Uncharged Dextran Research Articles

Effects of Electric Charge on the Interfacial Tension between Coexisting Aqueous Mixtures of Polyelectrolyte and Neutral Polymer

Upon demixing, an aqueous solution of a polyelectrolyte and an incompatible neutral polymer yields two phases separated by an interface with an ultralow tension. Here, both in theory and experiment, we study this interfacial tension in detail: how it scales with the concentrations of the polymers in the two phases and how it is affected by the interfacial difference in the electrical potential. Experiments are performed on an aqueous model system of uncharged dextran and charged nongelling gelatin. The experimental tension scales to the power ∼3 with the tie-line length in the phase diagram of demixing, in agreement with mean-field theory where space is filled with a binary mixture of polymer blobs. The interfacial electrical potential difference is experimentally found to decrease the interfacial tension in a way that is consistent with Poisson–Boltzmann theory inspired from Frenkel and Verwey–Overbeek.

Effect of dextran and dextran sulfate on the structural and rheological properties of model acid milk gels

Various types of polysaccharides are widely used in cultured dairy products. However, the interaction mechanisms, between milk proteins and these polysaccharides, are not entirely clear. To explore the interactions between uncharged and charged polysaccharides and the caseins, we used a model acid-milk-gel system, which allowed acidification to occur separately from gelation. The effect of adding uncharged dextran (DX; molecular weight ~2.0×106 Da) and negatively charged dextran sulfate (DS; molecular weight ~1.4×106 Da) to model acid milk gels was studied. Two concentrations (0.075 and 0.5%, wt/wt) of DX or DS were added to cold milk (~0°C) that had been acidified to pH values 4.4, 4.6, 4.8, or 4.9. Acidified milks containing DX or DS were then quiescently heated at the rate of 0.5°C/min to 30°C, which induced gelation, and gels were then held at 30°C for 17 h to facilitate gel development. Dynamic small-amplitude-oscillation rheology and large-deformation (shear) tests were performed. Microstructure of gels was examined by fluorescence microscopy. Gels made with a high concentration of DX gelled at a lower temperature, but after 17 h at 30°C, these gels exhibited lower storage moduli and lower yield-stress values. At pH 4.8 or 4.9 (pH values greater than the isoelectric point of caseins), addition of 0.5% DS to acidified milk resulted in lower gelation temperature. At pH 4.4 (pH values less than the isoelectric point of caseins), addition of 0.5% DS to acidified milk resulted in gels with very high stiffness values. Gels made at pH 4.8 or 4.9 with both concentrations of DS had much lower stiffness and yield-stress values than control gels. Microstructural analysis indicated that gels made at pH 4.4 with the addition of 0.5% DX exhibited large protein strands and pores, whereas gels made with 0.075% DX or the control gels had a finer protein matrix. At higher pH values (>4.4), gels made with 0.5% DX had a finer structure. At all pH values, gels made with 0.5% DS exhibited larger pores than the control gels. This study demonstrated that low concentrations of uncharged DX did not significantly affect the rheological properties of model acid milk gels; high concentrations of DX resulted in earlier gelation, possibly caused by depletion-induced attractions between casein particles, which altered the microstructure and created weaker gels. At pH values <4.6, negatively charged DS produced stiff casein gels, which might be due to attractive crosslinking by electrostatic interactions between DS and caseins at pH values below the isoelectric pH of casein (i.e., positively charged casein regions interacted with negatively charged DS molecules).

Biophysical effects in off-resonant gold nanoparticle mediated (GNOME) laser transfection of cell lines, primary- and stem cells using fs laser pulses.

Gold nanoparticle mediated (GNOME) laser transfection is a powerful technique to deliver small biologically relevant molecules into cells. However, the transfection of larger and especially negatively charged DNA remains challenging. The efficiency for pDNA was 0.57% using parameter that does not influence the endo- and exogenous DNA. In order to gain a deeper understanding of the actual molecule uptake process, the uptake efficiency was determined using molecules of different sizes. It was evaluated that uncharged dextran molecules (2000 kDa) were delivered with an efficiency of 68%. The intracellular distribution of injected molecules was visualized and larger molecules were primary found in the cytoplasm. Patch clamp measurements suggested a permeabilization time up to 15 minutes. The uptake efficiency depended on the size and charge of the molecule to deliver as well as the cell size. A minor role for transfection plays the cell type since primary stem cells were successfully transfected. The perforation efficiency of semi-adherent and suspension cells is influenced by the cell and molecule size.

