Macromolecular sieving by glomerular basement membrane in vitro: effect of polycation or biochemical modifications

To evaluate the effect of biochemical modifications not possible in vivo, filters of dog glomerular basement membrane (GBM) were constructed in ultrafiltration cells in vitro. The sieving coefficients (SCs) of three protein markers of differing size and charge (native, anionic bovine albumin-BSA; cationized BSA-cBSA; and immunoglobulin G-IgG) were determined using filters of differing amounts of control GBM, and under varying transmembrane pressures (delta P). Flow rates did not increase proportionately with increasing delta P, indicating filter compressibility. Protein SCs did not change with changing delta P, but did decrease with increasing filter thickness. Control filters showed a small but definite charge selectivity (SCcBSA++ - SCBSA greater than 0); a much greater degree of size selectivity (SCcBSA - SCIgG) was observed. Hexadimethrine (HDM), a polycation which causes proteinuria in vivo, led to marked increases in protein SCs. In contrast, removal of the major population of intrinsic GBM negative charges by carboxyl group methylation only produced a small increase in the filtration of BSA, with no change in filtration of cBSA or IgG. Other biochemical modifications (heparinase or neuraminidase treatment) had no effect on filter permselectivity. Carboxyl group methylation essentially abolished filter binding of cationized ferritin, which showed substantial binding to control filters. These in vitro studies provide confirmatory evidence for a direct effect of HDM on the permselective properties of GBM. In addition, biochemical modification studies suggest a fundamental difference between the binding of an exogenous polycation to GBM anionic sites and the removal of intrinsic charges.