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Effects of glomerular filtration rate on Ficoll sieving coefficients (θ) in rats

2006; Elsevier BV; Volume: 69; Issue: 8 Linguagem: Inglês

10.1038/sj.ki.5000027

1523-1755

Catarina Rippe, Daniel Asgeirsson, Daniele Venturoli, Anna Rippe, Bengt Rippe,

Renal Diseases and Glomerulopathies

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 (θ) to neutral fluorescein isothiocyanate (FITC)-Ficoll 70/400 obtained at low (hydropenic) vs raised (normal) glomerular filtration rates (GFRs). The θ 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 51Cr-ethylenediaminetetraacetic acid. FITC-Ficoll 70/400 was infused intravenously (i.v.) following an initial bolus dose. To raise GFR, to an average of ∼2 ml/min, 5 ml of serum together with glucagon (3 μg/min) was given i.v. FITC-inulin and FITC-Ficoll were determined in plasma and urine using size-exclusion high-performance liquid chromatography. The θ for Ficoll as a function of Stokes–Einstein radius was significantly reduced in the range of 13–43 Å when GFR was raised. The maximal θ lowering effect, in relative terms, of raising GFR was obtained for a Ficoll ae of ∼32 Å. For Ficoll36 Å (cf. albumin), θ 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 θ 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 Å in radius. 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 (θ) to neutral fluorescein isothiocyanate (FITC)-Ficoll 70/400 obtained at low (hydropenic) vs raised (normal) glomerular filtration rates (GFRs). The θ 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 51Cr-ethylenediaminetetraacetic acid. FITC-Ficoll 70/400 was infused intravenously (i.v.) following an initial bolus dose. To raise GFR, to an average of ∼2 ml/min, 5 ml of serum together with glucagon (3 μg/min) was given i.v. FITC-inulin and FITC-Ficoll were determined in plasma and urine using size-exclusion high-performance liquid chromatography. The θ for Ficoll as a function of Stokes–Einstein radius was significantly reduced in the range of 13–43 Å when GFR was raised. The maximal θ lowering effect, in relative terms, of raising GFR was obtained for a Ficoll ae of ∼32 Å. For Ficoll36 Å (cf. albumin), θ 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 θ 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 αLGlomerular oncotic pressure (Δπ) (mmHg)(a) Two-pore model analysis1.16±0.1750.3±0.38136±8.63.73±0.85 × 1052.67±0.45 × 10-46.04±1.5 × 10-528.0±1.12.03±0.1748.9±0.77140±113.96±0.73 × 1052.51±0.45 × 10-47.45±1.5 × 10-526.7±0.9GFR (ml/min)Mean pore radius (ul) (Å)Distribution parameter (sl)A0/ΔX (cm)JvL/GFRαLGlomerular oncotic pressure (Δπ) (mmHg)(b) Log-normal distributed+shunt analysis of average θ vs ae curves1.16±0.1738.8±0.791.19±0.021.04±0.12 × 1067.20±2.6 × 10-51.14±0.41 × 10-528.0±1.12.03±0.1738.8±0.701.18±0.011.21±0.22 × 1066.98±1.5 × 10-51.83±0.21 × 10-526.7±0.9a Analysis of Ficoll sieving data at the two GFR levels investigated using the two-pore model analysis of individual sieving curves. The oncotic pressures for the two GFRs are calculated as described in Lund et al.,5.Lund U. Rippe A. Venturoli D. et al.Glomerular filtration rate dependence of sieving of albumin and some neutral proteins in rat kidneys.Am J Physiol Renal Physiol. 2003; 284: F1226-F1234Crossref PubMed Scopus (115) Google Scholar assuming that the (afferent arteriole) plasma oncotic pressure is 22 mmHg. αL is calculated for LpS=0.22 (ml/min/mmHg), compatible with A0/ΔX 1 × 106 cm. For JVL/GFR and αL, n=7+3. For all other parameters, n=7.b Parameters for the log-normal distributed model+shunt analysis. The calculations are based on Rippe and Haraldsson,11.Rippe B. Haraldsson B. Transport of macromolecules across microvascular walls. The two-pore theory.Physiol Rev. 1994; 74: 163-219Crossref PubMed Scopus (9) Google Scholar Deen et al.20.Deen W.M. Bridges C.R. Brenner B.M. et al.Heteroporous model of glomerular size selectivity: application to normal and nephrotic humans.Am J Physiol. 