The Role And Mechanism Of BDNF/XBP1/Kalirin7 Signaling Pathway In The Improvement Of Depression Outcomes After Cerebral Ischemia By Intensive Training

Post-stroke depression(PSD)is one of the most important factors affecting patients’ quality of life.Studies have found that PSD patients lack of interest in the surrounding things,distracted attention,and chornic depression can lead to patients with varying degrees of cognitive and executive dysfunction,interfere with the effect of rehabilitation therapy,and even lead to some patients suicide,a serious threat to the quality of life of stroke survivors,and increase family and social burdens.Abnormal neuronal structure and function are closely linked to the occurrence of PSD.Synapses provide information transfer between neurons.BDNF(brain-derived neurotrophic factor)improves synaptic remodeling,but its regulatory mechanisms are unclear.XBP1(alkaline leucine zipper is an important transcription factor)regulates Kalirin7(Kalirin is a neuro-specific guanine nucleotide transforming factor in the Rho protein family,and Kalirin7 is the most expressed isomer)in neurodegeneration and promotes synaptic remodeling.Previous studies have found that BDNF can regulate XPB1 expression by activating CREB(cyclic adenosine effect element-binding protein).Previous studies have found that chronic random stimulation can cause depression in rats with cerebral infarction,and the expression of BDNF and Kalirin7 in the brain is decreased.Intensive training can improve PSD and increase the expression of BDNF,XBP1 and Kalirin7.Therefore,we hypothesized that reinforcement training might regulate synaptic remodeling through activation of BDNF/XBP1/Kalirin7 signaling pathway,so as to elucidate the effect and mechanism of reinforcement training on PSD.Results from this study will provide an experimental basis for furthering our understanding of the pathogenesis of PSD and for further optimization of its intervention strategies.Part I Neuroprotective effect of intensive training on PSD mice and its mechanism studyPurposeIn this study,different levels of exercise training intensity were set to compare and observe the improvement degree of depression in poststroke depression mice by intensive training,and observe the expression changes of BDNF,XBP1 and Kalirin7 in the hippocampus of poststroke depression mice after intensive training,so as to explore the neuroprotective effect and protective mechanism of intensive exercise training in mice with post-stroke depression.MethodsForty male C57BL/6 mice(7-8 weeks of age)were randomly assigned to the sham surgery group,post-stroke depression(PSD)group,PSD+ regular training(RT)group,and PSD+ intensive training(IT)group,with 10 mice in each group.In the sham surgery group,the general carotid artery,internal carotid artery and external carotid artery were the only arteries to be separated,and no line insertion was done.The rest groups were all treated with the Longa external carotid artery occlusion method in order to establish a model of permanent middle cerebral artery occlusion model(p MCAO).After the p MCAO model was successfully constructed,the single cage feeding plus chronic unpredictable stress stimulation(CUMS)induced PSD model,while the sham surgery group was not given CUMS.After successful modeling,the sham surgery group and the PSD group did not carry out exercise training,while the regular training group and the intensive training group carried out routine and intensive treadmill training respectively.After 14 days,the four groups of mice underwent behavioral experiments,including open field test(OF),elevated plus maze test(EPM)and tail suspension test(TST).The modified neurological severity scores(m NSS)performed at days 1,7 and 14,and after 14 days changes in the expression of BDNF,XBP1 and Kalirin7 were detected by Western blot analysis.ResultsAfter the end of 14 days of training,the neurological function defect was significantly improved after cerebral ischemia injury in both regular and intensive training groups.m NSS method was used to evaluate the motor function,balance and reflexes of the mice after cerebral ischemia.Compared to the sham surgery group,there was a significant increase in the neurological deficit score of mice in the PSD group.Compared to the PSD group,the neurological deficit score after training was significantly decreased,and the m NSS score of intensive training was lower than that of regular training.The open field experimental behavior results showed that after the modeling intervention,residence time in the core area of the PSD group was reduced compared to the sham surgery group,the central shortened,compared with the PSD group,the residence time in the PSD+RT group and the PSD+IT group was increased,and residence times in the PSD+IT group were higher than those in the PSD+RT group,statistically significant differences were observed(P<0.05).The results of the elevated plus maze experiment showed that after modeling intervention,the open arm residence time in the PSD group was shortened compared to the sham surgery group.And compared to the PSD group,the open arm residence time in the PSD+RT group and the PSD+IT group were increased,meanwhile the residence time in the PSD+IT group was better than that in the PSD+RT group,statistically significant differences were observed(P<0.05).The results of the tail suspension test showed that after the modeling intervention,the freezing time was shortened for the first time in the PSD group compared to the sham surgery group,and the total freezing time was increased within 2-6min.Compared to the PSD group,the freezing time was prolonged for the first time in the PSD+RT group and the PSD+IT group.The total freezing time decreased within 2-6min,and the freezing time was longer for the first time in the PSD+IT group,and the total freezing time was less within 2-6min.Western Blot results showed that after the modeling intervention,the protein expressions of BDNF,XBP1 and Kalirin7 in the PSD group were decreased which were compared to the sham surgery group.But compared to the PSD group,BDNF,XBP1 and Kalirin7 protein expressions in hippocampus of PSD+RT group and