Study Of ITRAQ-based Quantitative Proteomics In Synovial Fluid Of Temporomandibular Joint Disease

Background : Temporomandibular joint disorder(TMD)ranks fourth in the ranking of common diseases in the Stomatology,which may accompany by snapping joint,joint dysfunction and joint pain.Anterior disc displacement without reduction(ADDWR)and temporomandibular joint osteoarthrosis(TMJOA)are the two subtypes with the highest clinical visit rates.Clinically,there is one another disease that named for progressive condylar resorption with unknown etiology—idiopathic condyle absorption(ICR),which shows many overlaps and similarities with the symptoms,signs and imaging characteristics of TMD.Due to the complexity of the pathogenesis and the limitation of previous research methods,the etiology and pathogenesis of temporomandibular joint diseases are still unclear,and there is a lack of diagnostic and differential methods at the molecular level,as well as precise managements.These diseases seriously affect patients' quality of life till now.At present,quantitative proteomics,represented by isotope-labeled relative and absolute quantification(iTRAQ)technology,provides important ideas for the for the diagnosis,stage or activity monitoring,prognosis evaluation and the study of pathogenesis of joint diseases.Objective: The purpose of our study was to validate and analyze the differently expressed proteins(DEPs)in the synovial fluid(SF)of ADDWR,TMJOA and ICR patients by iTRAQ-based quantitative proteomics technology.We analyze the biological process and signal pathways by bioinformatic method,and screening the candidate biomarkers in SF.It is expected to provide a theoretical basis for the preliminary elaboration of the pathogenesis,the search for clinical diagnosis indicators and treatment targets.Methods: A total of 22 patients diagnosed as ADDWR,22 as TMJOA,15 as ICR and 10 healthy volunteers were recruited from May 2019 to October 2019 in stomatology of Chongqing medical university.SF samples were collected by conventional supraarticular puncture.Bradford method was used to quantify the total protein in samples.iTRAQ technology combined with Liquid chromatography-mass/mass(LC-MS/MS)spectrometry was used to isolate and identify the protein in the SF.Bioinformatics analysis such as GO,KEGG and PPI were performed for these DEPs.The potential biomarkers were screened by integrating protein differential expression,bioinformatics analysis results and literature reports.Result:(1)Totally,370 proteins were identified by iTRAQ technique.Compared with the healthy controls,56 proteins showed differently expressed,of which 14 were up-regulated and 42 were down-regulated in ADDWR group;in TMJOA group,45 proteins were up-regulated and 37 proteins were down-regulated in the 82 DEPs;in ICR group,38 proteins were up-regulated and 17 proteins were down-regulated in the 48 DEPs.(2)GO function analysis results showed that 25 biological processes,9 cell components and 8 molecular functions were annotated in ADDWR group DEPs;25 biological processes,9 cell components and 8 molecular functions were annotated in TMJOA group DEPs;25 biological processes,10 cell components and 8 molecular functions were annotated in ICR group DEPs.The main GO terms of DEPs were involved in biological processes of metabolic process,biological regulation,cellular process,response to stimulus;molecular functions of binding,enzyme regulator activity,catalytic activity;cell components of extracellular region part and organelle.(3)Comprehensive analysis of KEGG pathway results of the three groups of differential proteins showed that complement and coagulation cascade was the most important metabolic pathway.DEPs involved in this pathway included FIBB,FIBG,C4 BPA,C1R,PEDF,KAIN,IC1,et al.(4)Up-regulated proteins in the three disease groups included FIBB,APO,et al;down-regulated proteins in the three disease groups included PRDX2,CAH1,et al;HPT,S100A8,MMP3 showed specific up-regulated in TMJOA group;CH3L1 showed specific up-regulated,while PRG4 showed specific down-regulated in ICR group.Conclusions: Coagulation and complement cascade could play an important role in the occurrence of ADDWR,TMJOA,and ICR.The disorder of lipid metabolism may closely relate to TMJ diseases,especially TMJOA.HPT and S100A8 may be used as candidates for TMJOA diagnosis,CH3L1 and PRG4 may be used as candidates for ICR diagnosis,and POSTN may be a candidate for distinguishing TMD from ICR.This article provided a reference for screening the treatment targets of three temporomandibular joint diseases.