Glomerular filtration barrier charge and size selectivity
Techniques that explore charge and size selectivity of the glomerular filtration barrier in vivo rely on examining the differential clearance of circulating proteins of varying charges and
sizes or of neutral molecules of differing sizes. The clearance of albumin (Stokes-Einstein radius 36 A, isoelectric point 4-5) is compared to that of IgG (radius 55 A, isoelectric point 4-10) as an index of size selectivity, while the clearance of the most anionic species of albumin (glycated albumin) or IgG (IgG4) relative to those of less anionic or cationic species provides an indicator of charge selectivity. A selective increase in albumin compared to IgG indicates preservation of size selectivity, while an increase in fractional clearances of IgG4 and albumin (both anionic) represents a decrease in anionic pore charge (Deckert et al. 1988). An increase in the ratio of urinary IgG4/IgG also supports a decrease in charge selectivity (DiMario et al. 1989). An increase in excretion of glycated albumin, which is more anionic than native albumin, supports selective proteinuria (Ghiggeri et al. 1985; Kverneland et al. 1986).
Assessment of the pore size of the GBM in vivo is a much more demanding task. Neutral molecules of varying sizes, such as the neutral polymer dextran 40 (Nakamura & Myers 1988; Scandling & Myers 1992; Myers et al. 1995), are given by intravenous infusion, along with inulin. Dextran 40 is neither reabsorbed nor excreted by the tubule, so that the fractional clearance of a given dextran is the same as the ratio of its concentration in Bowman's Space fluid to plasma, i.e. its sieving coefficient. Clearance studies are performed and dextrans separated, usually by gel permeation chromatography, before quantification. The sieving coefficient is calculated as:
(urine concentration dextran ^ plasma concentration dextran)
(urine concentration inulin ^ plasma concentration inulin)
Dextrans behave as random coils in solution but become uncoiled when subjected to shear stress. Thus their transport across the glomerulus is facilitated and effective pore size may be overestimated (Bohrer et al. 1984; Davidson & Deen 1988). Several groups have therefore used ficoll 70, which behaves as a rigid sphere during transglomerular permeation (Oliver et al. 1992; Blouch et al. 1997; Anderson et al. 2000). Tritiated ficoll 70, spanning a radius range of 10-90 A, is infused along with inulin and para-aminohippurate (PAH). Ficoll fractions in urine and blood were separated by HPLC and the relative amounts in each fraction were determined by the radioactivity. Sieving coefficients for each discrete fraction are then calculated. Curves of sieving coefficients by size are constructed, assuming that the glomerular capillary wall is a heteroporous membrane perforated by a lower distribution of restrictive pores with a log-normal distribution and a parallel upper distribution of shunt-like pores (Blouch et al. 1997). A number of other assumptions must be made, including the glomerular transcapillary hydraulic pressure difference (AP).
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