-treated group at dose of 60 mg·kg-1,vehicle with IGRS-treated group at dose of 120 mg·kg-1,vehicle with Edaravone-treated group at dose of 3 mg·kg-1,the corresponding drugs were given for each group once a day for7 days.And all the rats were administrated before 1 h of operation.The cerebral ischemia reperfusion model in rats was established by blocking the right middle cerebral artery for 1.5h,then reperfusing for 24 h.And all the rats were administrated after 6 h of operation.After24 h of reperfusion,the neurological deficits,brain water content,brain index were observed,the cerebral infarct volume was measured by 2,3,5-triphenyltetrazolium chloride（TTC）staining,pathological changes in the hippocampus and cortex were observed by hematoxylin and eosin（HE）,and the normal nerve cells were counted.These experiments were employed to evaluate the protective effects of IGRS on cerebral ischemia reperfusion in rats.（2）The apoptotic cells were counted by TUNEL assay in brain cortex.The Caspase-3expression in brain cortex induced by cerebral ischemia reperfusion in rats was evaluated by immunohistochemistry.And the transmission electron microscope（TEM）was employed to observe the ultrastructure change of the endoplasmic reticulum induced by cerebral ischemia reperfusion injury.（3）The methods of q RT-PCR and Western-blotting were employed to determine the expression of GRP78,Caspase-12,and CHOP in the hippocampus induced by cerebral ischemia reperfusion injury.Results（1）After 24 h of reperfusion,compared with the sham group,the neurological deficits,brain water content,brain index and the cerebral infarct volume of the vehicle group were significantly increased（P<0.01）,and the brain tissue in rats of the vehicle group appeared pathological damage,nucleus pycnosis,dissolved in neurons,hippocampal structure was loose,and the number of the normal nerve cells significantly decrease（P<0.01）.Compared with the vehicle group,IGRS(120 mg·kg-1)could significantly decrease the increased neurological deficits,brain water content,brain index and the cerebral infarct volume,and it could significantly increase the normal nerve cells（P<0.05,P<0.01）.IGRS(60 mg·kg-1)could significantly decrease the increased neurological deficits,brain water content and the cerebral infarct volume,and it could significantly increase the normal nerve cells as well（P<0.05,P<0.01）.（2）After 24 h of reperfusion,compared with the sham group,the apoptosis rate and the Caspase-3 expression in brain cortex of the vehicle group was significantly increased（P<0.01）.The endoplasmic reticulum ultrastructure of hippocampal neuron was severely damaged,loosely arranged,obviously swollen,and the ribosome fell off with polyribosome depolymerization.Compared with the vehicle group,IGRS(60,120 mg·kg-1)could significantly decrease the increased apoptosis rate and the Caspase-3 expression in brain cortex（P<0.05,P<0.01）.The swelling degree and the loss of ribosomes in hippocampal nerve cells were significantly alleviated,and the number of ribosomes in the cytoplasmic matrix was significantly reduced.（3）After 24 h of reperfusion,compared with the sham group,the m RNA expressions of GRP78,CHOP and Caspase-12 in the vehicle group were significantly up-regulated（P<0.01）.And the protein expressions of GRP78,Caspase-12,and CHOP were significantly increased（P<0.01）.Compared with the vehicle group,IGRS(60,120 mg·kg-1)could significantly decrease the m RNA expressions of GRP78,Caspase-12,and CHOP（P<0.05,P<0.01）.IGRS(30 mg·kg-1)could significantly decrease the m RNA expressions of CHOP（P<0.05）.And the m RNA expression of GRP78 and Caspase-12 showed a downtrend.IGRS(120 mg·kg-1)could significantly decrease the protein expressions of GRP78,Caspase-12,and CHOP（P<0.05,P<0.01）.IGRS(60 mg·kg-1)could significantly decrease the protein expressions of GRP78 and CHOP（P<0.05）,and the protein expressions of Caspase-12 showed a downtrend.IGRS(30 mg·kg-1)could significantly decrease the protein expressions of CHOP（P<0.05）,and the protein expressions of GRP78 and Caspase-12 showed a downtrend.Conclusion（1）IGRS could significantly decrease the increased neurological deficits score,brain water content,brain index and the cerebral infarct volume,and also could obviously improve the histopathological damage of ischemic cerebral tissue,which indicated the protective effect of IGRS on cerebral ischemia reperfusion injury.（2）IGRS could obviously decrease the increased apoptosis rate and the Caspase-3expression of cortical neuron and improved the endoplasmic reticulum ultrastructure of hippocampal neuron,which indicate that the protective effect of IGRS on cerebral ischemia reperfusion injury maybe related to the inhibition of neuronal apoptosis and the protection of the endoplasmic reticulum ultrastructure.（3）IGRS could obviously down-regulated the m RNA expressions of GRP78,Caspase-12,and CHOP,and obviously decrease the protein expressions of GRP78,Caspase-12,and CHOP.It indicated that the protective effect of IGRS on cerebral ischemia reperfusion injury maybe related to the inhibition of neuronal apoptosis induced by endoplasmic reticulum stress. |
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