Glomerular Disease Rat Model, Differences In Urine Proteomics

Patients suffered from end-stage renal disease (ESRD) have to receive expensive renal replacement therapy to survive, among whom, more than half were caused by glomerular diseases. Effective treatments based on early definitive diagnosis may prevent or delay glomerular diseases deteriorating into ESRD. However, effect of the present therapeutics is not satisfying, and the development of new clinical therapeutics is hampered, because glomerular diseases are often poorly characterized by subtle histological changes on renal biopsy. The following limitations of renal biopsy bring issues such as either the correct patients are not enrolled or the outcome measures are inaccurate, and thus add complexity and risk to a clinical trial:1) it is not suitable for the patients with contraindications; 2) it is inconvenient for continuous monitoring; 3) it is not sensitive at the early stage of the disease; 4) it harvests such a small renal portion that it may not accurately portray the disease if the sampled portion is not affected. Urine test is expected to replace renal biopsy, for it is a simple, noninvasive test and can give the disease a more sensitive, unbiased and general portray.Glomerular diseases are considered to be consequences of injury primarily to the three cell types existing within the glomerular tuft:mesangial cells, podocytes (visceral epithelial cells), and endothelial cells. Not only does the form of injury but also the cell-specific response to injury govern the histological and clinical manifestations of glomerular diseases. Thus we expected that injuries of different glomerular cells would cause different changes in urinary proteome before the changes of cellular structure. The goal of this study was to identify differential urinary proteins distinguishing between injuries of different glomerular cells before significant histopathologic changes. For urinary proteomic analysis of glomerular diseases, high abundant proteins especially albumin in proteinuria had a strong suppression effect on identification of low abundant proteins, which in turn hampers in-depth analysis of urinary protein profiles. Therefore, ConA-enriched urinary proteome were profiled in this study.Adriamycin nephropathy and Thy1.1 glomerulonephritis were employed as models with different primary impaired cells. Adriamycin nephropathy has a progressive course with podocytes as the primary impaired cells. At the early phase, no apparent changes under light microscopy (LM), and only some focal "fusion" of foot processes by electron microscopy (EM). While Thy 1.1 glomerulonephritis is a self-limited process resembling mesangioproliferative glomerulonephritis (MsPGN) with mesangial cells as the primary impaired cells. Thy1.1 antigen is expressed on the surface of rat mesangial cells. The administration of a single iv injection of monoclonal antibodies (ox-7) directed against Thy1.1 antigen predictably leads to a brisk complement-dependent lysis of the mesangial cells within the first 48h; followed by the development of MsPGN. At the early phase, these two experimental glomerular disease models had similar histological appearance by LM. Then ConA-enriched urinary glycoproteome were profiled by gel-free shotgun tandem mass spectrometry, and compared with self-healthy controls to identify differential urinary proteins for each model. By comparing the changes of the differential proteins between these two models, we identified 39 proteins with different directions of changes, which may potentially be useful in differentiation; and 7 proteins with the same direction of changes, which may be potential indicators of early renal damage. They were of several origins:plasma proteins, proteins with urine or kidney specificity, proteins without tissue-specificity (mainly inflammatory mediators) etc. We suggest that a panel which contains proteins from several origins may provide glomerular diseases a more comprehensive view. Our results may help better understand the effects of injuries of different glomerular cells at the initial stage, differentiate different glomerular cell injuries, and finally achieve target therapy.