Chronology of renal scarring in males with Alport syndrome.

We investigated the onset of renal scarring in 62 males (aged 4-26 years) with Alport syndrome by measuring cortical interstitial volume fraction [Vv (interstitium/cortex)] and percentage global glomerular sclerosis in kidney biopsies. Male pediatric (n = 9) and adult (n = 7) donor kidneys served as controls. Creatinine clearance at the time of biopsy was available for 43 Alport patients. A statistically insignificant correlation between age and Vv (interstitium/cortex) was observed in normal subjects (r = +0.47, slope = 0.0009, P = 0.07). In the Alport patients, age was significantly correlated with Vv (interstitium/cortex (r = +0.49, slope = 0.01, P = 0.001) and global glomerular sclerosis (r = +0.41, P = 0.01), and inversely correlated with creatinine clearance (r = -0.33, P = 0.04). Creatinine clearance was inversely correlated with Vv (interstitium/cortex) (r = -0.78, P = 0.001) and global glomerular sclerosis (r = -0.74, P = 0.001). The correlation with creatinine clearance was especially strong for Vv (interstitium/cortex) values above the normal range, i.e., > 0.2 (r = -0.82, P = 0.001), and was absent for Vv (interstitium/cortex) < 0.2 (r = -0.119, P = 0.55). Creatinine clearance values less than 80 ml/min per 1.73 m2 occurred more frequently in patients with Vv (interstitium/cortex) values > 0.2 (P < 0.0001) and in patients with > 10% globally sclerosed glomeruli (P < 0.001). Patients < or = or > 10 years of age differed in Vv (interstitium/cortex) [0.13 +/- 0.09 (mean +/- SD) vs. 0.24 +/- 0.026, P < 0.001], the frequency of Vv (interstitium/cortex) > 0.2 (3/32 vs. 15/31, P < 0.0001), the frequency of > 10% globally sclerosed glomeruli (3/33 vs. 11/30, P < 0.05), mean creatinine clearance (113 +/- 7 vs. 84 +/- 10 ml/min per 1.73 m2, P = 0.057), and the frequency of creatinine clearance < 80 ml/min per 1.73 m2 (1/20 vs. 11/23, P < 0.01). Thus, reduced creatinine clearance in males with Alport syndrome is associated with Vv (interstitium/cortex) > 0.2 and > 10% globally sclerosed glomeruli. These are frequently detectable in the 2nd decade. We hypothesize that most Alport males will require intervention during the 1st decade for optimal preservation of kidney function.

Proximal tubular basement membrane width in insulin-dependent diabetes mellitus.

Although glomerular structure has been studied, careful evaluation of tubular basement membrane (TBM) structure in diabetes in humans has not been done. We measured proximal TBM width, glomerular basement membrane (GBM) width, mesangial fractional volume [Vv(Mes/glom)], mesangial matrix fractional volume [Vv(MM/glom)], and cortical interstitial fractional volume [Vv(Int/cortex)] in 35 insulin-dependent diabetic (IDDM) patients and 20 controls. The patients' mean age was 28 +/- 10 years (X +/- SD) and IDDM duration was 17 +/- 8 years. Twenty-five patients were normoalbuminuric, four microalbuminuric, and six had overt proteinuria. Tubular basement membrane and GBM widths were measured by the orthogonal intercept method and mesangial and interstitial parameters by point counting. The TBM width was 915 +/- 320 nm in IDDM patients and 558 +/- 116 nm in controls (P = 0.0005); the TBM width was also increased in normoalbuminuric patients (849 +/- 297 nm, P = 0.0005). The TBM width was strongly directly related to GBM width (r = 0.67, P < 0.001), Vv(Mes/glom) (r = 0.52, P < 0.01), and Vv(MM/glom) (r = 0.61, P < 0.001), but only weakly to Vv(Int/cortex) (r = 0.29, NS). The TBM width (r = 0.65, P < 0.001) and GBM width (r = 0.65, P < 0.001) were strongly related to hemoglobin A1C (HbA1C), while the Vv(Mes/glom) (r = 0.35, P < 0.05) and Vv(Int/cortex) (r = 0.30, NS) were only weakly related to HbA1C. Thus, increased proximal TBM width is an integral component of early nephropathology in IDDM patients. This study suggests that the metabolic disturbances of diabetes are strong determinants of the constellation of structural abnormalities occurring in human diabetic nephropathy.

Heparan sulfate proteoglycan in the glomerular basement membrane in type 1 diabetes mellitus.

