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W D Comper

Publications and source records attributed to W D Comper.

At least 37 records · Page 2Linked to original sources
PubMed1997

Protein degradation during renal passage in normal kidneys is inhibited in experimental albuminuria.

1. Tritium labelled proteins, namely bovine serum albumin ([3H]BSA), rat serum albumin ([3H]RSA), anionic horseradish peroxidase ([3H]aHRP) and immunoglobulin present in urine fractions from rat filtration studies in vivo and isolated perfused rat kidneys (IPKs) have been shown by gel chromatographic analysis to be severely degraded to small peptides. The degradation of RSA and BSA in vivo has been shown to be similar. 2. Degradation of proteins in the urine from IPK experiments was inhibited by including 150 mmol/l lysine in the perfusate. Similarly, [3H]BSA and [3H]aHRP excreted from rats with puromycin aminonucleoside nephrosis was again essentially intact for both IPK and in vivo experiments. 3. It appears that the degradation of proteins observed in urine obtained from control kidneys is due, in part, to proteolytic activity associated with the proximal tubule. Inhibition of proximal tubule function, which occurs for both lysine and puromycin aminonucleoside treatments (as calibrated by lysozyme uptake), results in inhibition of the degradation observed. Glomerular epithelial cells could also contribute to the degradation. 4. There was no generation of low-molecular-weight material in the perfusate or plasma arising from breakdown of circulating proteins or recycling of potential degradation products from the tubules.

Albuminuria↗
PubMed1997

Fractional clearance of albumin is influenced by its degradation during renal passage.

1. The fractional clearance of intact albumin as determined by fractionation of urine by gel chromatography gave a value of 3.9 +/- 1.6 x 10(-4) for the isolated perfused kidney and 2.1 +/- 0.6 x 10(-4) in vivo using ALZET osmotic pumps. 2. Albumin fractional clearance as measured by detection of the tritium label on the albumin molecule by radioactivity analysis gave a value of 7.5 +/- 3.9 x 10(-3) for the isolated perfused kidney and 2.3 +/- 0.9 x 10(-3) in vivo. 3. The major differences between assays that detect intact albumin compared with non-specific assays in the estimates of the fractional clearance of albumin can be explained by the degradation of approx. 90% of albumin to small peptides during its renal passage. This has been demonstrated by size exclusion chromatography of urine samples from experiments where (i) exogenous tritium-labelled albumin was used in isolated perfused kidneys, (ii) exogenous tritium-labelled albumin was administered intravenously and (iii) analysis was made with metabolically labelled endogenous albumin in vivo.

Albumins↗
PubMed1997

Why is proteinuria such an important risk factor for progression in clinical trials?

There are strong reasons to justify the concept that proteinuria is a major risk factor for progression in clinical trials. The evidence is strongest where therapeutic intervention has been focused on established renal disease, when changes in albumin excretion rate (AER) and glomerular filtration rate (GFR) occur within a short time span. Proteinuria is also important in emerging renal disease, such as incipient diabetic nephropathy (DN), since natural history studies show that small increases in AER predict clinical nephropathy and, ultimately, a decline in GFR. However, the absence of concurrent changes in GFR in incipient DN complicates the evaluation of clinical trials in this condition. It is also not certain that the degree of coupling of changes in AER and GFR is the same during intervention as during natural history studies. The importance of proteinuria as a risk factor for progression has been strengthened by recent evidence showing that proteinuria itself causes renal damage. Traditional concepts of the damaging effects of proteinuria have focused on the glomeruli, where mesangial expansion induced by transcapillary passage of proteins has been considered to lead to a decrease in glomerular filtration surface and a decline in GFR. New evidence suggests that interaction of albumin with proximal renal tubules may not only lead to renal damage but may also be causally related to increases in AER.

Albuminuria↗
PubMed1996

A codiffusing system as a novel model for capillary wall charge selectivity.

The electrostatic interaction associated with polyion-polyion interaction has been thought to be the basis for the differential transport of charged transport probes across the capillary wall. The charge of the transport probe may interact with the negative charges of capillary wall cell surfaces and the intercellular glycocalyx or extracellular matrix. This study has set out to quantitate the exact nature of the polyion-polyion interaction through the theoretical and experimental study of the partition-diffusion of albumin in solutions of anionic polysaccharides (dextran sulfate or heparin) at concentrations up to 250 meq 1(-1) or at an average intercharge distance of 2.2 nm, as compared with solutions of uncharged dextran at the same volume concentration. The results demonstrate that the albumin partition-diffusion is exactly the same in dextran sulfate, heparin and uncharged dextran matrices of the same polymer volume fraction. These results confirm previous studies from this laboratory that the charge effect through polyion-polyion interaction under physiological conditions is negligible.

