Permeation of macromolecules into the renal glomerular basement membrane and capture by the tubules

How the kidney prevents urinary excretion of plasma proteins continues to be debated. Here, using unfixed whole-mount mouse kidneys, we show that fluorescent-tagged proteins and neutral dextrans permeate into the glomerular basement membrane (GBM), in general agreement with Ogston's 1958 equation describing how permeation into gels is related to molecular size. Electron-microscopic analyses of kidneys fixed seconds to hours after injecting gold-tagged albumin, negatively charged gold nanoparticles, and stable oligoclusters of gold nanoparticles show that permeation into the lamina densa of the GBM is size-sensitive. Nanoparticles comparable in size with IgG dimers do not permeate into it. IgG monomer-sized particles permeate to some extent. Albumin-sized particles permeate extensively into the lamina densa. Particles traversing the lamina densa tend to accumulate upstream of the podocyte glycocalyx that spans the slit, but none are observed upstream of the slit diaphragm. At low concentrations, ovalbumin-sized nanoparticles reach the primary filtrate, are captured by proximal tubule cells, and are endocytosed. At higher concentrations, tubular capture is saturated, and they reach the urine. In mouse models of Pierson’s or Alport’s proteinuric syndromes resulting from defects in GBM structural proteins (laminin β2 or collagen α3 IV), the GBM is irregularly swollen, the lamina densa is absent, and permeation is increased. Our observations indicate that size-dependent permeation into the lamina densa of the GBM and the podocyte glycocalyx, together with saturable tubular capture, determines which macromolecules reach the urine without the need to invoke direct size selection by the slit diaphragm.

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Electron microscopic radioautographic studies on the incorporation of 3H proline in the glomerular basement membrane (GBM) in antistreptococcal-cell-membrane (SCM) injected mice

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100111057 ◽

1984 ◽

Vol 42 ◽

pp. 188-189

Author(s):

R.P. Nayyar ◽

C.F. Lange ◽

J. L. Borke

Keyword(s):

Body Weight ◽

Cell Membrane ◽

Basement Membrane ◽

Glomerular Basement Membrane ◽

Mesangial Matrix ◽

Electron Microscopic ◽

Group A ◽

Injection Protocol

Streptococcal cell membrane (SCM) antiserum injected mice show a significant thickening of glomerular basement membrane (GBM) and an increase in mesangial matrix within 4 to 24 hours of antiserum administration (1,2,3). This study was undertaken to evaluate the incorporation of 3H proline into glomerular cells and GBM under normal and anti-SCM induced conditions. Mice were administered, intraperitoneally, 0.1 ml of normal or anti-SCM serum followed by a 10 µC/g body weight injection of 3H proline. Details of the preparation of anti-SCM (Group A type 12 streptococcal pyogenes) and other sera and injection protocol have been described elsewhere (2). After 15 minutes of isotope injection a chase of cold proline was given and animal sacrificed at 20 minutes, 1,2,4,8,24 and 48 hours. One of the removed kidneys was processed for immunofluorescence, light and electron microscopic radioautographic studies; second kidney was used for GBM isolation and aminoacid analysis.

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Localization and quantitation of anionic charge sites in fetal and neonatal alveolar basement membranes.

Journal of Histochemistry & Cytochemistry ◽

10.1177/33.6.3158699 ◽

1985 ◽

Vol 33 (6) ◽

pp. 611-614 ◽

Cited By ~ 7

Author(s):

T B Ferrara ◽

S P Sisson ◽

R L Vernier

Keyword(s):

Basement Membrane ◽

Microscopic Analysis ◽

Basement Membranes ◽

Lamina Densa ◽

Human Lung Tissue ◽

Charge Site ◽

Electron Microscopic ◽

Anionic Charge ◽

Site Localization ◽

Standard Techniques

A method utilizing biopsy sized samples of lung for anionic charge site localization in alveolar and capillary basement membranes in human tissue is discussed. Tissue fixed in either paraformaldehyde-lysine-periodate or 1% paraformaldehyde with 0.05% glutaraldehyde, cut into 30 mu sections, and incubated with the cationic probe, polyethyleneimine, was processed for electron microscopic analysis using standard techniques. Anionic charge sites were identified and regularly distributed in increments of approximately 40-50 nm in the lamina rara externa of the alveolar basement membrane, with lesser amounts found in the lamina rara interna and lamina densa. Anionic charge sites were also demonstrated in the interstitial portion of the capillary basement membrane and on cell surfaces. These methods can be used to more broadly define the localization of anionic charge sites in human lung tissue in both normal and pathologic states.

