Preliminary Exploration On The Underlying Pathogenesis Of Chronic Prostatitis/chronic Pelvic Pain Syndrome(CP/CPPS)Based On Multi-omics Techniques

Chronic Prostatitis/Chronic Pelvic Pain Syndrome(CP/CPPS)accounts for more than 90%of all types of prostatitis.Despite this,global attention to this type of non-fatal complex disease is far less than other types of prostate diseases,such as prostate cancer(PCa)and benign prostatic hyperplasia(BPH),leading to the current deficiency of the understating of pathogenesis of CP/CPPS.It is obviously difficult to find breakthroughs in such diseases with complex pathogenesis,clinical phenotypes and heterogeneity by relying on the traditional single method of research.The overall progress of CP/CPPS is long-playing.Lacking accurate classification method based on biological mechanism results in slow development of clinical treatment,which is a choke point for CP/CPPS to break through.Traditionally,the research mode using a single method and targeting a single factor,can only pry into the tip of the iceberg of CP/CPPS.With the development of Multi-Omics techniques and the popularization of the concept of system biology,researchers are able to conceive deeply about complex diseases from a variety of perspectives.Using the research methods and techniques of multi-omics,the multi-dimensional display of CP/CPPS can grant researchers a comprehensive and novel understanding of such complex diseases,which is more conducive to the investigation of CP/CPPS personality,snagging and discovering the core problems of CP/CPPS.Unfortunately,although the concept of systems biology has been raised up for many years,the current research of CP/CPPS is still based on monotonous methods,and the incomplete information hinders the researchers' accurate interpretation of the internal mechanism of CP/CPPS.Therefore,for the first time,the current study used multi-omics research methods to primarily investigate the biological mechanism of CP/CPPS,aiming at constructing a multi-dimensional stereoscopic CP/CPPS road map to provide a reference for accurate classification based on biological mechanism and individualized treatment of CP/CPPS.The current study is mainly consisted of the following three parts:In the first part of the work,we,for the first time,obtained the differentially expressed gene profile of CP/CPPS tissue using the technique of whole genome transcriptome sequencing.Through transcriptome sequencing,we observed that the overall gene expression profile of CP/CPPS patients was significantly distinguishable from that of non-CP/CPPS controls,and as many as 1,941 differentially expressed genes underwent remarkable changes.In the personalized analysis,we found that the expression levels of key immune-related genes and metabolic-related genes in CP/CPPS patients varied evidentially.We also found that the related genes of B cells,CD4+T cells,CD8+T cells,neutrophils,macrophages and dendritic cells(DCs)were highly expressed.This is the first time for us to construct immune and metabolic gene differential expression profiles for clinical CP/CPPS patient.According to system biology theory,we propose that immune factors are potentially to occupy an essential position in the pathogenesis of CP/CPPS.The CP/CPPS differentially expressed genotype data constructed in this part will provide a reference for the future study of CP/CPPS mechanism.In the previous part of the study,we found the characteristic changes of critical immune genotypes in local CP/CPPS prostate tissues.According to system biology theory,will the changes in local immune factors be reflected in the overall immune level of the body?Therefore,in the second part of the work,we first verified the hypothesis by using proteomics technology,the world's advanced mass cytometry to detect peripheral blood immune cells of CP/CPPS.With mass cytometry,we found for the first time that CP/CPPS patients had distinct immunophenotypic characteristics,mainly natural killer cells(NK),myeloid dendritic cells(mDCs),and classical monocytes(Classical monocytes,Regulatory T cells(Treg)and naive CD8+T Cells(Naive CD8+T Cells)in CD45+cells were significantly higher in CP/CPPS than those in controls,while helper T cells 1(Thl)and effector CD8+ T Cells were significantly lower.Furthermore,the cytokines in these immune cells,such as MIP-1?,TNF?,IL-1?and MCP-1,also changed apparently between CP/CPPS and controls.Then we further exploited a proteomics technology-Multiplex Assays-to detect cytokines in serum and expressed prostatic secretions(EPS)of CP/CPPS patients,and found that cytokines such as MIP-1?,TNF?,IL-1? and MCP-1 also changed significantly,which was in line with the findings from mass spectrometry.In addition,the expression levels of cytokines from serum and EPS were remarkably correlated.Hence,we believe that both the proportion of global immune cells and the secretion of local cytokines would be changed during the development of CP/CPPS,and the global immunity is connected with local immunity by the way of immune cells-cytokines.To the best of our knowledge,this is the first study in the world to utilize proteomics approaches to reveal the accordant relationships between CP/CPPS local immunity and overall immunity.In the third part of the work,inspired by the variations in the metabolism-related gene expression profile of prostate tissue of CP/CPPS in the first part,we used metabonomics methods to detect the metabolites in the urine of CP/CPPS patients for the first time.The technique of liquid chromatography mass spectrometry(LC-MS)provided us with a better understanding of the metabolic status of CP/CPPS patients.Metabolomics studies can help understand the final consequences of small molecule production and metabolism in urine of CP/CPPS patients.Based on the premise of additive and compensatory changes in the first two parts of the genomic and proteomic status,metabolomics is useful for a more detailed understanding of the effective small changes caused by disease state and body compensation,which are magnified in metabolites.Therefore,the identification of metabolites is easier and more accurately reflects the state of the biological system.In the part,we found 314 differential metabolites in positive ion mode and 222 differential metabolites in negative ionization mode by LC-MS technology.A new model of combined diagnosis for CP/CPPS based on Methylguanidine,Chloroxylenol and Methyl 3,4,5-trimethoxycinnamate from positive ionization mode showed a good diagnostic efficiency with AUC value of 0.880.And a new model of combined diagnosis constructed by N-Tridecanoylglycine,N-Undecanoylglycine,Dodecanedioic acid,Hydrocinnamic acid and 3-tert-Butyl-4-hydroxyanisole from negative ionization mode exhibited a favorable clinical diagnostic value of CP/CPPS with AUC value of 0.909.The KEGG pathway enrichment results showed that the differential metabolites in the positive ionization mode were mainly enriched in the pathways of Riboflavin metabolism,Flavone and flavonol biosynthesis,Biosynthesis of type ? polyketide products,Penicillin and cephalosporin biosynthesis and Histamine H2/H3 receptor agonists/antagonists.For the negative ionization mode,the differential metabolites were mostly enriched in the pathways of Purine metabolism,Metabolism of xenobiotics by cytochrome P450,Naphthalene family,Microbial metabolism in diverse environments and Biosynthesis of phenylpropanoids.We believe that during the development of CP/CPPS,metabolism-related pathways would be changed,leading to the changes of the metabolite expression profile.There has been currently no reference to the metabolome measurement data of CP/CPPS patients,hence,our work in this part can provide a wide range of information for future researchers.In summary,we have used a multi-omics research method in the current project taking advantages of the clinical CP/CPPS tissue specimens,prostate fluid specimens,peripheral blood specimens and urine specimens.From local to systemic,the potential pathogenesis of CP/CPPS,a comprehensive study of transcriptomics,proteomics and metabolomics has been carried out.Of special interest are the current research models and findings to be taken as the important foundation and reference for further research on CP/CPPS in the future.