Compare Hormone Therapy With That Before On Urine Proteomics Of Immunoglobulin A Nephropathy

Objective: To discuss the polypeptide pattern in urine of patients with Immunoglobulin A nephropathy(IgA nephropathy)，comparing the effect in hormone therapy before and after treatment. In order to explore the sensitive and specificprotein in Immunoglobulin A nephropathy.Probe into the purpose and significance of dieases dynamic monitoring in urine proteomics.Methods: Morning first medistream urine sample form6healthy people and6patients were collected. So as to avoid bacterial growth and protein enzymolysis, stored samples at-80℃in1h.First, melted the18samples,and then centrifugated at4℃1500×g for10minutes，removing the cellular debris. After that, centrifugated at4℃10000×g for10minutes again,in order to remove other particlesin urine.The urine samples were fractionated by pre cold organic solvent(the volume ratio was4:1),matching into by acetone and ethanol, at-20℃overnight.Then centrifugated at4℃2500×g for30minutes, and abandoned supernatant fluid, and obtained the precipitate which contained urine protein, then dried downin a vacuum centeifuge,kept in-20℃for later use. After that, the protein concentration of each sample was determined by Bradford (Commassie Blue Fast Staining Solution) assay after dissolved fully with PBS (phsosphate buffer). Put1mg of samples to enzymolysis, and then took it to desalt at HPLC. After that,dried and concentrate the liquid mixture which contained peptides. Analysised with HPLC-ESI-MS/MS.The protein identification was searched in the database ofhuman_v3.71_reverse.faster with the help of MASCOT.Results:1.98proteins were identified from6urine samples of healthypeople.After the manual validation,29proteins were commomly expressed.2.165proteins were identified from IgA nephropathy before hormone therapy,.Through the manual validation,11proteins were commomly expressed. Compri sion them with the healthy,there were10specific proeins were expressed.Theywere Ig heavy chain V-III region TEI、Ig gamma-2chain C region、Alpha-1-antichymotrypsin、Alpha-1-antitrypsin、Vitamin D-binding protein、Hemoglobin subunit beta、IGL@protein、Ceruloplasmin、IGK@protein、Anti-(ED-B) scFV andSerotransferrin.Conclusion:1. Result from HPLC-ESI-MS/MS,165proteins were identifiedfrom IgA nephropathy,and we found the polypeptide pattern in urine of patientswith Immunoglobulin A nephropathy(IgA nephropathy) primarily. Ig heavy chainV-III region TEI、Ig gamma-2chain C region、Alpha-1-antichymotrypsin、Alpha-1-antitrypsin、Vitamin D-binding protein、Hemoglobin subunit beta、IGL@protein、Ceruloplasmin、IGK@protein、Anti-(ED-B) scFV and Serotransferrin may become the landmarks of IgA nephropathy.2. Compared hormone therapy with that before on Urine proteomics of Immunoglobulin A nephropathy,some kinds of the proteins may loss.