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Saudi Journal of Kidney Diseases and Transplantation
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ORIGINAL ARTICLE  
Year : 2013  |  Volume : 24  |  Issue : 2  |  Page : 286-291
Glomerular filtration barrier in pediatric idiopathic nephrotic syndrome


1 Department of Pathology, All India Institute of Medical Sciences, Ansari Nagar, New Delhi, India
2 Department of Pediatrics, All India Institute of Medical Sciences, Ansari Nagar, New Delhi, India

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Date of Web Publication26-Mar-2013
 

   Abstract 

Nephrotic syndrome (NS) is a common proteinuric disorder with defect in the perm-selectivity of the glomerular filtration barrier (GFB). Ultrastructural morphometric evaluation of the GFB in pediatric NS has been attempted in only a few studies. This study was aimed at qualitative and quantitative evaluation of the alterations involving the GFB in pediatric idiopathic NS with an attempt to correlate these alterations with the clinico-laboratory data. For this study, renal biopsies from nine patients with NS and two children with interstitial nephritis were included. Relevant clinical and laboratory data, including degree of 24-h proteinuria and renal function tests, were recorded. Renal biopsies were reviewed for morphologic and electron microscopic diagnosis. Ultrastructural morphometry of the GFB was performed using image analysis software. The age at onset of NS, duration of illness, presence of hypertension, and renal function tests were comparable between the group of patients with minimal change disease (MCD) and those with mesangioproliferative glomerulonephritis (mesPGN)/focal segmental glomerulosclerosis (FSGS). However, the latter group showed higher 24-h proteinuria compared with the group with MCD. Among the detected ultra-structural changes, glomerular basement membrane thickness and foot process width were significantly different between the MCD and the mesPGN/FSGS groups. The slit pore diameter in the glomeruli showed a positive correlation with the degree of proteinuria. We conclude that our study demonstrated remarkable differences in certain parameters and the glomerular ultrastructural alterations in the various categories of NS. These differences might underlie the observed variation in response of these entities to various therapies.

How to cite this article:
Sharma A, Gupta R, Bagga A, Dinda AK. Glomerular filtration barrier in pediatric idiopathic nephrotic syndrome. Saudi J Kidney Dis Transpl 2013;24:286-91

How to cite this URL:
Sharma A, Gupta R, Bagga A, Dinda AK. Glomerular filtration barrier in pediatric idiopathic nephrotic syndrome. Saudi J Kidney Dis Transpl [serial online] 2013 [cited 2019 Nov 22];24:286-91. Available from: http://www.sjkdt.org/text.asp?2013/24/2/286/109577

   Introduction Top


Nephrotic syndrome (NS) is a frequent pediatric renal disorder with variable response to therapy with steroids. Various animal experiments have outlined the basic defect to be an alteration in the glomerular filtration barrier (GFB) leading to an increased permeability to proteins and solutes. [1] However, the exact mechanisms underlying this defect are to a large extent unclear. Renal biopsy findings in pediatric NS commonly include minimal change disease (MCD), focal segmental glomerulosclerosis (FSGS), and mesangioproliferative glomerulonephritis (mesPGN). FSGS and mesPGN are associated with relatively poor response to steroids and unfavorable long-term prognosis. [2]

Electron microscopic examination of renal biopsies is considered essential in the diagnosis of pediatric NS to assess ultrastructural changes such as podocyte foot process effacement (FPE), basement membrane changes, and absence of electrondense deposits. [3] Morphometric evaluation of the GFB on ultrathin sections in pediatric NS has been attempted in very few studies, which have focused on only one or the other ultrastructural parameter of GFB. Studies on glomerular basement membrane (GBM) thickness did not show a significant difference between proteinuric patients and in those in remission, or between the various histomorphological patterns in NS. [4],[5] Other studies have evaluated slit diaphragm to show a reduction in the number of filtration slits in MCD. [6],[7] However, the results of ultrastructural morphometry are largely at variance in the various studies published so far.

We aim in this study to evaluate the ultrastructural morphometric alterations of the GFB in pediatric idiopathic nephrotic syndrome and compare the major histomorphologic diagnoses with regards to these alterations.


   Patients and Methods Top


This retrospective study included nine patients with idiopathic nephrotic syndrome (INS): four were diagnosed as MCD, four as mesPGN, and one as FSGS. For comparison of the ultrastructural features, two pediatric renal biopsies with interstitial nephritis were also included as controls.

In the nephrotic patients, age at onset of NS, duration of illness, and presence of hypertension were recorded. Relevant investigations including renal function tests (serum urea and creatinine), 24-h urinary protein excretion, and serum albumin were obtained.

