| OBJECTIVE: To explore the clinical and pathological heterogeneity of Ig A nephropathy through the retrospective analysis about the clinical and pathological characteristics of IgA nephropathy in Xinjiang Uygur region.;to optimize the urinary proteomics experiment program by exploring the application of iTRAQ labelled and Label free in urinary proteomics of IgA nephropathy;After the iTRAQ labelled with the application of liquid chromatography-mass spectrometry(LC-MS)and database search,to compare urinary proteome spectrum of the healthy population of Uygur and Han nationality,and to explore the differences between the urine protein;to compare the urinary proteome of Uygur healthy population and the patients with Ig A nephropathy,to explore the IgA nephropathy urine related differential protein spectrum in Xinjiang,with the use of protein database,to carry out search and analyze,to understand the pathogenesis of IgA nephropathy;through verifying the four different urine proteins,to screen candidate biomarkers of IgA nephropathy for Xinjiang.Methods: 1)Retrospectively analyze the clinical and pathological characteristics of 354 cases of Uygur and Han patients with primary Ig A nephropathy diagnosed in renal biopsy in Xinjiang,and analyze the similarities and differences between the clinical and pathological manifestations of IgA nephropathy in Uygur and Han,and the intrinsic relationship inconsistencies between Uygur and Han Ig A nephropathy clinical manifestations of proteinuria,hematuria,serum creatinine,glomerular filtration rate with pathological manifestations,and through the analysis about 22 cases of Uygur patients' clinical indicators,pathological indicators and survival of the kidneys after clinical intervention with poor efficacy and repeated renal biopsy,understand the disease incidence,development,intervention treatment effect and regression,understand the inconsistency of clinical manifestations and pathological manifestations of IgA nephropathy once again;2)through the research about i TRAQ labelled quantitative proteomics and Label free quantitative proteomics in Ig A nephropathy urine differentialproteomics,compare the advantages and disadvantages of both,optimize the experimental program for the study of urinary proteomics,select proper proteomics research methods according to different purpose;3)In this study,the urine of Uygur,Han healthy people and patients with IgA nephropathy diagnosed by renal biopsy in Xinjiang were taken as the research objective,albumin/Ig G antibody clearance and isotope relative labeling and absolute quantitative technique labeled two-dimensional LC-MS were used to establish IgAN urine protein mass spectrometry analysis methods for the detection method of urinary proteomics,to carry out racial and disease differential urinary proteome analysis,to obtain differentially expressed proteins,to conduct Gene ontology(GO)analysis for the identification of proteins and differential proteins,to detect the abundant protein of urine differential proteome of kidney between races and diseases,the differential protein screened between healthy people and IgA nephropathy patients was defined as Ig A nephropathy related differential proteins of Xinjiang Uygur,and to explore the IgA nephropathy-related urine protein mass spectrometry of Xinjiang region;four kinds of urine differential proteins were filtered from the urine of 6 cases of Uygur primary Ig A patients diagnosed by renal biopsy,6 cases of idiopathic membranous nephropathy,6 cases of healthy controls according to possible pathogenesis of IgA nephropathy,differential protein enrichment condition,to verify by differential expression protein with the use of Western blot,and to search candidate biomarkers for diagnosis of IgA nephropathy urine in Xinjiang.Results: 1)As for the clinical and pathological characteristics of IgA nephropathy in Han and Uygur,it was suggested that Uygur Ig A nephropathy had no special difference in clinical manifestations,but in pathological manifestation,Uygur IgA nephropathy patients have heavier endothelial cell proliferation,tubular-interstitial lesions and vascular lesions than Han patients.It was found after analyzing the related clinical and pathological indication of 22 cases of Uygur patients after repeated renal biopsy that except a small amount of Xinjiang Uygur IgA nephropathy patients' pathological indicator progression before and after biopsy,tubular-interstitial lesion,the proportion of spherical hardening and small artery lesions,there were no significant transformation;2)iTRAQ Label free studied urinary proteomics of IgA nephropathy,in the aspect of technical repetitive variation and biological individual variation,the iTRAQ could detect more protein,with relatively good technical repetition,based on the purpose of the experiment,iTRAQ quantitative proteomics is more appropriate in finding Xinjiang Ig A nephropathy urinary proteome non-invasive biomarkers,on such basis,optimization experimental program of XinjiangIgA nephropathy urinary proteomics spectrum was constructed initially;3)Through the LC-MS analysis about Uygur IgA nephropathy urinary proteome,a total of 640 kinds of proteins were identified,including 219 kinds of differential proteins,it could be found through the IPA functional analysis that acute stress response and NO related pathway function was reduced,vitamin C antioxidant pathway function was increased,the changes in pathway reflected the decompensated state of renal function processing.The functional analysis suggested that cell immune response,lipid metabolism abnormalities,coagulation system dysfunction,complement system activation play a vital role in the oncome and development of Uygur Ig A nephropathy.The findings from protein interactions network analysis were consistent with the founding above,and ADIPOQ,ICAM1,SERPINC-1,and TIMP1 could be used as an alternative protein for further functional validation and analysis.Four candidate biomarkers were selected from the differential urine proteins and four possible biomarkers were validated.They were respectively ADIPOQ involved in the inflammatory response,renal kidney tubular interstitial injury ADIPOQ and ICAM1,SERPINC-1 participated and affected the clotting process,as well as TIMP1 that affects the formation of extracellular matrix.The four biomarkers selected were validated by Western-bolt,suggesting that the AUC area of ADIPOQ and TIMP1 indicates that the two biomarkers have a higher diagnostic value and may serve as a candidate noninvasive biomarker for the diagnosis of Ig AN.CONCLUSION: There was no definite intrinsic relationship between Uygur IgA nephropathy clinical manifestation and pathology,showing heterogeneity,and there was an urgent need to find noninvasive biological indicators that can replace invasive renal biopsy,while it is possible to find noninvasive urine biomarkers for Xinjiang Uygur IgA nephropathy through the differential proteome analysis of optimized iTRAQ labeled quantitative urinary proteome,and differential protein signal transduction pathways,functional analysis suggested that cell immune response and lipid metabolism abnormality,coagulation system dysfunction,complement system activation play a vital role in the oncome and development of Uygur Ig A nephropathy.ADIPOQ and TIMP1 A may be used as a candidate noninvasive biomarker for the diagnosis of Ig AN;Urine protein mass spectrometry could be used to determine the degree of renal interstitial fibrosis and glomerular sclerosis,etc.by detecting urine differential proteins between different clinical manifestations and pathologic parameters of IgA nephropathy for the diagnosis,evaluation of IgA nephropathy to find new non-invasive biological markers,with a view to replace invasive renal biopsy paracentesis with urine proteomics analysisin future clinical practice. |
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