Biomedical subjects

M L Zeidel

Publications and source records attributed to M L Zeidel.

At least 37 records · Page 2Linked to original sources

PubMed1998

Water and solute permeabilities of medullary thick ascending limb apical and basolateral membranes.

The medullary thick ascending limb (MTAL) reabsorbs solute without water and concentrates NH4+ in the interstitium without a favorable pH gradient, activities which require low water and NH3 permeabilities. The contributions of different apical and basolateral membrane structures to these low permeabilities are unclear. We isolated highly purified apical and basolateral MTAL plasma membranes and measured, by stopped-flow fluorometry, their permeabilities to water, urea, glycerol, protons, and NH3. Osmotic water permeability at 20 degrees C averaged 9.4 +/- 0.8 x 10(-4) cm/s for apical and 11.9 +/- 0.5 x 10(-4) cm/s for basolateral membranes. NH3 permeabilities at 20 degrees C averaged 0.0023 +/- 0.00035 and 0.0035 +/- 0.00080 cm/s for apical and basolateral membranes, respectively. These values are consistent with those obtained in isolated perfused tubules and can account for known aspects of MTAL function in vivo. Because the apical and basolateral membrane unit permeabilities are similar, the ability of the apical membrane to function as the site of barrier function arises from its very small surface area when compared with the highly redundant basolateral membrane.

Ammonia↗

PubMed1998

The cost of caring for end-stage kidney disease patients: an analysis based on hospital financial transaction records.

The costs of care for end-stage renal disease patients continue to rise because of increased numbers of patients. Efforts to contain these costs have focused on the development of capitated payment schemes, in which all costs for the care of these patients are covered in a single payment. To determine the effect of a capitated reimbursement scheme on care of dialysis patients (both hemodialysis [HD] and peritoneal dialysis [PD]), complete financial records (all reimbursements for inpatient and outpatient care, as well as physician collections) of dialysis patients at a single medical center over 1 year were analyzed. For the period from July 1994 to July 1995, annualized cost per dialysis patient-year averaged $63,340, or 9.8% higher than the corrected estimate from the U.S. Renal Data Service (USRDS; $57,660). The "most expensive" 25% of patients engendered 44 to 48% of the total costs, and inpatient costs accounted for 37 to 40% of total costs. Nearly half of the inpatient costs resulted from only two categories (room charges and inpatient dialysis), whereas other categories each made up a small fraction of the inpatient costs. PD patients were far less expensive to care for than HD patients, due to reduced hospital days and lower cost of outpatient dialysis. Care for a university-based dialysis population was only slightly more expensive than estimates predicted from the USRDS. These results validate the USRDS spending data and suggest that they can be used effectively for setting capitated rates. Efforts to control costs without sacrificing quality of care must center on reducing inpatient costs, particularly room charges and the cost of inpatient dialysis.

Accounts Payable and Receivable↗

PubMed1998

Recent advances in water transport.

Complex organisms regulate the osmolalities of their compartments by limiting water flow across some membranes and promoting rapid water flow across others via proteins called aquaporins. Barrier epithelia limit water flow by reducing the mobilities of fatty acid chains in their apical membranes, especially in the outer leaflets of these membranes. Aquaporins are 28 to 30 kDa, 6 membrane-spanning proteins that are expressed in a wide variety of organisms from bacteria to plants to mammals. The structural and biophysical data are summarized to develop our best understanding of water pore function. In addition, the regulation of trafficking of AQP 2 into and out of the apical membranes of collecting duct principal cells is described.

Aquaporin 2↗

PubMed1997

Functional analysis of nodulin 26, an aquaporin in soybean root nodule symbiosomes.

