Venturoli, Daniele, and Bengt Rippe. Ficoll and dextran vs. globular proteins as probes for testing glomerular permselectivity: effects of molecular size, shape, charge, and deformability. Am J Physiol Renal Physiol 288: F605-F613, 2005; doi:10.1152/ajprenal.00171.2004.-Polydisperse mixtures of dextran or Ficoll have been frequently used as molecular probes for studies of glomerular permselectivity because they are largely inert and not processed (reabsorbed) by the proximal tubules. However, dextrans are linear, flexible molecules, which apparently are hyperpermeable across the glomerular barrier. By contrast, the Ficoll molecule is almost spherical. Still, there is ample evidence that Ficoll fractional clearances (sieving coefficients) across the glomerular capillary wall (GCW) are markedly higher than those for neutral globular proteins of an equivalent in vitro Stokes-Einstein (SE) radius. Physical data, obtained by "crowding" experiments or measurements of intrinsic viscosity, suggest that the Ficoll molecule exhibits a rather open, deformable structure and thus deviates from an ideally hard sphere. This is also indicated from the relationship between (log) in vitro SE radius and (log) molecular weight (MW). Whereas globular proteins seem to behave in a way similar to hydrated hard spheres, polydisperse dextran and Ficoll exhibit in vitro SE radii that are much larger than those for compact spherical molecules of equivalent MW. For dextran, this can be partially explained by a high-molecular-size asymmetry. However, for Ficoll the explanation may be that the Ficoll molecule is more flexible (deformable) than are globular proteins. An increased compressibility of Ficoll and an increased deformability and size asymmetry for dextran may be the explanation for the fact that the permeability of the GCW is significantly higher when assessed using polysaccharides such as Ficoll or dextran compared with that obtained using globular proteins as molecular size probes. We suggest that molecular deformability, besides molecular size, shape, and charge, plays a crucial role in determining the glomerular permeability to molecules of different species. capillary permeability; polysaccharides; macromolecules; reflection coefficient; transport POLYDISPERSE MIXTURES OF DEXTRAN, and more recently Ficoll, are frequently used as molecular probes in studies of glomerular permselectivity. After being filtered through the glomerular capillary wall (GCW), polysaccharides, unlike proteins, are left unreabsorbed by the proximal tubules. This implies that their sieving coefficients (), i.e., their filtrate-to-plasma concentration ratios, can be determined directly from their urinary clearances relative to that of a glomerular filtration rate marker (e.g., inulin). Infusing polydisperse mixtures of dextran or Ficoll and using chromatographic techniques to fractionate plasma and urine samples make it possible to simultaneously determine the for a wide spectrum of different-sized molecular probes, provided that proper size calibrations ...

filtration rate dependence of sieving of albumin and some neutral proteins in rat kidneys.

In the present review, we summarize the principles governing the transport of fluid and electrolytes across the peritoneum during continuous ambulatory peritoneal dialysis (CAPD) in “average” patients and during ultrafiltration failure (UFF), according to the three-pore model of peritoneal transport. The UF volume curves as a function of dwell time [V( t)] are determined in their early phase by the glucose osmotic conductance [product of the UF coefficient (LpS) and the glucose reflection coefficient (σg)] of the peritoneum; in their middle portion by intraperitoneal volume and glucose diffusivity; and in their late portion by the LpS, Starling forces, and lymph flow. The most common cause of UFF is increased transport of small solutes (glucose) across the peritoneum, whereas the LpS is only moderately affected. Concerning peritoneal ion transport, ions that are already more or less fully equilibrated across the membrane at the start of the dwell, such as Na+ (Cl–), Ca2+, and Mg2+, have a convection-dominated transport. The removal of these ions is proportional to UF volume (approximately 10 mmol/L Na+ and 0.12 mmol/L Ca2+ removed per deciliter UF in 4 hours). The present article examines the impact on fluid and solute transport of varying concentrations of Ca2+ and Na+ in peritoneal dialysis solutions. Particularly, the effect of “ultralow” sodium solutions on transport and UF is simulated and discussed. Ions with high initial concentration gradients across the peritoneum, such as K+, phosphate, and bicarbonate, display a diffusion-dominated transport. The transport of these ions can be adequately described by non-electrolyte equations. However, for ions that are in (or near) their diffusion equilibrium over the peritoneum (Na+, Ca2+, Mg2+), more complex ion transport equations need to be used. Due to the complexity of these equations, however, non-electrolyte transport formalism is commonly employed, which leads to a marked underestimation of mass transfer area coefficients (PS). This can be avoided by determining the PS when transperitoneal ion concentration gradients are steep.