Fundamental differences in model cell-surface polysaccharides revealed by complementary optical and spectroscopic techniques

The propensity of two classes of polysaccharides, charged heparins and uncharged dextrans serving as models of those found in the extracellular matrix, to form ordered arrangements in solids and in solution, were explored employing polarising optical microscopy and reflection anisotropy spectroscopy. The fundamental modes of molecular vibration in the solid state, which relate to the occupancy of conformational states at ambient temperatures, were also investigated using terahertz (1–15 THz) spectroscopy on purpose built beam lines at SRS, Daresbury and ANKA, Karlsruhe. In the solid state, evidence for the anisotropic arrangement of ion centres was observed for the Na, Ca and Mg ion forms of heparin by RAS but, this was absent in aqueous solution at 100 mg ml−1 and the absence of molecular ordering in solution was confirmed by polarised optical microscopy. The ion charged heparin specimens showed very strong absorption of THz radiation indicating the accessibility of unoccupied higher energy conformational states in these materials whereas the uncharged dextrans showed only weak THz absorption, implying that the majority of conformational states are occupied and there are few unoccupied higher energy states in these materials. This suggests the two classes of material will have very different conformational properties, particularly in response to temperature changes in the extracellular matrix.

3d localization and diffusion of proteins in polyelectrolyte multilayers

The interaction of diverse biomaterials with surfaces is more crucial than ever for biomedical applications to ensure efficiency and reproducibility. Very interesting surface materials are micrometer-thick polyelectrolyte multilayers. Not only their surface but also the bulk can be loaded with biomaterials like proteins or DNA for various purposes. Therefore, we established a method to analyze the lateral and vertical distribution of fluorescently labelled proteins of various size and charge in polyelectrolyte films composed of poly(L-lysine) and hyaluronic acid by confocal laser scanning microscopy. This approach enables us to measure the diffusion coefficients of the proteins via fluorescence recovery after photobleaching as a function of their vertical position in the film and facilitates the understanding of molecular interactions in the film with a high resolution in both space and time. As a result, we confirm that protein loading in the film is driven by electrostatic interactions – uncharged dextran molecules of 10 and 500 kDa do not diffuse into the film. Proteins of different sizes (3–11 nm) can diffuse relatively fast (D = 2–4 μm2 s−1) independent of their net charge, indicating complex interpolymer interactions. This approach is a new powerful experimental tool to design the polyelectrolyte multilayers for bio-applications by finding a relationship between intermolecular interactions and mobility and availability of biomolecules to biological samples (e.g. cells) or detection units (e.g. biosensors).

Interleukin-6 (IL-6) Regulates Claudin-2 Expression and Tight Junction Permeability in Intestinal Epithelium

In inflammatory bowel diseases (IBD), intestinal barrier function is impaired as a result of deteriorations in epithelial tight junction (TJ) structure. IL-6, a pleiotropic cytokine, is elevated in IBD patients, although the role of IL-6 in barrier function remains unknown. We present evidence that IL-6 increases TJ permeability by stimulating the expression of channel-forming claudin-2, which is required for increased caudal-related homeobox (Cdx) 2 through the MEK/ERK and PI3K pathways in intestinal epithelial cells. IL-6 increases the cation-selective TJ permeability without any changes to uncharged dextran flux or cell viability in Caco-2 cells. IL-6 markedly induces claudin-2 expression, which is associated with increased TJ permeability. The colonic mucosa of mice injected with IL-6 also exhibits an increase in claudin-2 expression. The claudin-2 expression and TJ permeability induced by IL-6 are sensitive to the inhibition of gp130, MEK, and PI3K. Furthermore, expression of WT-MEK1 induces claudin-2 expression in Caco-2 cells. Claudin-2 promoter activity is increased by IL-6 in a MEK/ERK and PI3K-dependent manner, and deletion of Cdx binding sites in the promoter sequence results in a loss of IL-6-induced promoter activity. IL-6 increases Cdx2 protein expression, which is suppressed by the inhibition of MEK and PI3K. These observations may reveal an important mechanism by which IL-6 can undermine the integrity of the intestinal barrier.

