Size and charge selectivity of the glomerular filter in early experimental diabetes in rats

Microalbuminuria is an early sign of diabetic nephropathy. The aim of the present study was to investigate whether the changes of the glomerular filtration barrier in early experimental diabetes are due to size- or charge-selective alterations. Wistar rats, made diabetic by streptozotocin (STZ) and having their blood glucose maintained at ∼20 mM for 3 or 9 wk, were compared with age-matched controls. Glomerular clearances of native albumin (Cl-HSA) and neutralized albumin (Cl-nHSA) were assessed using a renal uptake technique. Glomerular filtration rate and renal plasma flow were assessed using 51Cr-EDTA and [125I]iodohippurate, respectively. In a separate set of animals, diabetic for 9 wk, and in controls, glomerular sieving coefficients (θ) for neutral FITC-Ficoll (molecular radius: 15–90 Å) were assessed using size exclusion chromatography. At 3 wk of diabetes, Cl-HSA and Cl-nHSA remained unchanged, indicating no alteration in either size or charge selectivity. By contrast, at 9 wk of diabetes, there was a twofold increase of Cl-HSA, whereas Cl-nHSA remained largely unchanged, at first suggesting a glomerular charge defect. However, according to a two-pore model, the number of large pores, assessed from both Ficoll and Cl-HSA, increased twofold. In addition, a small reduction in proximal tubular reabsorption was observed at 3 wk, which was further reduced at 9 wk. In conclusion, no functional changes were observed in the glomerular filtration barrier at 3 wk of STZ-induced diabetes, whereas at 9 wk there was a decrease in size selectivity due to an increased number of large glomerular pores.

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Dynamic, size-selective effects of protamine sulfate and hyaluronidase on the rat glomerular filtration barrier in vivo

AJP Renal Physiology ◽

10.1152/ajprenal.00181.2014 ◽

2014 ◽

Vol 307 (10) ◽

pp. F1136-F1143 ◽

Cited By ~ 4

Author(s):

Kristinn Sverrisson ◽

Josefin Axelsson ◽

Anna Rippe ◽

Daniel Asgeirsson ◽

Bengt Rippe

Keyword(s):

Glomerular Filtration ◽

High Performance ◽

Protamine Sulfate ◽

Size Exclusion ◽

Endothelial Glycocalyx ◽

Glomerular Filtration Barrier ◽

Filtration Barrier ◽

Charge Selectivity ◽

The Impact

The proteinuric actions of protamine sulfate (PS) have classically been, at least partly, attributed to alterations of the negatively charged glomerular endothelial glycocalyx. To investigate whether the charge-selective properties of the glomerular filtration barrier (GFB) would be altered by PS, we assessed the glomerular sieving of conventional, uncharged, polydispersed Ficoll (n-Ficoll) compared with charge modified, conformationally intact, anionic (carboxymethylated) Ficoll (a-Ficoll) before and after systemic infusions of PS in rats. For comparison, we also investigated the impact of hyaluronidase (hyase), which partially degrades the glycocalyx, on GFB permeability. In anaesthetized Wistar rats, blood access was achieved, and the left ureter was cannulated for urine collection. Rats were infused with either n-Ficoll or a-Ficoll before and during systemic infusions with either PS or hyase. Plasma and urine samples were taken repeatedly and analyzed by high-performance size exclusion chromatography to assess glomerular sieving coefficients (θ) for Ficoll (radius 10–80 Å). The GFB showed a significant glomerular charge selectivity for Ficoll molecules of radius 20–35 Å. PS and hyase infusions reversibly increased θ for large Ficoll molecules (Ficoll molecules of radius 50–80 Å). Thus, for PS, θ for a-Ficoll molecules of radius 70 Å increased from 2.47 × 10−5± 1.1−5to 7.25 × 10−5± 1.1−5( P < 0.05) at 15 min. For hyase, changes in a-Ficoll molecules of radius 50–80 Å were, however, not statistically significant. Neither PS nor hyase had any effect on θ for n-Ficoll molecules of radius 20–45 Å or a-Ficoll molecules of radius 20–45 Å. It is concluded that systemically administered PS and hyase in moderate doses dynamically decreased the size selectivity of the rat GFB without affecting its charge selective properties.