Albumins↗
PubMed1996

Glomerular processing of dextran sulfate during transcapillary transport.

The fractional clearance of dextran sulfate across the glomerular capillary wall in the isolated perfused kidney is shown to be dependent on the concentration of dextran sulfate in the perfusate and the degree of sulfate substitution on dextran sulfate. While normal charge selectivity is apparent at low dextran sulfate perfusate concentrations, the nature of the concentration dependence of the fractional clearance clearly eliminates charge-electrostatic interactions as being responsible. A strong correlation has been experimentally established between fractional clearance and the degree of desulfation of dextran sulfate found in the urine. The correlation was further established through the demonstration that the lysosomotropic agent NH4Cl partially inhibits charge selectivity as well as desulfation. The glomerular cellular processing of the dextran sulfate was further studied through dextran sulfate-mediated release of metabolically labeled glomerular heparan sulfate. A model of glomerular capillary wall transport, invoking endothelial cell processing of dextran sulfate, is proposed to explain the differences between the fractional clearance of dextran and dextran sulfate.

Ammonium Chloride↗
PubMed1995

Uptake of dextran sulphate by glomerular intracellular vesicles during kidney ultrafiltration.

Dextran sulphate is often used as a model for albumin in understanding capillary-tissue exchange and the charge selective nature of the capillary wall, particularly in kidney ultrafiltration. In investigating the mechanism of transport in the kidney, autoradiographic analysis of the distribution of iodinated dextran sulphate in perfused rat kidneys demonstrates the preferential accumulation of the probe in the glomerular capillary wall. Tritium-labeled dextran sulphate is found to be specifically taken up by intracellular 20 to 40 nm vesicles that can be isolated post-perfusion. The molecular weight profile of vesicular dextran sulphate demonstrates that the dextran sulphate containing vesicles are from vascular glomerular cells. Dextran is not taken up by the vesicles. These results suggest that the apparent charge selectivity associated with the transglomerular transport of the dextran sulphate is associated, in part, with cell-mediated processes at the glomerular level.

Animals↗
PubMed1995

Albumin interaction with the glomerular capillary wall in vitro.

The binding of albumin to the glomerular capillary wall was studied using albumin-gold in perfused kidneys, the interaction of [3H]albumin with isolated glomeruli at 37 degrees C and 4 degrees C and the interaction at [3H]albumin with purified basement membrane. The albumin-gold was found to bind predominantly to the basement membrane and this interaction could be dissociated with high concentrations of albumin. There was binding of albumin to isolated rat glomeruli which exhibited temperature dependence. Glomeruli exhibited a binding site at both 37 degrees C and 4 degrees C with an association constant in the range of 1 to 3 x 10(4) M-1 that bound 7 x 10(13) molecules/glomerulus. At 37 degrees C, however, there was anomalous Scatchard binding behaviour at relatively higher concentrations of albumin (30 to 50 mg/ml) which could be due to either glomerular cell uptake or the appearance of multiple binding sites or both. The binding of albumin to isolated glomeruli and the glomerular albumin levels in isolated kidney perfusion could largely be accounted for by the binding of albumin to the glomerular basement membrane. The albumin binding to glomeruli at 37 degrees C was enhanced by Pronase digestion and heparinase digestion, but remained unchanged following trypsin treatment or neuraminidase treatment. Similarly, albumin was shown to bind to purified basement membrane preparations. This binding was also enhanced (approximately 80 times) by heparinase digestion but remained unchanged after digestion with chondroitinase ABC or hyaluronidase.(ABSTRACT TRUNCATED AT 250 WORDS)

Albumins↗
PubMed1995

Glomerular charge selectivity for anionic and neutral horseradish peroxidase.