Rapid monoclonal antibody adsorption on dextran-grafted agarose media for ion-exchange chromatography

The binding capacity and adsorption kinetics of a monoclonal antibody (mAb) are measured for experimental cation exchangers obtained by grafting dextran polymers to agarose beads and compared with measurements for two commercial agarose-based cation exchangers with and without dextran grafts. Introduction of charged dextran polymers results in enhanced adsorption kinetics despite a dramatic reduction of the accessible pore size as determined by inverse size-exclusion chromatography. Incorporation of neutral dextran polymers in a charged agarose bead results instead in substantially lower binding capacities. The effective pore diffusivities obtained from batch uptake curves increase substantially as the protein concentration is reduced for the resins containing charged dextran grafts, but are much less dependent on protein concentration for the resins with no dextran or uncharged dextran grafts. The batch uptake results are corroborated by microscopic observations of transient adsorption in individual particles. In all cases studied, the adsorption kinetics is characterized by a sharp adsorption front consistent with a shell-progressive, diffusion limited mechanism. Greatly enhanced transport rates are obtained with an experimental resin containing charged dextran grafts with effective pore diffusivities that are 1–9 times larger than the free solution diffusivity and adsorption capacity approaching 300 mg/cm 3 of particle volume.

Triphase Equilibria in Cellulose Nanocrystal Suspensions Containing Neutral and Charged Macromolecules

The effect of added nonadsorbing macromolecules on the phase behavior of suspensions of electrostatically stabilized cellulose nanocrystals is examined. Triphasic isotropic−isotropic−nematic equilibria develop in cellulose nanocrystal suspensions to which a mixture of dextran and blue dextran is added. Phase diagrams for different combinations of uncharged dextran and anionically charged blue dextran are presented. The range of dextran concentrations at which the triphase equilibrium is produced is strongly influenced by the molecular weight of the blue dextran. Reentrant phenomena as well as regions of biphase coexistence are present in the phase diagrams. A threshold difference in charge density between two blue dextrans of varying degree of dye substitution is necessary for triphase equilibrium to occur at a given number density of each polymer species. Different repulsive electrostatic and attractive entropic forces may dominate at different concentrations of the dextrans, contributing to the rich phase behavior of these suspensions.

Branched biodegradable polyesters for parenteral drug delivery systems

Continuous, ‘infusion-like’ drug release profiles from biodegradable parenteral delivery systems are difficult to achieve for proteins and other hydrophilic macromolecular drugs with commonly used linear polyesters from lactic acid (PLA) and its random copolymers with glycolic acid (PLG). Drug release rates can be modified either by increasing the hydrophilicity of polyesters or by manipulating the polymer architecture to adjust polymer degradation rates and thus drug release. Therefore, we investigated different branching concepts for biodegradable polyesters of PLA and PLG. For one four- and eight-arm poly(ethylene oxide)s (PEO) were grafted with shorter polyester chains leading to star-branched structures. Secondly we obtained comb-like polyesters using both charged and uncharged dextrans or poly(vinyl alcohol)s (PVA) as hydrophilic backbones. The star-shaped and brush-like grafted polymers were intensively characterized by methods, such as NMR, IR, SEC-SLS, DSC and viscosity measurements. Tailor-made properties make these novel biodegradable polyesters promising candidates for parenteral protein delivery systems. While the star-branched polyesters have shown some interesting properties with respect to their degradation behavior, retaining the PEO blocks longer than ABA triblock copolymers, their release properties need further optimization. Brush-like branched polyesters on the other hand seem to possess both degradation and release properties meriting further investigations for parenteral protein delivery systems.