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Loss of size selectivity of the glomerular filtration barrier in rats following laparotomy and muscle trauma

AJP Renal Physiology ◽

10.1152/ajprenal.00246.2009 ◽

2009 ◽

Vol 297 (3) ◽

pp. F577-F582 ◽

Cited By ~ 19

Author(s):

Josefin Axelsson ◽

Irma Mahmutovic ◽

Anna Rippe ◽

Bengt Rippe

Keyword(s):

Glomerular Filtration ◽

High Performance ◽

Size Exclusion ◽

Blunt Injury ◽

Abdominal Muscles ◽

Glomerular Filtration Barrier ◽

Glomerular Permeability ◽

Filtration Barrier ◽

Muscle Trauma ◽

Exclusion Chromatography

Posttraumatic microalbuminuria may be caused by either charge- or size-selective alterations in the glomerular filtration barrier, or both, and/or to a reduction in proximal tubular protein reabsorption. This study was performed to elucidate the pathophysiology of the increases in glomerular permeability occurring in rats exposed to a laparotomy or to a laparotomy and muscle trauma. In anesthetized Wistar rats (250–280 g), the left ureter was cannulated for urine collection, while simultaneously blood access was achieved. Rats were exposed to trauma by a laparotomy (L; n = 8), or by a combination of L and muscle trauma (MT; L+MT) induced by topical blunt injury of the abdominal muscles bilaterally. After L, muscles were crushed using hemostatic forceps at either 2 × 2 sites (“small” MT; n = 9), or at 2 × 5 sites (“large” MT; n = 9). Sham groups ( n = 16), not exposed to a laparotomy, were used as controls. The glomerular sieving coefficients (θ) to polydisperse FITC-Ficoll-70/400 (molecular radius 13–80 Å) were determined at 5 or 60 min after L and L+MT, respectively, from plasma and urine samples, and analyzed by high-performance size-exclusion chromatography. A tissue-uptake technique was used to assess θ for 125I-labeled serum albumin. L, with or without MT, increased θ for Ficoll55–80Å and albumin rapidly and markedly. θ-Ficoll70Å thus increased approximately threefold, and θ for albumin significantly, for all trauma groups. According to the “two-pore model” of glomerular permeability, these changes mainly reflect an increase in the number of large pores in the glomerular filter without any primary changes in the charge-selective properties of the filter.

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The Rat Glomerular Filtration Barrier Does Not Show Negative Charge Selectivity

Microcirculation ◽

10.1038/sj.mn.7800150 ◽

2002 ◽

Vol 9 (5) ◽

pp. 329-342 ◽

Cited By ~ 38

Author(s):

RICHARD C. SCHAEFFER ◽

MAX L. GRATRIX ◽

DAVID R. MUCHA ◽

JOSÉ M. CARBAJAL

Keyword(s):

Glomerular Filtration ◽

Negative Charge ◽

Glomerular Filtration Barrier ◽

Filtration Barrier ◽

Charge Selectivity

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A new function for parietal epithelial cells: a second glomerular barrier

AJP Renal Physiology ◽

10.1152/ajprenal.00214.2009 ◽

2009 ◽

Vol 297 (6) ◽

pp. F1566-F1574 ◽

Cited By ~ 47

Author(s):

Takamoto Ohse ◽

Alice M. Chang ◽

Jeffrey W. Pippin ◽

George Jarad ◽

Kelly L. Hudkins ◽

...

Keyword(s):

