Transport in liquid-filled pores of molecular dimensions plays an important role in membrane separations, in various forms of chromatography, and in catalysis, to name a few examples. A frequent observation is that if the pore dimensions are of the same order as those of a solute molecule, the apparent diffusion coefficient of that solute is much lower than in bulk solution. Likewise, rates of convective transport of such solutes are generally lower than the product of bulk concentration and volume flow rate. Thus, solute transport is typically "hindered" or restricted. A key objective of research on hindered transport is to be able to predict the applicable transport coefficients from such fundamental information as the size, shape, and electrical charge of the solutes and pores. The present status of this research is reviewed.

This review focuses on the intricate properties of the glomerular barrier. Other reviews have focused on podocyte biology, mesangial cells, and the glomerular basement membrane (GBM). However, since all components of the glomerular membrane are important for its function, proteinuria will occur regardless of which layer is affected by disease. We review the properties of endothelial cells and their surface layer, the GBM, and podocytes, discuss various methods of studying glomerular permeability, and analyze data concerning the restriction of solutes by size, charge, and shape. We also review the physical principles of transport across biological or artificial membranes and various theoretical models used to predict the fluxes of solutes and water. The glomerular barrier is highly size and charge selective, in qualitative agreement with the classical studies performed 30 years ago. The small amounts of albumin filtered will be reabsorbed by the megalin-cubulin complex and degraded by the proximal tubular cells. At present, there is no unequivocal evidence for reuptake of intact albumin from urine. The cellular components are the key players in restricting solute transport, while the GBM is responsible for most of the resistance to water flow across the glomerular barrier.

The molecular weight cutoff for glomerular filtration is thought to be 30-50 kDa. Here we report rapid and efficient filtration of molecules 10-20 times that mass and a model for the mechanism of this filtration. We conducted multimodal imaging studies in mice to investigate renal clearance of a single-walled carbon nanotube (SWCNT) construct covalently appended with ligands allowing simultaneous dynamic positron emission tomography, near-infrared fluorescence imaging, and microscopy. These SWCNTs have a length distribution ranging from 100 to 500 nm. The average length was determined to be 200-300 nm, which would yield a functionalized construct with a molecular weight of ∼350-500 kDa. The construct was rapidly (t 1/2 ∼ 6 min) renally cleared intact by glomerular filtration, with partial tubular reabsorption and transient translocation into the proximal tubular cell nuclei. Directional absorption was confirmed in vitro using polarized renal cells. Active secretion via transporters was not involved. Mathematical modeling of the rotational diffusivity showed the tendency of flow to orient SWCNTs of this size to allow clearance via the glomerular pores. Surprisingly, these results raise questions about the rules for renal filtration, given that these large molecules (with aspect ratios ranging from 100:1 to 500:1) were cleared similarly to small molecules. SWCNTs and other novel nanomaterials are being actively investigated for potential biomedical applications, and these observations-that high aspect ratio as well as large molecular size have an impact on glomerular filtration-will allow the design of novel nanoscalebased therapeutics with unusual pharmacologic characteristics.multimodal imaging | nanotechnology | renal C arbon nanotubes (CNTs) have interesting properties and have been proposed as novel components of drugs and devices in pharmaceutical and biomedical applications (1). CNTs have unique intrinsic physical, chemical, electronic, thermal, and optical properties and can be chemically modified (with, e.g., targeting ligands, magnetic, radioactive, fluorescent, and chemotherapeutic moieties) to exhibit additional extrinsic properties (2-4). Pharmacokinetic (PK) studies of covalently functionalized single-wall CNTs (SWCNTs) (5-9) and multiwall (MWCNTs) (10-12) have reported a short blood compartment half-life (1-3 h) and limited tissue (kidneys, liver, and spleen) accumulation and renal excretion. Clearance via renal mechanisms is significant (13), because it provides the opportunity for the host to eliminate SWCNTs, allowing potential therapeutic and diagnostic applications in vivo. The elimination of noncovalently modified SWCNTs has been reported to favor the hepatobiliary route, with evidence of a minor role for the renal route (14).Renal clearance of solutes occurs through a combination of glomerular filtration, active tubular secretion, and passive tubular reabsorption (15). In previous work, we reported radioactivity in the renal cortex and in the urine within 1 h of administration of radiolabeled...