Polycations induce calcium signaling in glomerular podocytes

The neutralization of the polyanionic surface of the podocyte by perfusion of kidneys with polycations, such as protamine sulfate, leads to a retraction of podocyte foot processes and proteinuria. This study investigates the effects of protamine sulfate or anionic, neutral, or cationic dextrans on the cytosolic calcium activity ([Ca2+]i) in podocytes. [Ca2+]i was measured in single cultured differentiated mouse podocytes with the fluorescence dye fura-2/AM. Protamine sulfate caused a concentration-dependent and partially reversible increase of [Ca2+]i (EC50 approximately 1.5 micromol/liter). Pretreatment of the cells with heparin (100 U/liter) inhibited the protamine sulfate-mediated increase of [Ca2+]i. Like protamine sulfate, diethylaminoethyl dextran (DEAE-dextran) concentration dependently increased [Ca2+]i in podocytes (EC50 approximately 20 nmol/liter), whereas dextran sulfate or uncharged dextran (both 10 micromol/liter) did not influence [Ca2+]i. A reduction of the extracellular Ca2+ concentration (from 1 mmol/liter to 1 micomol/liter) partially inhibited the protamine sulfate and the DEAE-dextran-induced [Ca2+]i response. Flufenamate (100 micromol/liter) or Gd3+ (10 micromol/liter), which are known to inhibit nonselective ion channels, did not influence the [Ca2+]i increase induced by protamine sulfate. In the presence of thapsigargin (50 nmol/liter), an inhibitor of the endoplasmic reticulum Ca2+-ATPase, both protamine sulfate and DEAE-dextran increased [Ca2+]i. The data indicate that polycations increase podocyte [Ca2+]i. The increase of [Ca2+]i may be an early event in the pathogenesis of protamine sulfate-mediated retraction of podocyte foot processes.

A codiffusing system as a novel model for capillary wall charge selecivity

The electrostatic interaction associated with polyion-polyion interaction has been thought to be the basis for the differential transport of charged transport probes across the capillary wall. The charge of the transport probe may interact with the negative charges of capillary wall cell surfaces and the intercellular glycocalyx or extracellular matrix. This study has set out to quantitate the exact nature of the polyion-polyion interaction through the theoretical and experimental study of the partition-diffusion of albumin in solutions of anionic polysaccharides (dextran sulfate or heparin) at concentrations up to 250 meq l −1 or at an average intercharge distance of 2.2 nm, as compared with solutions of uncharged dextran at the same volume concentration. The results demonstrate that the albumin partition-diffusion is exactly the same in dextran sulfate, heparin and uncharged dextran matrices of the same polymer volume fraction. These results confirm previous studies from this laboratory that the charge effect through polyion-polyion interaction under physiological conditions is negligible.

Control of endothelial cell gene expression by flow

The vessel wall is constantly subjected to, and affected by, the stresses resulting from the hemodynamic stimuli of transmural pressure and flow. At the interface between blood and the vessel wall, the endothelial cell plays a crucial role in controlling vessel structure and function in response to changes in hemodynamic conditions. Using bovine aortic endothelium monolayers, we show that fluid shear stress causes simultaneous differential regulation of endothelial-derived products. We also report that the downregulation of endothelin-1 mRNA by flow is a reversible process, and through the use of uncharged dextran supplementation demonstrate it to be shear stress-rather than shear rate-dependent. Recent work on the effect of fluid shear stress on endothelial cell gene expression of a number of potent endothelial products is reviewed, including vasoactive substances, autocrine and paracrine growth factors, thrombosis/fibrinolysis modulators, chemotactic factors, surface receptors and immediate-early genes. The encountered patterns of gene expression responses are classified into three categories: a transient increase with return to baseline (type I), a sustained increase (type II) and a biphasic response consisting of an early transient increase of varying extent followed by a pronounced and sustained decrease (type III). The importance of the dynamic character of the flow stimulus and the magnitude dependence of the response are presented. Potential molecular mechanisms of shear-induced gene regulation, including putative shear stress response elements (SSRE), are discussed. These results suggest exquisite modulation of endothelial cell phenotype by local fluid shear stress and may offer insight into the mechanism of flow-dependent vascular remodeling and the observed propensity of atherosclerosis formation around bifurcations and areas of low shear stress.

Glomerular barrier function following conversion from cyclosporine to azathioprine in renal transplant recipients.