1985; 249: F374-F389PubMed Google Scholar and McNamee and Wolf32.McNamee J.E. Wolf M.B. Prediction of permeability–surface area product data by continuous-distribution pore models.Microcirculation. 1998; 5: 275-280Crossref PubMed Scopus (4) Google Scholar and are described in detail in Appendix A1. Open table in a new tab Raising GFR from 1.2 to 2.0 ml/min (n=7), according to the two-pore model, implies a 15–20% reduction in θ for Ficoll fractions of radius 13–45 Å. Our measured sieving data for Ficoll reflected an even larger drop in θ than expected when simulated for the two-pore model (setting A0/ΔX at 390 000 cm). For Ficoll25 Å, the theoretically predicted reduction in θ from the two-pore theory, when GFR is raised from 1.2 to 2.0 ml/min, is from 0.65 to 0.55 using the parameters listed in Table 1a. However, measured values were 0.68±0.038 and 0.52±0.034, respectively. For Ficoll36 Å (cf. albumin), predicted θ values are 0.11 vs 0.10, whereas the measured data were 0.111±0.009 vs 0.081±0.012. The negative trend of the relationship between θ vs GFR is illustrated in Figure 4 for four different molecular radii, namely 20, 25, 30, and 36 Å. Raising GFR from 1.2 to 1.9 ml/min (n=7+3; see Materials and Methods) did not significantly affect θ for molecules in the range of 50–70 Å. However, for molecules larger than 70 Å (in ae), there was a tendency toward a reduced θ, even though this tendency did not reach statistical significance. This is illustrated in Figure 2b, showing that the 95% confidence interval of the θ ratio for low vs high GFRs was not significantly different from unity for ae>43 Å. Furthermore, the 95% confidence interval widened considerably for ae values >70 Å, reflecting the high variability in θ measurements for the largest molecules. This in vivo study demonstrates that the fractional clearances (θ) of Ficoll fractions in the radius range of 13–43 Å are reduced when GFR is raised from initially low values to higher (normal range) levels in hydropenic rats. The data agree with previous in vivo studies of θ vs GFR using either proteins or dextran as probes for testing the glomerular size selectivity.5.Lund U. Rippe A. Venturoli D. et al.Glomerular filtration rate dependence of sieving of albumin and some neutral proteins in rat kidneys.Am J Physiol Renal Physiol. 2003; 284: F1226-F1234Crossref PubMed Scopus (115) Google Scholar, 8.Chang R.L. Ueki I.R. Troy J.L. et al.Permselectivity of the glomerular capillary wall to macromolecules.Biophys J. 1975; 15: 887-906Abstract Full Text PDF PubMed Scopus (197) Google Scholar Theoretically, if the main sieving barrier in the glomerular capillary wall were close to the plasma side, the effect of an increased GFR would be to reduce the diffusional component of transport, so that, eventually, at high GFRs, θ will fall to approach (1-σ).11.Rippe B. Haraldsson B. Transport of macromolecules across microvascular walls. The two-pore theory.Physiol Rev. 1994; 74: 163-219Crossref PubMed Scopus (9) Google Scholar, 12.Taylor A.E. Granger D.N. Exchange of macromolecules across the microcirculation.in: Renkin E.M. Michel C.C. Handbook of Physiology. The Cardiovascular System. Microcirculation. American Physiological Society, Bethesda, MD1984: 467ppGoogle Scholar However, if the major sieving barrier were located at the PSD, then the reduction in θ will be counteracted by concentration polarization effects. The two opposing effects would largely cancel at low GFR levels, leaving θ unchanged when GFR is raised. However, at high GFRs, concentration polarization would dominate, tending to cause a rise in θ for macromolecules at high levels of GFR.1.Deen W.M. Lazzara M.J. Myers B.D. Structural determinants of glomerular permeability.Am J Physiol Renal Physiol. 