The purpose of the present study was to assess the role of diffusion and convection during filtration of Ficoll across the glomerular filter by comparing glomerular sieving coefficients (theta) to neutral fluorescein isothiocyanate (FITC)-Ficoll 70/400 obtained at low (hydropenic) vs raised (normal) glomerular filtration rates (GFRs). The theta for FITC-Ficoll was determined in anesthetized Wistar rats (304 +/- 18 g) following laparotomy and cannulation of the ureters, used for urine sampling. After surgery, GFR was 1.2 +/- 0.16 ml/min (+/- s.e.), assessed using the plasma to urine clearance of FITC-inulin and (51)Cr-ethylenediaminetetraacetic acid. FITC-Ficoll 70/400 was infused intravenously (i.v.) following an initial bolus dose. To raise GFR, to an average of approximately 2 ml/min, 5 ml of serum together with glucagon (3 microg/min) was given i.v. FITC-inulin and FITC-Ficoll were determined in plasma and urine using size-exclusion high-performance liquid chromatography. The theta for Ficoll as a function of Stokes-Einstein radius was significantly reduced in the range of 13-43 A when GFR was raised. The maximal theta lowering effect, in relative terms, of raising GFR was obtained for a Ficoll a(e) of approximately 32 A. For Ficoll(36 A) (cf. albumin), theta was reduced from 0.111+/- 0.009 to 0.081+/- 0.012 (P < 0.05; n = 7) for the GFR increment imposed. The reduction in theta for Ficoll after raising GFR indicates the presence of a high diffusive component of glomerular Ficoll filtration in rats in vivo and contradicts the notion of a significant concentration polarization effect in the glomerular filter upon Ficoll molecules < 50 A in radius.

The relative contribution of transcytosis vs. large pore transport to the passage of macromolecules across microvascular endothelia has been a controversial issue for nearly half a century. To separate transcytosis from ‘porous’ transport, the transcytosis inhibitors N-ethylmaleimide (NEM) and filipin have been tested in in situ or ex vivo perfused organs with highly conflicting results. In continually weighed isolated perfused organs, where measurements of pre- and post-capillary resistances, capillary pressure and capillary filtration coefficients can be repeatedly performed, high doses of NEM and filipin increased the bulk transport of macromolecules from blood to tissue, despite producing vasoconstriction. By contrast, in in situ perfused organs, marked reductions in the tissue uptake of albumin tracer have been observed after NEM and filipin. When tissue cooling has been employed as a means of inhibiting (active) transcytosis, results have invariably shown a low cooling sensitivity of albumin transport, compatible with passive transendothelial passage of albumin. This observation is further strengthened by the commonly observed dependence of albumin transport upon the capillary pressure and the rate of transcapillary convection. For low-density lipoprotein (LDL), a cooling-sensitive, non-selective transport component has been discovered, which may be represented by filtration through paracellular gaps, lateral diffusion through transendothelial channels formed by fused vesicles, or by transcytosis. From a physiological standpoint there is little evidence supporting active transendothelial transport of most plasma macromolecules. This seems to be supported by studies on caveolin-1-deficient mice lacking plasmalemmal vesicles (caveolae), in which there are no obvious abnormalities in the transendothelial transport of albumin, immunoglobulins or lipoproteins. Nevertheless, specific transport in peripheral capillaries of several hormones and other specific substances, similar to that existing across the blood-brain barrier, still remains as a possibility.