The diffusivities of uncharged macromolecules in gels (D) are typically lower than in free solution (D infinity), because of a combination of hydrodynamic and steric factors. To examine these factors, we measured D and D infinity for dilute solutions of several fluorescein-labeled macromolecules, using an image-based fluorescence recovery after photobleaching technique. Test macromolecules with Stokes-Einstein radii (rs) of 2.1-6.2 nm, including three globular proteins (bovine serum albumin, ovalbumin, lactalbumin) and four narrow fractions of Ficoll, were studied in agarose gels with agarose volume fractions (phi) of 0.038-0.073. The gels were characterized by measuring the hydraulic permeability of supported agarose membranes, allowing calculation of the Darcy permeability (kappa) for each gel sample. It was found that kappa, which is a measure of the intrinsic hydraulic conductance of the gel, decreased by an order of magnitude as phi was increased over the range indicated. The diffusivity ratio D/D infinity, which varied from 0.20 to 0.63, decreased with increases in rs or phi. Thus as expected, diffusional hindrances were the most severe for large macromolecules and/or relatively concentrated gels. According to a recently proposed theory for hindered diffusion through fibrous media, the diffusivity ratio is given by the product of a hydrodynamic factor (F) and a steric factor (S). The functional form is D/D infinity = F(rs/k1/2) S(f), where f = [(rs+rf)/rf]2 phi and rf is the fiber radius. Values of D/D infinity calculated from this effective medium theory, without use of adjustable parameters, were in much better agreement with the measured values than were predictions based on other approaches. The strengths and limitations of the effective medium theory for predicting diffusivities in gels are discussed.

It is well-known that solutes in liquid-filled pores of molecular dimensions have reduced diffusivities and are sieved during filtration. For solute molecules that are large enough to act as hydrodynamic particles, these phenomena can be explained by a combination of particle-wall hydrodynamic interactions and steric restrictions. Theoretical expressions that include those effects have been available for many years, but, even for spheres in pores of constant cross-section, certain hydrodynamic information has been lacking until recently. In particular, the local enhanced drag and local lag coefficient for off-axis positions had not been fully characterized, requiring that results for symmetrically positioned particles ("centerline approximations") be employed in predicting diffusional and convective hindrances. In this paper the current status of hindered transport theory is reviewed for neutral spheres in long cylindrical pores or slits, and it is shown that such approximations are no longer necessary. New expressions are presented for diffusive and convective hindrance factors that are properly averaged over the pore cross-section. The root-mean-square errors in the centerline approximations are 20% and 6%, respectively, for diffusive and convective hindrance factors in cylindrical pores; for slit pores the corresponding errors are 16% and 10%. Comparisons are made between the predictions and recent data obtained by tracking particle positions in microchannels.

Recent progress in relating the functional properties of the glomerular capillary wall to its unique structure is reviewed. The fenestrated endothelium, glomerular basement membrane (GBM), and epithelial filtration slits form a series arrangement in which the flow diverges as it enters the GBM from the fenestrae and converges again at the filtration slits. A hydrodynamic model that combines morphometric findings with water flow data in isolated GBM has predicted overall hydraulic permeabilities that are consistent with measurements in vivo. The resistance of the GBM to water flow, which accounts for roughly half that of the capillary wall, is strongly dependent on the extent to which the GBM surfaces are blocked by cells. The spatial frequency of filtration slits is predicted to be a very important determinant of the overall hydraulic permeability, in keeping with observations in several glomerular diseases in humans. Whereas the hydraulic resistances of the cell layers and GBM are additive, the overall sieving coefficient for a macromolecule (its concentration in Bowman's space divided by that in plasma) is the product of the sieving coefficients for the individual layers. Models for macromolecule filtration reveal that the individual sieving coefficients are influenced by one another and by the filtrate velocity, requiring great care in extrapolating in vitro observations to the living animal. The size selectivity of the glomerular capillary has been shown to be determined largely by the cellular layers, rather than the GBM. Controversial findings concerning glomerular charge selectivity are reviewed, and it is concluded that there is good evidence for a role of charge in restricting the transmural movement of albumin. Also discussed is an effect of albumin that has received little attention, namely, its tendency to increase the sieving coefficients of test macromolecules via steric interactions. Among the unresolved issues are the specific contributions of the endothelial glycocalyx and epithelial slit diaphragm to the overall hydraulic resistance and macromolecule selectivity and the nanostructural basis for the observed permeability properties of the GBM.