PSD+IT group were significantly higher than those of PSD+IT group at the same time,and the protein expressions of BDNF,XBP1 and Kalirin7 in hippocampus of PSD+IT group were significantly higher than those of PSD+RT group.Conclusions1.Exercise training significantly improved the neurological deficits of depressed mice after cerebral ischemia injury,and the effect of intensive training was better than that of regular training.2.Through behavioral tests,intensive training effectively improved the degree of depression in post-stroke depressed mice,and the effect of intensive training was better than that of regular training.3.WB results showed that exercise training could increase the expression of BDNF,XBP1 and Kalirin7 in hippocampus and promote the recovery of neural function,and the effect of intensive training was better than that of regular training.Part II Study on the effect and mechanism of BDNF/XBP1/Kalirin7 signaling pathway in improving PSD by intensive trainingPurposeTo investigate whether blocking or upregulating the expression of a single factor in the BDNF / XBP1 / Kalirin7 signaling pathway affects neurological dysfunction in PSD mice,we further investigated whether enhanced training reverses neuroprotective effects in mice through the BDNF / XBP1 / Kalirin7 signaling pathway.MethodsSixty male C57BL/6(7-8 weeks of age)mice were randomly divided into sham surgery group,XBP1 inhibitor negative control group,Trk B receptor agonist negative control group,PSD+IT+XBP1 inhibitor group,PSD+IT+Trk B receptor agonist group by random number method.Each group consisted of 10 animals.In the sham surgery group,the general carotid artery,internal carotid artery and external carotid artery were the only arteries to be separated,and no line insertion was done.The rest groups were all treated with the Longa external carotid artery occlusion method in order to establish a p MCAO model.After the model was successfully constructed,the single cage feeding plus CUMS induced PSD model,while the sham surgery group was not given CUMS.After successful modeling,the sham surgery group、XBP1 inhibitor negative control group and Trk B receptor agonist negative control group did not carry out exercise training,while the intensive training group carried out intensive treadmill training respectively.The XBP1 inhibitor was injected at 0.25mg/kg twice a week for two weeks and the Trk B receptor agonist was injected at 10mg/kg once a day for two weeks.The six groups of mice were subjected to behavioral experiments 14 days later,including open field experiment(OF),elevated cross maze experiment(EPM)and tail suspension experiment(TST).The modified neurologic severity scores were performed on day 1,7 and 14,and the expression changes of BDNF,XBP1 and Kalirin7 were detected by Western blot after 14 days.ResultsAfter 14 days’ training,m NSS scores of mice in each group decreased significantly.The mice in the unmodeled group only injected with XBP1 inhibitor and Trk B receptor agonist had no effect on the scores.Compared to the PSD+IT group,the scores in the PSD+IT+XBP1 inhibitor group were higher after 14 days of exercise,but the PSD+IT+Trk B agonist group scores were decreased,P<0.05.In the open field experiment,we found that compared to the sham surgery group,the non-modelled group only injected XBP1 inhibitor and Trk B receptor agonist had no effect on the behavior of mice.Compared with the PSD+IT+XBP1 inhibitor group,the residence time in the central region was shortened,while the residence time in the PSD+IT+Trk B receptor agonist group was increased,P<0.05.Results from the elevated plus maze experiment showed that the unmodeled group only injected XBP1 inhibitor and Trk B agonist had no effect on the behavior of mice compared to the sham surgery group.In the PSD+IT+XBP1 inhibitor group,the residence time in the open arm decreased while those increased in the PSD+IT+Trk B agonist group both compared to the PSD+IT group,P<0.0001.The results of the tail suspension experiment showed that,the unmodeled group only injected XBP1 inhibitor and Trk B receptor agonist had no effect on the behavior of mice compared to the sham surgery group.According to compared with the PSD+IT group,the first freezing time of the PSD+IT+XBP1 inhibitor group was shortened and the total freezing time increased within 2-6min.The first freezing time of the PSD+IT+Trk B agonist group was increased and the total freezing time decreased within 2-6min,P<0.05.The results of the Western Blot analysis showed that compared to the sham operation group,the expression of XBP1 decreased in the unmodeled group with XBP1 inhibitor,increased in the Trk B agonist group,and decreased in the PSD+IT+XBP1 inhibitor group compared with the PSD+IT group.The expression of XBP1 was increased in PSD+IT+Trk B agonist group.The expression of Kalirin7 decreased in the unmodeled group with XBP1 inhibitor,and increased in the Trk B agonist group compared to the sham surgery group.As for comparing to the PSD+IT+XBP1 inhibitor group,the expression of Kalirin7 decreased in the PSD+IT+XBP1 inhibitor group but it was increased in PSD+IT+Trk B agonist group.Conclusions1.XBP1 inhibitor reversed the neural recovery effect of intensive training in mice;2.Trk B receptor agonists promoted the recovery of neural function in intensive training mice;3.Intensive training improves PSD through BDNF/XBP1/Kalirin7 signaling pathway.Significance of this studyThis study elucidated the relationship between stroke depression and synaptic remodeling mediated by BDNF/XBP1/Kalirin7 signaling pathway,providing evidence for the pathogenesis of poststroke depression.At the same time,it reveals the relationship between the activity state of brain functional areas and the changes of post-stroke depression behavior,which provides supporting evidence for the rehabilitation theory of regulating the overall functional state of the cortical and subcortical structures after stroke.Intensive training can better improve post-stroke depression,and then better improve post-stroke dysfunction.Molecular mechanisms provide the basis for translational medicine and important theoretical support for stroke rehabilitation.