Studies in isolated perfused rat kidney have demonstrated that it exhibits apparently normal charge selectivity and tubular uptake of anionic horseradish peroxidase (aHRP; pI < 4.0) and neutral horseradish peroxidase (nHRP; pI = 7.5) when these proteins are measured for their enzyme activity. The absolute fractional clearance values for aHRP and nHRP were 0.006 +/- 0.002 and 0.041 +/- 0.007, respectively. It is evident, however, that the enzyme assay for horseradish peroxidase severely underestimates the quantity of protein in urine as compared to measurement of its tritium labeled form through radioactivity. Fractional clearances estimated by radioactivity and corrected for tubular reabsorption for [3H]aHRP and [3H]nHRP were 0.040 +/- 0.029 and 0.099 +/- 0.043, respectively, compared to those estimated by enzyme activity which were 0.012 +/- 0.004 and 0.070 +/- 0.037, respectively. While charge selectivity between the anionic and neutral forms of HRP was still evident, albeit significantly reduced, the major feature of this type of analysis is that the clearance of the aHRP protein is significantly increased compared to that determined by enzyme assay. This difference correlates with the observation that the aHRP protein is markedly degraded (61 to 65%), as determined by gel chromatography, during filtration. Similar degradation was seen in urine fractions collected after the aHRP protein was administered in vivo. Degradation also occurred for the nHRP protein in both the perfused kidney and in vivo but to a far lesser extent (approximately 14 to 21%). These studies demonstrate that the anionic form of HRP was preferentially degraded during filtration and that charge selectivity for differently charged proteins is not as marked as originally thought.

Animals↗
PubMed1994

Dextran sulfate binding to isolated rat glomeruli and glomerular basement membrane.

The binding of dextran sulfate to isolated glomeruli and glomerular basement membrane has been studied and compared to the glomerular uptake of dextran sulfate during isolated kidney perfusion. Two binding sites for [3H]dextran sulfate to isolated glomeruli could be identified at 37 degrees C; an high affinity site (KA = 4.76 x 10(6) M-1) (4.4 x 10(10) molecules/glomerulus) and a low affinity site (KA = 5.8 x 10(4) M-1) (2.0 x 10(11) molecules/glomerulus) whereas at 4 degrees C there was only an high affinity binding site (KA = 1.43 x 10(6) M-1) (7 x 10(10) molecules/glomerulus). The glomerular binding of dextran sulfate appears to be to cellular elements as the binding sites were not present in purified glomerular basement membrane which effectively did not bind dextran sulfate. The binding of dextran sulfate to isolated glomeruli was far in excess of that found associated with glomerular uptake during isolated kidney perfusion. The difference has been discussed in terms of the different capillary wall surfaces exposed to the dextran sulfate and the concentration gradients that may exist in the two types of experiments. The structural integrity of the glomerulus is also important in governing the amount of binding as structural disintegration through freeze-thawing and sonication revealed more binding sites. These data may suggest a specific distribution of binding sites and specific transglomerular transport pathways where many of these sites are not exposed. The nature of binding [3H]dextran sulfate to isolated glomeruli was different to isolated perfused kidney glomerular uptake when studied in terms of the kinetics of exchange with unlabelled dextran sulfate. This exchange data suggests a passive binding mechanism in isolated glomeruli whereas in isolated kidney perfusion the nature of the exchange would suggest a glomerular intracellular uptake (Tay, M., et al. (1991) Am. J. Physiol. 260, F549-F554).

Animals↗
PubMed1994

The thermodynamic and hydrodynamic properties of macromolecules that influence the hydrodynamics of porous systems.

The water flow across porous, semipermeable membranes associated with osmosis and filtration under a variety of conditions is analysed and compared to macromolecular diffusion across free-liquid boundaries, diffusion and sedimentation in the ultracentrifuge, and tracer diffusion of water. This study establishes that osmosis can be explained in terms of the irreversible thermodynamics of diffusion. For macromolecular osmotically active solutes in the semidilute concentration regime the water flows across semipermeable porous membranes are interpreted in terms of a rate-limiting solute-solvent exchange layer that exists on the solution side of the membrane adjacent to the membrane pore; both osmosis and filtration will be governed by these exchange layers. These exchange layers also yield unique properties of their constituent molecules in systems where there is osmotic equilibration between solutions of different solutes. This study also establishes the need to consider the internal osmotic pressure of membranes in the pressure balance associated with the flow across the membrane. The complex situation of partially permeable membranes is analysed for the simple case where there are no mechanical gradients and there is only one osmotically active solution that creates a rate-limiting exchange layer. This treatment predicts that the flow will be governed primarily by the osmotic pressure difference associated with the partitioning of the solute at the membrane-solution interface.

Animals↗
PubMed1994

Desulphation of dextran sulphate during kidney ultrafiltration.