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Inactive matrix metalloproteinase 2 is a normal constituent of human glomerular basement membrane. An immuno-electron microscopic study

The Journal of Pathology ◽

10.1002/(sici)1096-9896(200005)191:1<61::aid-path565>3.0.co;2-6 ◽

2000 ◽

Vol 191 (1) ◽

pp. 61-66 ◽

Cited By ~ 4

Author(s):

Sawsan M. Jalalah ◽

Peter N. Furness ◽

Gordon Barker ◽

Mark Thomas ◽

Leon L. Hall ◽

...

Keyword(s):

Basement Membrane ◽

Matrix Metalloproteinase ◽

Microscopic Study ◽

Electron Microscopic Study ◽

Glomerular Basement Membrane ◽

Matrix Metalloproteinase 2 ◽

Electron Microscopic ◽

Normal Constituent

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Electron microscopic studies of glomerular basement membrane (GBM) challenged with antisera to GBM or streptococcal cell membrane (SCM)

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100076251 ◽

1983 ◽

Vol 41 ◽

pp. 516-517

Author(s):

Nayyar R. P. ◽

C. F. Lange ◽

F. I. Volini

Keyword(s):

Body Weight ◽

Cell Membrane ◽

Basement Membrane ◽

Glomerular Basement Membrane ◽

Binding Capacity ◽

Normal Serum ◽

Electron Microscopic ◽

Microscopic Studies

The main objective of this study is to evaluate the synthesis and turnover of the glomerular basement membrane under normal and induced pathologial conditions. Mice were challenged with rabbit antisera by intraperitoneal (IP) or intravenous (IV) injection of 0.1 ml of normal serum, anti-glomerular basement membrane (GBM) or anti-streptococcal cell membrane (SCM) sera followed by a 10 μC/g body weight of H-L-Proline. A cold chase of L-proline was also administered 15 minutes later. Some of the animals also received additional chases of normal serum and anti-sera 2 and 4 hours after the initial injections. The animals were sacrificed at 2, 4, 6, 8, 18, 24, 40 and 48 hours, kidneys removed and processed for light, immunofluorescent, electronmicroscopic and radioautographic studies.Immunofluorescent and electronmicroscopic studies did not reveal any significant differences in the binding capacity of anti-sera based on the route of challenge (IP or IV).

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The Degradation of Human Glomerular Basement Membrane with Purified Lysosomal Proteinases: Evidence for the Pathogenic Role of the Polymorphonuclear Leucocyte in Glomerulonephritis

Clinical Science ◽

10.1042/cs0540233 ◽

1978 ◽

Vol 54 (3) ◽

pp. 233-240 ◽

Cited By ~ 8

Author(s):

M. Davies ◽

A. J. Barrett ◽

J. Travis ◽

E. Sanders ◽

G. A. Coles

Keyword(s):

Basement Membrane ◽

Glomerular Basement Membrane ◽

Sodium Dodecyl ◽

Molecular Size ◽

Polymorphonuclear Leucocyte ◽

Cathepsin G ◽

Ph Values ◽

Leucocyte Elastase ◽

Lysosomal Proteinases

1. Human polymorphonuclear leucocyte elastase and cathepsin G were incubated with preparations of isolated human glomerular basement membrane at neutral pH and 37°C. 2. The ability of these enzymes to degrade glomerular basement membrane was followed by the release of hydroxyproline. Both proteinases released considerable amounts of hydroxyproline. 3. By using Sephadex G-100 it was shown that the solubilized basement membrane fragments appeared as a single peak and had a molecular weight of over 100 000. These proteins after reduction were analysed by sodium dodecyl sulphate-gel electrophoresis to examine their subunit pattern and determine their molecular size. 4. The released basement membrane proteins gave at least four precipitin lines with a rabbit anti-(glomerular basement membrane) antiserum. 5. These results support the concept that polymorphonuclear leucocyte neutral proteinases play an important role in the pathogenesis of glomerulonephritis. 6. At acid pH values cathepsin B also released hydroxyproline from human glomerular basement membrane but the lysosomal carboxyl proteinase, cathepsin D, had no action.

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