| [Background]Parkinson’s disease(PD)is a complicated disease,commonly diagnosed among the elderly,which leads to degeneration of the central nerve system.PD is characterized by trembling of hands,arms,legs,jaw and face at rest,stiffness of the arms,legs and trunk,slowness of movement,poor balance and coordination and the vegetative nerve function disorder.Together with dementia and stroke,PD is known as the major diseases of disability that affect the quality of life of patients seriously in the elderly.Its prevalence,morbidity,and mortality,as well as associated economic burdens,have been steadily increasing.There were more than 2 million PD patients over 55 years of age in China.As the elderly population in China increases,the number of patients with PD is expected to double by 2030.With high disability rate,low cure rate and a cost of more than ten thousand yuan per year.Thus long-term care of PD patients brings a great economic burden to families and to society.PD is a complicated disease and the pathogenic mechanisms are still unclear.It presently lacks an effective therapy for its complex pathogenesis.From the research on PD of internal and external,the Dopamine replacement therapy of Western medicine is regarded as a dominant treatment and mainly focused on the symptom.However,there are certain restrictions for this treatment.Madopar is regarded as the most effective therapy for this disease,is known as a "golden drug" for its ability to restore dopamine levels in the brain.However,with chronic administration of Madopar,PD patients experience drug resistance,and an increase in side affects and adverse reactions.Thus managing both PD symptoms with medication,and adverse drug reactions is difficult.The operation treatment can’t popularize in clinical due to the long-term effect is poor.In recent years,the stem cell transplantation and gene therapy has shown a good prospect from the deep reaserch.However,immature and expensive technology and user resistance limits its popularization.Traditional Chinese medicine(TCM)has been used to treat PD for many years and can be traced back to the period of The Yelllow Emperor’s C.anon of Internal Medicine,is an an irreplaceable treatment for this disease.Based on the TCM theory,a large number of researches on the efficacy and mechanism of Chinese herb monomer and compound had been carried on and the results are quite satisfactory.The current popularity of traditional Chinese medicine in the treatment of PD implies its potential advantage in improving symptoms,reducing the drug-induced side effects and improving patients’ quality of life.TCM has potential advantages for improving clinical symptoms of PD-especially for the non-motor symptoms.Thus,appropriate use of TCM can improve the quality of life for patients and shows a significant advantage.Although TCM has been shown to be effective in PD patients,quantitative clinical evidence for TCM treatment is still limited.Therefore,as the PD caseload grows the clinical study of PD treatment and prevention is becoming an important subject for contemporary integrative medicine.Further explore about the mechanism of PD to improve the clinical efficacy of TCM is an important clinal project.[Objective]To observe the the behavior of rats before and after modeled and expression of apoptosis related proteins and ubiquitin proteasome pathway related proteins in brain tissue of rats by establish the rat model of PD.To explore the effect of Xifeng Dingchan Pill(XFDC)on protein degradation pathway and the protective effect on dopaminergic neurons of PD model rats.The results of this study will provide evidence for TCM treat PD.[Methods]In this study,the SPF male SD rats were used to establish PD model by 6-hydroxydopamine(6-OHDA)injected oneside of rat brain from two points.All the successful modeled rats were divided into model group,Madopar group and XFDC high,middle and low dosage group randomly.Meanwhile,the sham operation control group was designed.Corresponding drugs were intragastric administered to rats once a day for 8 weeks.The experiment is divided into three parts:Experiment 1:To observe the effect of XFDC on the change of behavior after induced by apomorphine(Apo)by computer the total number of rotating rings of rats in 30 minutes.To observe the morphological changes of brain tissue and the expression of substantia nigra Tyrosine hydroxylase(TH)immunoreactive cell with immunohistochemistry method.Experiment 2:The expression of Bcl-2,Bax and Caspase-3 immunoreactive cells in brain tissue of rats were detected by immunohistochemical method and the changes of apoptosis were detected by TUNEL method to evaluate the effect of XFDC on the apoptosis in each group.Experiment 3:The expression of ubiquitin(UB),ubiquitin activating enzyme E1(UBE1),Parkin,ubiquitin C-terminal hydrolase-L1(UCH-L1)and alpha-synuclein(a-Syn)in the brain tissue of rats were detected by Western Blot method to evaluate the effect of XFDCP on the ubiquitin proteasome pathway.[Results]1 Behavioral testingThe results showed:The sham operation control group didn’t have the rotational behavior to the uninjured side after induced by Apo before and after treatment.Compared with the sham operation control group,the speed of rotational behavior to the uninjured side of rats in model control group was faster and more than 210r/30min.Compared with the model control group,the speed of rotational behavior to the uninjured side of rats in Madopa group and XFDC high,middle and low dose group were insignificant difference(P>0.05)before treatment.Compared with the model control group,the speed of rotational behavior to the uninjured side of rats in Madopa group and XFDC high,middle and low dose group were slower after intragastric administration,and the results were highly significant difference(P<0.05).Along with the time extension the rotational speed is more and more slow.2 Morphological observation of brain tissueThe TH immunohistochemistry results showed:There were many TH immunoreactive cells in the substantia nigra in the sham operation control group.TH immunoreactive cells were plumpn with round or oval sharp and the cytoplasm was brown.Compared with the sham operation control group,the number of TH immunoreactive cell in the model group was reduced significantly,and the results were highly significant difference(P<0.05).Most of the cells were not completely with uneven staining.There were mang proliferous glial cells.Compared with the model group,the number of TH immunoreactive cells was increased significantly in the Madopa group and XFDC high,middle and low dose group,and the results were highly significant difference(P<0.05).3 Neurons apoptosis testingThe immunohistochemistry results showed:There were a large number of Bcl-2 immunoreactive cells and a few numbers of Bax immunoreactive cells and Caspase-3 immunoreactive cells in the sham operation control group.Compared with the sham operation control group,the number of Bcl-2 immunoreactive cell was reduced significantly while the number of Bax and Caspase-3 immunoreactive cells was increased significantly in the model group,and the results were highly significant difference(P<0.05).Compared with the model group,the number of Bcl-2 immunoreactive cells were increased while the number of Bax and Caspase-3 immunoreactive cells was reduceed significantly in the Madopa group and XFDC high,middle and low dose group,and the results were highly significant difference(P<0.05).The TUNEL results showed:There were a few numbers of apoptotic cells in substantia nigra of rats in the sham operation control group.Compared with the sham operation control group,the apoptotic cells were increased significantly in the model group,and the results were highly significant difference(P<0.05).Compared with the model group,the apoptotic cells were reduced significantly in Madopa group and XFDC high,middle and low dose group,and the results were highly significant difference(P<0.05).4 The expression of UPP related protein testingCompared with the sham operation control group,in the model group the expression of α-Syn was increased while the the expression of UB,UBE1,Parkin and UCH-L1 were reduced,and the results were highly significant difference(P<0.05).Compared with the model group,in Madopa group and XFDC high,middle and low dose group the expression of α-Syn was reduced while the the expression of UB,UBE1,Parkin and UCH-L1 were increased,and the results were highly significant difference(P<0.05).[Conclusion]XFDC has functions of tonifying the liver and kidney as well as extinguishing wind.It can protect the neuronal cell of rats from the damage caused by 6-OHDA.It is effective in improve the behavior of rats of PD model.The use of XFDC can help to reduce the number of apoptotic cells and regulate the function of UPP to promote the degradation of abnormal proteins.It plays an important role in protecting the dopaminergic neuronal cells. |
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