The renal biopsy specimens for electron microscopy were fixed in 3% glutaraldehyde with 0.1 mol/L phosphate buffer, followed by post-fixation in buffered 1% osmium tetroxide and embedding in epoxy resin. Ultrathin sections (60-80 nm) were cut on copper grids and double-stained by uranyl acetate and lead citrate. Transmission electron microscopy was performed on Philips Morgagni 268 microscope (Philips, Eindhoven, The Netherlands). Digital images of a minimum of two glomeruli per biopsy were captured and ultrastructural morphometry was performed using the analySIS Pro TM software (Soft Imaging System, Muenster, Germany). For morphometry, a minimum of 15 measurements of endothelial fenestration width, GBM thickness, slit pore diameter, and podocyte foot process width were taken [Figure 1]a-d. At least 400 μm length of the GBM was measured to assess podocyte foot process width distribution.
Figure 1. Electron micrographs that show measurement of glomerular basement membrane thickness (a, ×880), podocyte slit pore diameter (b, ×3500) and foot process width (c, ×3500). One of the cases of minimal change disease shows a markedly thickened glomerular basement membrane (d, ×3500).

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Appropriate statistical tests were applied to analyze the difference between the groups with regard to clinical, laboratory, and ultrastructural morphometric parameters. For statistical analysis, mesPGN and FSGS were grouped together.


   Results Top


Of the nine patients with NS, five were girls. The mean age at onset of NS was comparable in both the groups of MCD and mesPGN/FSGS (31.5 ± 26.5 months vs. 34.7 ± 16.3 months, P = 0.82). Similarly, no significant difference was found in the duration of NS at biopsy (28.8 ± 16.4 months in the MCD vs. 21.8 ± 14.6 months in the mesPGN/FSGS groups, P = 0.51). Hypertension at the time of biopsy was detected in one patient with MCD and two in the mesPGN/ FSGS group.

There were no significant differences in the serum levels of urea, creatinine, or albumin between the MCD and mesPGN/FSGS groups. A significant difference was, however, found in the degree of 24-h proteinuria (4.47 ± 1.0 g/day in MCD vs. 6.9 ± 1.5 g/day in mesPGN/FSGS, P = 0.02), as shown in [Table 1].
Table 1: Clinical and laboratory data of children with idiopathic nephrotic syndrome.

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Ultrastructural features

In the MCD group, diffuse FPE, microvillous transformation of podocytes, increase in slit pore diameter, and mild diffuse thickening of GBM were the common ultrastructural changes [Figure 2]. In addition, the mesPGN/FSGS group showed frequent mesangial expansion on electron microscopy.
Figure 2. Uranyl acetate- lead citrate stained electron micrographs that show diffuse effacement (a, ×880) and microvillous transformation (b, ×1800) of podocyte foot processes. This case demonstrates podocyte hypertrophy, cytoplasmic vacuolization and microvillous transformation (c, ×880). Higher magnification micrograph shows cytoplasmic vacuolization of podocyte foot processes (d, ×2200)

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Ultrastructural morphometric evaluation

The GBM thickness in the MCD group was lower than that in mesPGN/FSGS group (P = 0.01). A significant difference was also seen in the GBM thickness between the control biopsies and the mesPGN/FSGS group (P = 0.01). However, though the GBM thickness was higher in the MCD group compared to the controls, the difference was not statistically significant.

Endothelial fenestration width was comparable in all the three groups (controls, MCD, and mesPGN/FSGS). Slit pore diameter was significantly increased in the mesPGN/FSGS group compared to the controls (P = 0.04), though no statistical difference was found between the MCD group and the controls, as well as between the MCD and the mesPGN/FSGS groups. The morphometry data are shown in [Table 2].
Table 2: Results of glomerular ultrastructural morphometry in patients with idiopathic nephrotic syndrome.

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In contrast to other morphometric parameters, the foot process width was highest in the MCD group with a significant difference from the controls (P = 0.005) and the mesPGN/FSGS group (P = 0.005); no significant difference was found between the controls and the mesPGN/ FSGS group.

There was a significant correlation between the degree of proteinuria with the slit pore widening (P = 0.01), while there was no correlation with the degree of podocyte FPE.


   Discussion Top


The basic functional defect in NS is increased permeability of the GFB, which is composed of a fenestrated endothelium, GBM, and visceral epithelial cells (podocytes). Various factors affecting the filtration of molecules across the GFB include physical properties of the particle (size larger than 40 Å or weight >200 kD is not filtered), charge on the surface of molecule (negatively charged molecules do not pass easily due to the anionic charge on GBM), and dynamics of renal blood flow. [1] However, the exact mechanisms of alterations in permselectivity of the GFB in either MCD or FSGS remain unknown.

The most common pathomorphologic forms of pediatric NS include MCD, FSGS, and diffuse mesangial proliferation (mesPGN). Recent studies have shown an increase in biopsy diagnosis of FSGS, partly due to the more liberal biopsy recommendations in the pediatric NS. [2] Various studies have also shown that mesPGN and FSGS are associated with frequent relapses, steroid dependence/resistance, and higher incidence of renal function derangement. [1] In the present study, patients with biopsy diagnosis of mesPGN/FSGS had significantly higher degree of proteinuria compared to those with MCD. Other clinical parameters including age at onset of disease, duration of illness, prevalence of hypertension, and serum urea and creatinine were not significantly different between the two groups.