Epithelial Cells ◽

Basement Membrane ◽

Glomerular Filtration ◽

Glomerular Filtration Barrier ◽

Impermeable Barrier ◽

Filtration Barrier ◽

Proximal Tubular ◽

Parietal Epithelial Cells ◽

Glomerular Barrier

The functional role of glomerular parietal epithelial cells (PECs) remains poorly understood. To test the hypothesis that PECs form an impermeable barrier to filtered protein through the formation of tight junctions (TJ), studies were performed in normal animals and in the anti-glomerular basement membrane (GBM) model of crescentic nephritis. Electron microscopy showed well-defined TJ between PECs in normal mice, which no longer could be identified when these cells became extensively damaged or detached from their underlying Bowman's basement membrane. The TJ proteins claudin-1, zonula occludens-1, and occludin stained positive in PECs; however, staining decreased in anti-GBM disease. To show that these events were associated with increased permeability across the PEC-Bowman's basement membrane barrier, control and diseased animals were injected intravenously with either Texas red-conjugated dextran (3 kDa) or ovalbumin (45 kDa) tracers. As expected, both tracers were readily filtered across the glomerular filtration barrier and taken up by proximal tubular cells. However, when the glomerular filtration barrier was injured in anti-GBM disease, tracers were taken up by podocytes and PECs. Moreover, tracers were also detected between PECs and the underlying Bowman's basement membrane, and in many instances were detected in the extraglomerular space. We propose that together with its underlying Bowman's basement membrane, the TJ of PECs serve as a second barrier to protein. When disturbed following PEC injury, the increase in permeability of this layer to filtered protein is a mechanism underlying periglomerular inflammation characteristic of anti-GBM disease.

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Unaltered size selectivity of the glomerular filtration barrier in caveolin-1 knockout mice

AJP Renal Physiology ◽

10.1152/ajprenal.00075.2009 ◽

2009 ◽

Vol 297 (2) ◽

pp. F257-F262 ◽

Cited By ~ 9

Author(s):

Gustaf Grände ◽

Catarina Rippe ◽

Anna Rippe ◽

Awahan Rahman ◽

Karl Swärd ◽

...

Keyword(s):

Glomerular Filtration ◽

High Performance ◽

Microvascular Permeability ◽

Size Exclusion ◽

No Production ◽

Glomerular Permeability ◽

Caveolin 1 ◽

Filtration Barrier ◽

Transcapillary Escape Rate ◽

Exclusion Chromatography

The transfer of albumin from blood to tissue has been found to be increased in caveolin-1 knockout (KO) mice. This has been considered to reflect increased microvascular permeability, conceivably caused by an increased endothelial production of nitric oxide (NO) in these mice. To investigate whether such an increase in NO production would also affect glomerular barrier characteristics, the glomerular sieving coefficients (θ) to neutral FITC-Ficoll 70/400 (molecular radius 13–90 Å) were determined in caveolin-1 KO mice vs. their wild-type counterparts. The θ for Ficoll were assessed using high-performance size-exclusion chromatography on blood and urine samples. Furthermore, the transcapillary escape rate (TER) of 125I-labeled albumin and plasma volume (PV) were determined in both types of mice. The kidney expressed low levels of caveolin-1 compared with the lung and bladder, but immunofluorescence associated with vascular structures was evident. Staining was lost in the caveolin-1 KO kidney, as was caveolin-1 expression in the lung and bladder. Despite an increase in the glomerular filtration rate in caveolin-1 KO mice (0.23 ± 0.04 vs. 0.10 ± 0.02 ml/min; both n = 7; P < 0.05), the glomerular Ficoll sieving curves were nearly identical. Furthermore, caveolin-1 KO mice showed an increased PV (6.59 ± 0.42 vs. 5.18 ± 0.13 ml/100 g; P < 0.01) but only a tendency toward an increased TER (14.69 ± 1.59 vs. 11.62 ± 1.62%/h; not significant). It is concluded that in caveolin-1 KO mice the glomerular permeability was not increased, despite the presence of glomerular hyperfiltration. The present data are in line with the concept that the increased transvascular albumin leakage previously found in mice lacking caveolin-1 may be due to an elevation in systemic microvascular pressure due to precapillary vasodilatation, rather than being a consequence of increased microvascular permeability per se.

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Acute reactive oxygen species (ROS)-dependent effects of IL-1β, TNF-α, and IL-6 on the glomerular filtration barrier (GFB) in vivo

AJP Renal Physiology ◽

10.1152/ajprenal.00111.2015 ◽

2015 ◽

Vol 309 (9) ◽

pp. F800-F806 ◽

Cited By ~ 19

Author(s):

Kristinn Sverrisson ◽

Josefin Axelsson ◽

Anna Rippe ◽

Daniel Asgeirsson ◽

Bengt Rippe

Keyword(s):

