A Preliminary Study On The Pathogenesis Of Neurosyphilis Based On Proteomics And Metabonomics

[Background]Neurosyphilis(NS)is a chronic infection of the central nervous system(CNS)caused by treponema pallidum(TP),which can occur in any stage of syphilis.The incidence of NS is variable in different stages of syphilis,with an unclear mechanism.A higher NS incidence occurs in the latent stage due to non-treatment or irregular treatment.10-25% of patients with irregularly treatment may develop into NS,with a variety of clinical manifestations which related to the affected lesions.However,asymptomatic neurosyphilis(ANS)was most common.Approximately35% patients with ANS may progress to symptomatic neurosyphilis(SNS)spontaneously.Persistent infection or without treatment are also the risks of developing to SNS.Most individuals can spontaneously suppress the inflammatory response associated with the early invasion of Tp into the central nervous system,even without treatment for cerebrospinal fluid(CSF)Tp infection.However,it is unclear why some patients fail to suppress this response,leading to further development of NS.Therefore,there is an urgent need to explore the mechanisms underlying the onset and progression of NS and the susceptibility to the development of symptoms in NS,and to provide possible new research directions for NS.Proteomics is based on mass spectrometry,bioinformatics and other techniques to acquire the cellular,tissue organ and organismal proteins,providing functional and informative information on a set of proteins that can complement genomics and transcriptomics.Metabolomics is the simultaneous qualitative and quantitative analysis of all small molecules(<1000 Da)of endogenous metabolites,providing biological endpoint information.Integrated proteomics and metabolomics enable to mutually validate and complement of differential proteins and metabolites,providing more comprehensive information for the identification of new biomarkers and pathogenesis.[Objective]Liquid phase secondary mass spectrometry(LC-MS/MS)was used to acquire the significantly differentially expressed CSF proteins and metabolites between the SNS,ANS and non-neurosyphilis(NNS)patients,then integrate the proteomics and metabolomics profiles through bioinformatics to screen the suspected proteins/metabolites and related signalling pathways,providing new research directions for the possible pathogenesis of the NS.[Methods]SNS(n=7),ANS(n=10)and NNS(n=13)patients who visited the outpatient department of Guangzhou Institute of Dermatology from April 2021 to April 2022,were included the present study.Protein and small molecule metabolites in CSF were detected based on LC-MS/MS platform.The Fold change(FC)> 2 and p <0.05 were used as the screening criteria for significant differential proteins.The statistically significant differential metabolites were screened based on the variable importance projection(Variable importance in projection,VIP)≥ 1 in OPLS-DA model,FC ≥ 1.2 or ≤ 0.83,and p < 0.05.Finally,univariate correlation,unsupervised model,supervised model and biological function conjoint analysis were conducted for differential proteins and metabolites.[Results]1.Based on lable-free data independent acquisition(DIA)quantitative proteomics and bioinformatics analysis,a total of 268 differential expression of proteins(82 up-regulated and 186 down-regulated)were identified between NS vs.NNS,which were mainly involved in purine metabolism,glycosaminoglycan biosynthesis-heparin sulfate/heparin,and autophagy.A total of 35 differential expression of proteins were identified between SNS vs.ANS,of which 18 were up-regulated and 17 were down-regulated,enriched in the ribosome biogenesis in eukaryotes,and osteoclast differentiation pathways.2.Based on LC-MS/MS untargeted metabolomics,we screened 17 metabolites(11 up-regulated and 6 down-regulated)between NS vs.NNS,mainly enriched in purine metabolism,folate biosynthesis,and bile secretion pathways.A total of 11metabolites(all up-regulated)were detected between SNS vs.ANS,which were enriched in prostate cancer,neuroactive ligand-receptor interactions,cortisol synthesis and secretion,galactose metabolism,and other pathways.3.Correlation analysis of differential proteins and metabolites were conducted.The co-annotated pathways of differential proteins and metabolites between NS vs.NNS were purine metabolism and bile secretion,of which inclued integrin β4,APOB-100,glutamate receptor 2,glutamate receptor 4,CXCL10,HGprt,ADA2,HS6ST-3,XYLT-1,XYLT-2,uric acid,and 5-Hiaa(all p<0.05)that may participate in the occurrence and development of NS by destroying the function of the blood-brain barrier and increasing the permeability of the blood-brain barrier,as well as promoting the inflammation of the central nervous system.The differential proteins and metabolites between SNS vs.ANS were co-annotated to neuroactive ligand-receptor interactions and TRP channel-mediated inflammatory responses.Dysregulation of DPP4,cortisol,CAMK-IIα,APP,D-Asp,FSTL5,and PTN(all p<0.05)may play a important role in the mechanism of SNS by impairing the function of calcium signalling channels,neurons,hippocampal and synapses transmission.[Conclusion]1.There were significant differences in expression of protein and metabolism between NS vs.NNS,SNS vs.ANS in CSF,respectively,which were involved in the regulation of various signaling pathways and metabolic pathways respectively.2.Differential proteins and metabolites in the CSF between NS and NNS may be involved in the development of NS by impairing blood-brain barrier function,increasing blood-brain barrier permeability,and causing central nervous system inflammation.3.Differential proteins and metabolites in the CSF of SNS and ANS may play a role in the pathogenesis of SNS by down-regulating the function of calcium signalling channel and synapses,and damaging neurons.