The renal clearance of [3H]dextran sulphate by the isolated perfused rat kidney was associated with desulphation of the molecule, as demonstrated by ion-exchange and affinity chromatography of material resident in both glomeruli and urine samples. This process also occurred in vivo. The molecular size distribution of glomerular dextran sulphate in the perfused kidney was indistinguishable from that in the perfusate, and although urinary material was smaller it remained macromolecular. Sulphatase activity was not detected in urine or in the perfusate of perfused kidneys, but was detected in glomerular and non-glomerular cortex fractions isolated by a sieving procedure. The identification of significant biochemical changes to dextran sulphate demonstrates that it does not function as an inert transport probe, and supports the concept of cellular involvement in the process of renal charge selectivity.

Animals↗
PubMed1994

Transglomerular transport of DEAE dextran in the isolated perfused kidney.

OBJECTIVE: The renal fractional clearance of [3H]DEAE dextran has been widely used to substantiate the charge selective model for renal permselectively, although there has only been one reported study on this type of clearance. This study sets out to examine the fractional clearance and glomerular processing of DEAE dextran. METHODS: Fractional clearance studies were performed using isolated perfused rat kidneys. The glomerular processing of DEAE dextran was assessed by examining the kinetics of DEAE dextran uptake in glomeruli isolated post perfusion. RESULTS: The fractional clearance of DEAE dextran used in the concentration range of 15-150 micrograms/ml in the perfusate of the isolated perfused kidney did not produce the classical in vivo facilitated transport of DEAE dextran as compared to dextran as observed by Bohrer et al. The fractional clearance curve displays retarded clearance of low molecular weight (small radii) DEAE dextran, giving the appearance of a 'flat curve'. Similar results were obtained when an oxygen free radical scavenger cocktail was included in the perfusate. These results may be due to the fact that DEAE dextran binds to the glomeruli (at an order of magnitude greater than dextran sulfate). Perfused kidneys with [3H]DEAE dextran for 1 h followed by a five minute perfusion with unlabelled DEAE dextran revealed no significant change in the glomerular levels of [3H]DEAE dextran (unlike dextran sulfate). Perfusion of rat kidneys with 15 micrograms/ml DEAE dextran produced no changes in the electron microscopical morphology of the glomerulus and no changes in the fractional clearance of dextran. CONCLUSIONS: These results do not support the glomerular charge selectivity model that involves a non-binding electrostatic interaction of the charged dextran with the fixed anion charges of the glomerular capillary wall.

Animals↗
PubMed1993

Anionic charge concentration of rat kidney glomeruli and glomerular basement membrane.

Estimates of levels of glomerular and glomerular-basement-membrane anion charge should serve as useful quantitative markers for the integrity of the tissues in health and disease. We have developed a simple, rapid, technique to measure this charge through the use of ion exchange with radioisotopes 22Na+ and 36Cl- at low ionic strengths in phosphate buffer. When this technique is used, normal glomeruli isolated from rat have a measured net anion charge concentration of 17.4 +/- 3.7 p-equiv. per glomerulus (n = 20). Perfused rat kidneys that lose approximately half of their glomerular heparan [35S]sulphate content (owing to oxygen-radical damage) exhibited a lower anion charge, of 7.5 +/- 1.6 p-equiv. per glomerulus (n = 5). Glomerular basement membranes prepared from rat glomeruli by a sonication-centrifugation procedure in the presence of enzyme inhibitors had a charge concentration of 6.3 +/- 0.7 mu-equiv./g wet wt. of tissue (n = 4), whereas membranes prepared by sonication, centrifugation, DNAse and detergent treatment had a charge concentration of 7.1 +/- 1.6 mu-equiv./g wet wt. (n = 4). Isotope-dilution experiments with 3H2O on these detergent-prepared glomerular basement membranes demonstrated that they had a water content of approx. 93%, which would then give a net anion charge concentration of 7.6 +/- 1.7 m-equiv./l (n = 4). These values are in good agreement with those obtained by others using titration techniques [Bray and Robinson (1984) Kidney Int. 25, 527-533]. The relatively low magnitude of glomerular anion charge in normal kidneys is consistent with other recent findings that glomerular anion charge is too low to affect the glomerular transport of charged molecules in a direct, passive, biophysical manner through electrostatic interactions.

Animals↗
PubMed1993

Non-electrostatic factors govern the hydrodynamic properties of articular cartilage proteoglycan.