The renal side effects are the major limitation of the use of cyclosporine in clinical transplantation. We studied the reversibility of changes in renal hemodynamics and glomerular barrier function in 17 patients with moderately impaired renal function at least 1 year after kidney transplantation. All patients were studied both during cyclosporine treatment and 3 months after conversion to azathioprine. During azathioprine treatment both glomerular filtration rate and effective renal plasma flow increased significantly (from 44.3 +/- 4.2 mL/min to 63.5 +/- 5.4 mL/min and from 192 +/- 12.8 mL/min to 260 +/- 14.6 mL/min, respectively). Despite the marked changes in renal hemodynamics, no significant changes were observed in the fractional clearances of uncharged dextrans. When calculating the characteristics of the filtration barrier, we observed a trend toward an increase in the ultrafiltration coefficient (Kt). This trend was abolished when an increase in net filtration pressure (delta P) was assumed to result from reduced prerenal vasoconstriction. We conclude that despite marked improvement of renal perfusion and glomerular filtration, conversion from cyclosporine to azathioprine did not significantly alter the permeability characteristics of the glomerular filtration barrier in renal transplant recipients with moderately reduced renal function. Improvement in renal function following conversion could result from an increase in either Kf or delta P. Since renal plasma flow was increased significantly, the observed improvement in glomerular filtration rate is likely to be, at least in part, due to an increase in glomerular capillary plasma flow.

Charge selectivity of the glomerular filtration barrier in healthy and nephrotic humans.

We used dextran sulfate (DS) to evaluate barrier charge selectivity in 11 nonproteinuric subjects and in 11 patients with the nephrotic syndrome due to either membranous nephropathy or minimal change nephropathy. The 3H-DS preparation spanned a molecular radius interval of 10-24 A and exhibited size-dependent protein binding in vitro. Urine and ultrafiltrates of plasma were separated by size into narrow fractions using gel permeation chromatography. The sieving coefficient (theta) for ultrafilterable DS of 15A radius averaged 0.68 +/- 0.03 in nonproteinuric vs. 0.95 +/- 0.05 in nephrotic subjects (P < 0.001). Uncharged dextrans of broad size distribution were used to evaluate barrier size-selectivity in separate groups of nonproteinuric subjects (n = 19) and nephrotic patients with either minimal change (n = 20) or membranous nephropathy (n = 27). The value of theta for an uncharged dextran of similarly small radius (approximately 18 A) was significantly larger than that observed for DS in nonproteinuric subjects, but was similar in nephrotic individuals. Further, impaired barrier size-selectivity, as assessed by the sieving profile for uncharged dextrans (18-60 A radius), failed to account fully for the observed level of albuminuria in almost half of the patients with either minimal change (9/20) or membranous nephropathy (12/27). Together these findings suggest that the human glomerular capillary wall normally provides an electrostatic barrier to filtration of negatively charged macromolecules such as albumin, and that impairment of this electrostatic barrier contributes to the magnitude of albuminuria in the nephrotic syndrome.

Investigation of the ability of several naturally occurring and synthetic polyanions to bind to and potentiate the biological activity of acidic fibroblast growth factor

The ability of several animal, plant, and bacterial derived polyanions (PAs) as well as synthetic PAs to compete with heparin for the binding of acidic fibroblast growth factor (aFGF) was correlated with their ability to potentiate the mitogenic and neurotrophic actions of this factor. Dextran sulphate, kappa-carrageenan, pentosan sulphate, polyanethole sulfonate, heparin, and fucoidin competed for the heparin binding site on aFGF at relatively low concentrations (< 50 micrograms/ml). lambda-carrageenan, iota-carrageenan, and polyvinyl sulphate exhibited lower affinity for aFGF, whereas hyaluronic acid, dermatan sulphate, chondroitin-6-sulphate, chondroitin-4-sulphate, and uncharged dextran displayed very low or no demonstrable affinity. Potentiation of the mitogenic action of aFGF for Balb/c 3T3 fibroblasts tended to be in general agreement with the aFGF binding affinity of the PAs. However, polyanethole sulfonate, the carrageenans, polyvinyl sulphate, fucoidin, and pentosan sulphate exerted a mitogenic action on the 3T3 cells that was independent of, and in addition to, the ability of these GAGs to potentiate the action of aFGF. The ability to potentiate the neurotrophic action of aFGF for E8 chick ciliary neurons was a general property of those PA with low or no activity in the mitogen assay. Thus hyaluronic acid, dermatan sulphate, chondroitin-4-sulphate, chondroitin-6-sulphate, and even unchanged dextran all potentiated aFGF induced neuronal survival. The differential effects of these PA in potentiating the biological activities of aFGF are discussed in relation to their ability to compete for the heparin-binding site of aFGF.