In anesthetized rabbits (n = 25) subject to slow intravenous saline loading (0.4 ml.min-1.kg-1) for 3 h, we measured pulmonary interstitial pressure (Pip) in intact in situ lungs with glass micropipettes inserted directly into the lung parenchyma via a "pleural window." Measurements were done in apneic animals at the end-expiratory volume with O2 delivered in the trachea. Pip was -10 +/- 1.5 (SD) cmH2O in control and increased to 0.6 +/- 3.8 and 5.7 +/- 3.3 cmH2O at 66 and 180 min, respectively. The wet-to-dry weight ratio (W/D) of the lung was 5.04 +/- 0.2 in the control group and 5.34 +/- 0.7 at 180 min (+6%); the corresponding W/D for intercostal muscles were 3.25 +/- 0.03 and 4.19 +/- 0.5 (+28%). Pulmonary interstitial compliance was 0.47 ml.mmHg-1.100 g wet wt-1. Pulmonary arterial and left atrial pressures were 18.4 +/- 2 and 3 +/- 1 cmH2O in control and increased to 19.5 +/- 2.9 and 4.6 +/- 1.7 cmH2O at 180 min, respectively. Aortic flow (cardiac output) increased from 103 +/- 35 to 131 +/- 26 ml/min; pulmonary resistance fell from 0.17 +/- 0.06 to 0.14 +/- 0.05 cmH2O.min.ml-1 (-18%), suggesting that the increase in Pip did not limit blood flow. The pulmonary capillary-to-interstitium filtration pressure gradient decreased sharply from a control value of 10 cmH2O to 0 cmH2O within 60 min because of the increase in Pip and remained unchanged for < or = 180 min. Data suggest that the pulmonary interstitial matrix can withstand fluid pressures above atmospheric, preventing the development of pulmonary alveolar flooding.

It is established that the glomerular filter sieves macromolecules based on their size, shape, and charge. Anionic proteins are thus retarded compared with their neutral or cationic counterparts. However, recent studies have indicated that charge effects are small, or even "anomalous," for polysaccharides. We therefore investigated the impact of charge on the glomerular permeability to polysaccharides by comparing sieving coefficients (theta; primary urine-to-plasma concentration ratio) for negatively charged, carboxymethylated (CM) FITC-Ficoll and FITC-dextran with their neutral counterparts. For these probes, theta were determined in anesthetized Wistar rats [269 +/- 2.7 g (+/-SE; n = 36)], whose ureters were cannulated for urine sampling. The glomerular filtration rate was assessed using FITC-inulin. Polysaccharides were constantly infused, and after equilibration, urine was collected and a midpoint plasma sample was taken. Size and concentration determinations of the FITC-labeled polysaccharides were achieved by size-exclusion HPLC (HPSEC). For CM-Ficoll, theta was significantly increased (32 times at 55 A) compared with that of uncharged Ficoll. A small increase in theta for CM-dextran compared with neutral dextran was also observed (1.8 times at 55 A). In conclusion, negatively charged Ficoll relative to neutral Ficoll was found to be markedly hyperpermeable across the glomerular filter. Furthermore, negatively charged Ficoll was observed to be larger on HPSEC compared with its neutral counterpart of the same molecular weight. It is proposed that the introduction of negative charges in the "dendrimeric," cross-linked Ficoll molecule may alter its configuration, so as to make it more extended, and conceivably, more flexible, thereby increasing its glomerular permeability.

Ultrafiltration failure (UFF) is a common complication of long-term peritoneal dialysis (PD). Functionally UFF is in most cases characterized by an enhanced peritoneal mass transfer area coefficient for glucose (PS(g)) combined with a largely unchanged peritoneal glucose osmotic conductance (L(p)Ssigma(g)). Morphologically, marked UFF occurs with fibrosis of the submesothelial zone in the peritoneum, combined with vasculopathy and vascular proliferation in deeper tissues. To computer simulate UFF, changes both in the vasculature and in the interstitium have to be taken into account. For that purpose, we used a three-pore membrane/fiber matrix serial barrier model, applying the three-pore model to the capillaries and the fiber-matrix model to the interstitium. The parameters of the three-pore model have been published previously. The interstitial fiber density was set at 0.5% (vol/vol) and the fiber radius (r(f)) at 6 A during control. If the interstitial fiber density was increased from 0.5 to 3%, and r(f) to 7.5 A (cf. collagen) while the capillary surface area was increased by 40% from control, then PS(g) increased from 9.3 to 11.5 ml/min, while the UF coefficient (L(p)S) was largely unchanged. Further increases in vascular surface area combined with further increases in fiber density caused further increments in PS(g), whereas L(p)S remained unchanged. It is concluded that a matrix of fibers coupled in series with a three-pore membrane may be used for simulating the pathophysiological alterations occurring in the peritoneum in UFF, explaining the commonly observed "uncoupling" of small solute transport (PS) from the peritoneal UF coefficient (L(p)S) in this condition.