Publications (2)2.34 Total impact
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Article: Quantitative immunoelectron-microscopic analysis of the type IV collagen alpha1-6 chains in the glomerular basement membrane in childhood thin basement membrane disease.
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ABSTRACT: Thin basement membrane disease (TBMD) is characterized histologically by diffuse thinning of glomerular basement membrane (GBM). Although recent genetic analysis has shown that TBMD might be included within type IV collagen disorders, conventional immunohistochemical studies demonstrated normal labeling of type IV collagen alpha chains in the GBM. We have, however, successfully used confocal laser scanning microscopy to demonstrate a significantly reduced signal of type IV collagen alpha5 chain (alpha5(IV)) along capillary walls in TBMD. In order to further understand the association of type IV collagen with TBMD, we used immunoelectron microscopy to examine renal biopsies from 6 children with TBMD and six control children with minimal change nephrotic syndrome. Ultrathin sections of LR gold resin were incubated with a rat monoclonal antibody against human alpha1(IV), alpha2(IV), alpha3(IV), alpha4(IV) alpha5(IV) or alpha6(IV) followed by colloidal gold conjugated goat anti-rat IgG. After taking electron micrographs, the labeling was quantitatively evaluated in the area occupied by the segments of basement membrane. The basement membrane was divided into three equal segments viz. subepithelial side, central portion and subendothelial side. In control subjects, the number of gold particles for alpha1(IV) or alpha2(IV) was significantly greater in the subendothelial side and central portion than in the subepithelial side of the GBM, whilst alpha3(IV), alpha4(IV) or alpha5(IV) labeling was significantly more prominent in the central portion compared to the subepithelial and subendothelial side of the GBM. TBMD samples showed a similar distribution pattern except that the subepithelial side and central portion of the GBM had a significantly reduced amount of alpha5(IV) antigen compared to control subjects. This is the first report demonstrating a diminished labeling intensity of alpha5(IV) in the central portion and subepithelial side of the GBM in renal biopsy specimens from patients with TBMD. These findings suggest that an abnormality of alpha5(IV) might possibly be associated with the pathogenesis of TBMD.Clinical nephrology 12/2005; 64(5):329-36. · 1.17 Impact Factor -
Article: Analysis of macrophages in urine sediments in children with IgA nephropathy.
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ABSTRACT: Although infiltrating macrophages found in renal biopsy specimens have been accepted as a useful marker for evaluating the activity of IgA nephropathy (IgAN), it is difficult to perform renal biopsies repeatedly, especially in children. To establish a more convenient and noninvasive method for estimating the degree of macrophage infiltration we examined the number of macrophages in urinary sediments. Ten ml of morning urine were collected from 30 children with IgAN, 10 with thin basement membrane disease (TBMD), 8 with idiopathic renal hemorrhage (IRH) which was defined as nonglomerular hematuria due to nutcracker phenomenon revealed on ultrasonography, and 10 healthy children as controls. Ten of the 30 children with IgAN were treated with combination therapy comprising prednisolone, warfarin and dipyridamole and urine samples were collected weekly during the period of treatment. Two microl of the urine sediment were smeared on glass slides, dried and stained with a monoclonal antibody to human macrophages (anti-CD68, PG-M1) followed by a FITC-conjugated secondary antibody. After staining with propidium iodide (PI), the cells were examined by fluorescence microscopy with cells stained with both FITC and PI being counted as macrophages. In addition, anti-CD68 staining was used to quantify macrophage infiltration in renal biopsies from the same group of IgAN patients. The number of urine macrophages in children with IgAN was significantly higher than in children with TBMD and IRH as well as the control group (p < 0.01), whereas that was similar among TBMD, IRH and healthy children. In IgAN, there was a significant correlation between urine macrophage number and the activity index (p < 0.01), proteinuria (p < 0.01) and urine WBC count (p < 0.01). In addition, there was also a significant correlation between urine macrophage number and glomerular (p < 0.05) as well as interstitial macrophage infiltration (p < 0.01). In children with IgAN who received combination therapy, urine macrophage number decreased significantly (p < 0.01) in the 1st week of treatment whilst the degree of proteinuria decreased significantly (p < 0.01) in the 4th week. Urinary macrophage number may represent a noninvasive and straightforward estimate of the pathological activity evident in renal biopsy specimens, and may also be a more sensitive indicator than proteinuria of the therapeutic effect of interventional treatments in childhood IgAN.Clinical nephrology 12/2004; 62(5):336-43. · 1.17 Impact Factor
