| ObjectiveIn vivo,clear Huangputongqiao Capsule has therapeutic effect on AD,has neuroprotective effect on AD animal model of Huangputongqiao Capsule on AD neuroprotective effect and its mechanism of action,and provide the basis for clinical application of modern experiment,also for the treatment of AD to provide new ideas and methods.Methods150 SD rats were fed for 7 days,from healthy female and male rats from the 96rats were randomly divided into 8 groups:control group,sham operation group,model group,Huangputongqiao Capsule high dose group,middle dose group,low dose group,donepezil group and nimodipine group.Male and female separately,and labeled with picric acid.In vivo,25-35 bilateral hippocampal injection combined with model to establish AD animal model by intraperitoneal injection of D-galactose and A beta,learn from the perspective of behavior through the Morris water maze experiment,HE staining was used to observe the morphology of hippocampal tissue of rats changed to validate the model successfully established and Huangputongqiao Capsule on the treatment effect from AD rats the perspective of Pathology,confirmed Huangputongqiao Capsule has therapeutic effect on AD.The AD animal model by using transmission electron microscope to detect Huangputongqiao Capsule on rat hippocampus Microstructure Effects and neuronal damage,confirmed Huangputongqiao Capsule has a neuroprotective effect on AD animal model.The establishment of AD animal model,using blot method to detect Western expression in hippocampus of rats Ca M,Ca MKK,Ca MKⅣ,CREB,tau protein,tau protein phosphorylation,confirmed Huangputongqiao Capsule on hippocampal Ca2+-Ca MKⅣ signaling pathway and tau protein hyperphosphorylation of AD rats can inhibit the possible mechanism of action.Results:1)Morris water maze test showed that:Compared with the blank group,sham operation group from first to fifth days to escape latency had no significant difference（P>0.05）;compared with sham operation group,model group from first days to fifth days,the escape latency of significant difference was statistically significant（P<0.01）;compared with the model group,Huang Pu Tong the capsule high low dose group,donepezil group and nimodipine group for first days without the escape latency difference was not statistically significant（P>0.05）,with the increase of the number of training days,rats in each treatment group showed different degrees of escape latency shortened trend.Compared with the model group,donepezil,Huang Puzhong dose group from the beginning of the second day incubation period,began to appear the difference（P<0.05）,third～5 days decreased significantly（P<0.01）;compared with the model group,nimodipine,Huang Pu high dose group rats at third days of escape latency there is statistical significance in the difference（P<0.05）,from fourth to 5,there was significantly difference between them（P<0.01）;compared with the model group,low dose group of Huang Pu in fourth,the escape latency began to appear statistical difference（P<0.05）.2)HE staining showed:In the sham operation group,pyramidal cells in the hippocampal CA3 area of the sham operated group were multilayered,orderly and closely arranged,and the cells were evenly distributed,regular in shape,clear in structure,homogeneous in cytoplasm,and red in color.The nuclei were round and large,and the positions were in the center.The nucleolus was clear and clear.Compared with the sham operation group,the number of pyramidal cells in the hippocampus CA3 area of model group was sparse and arranged in disorder.The pyramidal cell body contracted and deformed,with irregular shape,cytoplasm concentration,nucleus condensation,deep staining and marginalization,and nucleolus blurred.Treatment of pathological morphology in hippocampal CA3 region of rats donepezil and nimodipine group and Huangputongqiao Capsule has significantly improved,the number of pyramidal cells slightly reduced,arranged regularly,only individual cell nuclei pyknosis and deep staining,the morphology of the cells were close to normal.3)electron microscope ultrastructure results showed:The number of sham neurons was significantly higher than that in the model group.There were abundant organelles in the cytoplasm,many mitochondria,clear ridges,complete nuclear membrane,double layer distribution,visible nuclear pores and uniform chromatin distribution.In the model group,the rough endoplasmic reticulum expanded obviously,the mitochondrial degeneration,the crista partially broken,the nuclear chromatin distribution was still uniform,and the nuclear membrane was complete.A large number of microtubules were found in the medium dose treatment group.The density of the ribosome was uniform,the mitochondria were rich,and the nuclear chromatin was evenly distributed.There were two lysosomes in the donepezil group,and there were more ribosomes and few mitochondrial crista were broken.The number of organelles in Nimodipine group was significantly higher than that in the model group,and the nuclear membrane was complete,the chromatin of the nucleus was evenly distributed,and a few mitochondria were vacuolated.4)WB test results showed that:Compared with sham operation group,model group,Ca M protein expression was not obviously increased（P>0.05）;compared with the model group,Huangputongqiao Capsule treatment group,donepezil group and nimodipine group the expression of Ca M protein decreased significantly（P<0.05）.Compared with sham operation group,model group,Ca MKK protein expression increased significantly（P<0.05）;compared with the model group,Huangputongqiao Capsule treatment group,donepezil group and nimodipine group the expression of Ca MK protein decreased significantly（P<0.05）.Compared with sham operation group,model group,Ca MKⅣ protein expression increased significantly（P<0.05）;compared with the model group,Huangputongqiao Capsule treatment group,donepezil group and nimodipine group Ca MKⅣ protein expression decreased significantly（P<0.05）.Compared with sham operation group,model group,CREB protein expression increased significantly（P<0.05）;compared with the model group,Huangputongqiao Capsule treatment group CREB protein expression was not decreased obviously（P>0.05）,donepezil group and nimodipine group the expression of CREB protein was significantly reduced（P<0.05）.Compared with sham operation group,model group,tau protein expression increased significantly（P<0.05）;compared with the model group,Huangputongqiao Capsule treatment group,donepezil group and nimodipine group the expression of tau protein decreased significantly（P<0.05）.Compared with sham operation group,model group of phosphate tau protein expression increased significantly（P<0.05）;compared with the model group,Huangputongqiao Capsule treatment group,donepezil group of phosphorylated tau protein expression decreased obviously（P<0.05）,nimodipine group was not decreased obviously（P>0.05）.Conclusion:1)Huangputongqiao Capsule can obviously improve the learning and memory ability of AD rats;2)Huangputongqiao Capsule can relieve the neurons of rat hippocampus AD damage;3)Huangputongqiao Capsule could regulate Ca2+-Ca MKⅣ signaling pathway in the hippocampus plays a role in the treatment of AD. |
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