Application Value Of Skin Biopsy In Diagnosis Of Parkinson's Disease

Background:Parkinson disease(PD)is rank only second to the first common neurodegenerative disease Alzheimer disease,in the elderly,with a high prevalence of 1.7%.With the aging of the population,the burden of disease is getting heavier.Due to the lack of specific peripheral biomarkers,the diagnosis of PD is mainly based on clinical symptoms,and the early misdiagnosis rate is high,which seriously affects the treatment effect of patients.The manifestations of small fiber neuropathy(SFN),such as sensory abnormalities and autonomic nerve symptoms,are the common non-motor symptoms of PD patients,which often occurs years earlier than the motor symptoms.We can early know PD through the recognition of non-motor symptoms.Interepidermal nerve fiber density(IENFD)is the gold standard recommended in European and American guidelines for the diagnosis of SFN.Recent years,researchers found that the IENFD of PD patients was significantly reduced by skin biopsy,and the prevalence of SFN had reached to 56.9%.In the recent 6 years,researchers found that phosphorylated ?-synuclein(p-syn)deposited in skin nerves fibers in PD patients,but in none of healthy controls nor atypical Parkinson's disease.It's highly suggested that p-syn in skin nerve may be a peripheral biomarker for thediagnosis of PD.However,due to different sampling sites,amount of sampling and section thickness in different laboratories,the positive rate of detection varies greatly,which affects the diagnostic efficiency of this method and further affects its clinical application.Our study aimed to explore the status of SFN in population of PD in China and investigete the value of skin biopsy in the diagnosis of SFN.In addition,the detection for p-syn positive rate was optimized by comparing the sample from different site,different location and thickness of slice by skin biopsy.We further research the efficacy of the optimized method in the diagnosis of PD,and the influence of IENFD on the diagnostic efficiency of p-syn in skin.We aim tto improve the early recognition of PD,promote the clinical transformation of skin p-syn as a biomarker for early diagnosis of PD,and improve the accuracy of early diagnosis of PD.Objectives:1.Explore the current status of SFN in Chinese PD population and the value of skin biopsy in the diagnosis of SFN;2.Optimize the method of skin biopsy to detect p-syn by comparing the effect of the location,quantity and section thickness on the positive rate;3.To study the efficacy of skin p-syn in the diagnosis of PD and the effect of IENFD on the diagnostic efficacy of p-syn.Methods:We recruit 82 PD patients and 30 healthy controls from February 26,2019 to January 31,2020,who visited the First Affiliated Hospital of Zhengzhou University.This research was approved by the ethical committee of the First Affiliated Hospital of Zhengzhou University.All participants were informed the subjects of the skin biopsy process,benefits and possible risks,and signed informed consent.A 3mm skin punch biopsies were obtained from distal leg,proximal thigh,the cervical C7 paravertebral area and the forearm.Protein gene production 9.5(PGP9.5)was used for immunohistochemical staining as a neuron biomaker,and the number of nerve fibers acrossing the unit length basement membrane,that is interepidermal nerve fiber density(IENFD),was counted under bright field.Anti-PGP9.5 and phosphorylated?-synuclein(p-syn)antibodies were used to label neuron and phosphorylated?-synuclein,respectively by immunofluorescence staining,and p-syn deposition was observed under a fluorescent microscope.Results were analyzed by SPSS 25.0.Results:1.The distal leg IENFD was significantly decreased in PD compared with the healthy controls(6.32±3.27/mm vs 8.29±2.68/mm,P=0.005),and it was negatively correlated with Hoehn-Yahr stage(t=-2.624,P=0.011).2.55.1%of PD patients has SFN,which had higher scores of SFN-SIQ(7.67±3.28 vs 2.23± 1.41,P<0.001)and COMPASS-31(14.96±11.28 vs 4.44±5.61,P<0.001)than those without SFN.3.The sensitivity,specificity,positive predictive value and negative predictive value of IENFD by skin biopsy in diagnosing SFN in PD were 73.9%,73.3%,86.4%,and55.0%,respectively,and the area under the ROC curve(AUC)was 0.714.(95%CI:0.602-0.826,P=0.001).4.The positive rate of p-syn in the cervical of PD patients(69.4%)was significantly higher than that in the distal leg(41.2%),the proximal thighs(40%),and the forearms(21.4%).5.The positive rate of p-syn in cervical combined with distal leg,proximal thigh,and the forearm was 80.5%,80%,78.6%,respectively.The positive rate of p-syn in forearm and the distal leg and the cervical was 92.9%,but when compared with the single site of the cervical,there was no significant difference.6.The positive rate of p-syn in 2 samples taken from the cervical was 90%,and the positive rate of 2 samples taken from the distal leg was still 40%,with no significant statistical difference.7.The positive rate of p-syn in 50 ?m floating staining was significantly higher than that in 15 ?m patch staining(87.5%vs 56,3%,P=0.049).8.Skin p-syn appeared as punctate or linear in the subepidermal nerve plexus,dermal nerve bundle,sweat glands,blood vessels,arrector pili muscle in PD patients,but not in healthy controls.9.There was no significant difference in IENFD between patients with P-syn and without p-syn in PD(6.44±3.45fibers/mm vs 5.85±2.53fibers/mm,P=0.552),and there was no significant difference in SFN ratio between patients with p-syn deposition and non-deposition group in PD(76.4%vs 71.4,P=0.702).10.The sensitivity,specificity,positive predictive value and negative predictive value of p-syn for diagnosing PD were 79.7%(55/69),100%(30/30),100%(55/55)and 68.2%(30/44),respectively,and the Jordan index was 0.84.The sensitivity,specificity,positive predictive value and negative predictive value of p-syn combined with IENFD for diagnosing PD were 92.8%(64/69,P=0.026),73.3%(22/30,P=0.002),88.9%(64/72,P=0.011),and 81.5%(22/27,P=0.219),respectively.Conclusions:1.The incidence of SFN is high in Chinese patients with PD,and skin biopsy is a reliable method for diagnosing PD with SFN.2.The p-syn positive rate of cervical was highest in single-site sampling in PD.Multi-site sampling or single-site multi-tissue sampling could increase the positive rate of p-syn,but the difference was not significant.The slice thickness significantly affected the positive rate of p-syn.3.Skin p-syn detecting is a reliable method for the diagnosis of PD which could improve the sensitivity of IENFD and can be used as a peripheral biomarker for PD.