| Objective:Using the method of network pharmacology,this study found the relevant targets of Bupleurum Chuanxiong in the treatment of post-stroke depression（PSD）,and preliminarily predicted the mechanism of this treatment process.The compound model of PSD rats was established,and the improvement effect of Bupleurum Chuanxiong on depression related behavior of PSD rats was observed by behavioral test.Through the detection of the expression level of related inflammatory factors in the brain of PSD rats,the results of early network pharmacological prediction were verified,and the feasibility of this method was discussed.Methods:Use the method of network pharmacology to screen the drug components and drug properties of Bupleurum chuanxiong,that is,target prediction,construct the drug component target network,and use the corresponding database to screen the core targets.The PSD compound model was established by bilateral common carotid artery ligation（2-VO）combined with solitary culture and unpredictable chronic mild stimulation（cums）.The cerebral blood flow was measured by dual channel laser Doppler cerebral blood flow meter on the 7th and 14th days after ligation.Open field experiment,forced swimming experiment and tail hanging experiment were carried out on 28d and 42d after ligation,sucrose preference experiment and Morris water maze experiment were carried out on 42d after stroke,and the depression like symptoms,motor ability and learning and memory ability of rats were detected.The expression levels of TNF,PTGS2,IL6,IL1,ICAM1 and MMP2 in the brain of rats in each group were detected by ELISA and Western blot on the 42nd day after ligation.Results:The prediction results of network pharmacology showed that TNF-α,PTGS2,IL6,IL1β,ICAM1 and MMP2 were the top 20 targets in the protein interaction network.The results of KEGG enrichment analysis show that these targets can play a role in multiple top 10 pathways related to the treatment process at the same time,and are at the core of each pathway.These six targets are taken as the core targets of follow-up research.The results of cerebral blood flow measurement showed that compared with the blank group,the cerebral blood flow of the model group,the positive drug group and the low,medium and high dose groups of Bupleurum Chuanxiong decreased significantly at three time points of 0d,7d and 14d（P<0.01）,and there was no significant difference in the blood pressure measured by tail cannula method（P>0.05）.The behavioral results are as follows:（1）the results of open field experiment show that:compared with the blank group,there are significant differences in the number of standing times,latency,total number of exercise grids,total distance,speed,number of entering the center grid,time and number of entering the center in the model group on 28d and 42d（P<0.05,P<0.01）.Compared with the blank group,the number of standing times,latency and total number of exercise grids in the model group on 28d and 42d,There were significant differences in the total distance,speed,the number of cells entering the center,the time of entering the center and the number of times entering the center（P<0.05,P<0.01）.Compared with the model group,there were significant differences in the number of standing times,latency,total number of cells,total distance and speed of rats in the positive drug group and the Chaihu Chuanxiong high dose group（P<0.05,P<0.01）,and the number of cells entering the center on 42d in the Chaihu Chuanxiong medium dose group,There were significant differences in the time and times of entering the center（P<0.05）.（2）The results of forced swimming experiment showed that compared with the blank group,the immobility times and immobility time of rats in the model group on 28d and 42d were significantly different（P<0.05,P<0.01）.Compared with the model group,the immobility time of rats in the positive drug group,low,medium and high dose groups of Bupleurum Chuanxiong were significantly different on 42d（P<0.05,P<0.01）,The immobility times of rats in the positive drug group and the Chaihu Chuanxiong high dose group were significantly different from those in the model group at 42 days（P<0.05）.（3）The results of tail suspension experiment showed that compared with the blank group,the immobility times and immobility time of model group rats on 28d and 42d were significantly different（P<0.05,P<0.01）.Compared with the model group,the immobility time and times of rats in the positive drug group,bupleurum Chuanxiong low,medium and high dose groups were significantly different at 42 days（P<0.05,P<0.01）.（4）The results of sucrose preference test showed that compared with the blank group,the 42d sugar and water consumption rate of model group rats decreased significantly（P<0.01）,and compared with the model group,the 42d sugar and water consumption rate of positive drug group rats,bupleurum Chuanxiong medium and high dose group rats increased significantly（P<0.05,P<0.01）.（5）The results of water maze showed that there were significant differences in escape latency,movement speed,movement distance and platform crossing times between the model group and the blank group（P<0.05,P<0.01）.Compared with the model group,there were significant differences in escape latency,total distance of exercise and the number of platform crossing in the middle and high dose groups of Bupleurum Chuanxiong（P<0.05,P<0.01）.The detection results of inflammatory factors showed that compared with the blank group,the levels of TNF-α,PTGS2,IL6,IL1β,ICAM1 and MMP2 in the brain of the model group increased significantly at 42 days（P<0.01）.Compared with the model group,the levels of TNF-α,PTGS2,IL6,IL1β,ICAM1 and MMP2 in the brain of the positive drug group decreased significantly（P<0.01）,and the levels of MMP2 and IL6 in the brain of the Chaihu Chuanxiong low-dose group decreased significantly（P<0.05,P<0.01）,The levels of TNF-α,PTGS2,ICAM1,MMP2 and IL1β in brain of rats in Bupleurum Chuanxiong middle dose group（P<0.05,P<0.01）,and the levels of TNF-α,PTGS2,IL6,IL1β,ICAM1 and MMP2 in brain of rats in Bupleurum Chuanxiong high dose group（P<0.05,P<0.01）.Conclusion:Using the method of network pharmacology to predict the inhibition of inflammatory response may be the main mechanism of Bupleurum Chuanxiong intervening in the pathogenesis of PSD.The compound model of PSD rats was established by bilateral common carotid artery ligation combined with solitary rearing and unpredictable chronic mild stimulation.42 days after ligation,PSD rats showed obvious depression like symptoms,which were reflected in the decline of exercise ability in open field experiment and Morris water maze experiment,the prolongation of despair time in forced swimming experiment and tail suspension experiment,and the low sensitivity to reward stimulation in sucrose preference experiment,It shows that this method can replicate the compound model of PSD rats.At the same time,the results of water maze showed that PSD rats had a certain degree of decline in learning and memory ability at 42d,which was consistent with the emergence of depression like behavior,indicating that the lack of learning and memory ability in the pathogenesis of PSD was also one of the causes of depression.Stroke can lead to a significant increase in the levels of TNF-α,PTGS2,IL6,IL1β,ICAM1 and MMP2 in the brain of PSD rats,which is correlated with the results of behavioral tests.PSD rats have increased inflammatory response and brain injury due to cerebral ischemia and long-term external environmental stimulation,resulting in a series of depression like behaviors.The results show that Bupleurum Chuanxiong has a certain intervention effect on the expression of related inflammatory factors in the brain of PSD rats.At 42 days after stroke（28 days stimulated by CUMS）,the behavioral parameters of rats in Bupleurum Chuanxiong group were significantly improved,and the level of related inflammatory factors in the brain also decreased significantly at 42 days,and the expression level of inflammatory factors decreased significantly with the increase of administration concentration.The effect of Bupleurum Chuanxiong high dose group（4.32g/kg/d）was the most significant.This experimental result is consistent with the prediction results of early network pharmacology,which proves that Bupleurum Chuanxiong can achieve the purpose of treating post-stroke depression by inhibiting the inflammatory reaction in the brain. |
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