Ultrastructural examination of renal biopsies forms an integral part of evaluation in patients with medical renal diseases. In a study of electron microscopy in the pediatric NS, ultrastructural examination was essential for diagnosis in 73% of the cases and supported the light microscopic findings in 27% by ruling out additional features. [3] However, morphometric analysis of renal biopsies in the pediatric NS has been performed in a few studies. Morita et al evaluated GBM thickness in children with MCD, including those at initial presentation and those in remission. The authors showed a trend toward a thicker GBM in young males, while no significant difference was found between the proteinuric patients and those in remission. [4] Another study compared the GBM thickness among the patients with MCD and those with FSGS and found no significant difference between them. [5] In our study, the GBM was significantly thicker in the mesPGN/FSGS group compared to the MCD group (473.82 ± 77.62 vs. 352.77 ± 23.78).

Podocyte FPE, as a reaction of podocytes to injury, has been documented in the various pediatric NS. Studies evaluating the correlation between FPE and proteinuria in the patients with MCD have yielded conflicting results. Few studies have shown a weak but a significant correlation of FPE and proteinuria, while one study including nephrotic patients with MCD (not in remission) did not find a similar correlation. [8],[9],[10] In our study, the podocyte FPE did not correlate with the degree of proteinuria.

The slit diaphragm is an intercellular junction that forms a functional filter and establishes contact between two adjacent foot processes. Earlier ultrastructural studies have shown a reduction in the number of filtration slits in patients with MCD. [6],[7],[11] In our study, the slit pore diameter was measured, and it was significantly longer in the mesPGN/FSGS (P = 0.04). In addition, foot process width was more in the MCD compared with both controls (P = 0.006) and the mesPGN/FSGS group (P = 0.005). In a study by Bohman et al, the mean foot process width was shown to correlate with glomerular filtration rate. [12] In the present study, slit pore widening showed positive correlation with the degree of proteinuria.

In conclusion, this study highlights several ultrastructural alterations in the renal biopsies of the pediatric nephrotic syndrome. Significant differences were observed in the GBM thickness and foot process width, and slit pore widening, rather than FPE, correlated with the degree of proteinuria. These ultrastructural changes need to be evaluated in larger studies and in greater detail to highlight the difference between various groups of NS.

 
   References Top

1.McBryde KD, Kershaw DB, Smoyer WE. Pediatric steroid resistant nephrotic syndrome. Curr Prob Pediatr 2001;31:280-307.   Back to cited text no. 1
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2.Bonilla-Felix M, Parra C, Dajani T, et al. Changing patterns in the histopathology of idiopathic nephrotic syndrome in children. Kidney Int 1999;55:1885-90.   Back to cited text no. 2
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3.Rivera A, Magliato S, Meleg-Smith S. Value of electron microscopy in the diagnosis of childhood nephrotic syndrome. Ultrastruct Pathol 2001;25:313-20.   Back to cited text no. 3
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4.Morita M, White RH, Raafat F, Barnes JM, Standring DM. Glomerular basement membrane thickness in children: A morphometric study. Pediatr Nephrol 1988;2:190-5.   Back to cited text no. 4
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5.Shindo S, Yoshimoto M, Kuriya N, Bernstein J. Glomerular basement membrane thickness in recurrent and persistent hematuria and nephrotic syndrome: Correlation with sex and age. Pediatr Nephrol 1988;2:196-9.   Back to cited text no. 5
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6.Guasch A, Myers BD. Determinants of glomerular hypofiltration in nephrotic patients with minimal change nephropathy. J Am Soc Nephrol 1994;4:1571-81.   Back to cited text no. 6
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7.Patrakka J, Lahdenkari AT, Koskimies O, Holmberg C, Wartiovaara J, Jalanko H. The number of podocyte slit diaphragms is decreased in minimal change nephrotic syndrome. Pediatr Res 2002;52:349-55.   Back to cited text no. 7
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8.Powell HR. Relationship between proteinuria and epithelial cell changes in minimal lesion glomerulopathy. Nephron 1976;16:310-7.   Back to cited text no. 8
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9.Koop K, Eikmans M, Baelde HJ, et al. Expression of podocyte-associated molecules in acquired human kidney diseases. J Am Soc Nephrol 2003;14:2063-71.   Back to cited text no. 9
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10.van den Berg JG, van den Bergh Weerman MA, Assmann KJ, Weening JJ, Florquin S. Podocyte foot process effacement is not correlated with the level of proteinuria in human glomerulopathies. Kidney Int 2004;66:1901-6.   Back to cited text no. 10
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11.Lahdenkari AT, Lounatmaa K, Patrakka J, et al. Podocytes are firmly attached to glomerular basement membrane in kidneys with heavy proteinuria. J Am Soc Nephrol 2004;15:2611-8.  Back to cited text no. 11
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12.Bohman SO, Jaremko G, Bohlin AB, Berg U. Foot process fusion and glomerular filtration rate in minimal change nephrotic syndrome. Kidney Int 1984;25:696-700.  Back to cited text no. 12
[PUBMED]    

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Correspondence Address:
Amit K Dinda
Department of Pathology, All India Institute of Medical Sciences, Ansari Nagar, New Delhi
India
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DOI: 10.4103/1319-2442.109577

PMID: 23538351

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