Charge Selectivity is a Concept That Has Yet to be Demonstrated

The following is the abstract of the article discussed in the subsequent letter:

	ABSTRACT

Ohlson M, Sörensson J, and Harraldsson B. Glomerular size and charge selectivity in the rat as revealed by FITC-Ficoll and albumin. Am J Physiol Renal Physiol 279: F992-F993, 2000. Fractional clearances () for FITC-Ficoll and albumin were estimated in isolated perfused rat kidneys in which the tubular activity was inhibited by low temperature (8°C) and/or 10 mM NH₄Cl. The Ficoll data were analyzed according to a two-pore model giving small and large pore radii of 46 Å and 80-87 Å, respectively. The estimated negative charge density was 35-45 meq/l at 8°C. Perfusion with erythrocyte-free solutions of kidneys at 37°C reduced glomerular size and charge permselectivity. Thus the large pore fraction of the glomerular filtrate (f_L) was 1.64% at 37°C compared with 0.94% at 8°C. The for albumin was four times higher at 37°C than at 8°C (0.86% vs. 0.19%, respectively). NH4Cl caused further irreversible damage to the glomerular barrier. We conclude that there are no deleterious effects on the glomerular barrier of a reduction in temperature from 37°C to 8°C. Therefore our data seem to disprove the hypothesis of low glomerular permselectivity and transtubular uptake of intact albumin and support the classic concept of a highly selective glomerular barrier.

	LETTER

Charge Selectivity is a Concept That Has Yet to be Demonstrated

To the Editor: I would like to draw to the attention of readers that differential excretion rates of charged proteins are still being unjustifiably interpreted in terms of charge selectivity (that is the result of electrostatic repulsion of negatively charged protein such as albumin with the fixed negative charges on the glomerular capillary wall) as in the recent paper of Ohlson et al. (2). These investigators examined albumin excretion in kidneys maintained in anesthetized rats, whose body temperature was maintained at 37°C via a thermostatically controlled heat pad, and perfused with a solution at 8°C containing 18 mg/ml human serum albumin. The issue is that charge selectivity in extracellular physiological systems is a concept that has been frequently proposed but has never been demonstrated. Studies performed in vivo have demonstrated that probes, previously used to measure "charge selectivity" including dextran sulfate and charged proteins, are found to be excreted in degraded form (Refs. 3 and 6 and earlier references cited therein) so any conclusions about charge effects cannot be made. There are also important and elegant physicochemical studies that have been overlooked but have demonstrated that interaction of albumin with highly charged polyanions under physiological conditions is simply one of excluded volume effects or size exclusion (1, 4). No charge interactions were evident. Albumin distribution in nonrenal tissues that have far higher fixed negative charge concentration than the glomerular capillary wall is also governed by non-electrostatic-based excluded volume effects (5). In the studies of Ohlson et al. (2) it is simply not enough to obtain empirical data that has the same pattern as purported "charge selectivity" to justify that it is charge selectivity. It is apparent that many types of anomalous interactions may occur in their unusual perfusion system that may lead to a particular excretion pattern.

	REFERENCES

1.   Ogston, AG, and Preston BN. The exclusion of protein by hyaluronic acid. J Biol Chem 241: 17-19, 1966[Abstract/Free Full Text].

2.   Ohlson, M, Sörensson J, and Haraldsson B. Glomerular size and charge selectivity in the rat as revealed by FITC-Ficoll and albumin. Am J Physiol Renal Physiol 279: F84-F91, 2000[Abstract/Free Full Text].

3.   Osicka, TM, Houlihan CA, Chan JG, Jerums G, and Comper WD. Albuminuria in patients with type I diabetes is directly linked to changes in the lysosomal-mediated degradation of albumin during renal passage. Diabetes 49: 1579-1584, 2000[Abstract].

4.   Shaw, M, and Schy A. Exclusion in hyaluronate gels. Biophys J 17: 47-55, 1977[Web of Science][Medline].

5.   Snowden, J, and Maroudas A. Distribution of serum albumin in human normal and degenerate cartilage. Biochim Biophys Acta 428: 726-740, 1976[Medline].

6.   Vyas, SV, Burne MJ, Pratt LM, and Comper WD. Glomerular processing of dextran sulphate. Arch Biochem Biophys 332: 205-212, 1996[Web of Science][Medline].

Wayne D. Comper, Department of Biochemistry and Molecular Biology Monash University Clayton, Victoria, Australia 3800

	REPLY

To the Editor: Dr. Comper is concerned about the term "charge selectivity," but is it not more important to discuss the 100-fold difference in albumin clearance? Thus, the real issue is that Dr. Comper and co-workers have suggested the sieving coefficient for albumin to be close to 8% and that albumin is retrieved from the urine in intact form by the tubular cells (11). In humans this would imply a daily glomerular filtration of more than 700 g (!) of albumin (8). In our study, we found no experimental support for their hypothesis (8), and we proposed an alternative explanation of their experimental findings (8). Furthermore, recent studies of the cubulin-megalin complex (1) strongly support the classical notion (7) that all albumin molecules reabsorbed by the tubular cells are degraded.