Heparan sulfate proteoglycans (HSPG) are negatively charged constituents of the renal extracellular matrix including the glomerular basement membrane (GBM) and mesangial matrix. Biochemical and functional studies of patients with type-1 insulin dependent diabetes mellitus (IDDM) suggest that alterations of HSPG may occur in diabetic nephropathy. We have utilized a specific cytochemical method and electron microscopy to quantitate the distribution of HSPG in the GBM of 10 normal people and in 16 IDDM patients with a spectrum of clinical and structural changes. Enzyme incubation studies of normal infant kidney demonstrated that heparitinase removed 94% of the stainable anionic sites in the lamina rara externa (LRE) and 77% of the sites in the lamina rara interna (LRI) of the GBM. In contrast, incubation in the enzyme chondroitinase ABC did not reduce the number of sites in the LRE but reduced the number of sites in the LRI by 26%. The HSPG anionic sites in normal subjects were distributed in the LRE as 20.9 +/- 1.3, and in the LRI as 13.1 +/- 2.2 per micron GBM length. Anionic sites were slightly reduced (19.6 +/- 1.3, P less than 0.04) in the LRE of IDDM patients with normal urinary albumin excretion rates (UAE), or microalbuminuria, and were reduced in both the LRE and LRI of IDDM patients with clinical proteinuria (13.1 +/- 2.3, P less than 0.001 and 8.9 +/- 2.1, P less than 0.001, respectively). The number of anionic sites in the LRE and LRI, respectively, correlated with UAE (r = +0.78, P less than 0.001, r = +0.58, P less than 0.02), with GBM thickness (LRE, r = +0.81, P less than 0.001; LRI, r = +0.67, P less than 0.01) and with the volume fraction of mesangium (LRE, r = +0.59, P less than 0.02; LRI, r = +0.58, P less than 0.03). These data confirm earlier biochemical findings of a reduction of HSPG in the GBM in advanced diabetic nephropathy but do not provide evidence for the loss of HSPG in the GBM as a mechanism for early microalbuminuria.

Anionic sites in the human kidney: ex vivo perfusion studies.

Heparan sulfate-proteoglycan (HS-PG) anionic sites located in the glomerular basement membrane (GBM) are thought to contribute to glomerular permselectivity in man. The number and distribution of HS-PG anionic sites in the GBM and mesangial matrix of seven normal human kidneys and three kidneys from children with congenital nephrotic syndrome (CNS) were evaluated by ex vivo perfusion of polyethyleneimine (PEI; Mr 40,000 to 60,000). In the normal kidneys lamina rara externa (LRE) anionic sites (21.8 +/- 2.4 per 1000-nm actual GBM length) were well labeled and similar to those obtained by immersion staining of fixed tissue with Mr 1200 PEI. Lamina rara interna (LRI) anionic site number (22.0 +/- 2.6 per 1000-nm GBM) and appearance were better demonstrated by PEI perfusion than by the immersion technique. PEI perfusion also demonstrated regularly arranged (60 to 120 nm apart) mesangial matrix anionic sites (20- to 30-nm diameter) at 4.09 +/- 0.59 x 10(3) sites per nm2 matrix. PEI perfusion of three kidneys from children with CNS demonstrated decreased (16.3 +/- 1.9 per 1000-nm GBM) LRE anionic sites and normal LRI anionic sites (22.0 +/- 3.5 per 1000-nm GBM). Mesangial volume was increased, and mesangial anionic sites were less frequent (3.24 +/- 0.42 x 10(3) per nm2 matrix) and irregular in size in the children with CNS.(ABSTRACT TRUNCATED AT 250 WORDS)

Glomerular basement membrane anionic charge site changes early in aminonucleoside nephrosis.

Alterations of glomerular basement membrane (GBM) anionic (charge sites, CSs) in the development of proteinuria in a model of idiopathic nephrotic syndrome in man (puromycin aminonucleoside nephrotic syndrome [PAN] in the rat) were assessed quantitatively and sequentially early after disease induction. GBM CSs (known to consist mainly of heparan sulfate-rich proteoglycans) were stained in vivo and, in a separate group of animals by an in vitro method, with the cationic marker polyethyleneimine (PEI) studied by electron microscopic examination. Four hours after administration of PAN, there was a significant decrease in GBM lamina rara externa CSs: 18 +/- 0.7 versus 22.0 +/- 2.2 per 1000 nm GBM in controls by PEI injection and 17.2 +/- 2.7 versus 21.1 +/- 1.6 per 1000 nm GBM in controls by PEI in vitro staining. This CS alteration coincided with changes in glomerular epithelial cell morphologic characteristics (increased cytoplasmic organelles and rough endoplasmic reticulum) and preceded the detection of foot process broadening (at 24 hours) and increased urinary albuminuria (suggested at 12-24 hours, statistically significant at 36-48 hours). These results suggest that GBM CS-heparan sulfate proteoglycan alterations consisting of either decreased number and/or less anionic charge occur early in PAN and support a role for glomerular epithelial cell maintenance of GBM CS for normal glomerular function.