Comparative proteomic analysis of glomerular proteins in IgA nephropathy and IgA vasculitis with nephritis

Abstract

Background: IgA nephropathy (IgAN) and IgA vasculitis with nephritis (IgAVN) are related glomerular diseases characterized by marked similarities in immunological and histological findings. We herein performed a comparative proteomic analysis of glomerular proteins in IgAN and IgAVN.

Methods: We used renal biopsy specimens from 6 IgAN patients without nephrotic syndrome (NS) (IgAN-I subgroup), 6 IgAN patients with NS (IgAN-II subgroup), 6 IgAVN patients with 0-8.0% of glomeruli with crescent formation (IgAVN-I subgroup), 6 IgAVN patients with 21.2-44.8% of glomeruli with crescent formation (IgAVN-II subgroup), 9 IgAVN patients without NS (IgAVN-III subgroup), 3 IgAVN patients with NS (IgAN-IV subgroup), and 5 control cases. Proteins were extracted from laser microdissected glomeruli and analyzed using mass spectrometry. The relative abundance of proteins was compared between groups. An immunohistochemical validation study was also performed.

Results: More than 850 proteins with high confidence were identified. A principal component analysis revealed a clear separation between IgAN and IgAVN patients and control cases. In further analyses, 546 proteins that were matched with ≥ 2 peptides were selected. The levels of immunoglobulins (IgA, IgG, and IgM), complements (C3, C4A, C5, and C9), complement factor H-related proteins (CFHR) 1 and 5, vitronectin, fibrinogen chains, and transforming growth factor-β inducible gene-h3 were higher (> 2.6 fold) in the IgAN and IgAVN subgroups than in the control group, whereas hornerin levels were lower (< 0.3 fold). Furthermore, C9 and CFHR1 levels were significantly higher in the IgAN group than in the IgAVN group. The abundance of some podocyte-associated proteins and glomerular basement membrane (GBM) proteins was significantly less in the IgAN-II subgroup than in the IgAN-I subgroup as well as in the IgAVN-IV subgroup than in the IgAVN-III subgroup. Among the IgAN and IgAVN subgroups, talin 1 was not detected in the IgAN-II subgroup. This result was supported by immunohistochemical findings.

Conclusions: The present results suggest shared molecular mechanisms for glomerular injury in IgAN and IgAVN, except for enhanced glomerular complement activation in IgAN. Differences in the protein abundance of podocyte-associated and GBM proteins between IgAN and IgAVN patients with and without NS may be associated with the severity of proteinuria.

Keywords: Comparative proteomic analysis; Glomerular proteins; IgA nephropathy; IgA vasculitis with nephritis; Mass spectrometry.

Fig. 1

Principal component analysis. Data obtained from 6 IgAN patients without NS (IgAN-I), 6 IgAN patients with NS (IgAN-II), 6 IgAVN patients with 0–8.0% of glomeruli with crescent formation (IgAVN-I), 6 IgAVN patients with 21.2–44.8% of glomeruli with crescent formation (IgAVN-II), and 5 control cases (time 0 transplant biopsies) are shown. Data from the IgAN-I, IgAN-II, IgAVN-I, and IgAVN-II subgroups and the control group are represented by green, red, orange, blue, and purple circles, respectively. Variance is given as a percentage for both the first and second principal components (PC1 and PC2).

Fig. 2

Immunohistochemical staining for talin 1. a-c Three IgAN patients without NS. Glomerular talin 1 staining is observed in parietal epithelial cells and podocytes, along capillary walls, and in the urinary space (probable cell debris of detached parietal epithelial cells and podocytes). Staining for talin 1 is also detected in proximal tubular epithelial cells. d-f Three IgAN patients with NS. Severe mesangial expansion is observed. Staining for talin 1 in parietal epithelial cells and podocytes, along capillary walls, and in the urinary space is less intense than that in panels a-c. Enlarged images are inserted at the lower left. Bars represent 20 μm. Original magnification of each image: ×400