The hydrodynamic frictional resistance to water flow exerted by articular cartilage proteoglycan is shown to be similar to that of proteoglycan isolated from Swarm rat chondrosarcoma, and independent of the state of aggregation of the proteoglycan. Frictional resistance is dependent, however, on the chain segments of the constituent chondroitin-sulphate and keratan-sulphate chains of the proteoglycan. Frictional resistance offered by chondroitin sulphate was independent of pH over the range 3.2-8.7. This confirms previous studies, associated with varying ionic strength and chemical modification of ionic groups of chondroitin sulphate, which showed that the frictional resistance offered by this molecule is independent of electrostatic factors. Water-structure-breaking and hydrogen-bond-breaking solvents were also without major effects on the flow resistance offered by chondroitin sulphate. An overall secondary structure of chondroitin sulphate was not evident, as it showed no significant difference to dextran in terms of its temperature dependence of relative viscosity. Local regions of rigid secondary structure, as manifested through inter-residue hydrogen bonding between sugar residues, is likely to control flow resistance as periodate-oxidized chondroitin sulphate and periodate-oxidized and reduced preparations showed a significant decrease in their frictional resistance to water.

Animals↗
PubMed1993

Thermodynamic nonideality in macromolecular solutions: interpretation of virial coefficients.

Consideration is given to the interpretation of virial coefficients reflecting thermodynamic nonideality in incompressible solutions of a single macromolecular species for which there is no volume change on mixing. Expressions are presented for the concentration dependence of thermodynamic activity under conditions where either the chemical potential of solvent or the pressure is fixed, these two conditions being mutually exclusive. For the former situation, which applies to partition equilibrium procedures, the thermodynamic activity is most conveniently defined on the molar scale because the coefficients in polynomial expansions for the osmotic pressure and the activity coefficient in terms of molarity are then related to each other without the inclusion of partial molar volume terms. Under conditions of constant pressure a similar situation prevails provided that the osmotic pressure and the corresponding activity coefficient are expanded in powers of molality. In either case conversion of the virial expansions to the other concentration scale is possible, but requires the introduction of partial molar volume terms into the virial coefficients. The implications of these findings are discussed in relation to results obtained by osmometry, isopiestic measurements, equilibrium dialysis, gel chromatography, and sedimentation equilibrium.

Chromatography, Gel↗
PubMed1991

Charge selectivity in kidney ultrafiltration is associated with glomerular uptake of transport probes.

The isolated perfused kidney exhibits substantial charge selectivity, as in vivo, in relation to fractional clearance of [3H]dextran sulfate and [3H]dextran. When cycloheximide is present in perfusate, fractional clearance of dextran sulfate is increased and proteinuria becomes significant, but glomerular filtration rate remains essentially unchanged compared with control. The possible role of cells in affecting transglomerular transport was demonstrated when isolated glomeruli from control perfused kidneys showed a very significant resident concentration of [3H]dextran sulfate and [3H]albumin, whereas there was no corresponding accumulation of [3H]dextran or [3H]inulin. Glomerular concentration of dextran sulfate and albumin was significantly reduced by cycloheximide. Kinetics of uptake and release of glomerular dextran sulfate indicated that it had a half-life of glomerular residence of approximately 2-3 min and that this half-life was considerably extended in the presence of cycloheximide. The half-life for glomerular residence of albumin was in the range of 30-40 min. The conclusion from this work is that glomerular charge selectivity for dextran sulfate could be quantitatively rationalized on the basis of transient uptake and release by glomerular cells.

Animals↗
PubMed1990

Extracellular matrix interactions: sulfation of connective tissue polysaccharides creates macroion binding templates and conditions for dissipative structure formation.

Evidence is now accumulating that the post-polymer modification process of sulfation of connective tissue polysaccharides is primarily to provide an interactive macroion for enthalpic interactions rather than influence thermodynamic non-ideality which primarily affects water distribution in biological systems. Metabolic energy considerations also distinguish these physicochemical classifications. Thermodynamic non-ideality is embodied in the carboxyl group and polysaccharide chain which are energetically favoured in biosynthesis, whereas considerable energy input is required for sulfation. The sulfation process gives rise to macroions, with a wide variety of negative charge patterns, that may participate in heterotypic macromolecular interactions. This partial informational specificity is discussed in terms of evolutionary flexibility of the extracellular matrix as rationalized on the qualitative aspects of dissipative structure formation. The concept of multiple binding interactions of varying specificity associated with connective tissue polysaccharides raises the awareness of a more random, less highly ordered, extracellular matrix as compared to the tight machine-like organization generally found for processes in the cell. This is discussed in terms of the physiological adaptation and development of multicellular-tissue systems.

Animals↗