Slope of serial glomerular filtration rate and the progression of diabetic glomerular disease.

Glomerular function was evaluated longitudinally over a 24- to 48-month period in 18 patients with diabetic glomerular disease (DGD) manifested by proteinuria. GFR was determined by iothalamate clearance at 4-month intervals. The patients were divided into two groups: Group 1 (N = 9) had subnephrotic proteinuria and an initially normal GFR of 91 +/- 8 mL/min. Group 2 (N = 9) had nephrotic-range proteinuria, and initial GFR was reduced to 53 +/- 5 mL/min. Serial GFR fluctuated over time in Group 1, but no trend towards hypofiltration was evident. In contrast, GFR declined linearly in Group 2 at 1.1 +/- 0.3 mL/min per month. The transglomerular sieving of uncharged dextrans of graded size was analyzed and initially revealed a uniform reduction in glomerular pore density and an enhancement of shuntlike pores. Pore density was initially reduced by 80% and declined further after 24 months in nephrotic Group 2; corresponding pore density in subnephrotic Group 1 was reduced by half but remained constant. Renal biopsy of four members of Group 1 revealed a 22% prevalence of global glomerulosclerosis. Remaining open glomeruli exhibited hypertrophy, excessive extracellular matrix, and deformation of epithelial podocytes. The latter abnormality appeared to be the predominant determinant of lowered ultrafiltration capacity. It was inferred that trials of therapy to attenuate the progression of DGD should be initiated at a functional level similar to that in subnephrotic Group 1. Because GFR is unlikely to decline over a 2- to 4-yr period, it is suggested that such trials be extended for longer periods. Alternatively, morphometric analysis of serial renal biopsies may shorten the time needed to demonstrate effective renoprotection in DGD.

Effects of cyclosporin A on glomerular barrier function in the nephrotic syndrome.

1. To elucidate the mechanisms by which cyclosporin A diminishes proteinuria, we studied 20 patients with severe nephrotic syndrome. Biopsy-established pathologies included minimal change disease (n = 5), membranous glomerulopathy (n = 6), membranoproliferative glomerulonephritis (n = 5) and focal segmental glomerulosclerosis (n = 4). Before, at the end of a 90 day course of cyclosporin A, and finally 1 month after stopping cyclosporin A we determined 24 h protein excretion. Measurements of glomerular filtration rate, effective renal plasma flow, fractional clearance rates of albumin and immunoglobulins with different charges and the transglomerular sieving of uncharged dextrans of broad size distribution were used to study the effects of cyclosporin A on renal perfusion and the glomerular filtration barrier. The findings were analysed with a theoretical model of solute transport. 2. Among the different forms of glomerulopathy the response to low-dose cyclosporin A (trough levels 32.0-36.9 ng/ml) varied markedly. In minimal change disease, proteinuria decreased from 9.5 +/- 3.1 to 1.3 +/- 0.2 g/24 h (mean +/- SEM, P less than 0.01). This response was due to restoration of the charge selectivity of the glomerular barrier. The depressed value of the glomerular permeability coefficient also returned to normal. Glomerular filtration rate, effective renal plasma flow and renal vascular resistance did not change. Proteinuria returned after stopping cyclosporin A, although it did not reach pretreatment levels. In membranous glomerulopathy, proteinuria fell from 9.9 +/- 1.5 to 1.8 +/- 0.3 g/24 h (P less than 0.01). Changes in protein excretion and dextran sieving were compatible with an increase in glomerular permselectivity and a decrease in filtrate flow through the 'shunt' pathway. Glomerular filtration rate was maintained, although effective renal plasma flow fell significantly. Proteinuria relapsed after stopping cyclosporin A. In membranoproliferative glomerulonephritis and focal segmental glomerulosclerosis proteinuria did not respond to cyclosporin A, although cyclosporin A exerted important haemodynamic effects. 3. In minimal change disease and membranous glomerulopathy cyclosporin A exerts its beneficial effects on proteinuria through changes in the properties of the glomerular barrier, resulting in increased charge and size selectivity, respectively.

