| Objective:Parkinson's disease(PD)is the most common neurodegenerative disease,which is mainly manifested as progressive dyskinesia.The diagnosis and treatment are usually based on changes in motor signs such as tremor,motor retardation,muscle stiffness and postural instability.However,in some PD patients,non-motor symptoms such as anxiety,fatigue,depression,sleep disorder,gastrointestinal and sexual dysfunction have also been observed,especially intestinal dysfunction,which has been gradually concerned by scientists in recent years.Acupuncture treatment has been shown to improve the autoimmune immunity and disease resistance in many studies,especially for non-infectious chronic diseases.Some studies have shown that acupuncture can have a certain effect on neurodegenerative diseases.To study whether electroacupuncture stimulation can relieve the motor symptoms of mice with Parkinson 's disease,we observed the effects of electroacupuncture on neurobehavioral,neuroinflammation,intestinal microbes and Parkinson's disease related markers by stimulating acupoints under the guidance of traditional Chinese medicine theory,using MPTP/MPP+-induced cells and mouse model.From the perspective of acupuncture regulation of biological immunity and microorganism,this paper explored the treatment and biological mechanism of Parkinson's disease under the guidance of traditional Chinese medicine theory.Methods:This paper expounded the mechanism of acupuncture intervention for Parkinson's disease from two aspects:theory and experiment.In the theoretical research,through a lot of summaries of traditional Chinese medicine acupuncture treatment and Western medicine research of Parkinson's disease,we found that acupuncture is one of the effective treatment methods in Chinese medicine that should be promoted in the clinical practice.In the basic experimental,we applied MPTP subcutaneous sustained release to simulate the pathogenesis of human Parkinson's disease.Through continuous observation of symptoms and signs in Parkinson's disease mouse model and neurobehavioral tests at different times,it was found that MPTP subcutaneous sustained release can successfully replicate the Parkinson's disease model.And the expression of inflammatory factors in the intestinal tissue and the substantia nigra of Parkinson's disease mouse were detected at different time points to explore whether the onset of Parkinson's^disease started in the intestine.Secondly,we used electroacupuncture on the MPTP-induced Parkinson's mouse model,and observed the symptoms,neurobehavioral,intestinal microbes,proinflammatory cytokine expression in various tissues and Parkinson's disease-related proteins.From behavioral to intestinal microbes and molecular level,the mechanism of electroacupuncture intervention in Parkinson's disease mouse model was explored,and the biological mechanism of the electroacupuncture intervention for central nervous degeneration was explored.Finally,we used MPP+/MPTP to induce PC 12 cells(pheochromocytoma cells,an in vitro experiment for neurological disease research)to simulate the pathogenesis of Parkinson's^disease in vitro.In this part of the experiment,we confirmed that MPTP can cause neuronal apoptosis and oxidative stress by concentration-dependent experiments.It was also found that a certain concentration of glutamine can inhibit the apoptosis and the occurrence of oxidative stress of PC 12 cells induced by MPP+.Results:(1)The PD mouse model was successfully established by MPTP subcutaneous sustained-release injection.From the first week to the eighth week after the model was started,the symptoms and signs of PD mouse were gradually abnormal,and the non-motor symptoms of PD appeared.By the eighth week,neurobehavioral tests showed that Parkinson's^motor symptoms occurred.The results of this experiment can not only prove that MPTP can lead to the progressive exacerbation of motor symptoms in PD mouse model,but also that the neurotoxin of MPTP can cause PD mouse with intestinal dysfunction.(2)Applied molecular biology experiments show that MPTP-induced PD mouse model can not only increase the number of inflammatory factors in the intestinal system,but also many inflammatory factors in the cerebral substantia nigra with the advancement of modeling time(eighth week),indicating that neuroinflammation plays an important role in the MPTP-induced PD mouse model and is an important cause of PD development.(3)Using the acupuncture early intervention of MPTP-induced PD mouse model,compared with the blank control group by neurosurgery test,it was found that the PD model group had different degrees of dyskinesia at the 8th Week,accompanied by constipation symptoms.Compared with the PD model group,the acupuncture intervention group had reduced exercise or cognitive impairment.(4)The results of sequencing experiments showed that compared with the blank control group,the MPTP-induced PD mouse model was dysregulated,while the acupuncture intervention group had a regulatory effect on some dysbacteriosis.It also has a good inhibitory effect on the release of pro-inflammatory factors in the intestinal tract and the substantia nigra.Microbiological examination showed that acupuncture intervention has a good two-way regulation on intestinal microflora in PD mouse model.(5)Detection of GFAP,TH and oxidative stress related indicators in PD showed that MPTP-induced PD mouse model can lead to abnormal expression of GFAP,TH and oxidative stress.Early acupuncture intervention can slow down the occurrence and development of molecular biological indicators related to PD,which may also have a certain correlation with intestinal flora imbalance.(6)Apoptosis of PC 12 Cells Induced by MPP+.After 48 hours of MPP+intervention,nuclear morphology changes were analyzed by AO/EB staining.MPP+can increase apoptosis in PC 12 cells in a dose-dependent manner.The oxidative stress response induced by MPP+in PC 12 cells was determined by ELISA.The contents of SOD and GSH-Px were significantly lower than those in the blank control group.(7)The cells were pretreated with glutamine for 1 h,then the cells were incubated with MPP + for 48 h.The results showed that the viability of PC 12 cells was significantly increased after glutamine pretreatment,and glutamine 64pm had the strongest protective effect on PC 12 cells.In addition,AO/EB staining showed that glutamine intervention of MPP+induced PC 12 cells can significantly reduce the level of apoptosis,indicating that glutamine has protective effect on MPP+-induced PC 12 cell apoptosis.Conclusion:(1)The MPTP subcutaneous sustained release method successfully replicated the PD mouse model.(2)The PD mouse model induced by subcutaneous sustained release of MPTP showed a gradual trend,which could eventually lead to behavioral changes from non-motor symptoms to motor symptoms.MPTP-induced PD mouse model increased inflammatory factors first in the gastrointestinal system,followed by metastasis to the midbrain.MPTP-induced PD mouse models can cause gastrointestinal dysfunction and can lead to changes in the gastrointestinal microbial environment leading to massive release of inflammatory factors.(3)Early acupuncture intervention MPTP subcutaneous sustained-release injection induced PD mouse model can effectively change the symptoms and signs,neurobehavioral dysfunction,non-motor symptoms and motor symptoms of rats.The reasons for changing the signs and behavioral disorders of PD mouse model may be achieved by changing the gut microbiota,reducing the expression of gastrointestinal inflammatory factors and the expression of GFAP and TH.(4)MPP+can cause apoptosis in PC12 cells,and the cause of apoptosis may be oxidative stress response of cells;different concentrations of glutamine can inhibit MPP+-induced apoptosis of PC 12 cells,and may be achieved by inhibiting the oxidative stress response of cells. |
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