Reactive Oxygen Species ◽

Glomerular Filtration ◽

Reactive Oxygen ◽

Size Exclusion ◽

Oxygen Species ◽

Glomerular Filtration Barrier ◽

Glomerular Permeability ◽

Filtration Barrier ◽

Tnf Α ◽

Superoxide Scavenger

This study was performed to investigate the immediate actions of the proinflammatory cytokines IL-1β, TNF-α, and IL-6 on the permeability of the glomerular filtration barrier (GFB) in rats and to test whether these actions are dependent upon the release of reactive oxygen species (ROS). In anesthetized rats, blood access was achieved and the left ureter was cannulated for urine collection. Rats were continuously infused intravenously with either IL-1β (0.4 and 2 μg·kg−1·h−1), TNF-α (0.4 and 2 μg·kg−1·h−1), or IL-6 (4 and 8 μg·kg−1·h−1), together with polydisperse FITC-Ficoll-70/400 and inulin for 1 h. Plasma and urine samples were analyzed by high performance size exclusion chromatography (HPSEC) for determination of glomerular sieving coefficients (θ). The glomerular filtration rate (GFR) was also assessed (51Cr-EDTA). In separate experiments, the superoxide scavenger tempol (30 mg·kg−1·h−1) was given before and during cytokine infusions. IL-1β and TNF-α caused rapid, partly reversible increases in glomerular permeability to large molecules (Ficoll50–80Å), peaking at 5–30 min, while IL-6 caused a more gradual increase in permeability, leveling off at 60 min. Tempol almost completely abrogated the glomerular permeability effects of the cytokines infused. In conclusion IL-1β, TNF-α, and IL-6, when infused systemically, caused immediate and partly reversible increases in glomerular permeability, which could be inhibited by the superoxide scavenger tempol, suggesting an important role of ROS in acute cytokine-induced permeability changes in the GFB.

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Reduced diffusion of charge-modified, conformationally intact anionic Ficoll relative to neutral Ficoll across the rat glomerular filtration barrier in vivo

AJP Renal Physiology ◽

10.1152/ajprenal.00183.2011 ◽

2011 ◽

Vol 301 (4) ◽

pp. F708-F712 ◽

Cited By ~ 17

Author(s):

Josefin Axelsson ◽

Kristinn Sverrisson ◽

Anna Rippe ◽

William Fissell ◽

Bengt Rippe

Keyword(s):

Glomerular Filtration ◽

High Performance ◽

Size Exclusion ◽

Glomerular Filtration Barrier ◽

Glomerular Permeability ◽

Charge Effects ◽

Filtration Barrier ◽

A Charge

The glomerular filtration barrier (GFB) is commonly conceived as a negatively charged sieve to proteins. Recent studies, however, indicate that glomerular charge effects are small for anionic, carboxymethylated (CM) dextran vs. neutral dextran. Furthermore, two studies assessing the glomerular sieving coefficients (θ) for negative CM-Ficoll vs. native Ficoll have demonstrated an increased glomerular permeability for CM-Ficoll (Asgeirsson D, Venturoli D, Rippe B, Rippe C. Am J Physiol Renal Physiol 291: F1083–F1089, 2006; Guimarães M, Nikolovski J, Pratt L, Greive K, Comper W. Am Physiol Renal Physiol 285: F1118–F1124, 2003.). The CM-Ficoll used, however, showed a larger Stokes-Einstein radius ( ae) than neutral Ficoll, and it was proposed that the introduction of negative charges in the Ficoll molecule had made it more flexible and permeable. Recently, a negative FITC-labeled CM-Ficoll (CMI-Ficoll) was produced with a conformation identical to that of neutral FITC-Ficoll. Using these probes, we determined their θ:s in anesthetized Wistar rats (259 ± 2.5 g). After blood access had been achieved, the left ureter was cannulated for urine sampling. Either polysaccharide was infused (iv) together with a filtration marker, and urine and plasma were collected. Assessment of θ FITC-Ficoll was achieved by high-performance size-exclusion chromatography (HPSEC). CMI-Ficoll and native Ficoll had identical elugrams on the HPSEC. Diffusion of anionic Ficoll was significantly reduced compared with that of neutral Ficoll across the GFB for molecules of ae ∼20–35 Å, while there were no charge effects for Ficoll of ae = 35–80 Å. The data are consistent with a charge effect present in “small pores,” but not in “large pores,” of the GFB and mimicked those obtained for anionic membranes in vitro for the same probes.

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