Glomerular size-selectivity and microalbuminuria in early diabetic glomerular disease

Sieving coefficients of uncharged dextrans of graded size (radii 30 to 60 A) were used to characterize barrier size-selectivity in nonazotemic diabetic humans with microalbuminuria (Group 1, N = 11) or macroalbuminuria (Group 2, N = 21). Compared to a non-diabetic control group (N = 21) the low radius end of the sieving profile was depressed, whereas the high radius end was elevated in each diabetic group, more so in Group 2 than Group 1. A heteroporous membrane model revealed the major portion of the glomerular barrier to be perforated by restrictive pores of approximately 56 A radius in all three groups. However, in keeping with a parallel trend for GFR, the relative density of restrictive pores was control greater than Group 1 greater than Group 2. The remaining minor portion of the barrier was perforated by large, shunt-like pores, the relative prominence of which ranked Group 2 greater than Group 1 greater than control. Although the hypothetical, fractional clearance of macromolecules attributable to the shunt-like pores varied directly with fractional clearances of albumin and IgG, the progressive increment in the latter fractional protein clearances in the two diabetic groups was disproportionate. This raises the possibility that factors in addition to barrier size defects contribute to the development, magnitude and composition of proteinuria early in the course of diabetic glomerular disease.(ABSTRACT TRUNCATED AT 250 WORDS)

Angiotensin converting enzyme inhibition improves glomerular size-selectivity in IgA nephropathy

Clearances of uncharged dextrans of broad size distribution were used to evaluate the effects of a 30 day course of enalapril on glomerular barrier function in 10 patients with IgA nephropathy and proteinuria (from 1.4 to 5.6 g/day). Dextran clearance experiments were repeated three times: before enalapril therapy, after 30 days of enalapril and again 30 days after enalapril withdrawal. GFR, but not RPF, was significantly reduced by enalapril (basal 38.3 +/- 11.9, enalapril 30.2 +/- 12.6 ml/min/1.73 m2) and returned to basal values after enalapril withdrawal. Urinary protein excretion and fractional clearance of albumin were both significantly reduced by enalapril (basal 2.3 +/- 1.1 g/day and 102 +/- 90 x 10(-5), enalapril 1.2 +/- 0.6 g/day and 51 +/- 23 x 10(-5), respectively) and returned to basal values after enalapril withdrawal. Transglomerular passage of large dextrans (radii 54 to 62 A), but not of lower size (26 to 42 A) were significantly lowered by enalapril. When enalapril was withdrawn the dextran-sieving profile returned comparable to the baseline levels. A theoretical analysis of dextran-sieving profiles indicated that enalapril lowered the radius of largest membrane pores. This effect was independent from glomerular hemodynamic changes. We conclude that angiotensin converting enzyme inhibitors (CEI) in humans with IgA nephropathy reduces urinary protein excretion by a primary action on the intrinsic glomerular membrane properties enhancing barrier size-selective function. The hypofiltration associated with enalapril therapy in these patients, which was eliminated by its withdrawal, has to be taken into account as a possible undesired effect of CEI in long-term treatment.

Glomerular dysfunction in nephrotic humans with minimal changes or focal glomerulosclerosis

Fractional clearances (theta) of uncharged dextrans (radii 28-60 A) were used to characterize glomerular dysfunction in 34 nephrotic humans with either minimal-change nephropathy (MCN) or focal, segmental glomerulosclerosis (FSGS). A theoretical analysis of theta of dextran with a heteroporous membrane model revealed a similar alteration, more marked in FSGS than MCN. The number of restrictive pores perforating the major membrane component was reduced in parallel with the prevailing glomerular filtration rate (GFR). Parallel shuntlike pores in the remaining membrane component were more prominent, pointing to impaired size selectivity. However, the theta of large (60 A) dextrans attributable to these shunts exceeded control in FSGS only, suggesting that coexistent impairment of charge selectivity contributed importantly to the proteinuria in MCN. Membrane properties returned toward normal when MCN remitted. Glomerular morphometry revealed the frequency of epithelial filtration slits to vary with the extent of membrane dysfunction. Despite offsetting hypertrophy of remnant glomeruli in FSGS, a loss of filtration surface due to sclerosis likely contributed to the more marked reductions in pore number and GFR observed in this disorder than in MCN.