Investigation Of The Relationship Between Cathepsin B-Mediated Apoptosis And ERK1/2Signaling Pathway After Cerebral Ischemia-reperfusion In Rat

Background and ObjectiveNerve cell will be apoptosis in ischemic penumbra after cerebral ischemia andreperfusion,and it cause nerve apoptosis complex,involving many organelles andprotein signal pathway.According to the study at present,the lysosome organizationprotease B (Cathepsin B), ERK1/2signaling pathways involved nerve cell apoptosisprocess after the cerebral ischemia-reperfusion.But the role of different respectively,Cathepsin B play a role with cell apoptosis,and ERK1/2signal pathway is associatedwith promoting the cells survival.We deal with the rat model with the Cathepsin Bspecificity inhibitors CA-074,investigating the change of the protein expression ofCathepsin B and p-ERK1/2.This paper discusses the mutual relationship betweenCathepsin B and ERK1/2signaling pathways in cell apoptosis,and the cellantiapoptotic role of CA-074,riching cerebral ischemia and reperfusion injury neuralcell apoptosis mechanism,to ischemic stroke the prevention and cure of certainsignificance.Methods56male Sprague-Dawley rats (280±20g,10-12weeks old,CLA)were randomlydivided into nine groups: sham-operated control group (Sham,n=6),model group2h(M-2h,n=5), model group6h(M-6h,n=5), model group12h(M-12h,n=5) modelgroup24h (M-24h,n=10),intervention group2h(I-2h,n=5), intervention group6h(I-6h,n=5), intervention group12h(I-12h,n=5),intervention group24h (I-24h,n=10).And transient middle cerebral artery occlusion was performed as Longa described.The Intervention group was injected with CA-074through the lateral ventriclepuncture.The rats of other groups were injected with the same volume of DMSO at the same time points.We use Longa ’s of5levels grading evaluation neural functiondefect.TTC Staining,HE dyeing,immunohistochemistry and TUNEL were performedfor assessment of infarct volumes,Pathological changes,expressions of Cathepsin Band p-ERK1/2in the parietal lobe cortex ischemiaandapoptosis.Results1.Model for success rate:72.73%,The relative infarct volume of model group24his26.63±3.23%.2.Infarction was not observed in sham-operated group,whereas Corticalinfarctions(white color) were detected in model groups and intervention groups.Therelative infarct volume (P<0.05) and neurologic deficit score (P<0.05) of theintervention group were significantly decreased compared with the model groups.3.In rat ischemia side cortex,no Cathepsin B positive expression was detected insham-operated group.Cathepsin B positive expression was detected in the model ratsat various time points.A significant increase during the2-hour reperfusion,and a peakat6-hour reperfusion,and an graduallys decrease at12,24-hour,(P<0.001).Theintervention groups by CA-074were significantly decreased in the expression ofCathepsin B compared with the model groups at the various time points(P<0.05).4.In rat ischemia side cortex,no p-ERK1/2positive expression was detected insham-operated group.p-ERK1/2positive expression was detected in the model rats atvarious time points.A increase during the2-hour reperfusion,and were graduallyincreased at6-hour reperfusion,and a peak at12-hour reperfusion,and an decrease at24-hour,(P<0.001).The intervention groups by CA-074were significantly increased inthe expression of p-ERK1/2compared with the model groups at the various timepoints(P<0.05).5.In rat ischemia side cortex,sham-operated group can be observed in scatteredover the apoptotic cells.TUNEL-positive cells can be observed in the model rats,andwere gradually increased at6,12,24hours of reperfusion （P<0.001）.CA-074significantly reduced cell apoptosis induced ischemia-reperfusion at6,12,24hours(P<0.05),No change at2hours (P>0.05). ConclusionsLysosome Cathepsin B may mediate apoptosis through inhibiting ERK1/2signaling pathway after cerebral ischemia-reperfusion in rat.