The Study Of Brain Dopamine Transporter,The Level Of ProBDNF In Serum And Intestinal Flora’s Changes In Patients With Parkinson’s Disease

As one of the most common neurodegenerative diseases,Parkinson’s disease（PD）is still diagnosed based on the medical history and clinical manifestations.The clinical misdiagnosis rate is very high,especially in the early stage of the disease course.Three follow-up visits are needed to diagnose PD.In order to improve the accuracy of diagnose,many scholars research the related markers of PD.DAT-PET imaging is the most sensitive method to diagnose PD.It is of great clinical value to study the changes of dopamine in patients with PD,judging the condition of patients and guiding the treatment.The cause of PD is unknown at present.The typical pathological change is the mass death of dopaminergic neurons in the substantia nigra.The precursor of brain derived neurotrophic factor（proBDNF）is widely present in the central nervous system.It was proved that proBDNF is the endogenous gene product in human brains with or without PD.proBDNF is involved in the regulation of synaptic plasticity,and proBDNF may have effects on neurons.Previous studies have shown that the incidence of PD patients is related to the decrease of BDNF in serum.But as a precursor of BDNF,the changes of proBDNF in serum of PD patients have not been reported.Intestinal flora is closely related to human health.Increase with age,the function of gastrointestinal organs and HPA axis decrease,which will change the intestinal flora.In turn,intestinal flora can influence the health of the host by regulating immune,inflammatory and neuroendocrine systems.Therefore,intestinal flora may have an relationship with PD.It has been suggested that the imbalance of intestinal flora can promote the pathogenesis of PD.PD patients always have gastrointestinal problems and the intestinal symptoms can not be obviously improved after taking drugs for PD.Moreover,some drugs can trigger gastrointestinal discomfort of PD patients.Thus,studying the intestinal flora of PD patients and healthy people is meaningful,and it can help us understand the pathogenesis factor of PD.Therefore,this study inclues three parts:the changes of dopamine transporter in PD,the changes of BDNF and proBDNF in serum,the changes of intestinal flora in PD patients.It is hoped that the systematic changes of PD patients can be viewed from multiple perspectives,providing help to find the cause,early diagnosis and guiding treatment.Part Ⅰ:The changes of Brain dopamine transporter in patients with Parkinson’s diseaseMaterials and methods:the clinical data of 134 clinically diagnosed PD patients and¹¹C-CFT DAT PET imaging data were analyzed.The 134 PD patients were divided into two groups according to hoehn-yahr（H-Y）classification,namely,early stage,middle-late stage.The differences in DAT uptake indexes of the onset side and the contralateral caudate nucleus,anterior pupule nucleus and posterior pupule nucleus of the two groups were compared respectively.The relationship between uptake index and age,course of disease,H-Y grading,clinical symptom scores（UPDRSⅢ）,the movement symptom scores（NMSS）,daily life events rating scale（ADL）were analysized.Results:compared with the early PD group,the DAT uptake index in all regions was decreased in the middle-late stages PD group,except the posterior pupule nucleus.Compared with the early stages PD group,the DAT uptake index of the opposite side of the onset decreased,and the difference was statistically significant.There was no statistically significant difference between the other two groups.Striate body part of the brain regions DAT consume index and PD patients age,disease course,H-Y grading,UPDRSⅢ.The uptake index in bilateral caudate nucleus has negative correlation with onset age,course of diseases,H-Y grading,UPDRSⅢscores and ADL scores,NMSS scores（P<0.05）.And the relationship between before and after bilateral putamen uptake index intake index and onset age,course of diseases,H-Y grading,UPDRSⅢscores and ADL scores,NMSS scores was significantly negative correlation（P<0.05）.Conclusion:¹¹C-CFT PET imaging can provide medical evidence for different severity of PD and it is helpful to evaluate the severity of PD.Part Ⅱ:The changes of proBDNF in serum of patients with Parkinson’s diseaseMaterials and methods:PD patients were diagnosed in neurology department of Daping hospital,then their blood samples and matched controls were collected,and the levels of proBDNF and BDNF in all participants were measured with elisa kit.Results:compared with healthy control group,BDNF level in serum was decreased,proBDNF and proBDNF/mBDNF was increased in serum of PD patients.There was no significant correlation between the blood BDNF level,proBDNF level and proBDNF/mBDNF in PD patients and the severity of the disease.Conclusion:BDNF and proBDNF in serum of patients with PD are changed compared with the healthy control group,which could be used as a potential serological index for the diagnosis and treatment of PD.Part Ⅲ:The changes of intestinal flora in patients with Parkinson’s diseaseMaterials and methods:1.Fecal specimens were collected from PD patients with the same age and gender of healthy people in daping hospital of the army medical university.Finally,35 patients who did not take anti-parkinson’s drugs,45 patients who had taken anti-parkinson’s drugs and 80healthy people matched were recruited in this study.2.After getting the patient’s informed consent,stool specimens for research object,were immediately stored into the lab in a-80℃refrigerator.3.DNA was extracted from feces,and the v3-v4 region in the 16S rRNA of the extracted DNA was amplified by PCR.The amplified DNA was sequenced,and the sequencing data were analyzed according to the database.4.A total of 160 samples were eventually amplified with a total sequence number of16692904.The 14571892 qualdfied reads for subsequent analysis,according to the RDP database,sequences with similarity greater than 97%were defined as the same OUT,which was used to calculate the abundance of various bacteria at different classification levels.5.Alpha diversity analysis,Beta diversity analysis and LEfSe analysis were performed on intestinal flora between PD group and healthy group,and between the PD group taking drugs and not taking drugs.Results:1.We compared the composition of PD patients group and healthy control group at the level of phylum and genus classification.On the phylum classification level,Firmicutes,Bacteroidetes,Proteobacteria,and Actinobacteria were the main phyla.Compared with the normal control group,firmicutes,actinomycota and Verrucomicrobia were increased in the PD group.There was a decrease in bacteroidetes.At the level of genus classification,compared with the control group,the PD group showed decreased Bacteroides,prevotella,blautia and Fusobacterium.2.Intestinal bacteria analysis of PD patients and healthy control group At the level of phylum classification,we found that there were significant differences between bacteroidetes and actinomycetes.Bacteroidetes significantly decreased in PD group,while Firmicutes significantly increased.At the level of genus,the Parasutterella of PD group decreased,while the Alistipes of PD group increased.3.The analysis of the difference of intestinal flora between the patients who did not take drugs and those who had taken drugs in PD group.Alpha diversity analysis showed no significant difference in diversity and abundance between the two groups.PCoA community composition analysis showed no significant difference between the two groups.Metastats analysis showed that significantly increased at the Actinobacteria compared with the non-drug group at the phylum level,streptococcus and Anaerostipes increased at the genus level,while the bacilli level was lower in the drugs group.Conclusion:the intestinal flora of PD patients changed,and there was statistically difference between the abundance of various bacteria and the healthy control group.There was no significant change in the abundance of intestinal flora between PD patients taking drugs and those not taking drugs,but there was a difference in the flora.