2001; 281: F579-F596PubMed Google Scholar, 7.Edwards A. Daniels B.S. Deen W.M. Ultrastructural model for size selectivity in glomerular filtration.Am J Physiol. 1999; 276: F892-F902PubMed Google Scholar This was, however, not seen in the present study. The present data, demonstrating a fall in θ with GFR, are rather consistent with the glomerular barrier producing a size-separation effect 'early' in the filter. The measured reduction in θ for Ficoll (13–43 Å in radius) was slightly larger than predicted by the two-pore theory (Figure 4). A major reason is that fitting measured Ficoll θ data to the two-pore theory underestimates the diffusive component (A0/ΔX) of transport. The low A0/ΔX obtained for Ficoll (∼400 000 cm) is a major factor contributing to the discrepancy between the predicted and experimental values. If the value for A0/ΔX obtained using the log-normal distributed pore theory (∼1.2 × 106 cm) or A0/ΔX obtained using protein probes (∼1.8 × 106 cm) was used instead, predictions became more accurate. While the diffusion component was sufficiently high for θ of Ficoll fractions in the range of 13–43 Å, to be markedly reduced with GFR elevations, the diffusional component is negligible for molecules approaching the small pore radius. Any increases in GFR will thus leave θ unaffected for such Ficoll fractions. Our data thus clearly demonstrate that there was no reduction in θ for molecules of radius between 45 and 60 Å (Figure 2b). For molecules larger than 60 Å in radius, which, according to the two-pore model, are considered to pass through large pores only, convection would be the dominating transport mode. Here, increasing GFR will theoretically reduce the relative fluid flow occurring through large pores (JVL/GFR) and reduce θ, so that it, eventually, would approach αL(1-σL).16.Tencer J. Frick I.M. Öqvist B.W. et al.Size-selectivity of the glomerular barrier to high molecular weight proteins: upper size limitations of shunt pathways.Kidney Int. 1998; 53: 709-715Abstract Full Text PDF PubMed Scopus (114) Google Scholar In our study, we found a tendency toward a reduction in θ for very large Ficoll fractions (>70 Å in radius). However, because only very small amounts of high molecular radius material (θ immunoglobulin (Ig)G>α2-macroglobulin. IgM was not found in rat primary urine at all. Moreover, in the cooled isolated perfused kidney (cIPK), there is ample evidence for the presence of two populations of pores in the glomerular filter,22.Haraldsson B. Sörensson J. Why do we not all have proteinuria? An update of our current understanding of the glomerular barrier.News Physiol Sci. 2004; 19: 7-10Crossref PubMed Scopus (92) Google Scholar and application of the two-pore model makes it possible to compare the present results with those obtained in the cIPK. Instead of the concept of a wide pore distribution in the glomerular filter, one has to consider the fact that carbohydrates, such as Ficoll and dextran, exhibit an 'extended' molecular conformation, partly correlated to their larger SE radius (compared to that for globular proteins) for any given molecular mass.15.Venturoli D. Rippe B. 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. 2005; 288: F605-F613Crossref PubMed Scopus (318) Google Scholar Such an extended configuration, conceivably, renders most carbohydrates more flexible (compressible) than any globular protein. It is likely that an increased flexibility makes carbohydrates behave differently compared to rigid proteins in the permeable pathways (pores).13.Bohrer M.P. Patterson G.D. Carroll P.J. Hindered diffusion of dextran and Ficoll in microporous membranes.Macromolecules. 1984; 17: 1170-1173Crossref Scopus (147) Google Scholar, 15.Venturoli D. Rippe B. 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. 2005; 288: F605-F613Crossref PubMed Scopus (318) Google Scholar, 17.Lavrenko P.N. Mikriukova O.I. Okatova O.V. On the separation