Let us turn to the issue of terminology. Evidence for charge selectivity have been reported, not only in an "unusual perfusion system," but also in humans (2) and intact rats (3, 14). Dr. Comper implies that charge selectivity only occurs in the cooled isolated perfused kidneys (IPK), but their glomerular permeability characteristics resemble those reported for intact rats (3, 10). Moreover, charge effects have been studied, not only with albumin (8) but also using horseradish peroxidase (13), lactate dehydrogenase (6), myoglobin (14), ovalbumin (12), and orosomucoid (12). Dr. Comper cites neither of these studies. As to the theoretical background for charge interactions, there are modern theories (5) extending the work of Ogston. The cited model has experimental support (4), and it has been applied to glomerular sieving data for different solutes in the IPK (12). The description of charge interactions by Dr. Comper is therefore far from adequate. The exact functional pore dimensions and charge densities in the glomerular barrier (9) seem to be overestimated if dextran is used as a tracer. This fact does not, however, invalidate the classic view of glomerular size and charg eselectivity.

The observation by Dr. Comper's group that certain conditions may cause dramatic reductions of glomerular permselectivity (11) is an important contribution to the field. Their hypothesis that this reflects a normally "leaky" glomerular barrier has been experimentally rejected (1, 7, 8), but that does not diminish their findings; it merely calls for alternative explanations. We have suggested the delicate endothelial cell coat to be involved (9). Further analysis of the conditions causing proteinuria (11) will no doubt give us new insights about the intricate glomerular barrier.

	REFERENCES

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2.   Guasch, A, Deen WM, and Myers BD. Charge selectivity of the glomerular filtration barrier in healthy and nephrotic humans. J Clin Invest 92: 2274-2282, 1993.

3.   Hjalmarsson, C, Ohlson M, and Haraldsson B. Puromycin aminonuceleoside damages the glomerular size barrier with minimal effects on charge density. Am J Physiol Renal Physiol 281: F503-F512, 2001[Abstract/Free Full Text].

4.   Johnson, EM, Berk DA, Jain RK, and Deen WM. Diffusion and partitioning of proteins in charged agarose gels. Biophys J 68: 1561-1568, 1995[Web of Science][Medline].

5.   Johnson, EM, and Deen WM. Electrostatic effects on the equilibrium partitioning of spherical colloids in random fibrous media. J Colloid Interface Sci 195: 268-268, 1997.

6.   Lindström, KE, Johnsson E, and Haraldsson B. Glomerular charge selectivity for proteins larger than serum albumin as revealed by lactate dehydrogenase isoforms. Acta Physiol Scand 162: 481-488, 1998[Web of Science][Medline].

7.   Maunsbach, AB. Absorption of I-125-labeled homologous albumin by rat kidney proximal tubule cells. A study of microperfused single proximal tubules by electron microscopic autoradiography and histochemistry. J Ultrastruct Res 15: 197-241, 1966[Web of Science][Medline].

8.   Ohlson, M, Sörensson J, and Haraldsson B. Glomerular size and charge selectivity in the rat as revealed by FITC-Ficoll and albumin. Am J Physiol Renal Physiol 279: F84-F91, 2000.

9.   Ohlson, M, Sörensson J, and Haraldsson B. A gel-membrane model of glomerular charge and size selectivity. Am J Physiol Renal Physiol 280: F395-F405, 2001.

10.   Oliver, JD, III, Anderson S, Troy JL, Brenner BM, and Deen WM. Determination of glomerular size selectivity in the normal rat with Ficoll. J Am Soc Nephrol 3: 214-228, 1992[Abstract].

11.   Osicka, TM, Pratt LM, and Comper WD. Glomerular capillary wall permeability to albumin and horseradish peroxidase. Nephrology 2: 199-212, 1996.

12.   Sörensson, J, Ohlson M, and Haraldsson B. A quantitative analysis of the glomerular charge barrier in the rat. Am J Physiol Renal Physiol 280: F646-F656, 2001[Abstract/Free Full Text].

13.   Sörensson, J, Ohlson M, Lindström K, and Haraldsson B. Glomerular charge selectivity for horseradish peroxidase and albumin at low and normal ionic strengths. Acta Physiol Scand 163: 83-91, 1998[Web of Science][Medline].

14.   Wolgast, M, and Källskog Ö, and Wahlström H. Characteristics of the glomerular capillary membrane of the rat kidney as a hydrated gel. II. On the validity of the model. Acta Physiol Scand 158: 225-232, 1996[Web of Science][Medline].

Börje Haraldsson, Maria Ohlson, Jenny Sörensson, Department of Physiology Göteborg University Box 432 SE 405 30 Gothenburg, Sweden