S-Nitrosothiols have generated considerable interest due to their ability to act as nitric oxide (NO) donors and due to their possible involvement in bioregulatory systems-e.g., NO transfer reactions. Elucidation of the reaction pathways involved in the modification of the thiol group by S-nitrosothiols is important for understanding the role of S-nitroso compounds in vivo. The modification of glutathione (GSH) in the presence of S-nitrosoglutathione (GSNO) was examined as a model reaction. Incubation of GSNO (1 mM) with GSH at various concentrations (1-10 mM) in phosphate buffer (pH 7.4) yielded oxidized glutathione, nitrite, nitrous oxide, and ammonia as end products. The product yields were dependent on the concentrations of GSH and oxygen. Transient signals corresponding to GSH conjugates, which increased by one mass unit when the reaction was carried out with 15 N-labeled GSNO, were identified by electrospray ionization mass spectrometry. When morpholine was present in the reaction system, N-nitrosomorpholine was formed. Increasing concentrations of either phosphate or GSH led to lower yields of N-nitrosomorpholine. The inhibitory effect of phosphate may be due to reaction with the nitrosating agent, nitrous anhydride (N 2 O 3 ), formed by oxidation of NO. This supports the release of NO during the reaction of GSNO with GSH. The products noted above account quantitatively for virtually all of the GSNO nitrogen consumed during the reaction, and it is now possible to construct a complete set of pathways for the complex transformations arising from GSNO ؉ GSH.S-Nitrosothiols, RSNO, with certain exceptions, are unstable in aqueous solution. For example, S-nitrosoglutathione (GSNO) undergoes decomposition over hours, whereas Snitrosocysteine has a half-life of less than 2 min. The initial step in the decomposition of RSNO is believed to be homolytic cleavage of the SON bond to give nitric oxide (NO) and a thiyl radical (1, 2). These compounds are involved in many bioregulatory functions, including vasodilation and inhibition of platelet aggregation. The existence of more stable transport forms of NO has been postulated in view of the short half-life of authentic NO in vivo (3). Low molecular weight thiols such as cysteine, glutathione (GSH), and penicillamine are prime candidates for such carrier molecules, and they can form S-nitrosothiols on reaction with oxides of nitrogen (4). It has been assumed that the biological effects of these compounds are due to the spontaneous release of NO; however, this hypothesis is not supported by currently available data (5-7).Although a few studies have been carried out in an attempt to determine the reaction products and to deduce the mechanism of the modification of the thiol group by S-nitrosothiols, the experiments were purely qualitative and no clear mechanistic picture has emerged (8,9). In this report, we describe the reaction of GSNO with GSH, a tripeptide with intracelluclar concentrations as high as 10 mM (10). It is involved in the cell's antioxidant defens...

An understanding of the rate of reaction of nitric oxide (NO) with oxygen in aqueous solutions is needed in assessing the various actions of NO in the body. A novel approach was developed for studying the kinetics of this reaction, which permitted simultaneous and continuous measurements of the concentrations of NO and the principal product, nitrite (NO2-). Nitric oxide was measured using a chemiluminescence detector, with continuous sampling achieved by diffusion of NO through a membrane fitted into the base of a small, stirred reactor. The results with various initial NO and O2 concentrations confirmed that the rate of reaction is second-order in NO and first-order in O2. The rate of reaction of NO was described by the expression 4k1 [NO]2[O2], where k1 was (2.1 +/- 0.4) x 10(6) M-2 s-1 at 23 degrees C and (2.4 +/- 0.3) x 10(6) M-2 s-1 at 37 degrees C. The value of k1 was the same at pH 4.9 and 7.4. The rate of formation of NO2- equaled the rate of reaction of NO (within experimental uncertainty of a few percent), and there was no detectable formation of nitrate (NO3-). This confirmed that NO and NO2- were the only NOx species present in significant amounts and supported the validity of pseudo-steady-state assumptions for NO2 and N2O3, which are intermediates in the conversion of NO to NO2-.(ABSTRACT TRUNCATED AT 250 WORDS)