Upon infection of soybean roots, nitrogen-fixing bacteria become enclosed in a specific organelle known as the symbiosome. The symbiosome membrane (SM) is a selectively permeable barrier that controls metabolite flux between the plant cytosol and the symbiotic bacterium inside. Nodulin 26 (NOD 26), a member of the aquaporin (AQP) water channel family, is a major protein component of the SM. Expression of NOD 26 in Xenopus oocytes gave a mercury-sensitive increase in osmotic water permeability (Pf). To define the biophysical properties of NOD 26 water channels in their native membranes, symbiosomes were isolated from soybean root nodules and the SM separated as vesicles from the bacteria. Permeabilities were measured using stopped-flow fluorimetry in SM vesicles with entrapped carboxyfluorescein. Osmotic water permeability (Pf) of SM was high, with a value of 0.05 +/- 0.003 cm/s observed at 20 degrees C (mean +/- S.E.; n = 15). Water flow exhibited a low activation energy, was inhibited by HgCl2 (0.1 mM), and exhibited a unit conductance of 3.2 +/- 1.3 x 10(-15) cm3/s, a value 30-fold lower than that of AQP 1, the red blood cell water channel. Diffusive water permeability (Pd) was 0.0024 +/- 0.0002 cm/s, and the resulting Pf to Pd ratio was 18.3, indicating that water crosses the SM in single file fashion via the NOD 26 water channel. In addition to high water permeability, SM vesicles also show high mercury-sensitive permeability to glycerol and formamide, but not urea, suggesting that NOD 26 also fluxes these solutes. Overall, we conclude that NOD 26 acts as a water channel with a single channel conductance that is 30-fold lower than AQP 1. Because the solutes that permeate NOD 26 are far larger than water, and water appears to cross the channel via a single file pathway, solute flux across NOD 26 appears to occur by a pathway that is distinct from that for water.

Animals↗

PubMed1997

Interactions of ultrapure bovine hemoglobin with renal epithelial cells in vivo and in vitro.

Using an ultrapurified hemoglobin (Hb) solution, we investigated the physiological effects and cellular processing of Hb in rat kidneys and in cultured opossum kidney (OK) cells. Rats infused with < 5.0 g/kg Hb showed no change in baseline serum creatinine (SCr) values (0.58 +/- 0.05 mg/dl) over 48 h, whereas transient acute renal failure followed infusion of 7.5 g/kg Hb (SCr 3.4 +/- 1.02 mg/dl, P = 0.02). Histology of Hb-infused kidneys demonstrated tubular epithelial cell injury. Renal injury was not caused by volume or oncotic load, cardiovascular effect, or ATP depletion. After Hb infusion, heme oxygenase, the rate-limiting enzyme in Hb catabolism, was induced in an organ-specific fashion. Inhibiting heme oxygenase activity with cimetidine did not alter Hb renal injury. Using OK cells, we determined that renal epithelia process Hb by fluid-phase endocytosis. Proton permeability of fluorescein Hb endosomes was unaltered compared with fluorescein dextran controls, demonstrating that Hb does not alter endosomal membrane integrity. These data suggest that Hb renal injury in rats occurs following large doses of ultrapure Hb, does not alter early steps in Hb endosomal processing by renal epithelia, and involves a mechanism that is not heme oxygenase dependent.

Acute Kidney Injury↗

PubMed1997

Low permeabilities of MDCK cell monolayers: a model barrier epithelium.

Barrier epithelia such as the renal collecting duct (in the absence of antidiuretic hormone) and thick ascending limb, as well as the stomach and mammalian bladder, exhibit extremely low permeabilities to water and small nonelectrolytes. A cell culture model of such epithelia is needed to determine how the structure of barrier apical membranes reduce permeability and how such membranes may be generated and maintained. In the present studies, the transepithelial electrical resistance and isotopic water and urea fluxes were measured for Madin-Darby canine kidney (MDCK) type I and type II cells, as well as type I cells expressing the mucin protein, MUC1, in their apical membranes. Although earlier studies had found the unstirred layer effects too great to permit measurement of transepithelial permeabilities, use of ultrathin semipermeable supports in this study overcame this difficulty. Apical membrane diffusive water permeabilities were 1.8 +/- 0.4 x 10(-4) cm/s and 3.5 +/- 0.5 x 10(-4) cm/s in MDCK type I and type II cells, respectively, at 20 degrees C. Urea permeability in type I cells at the same temperature was 6.0 +/- 0.9 x 10(-6) cm/s. These values resemble those of other barrier epithelial apical membranes, either isolated or in intact epithelia, and the water permeability values are far below those of other epithelial cells in culture. Transfection of MDCK type I cells with the major human urinary epithelial mucin, MUC1, led to abundant expression of the fully glycosylated form of the protein on immunoblots, and flow cytometry revealed that virtually all the cells expressed the protein. However, MUC1 had no effect on water or urea permeabilities. In conclusion, MDCK cells grown on semipermeable supports form a model barrier epithelium. Abundant expression of mucins does not alter the permeability properties of these cells.

Animals↗

PubMed1996

Effects of submicellar bile salt concentrations on biological membrane permeability to low molecular weight non-ionic solutes.

Bile salts have been hypothesized to mediate cytotoxicity by increasing membrane permeability to aqueous solutes. We examined whether submicellar bile salt concentrations affect model and native membrane permeability to small uncharged molecules such as water, urea, and ammonia. Osmotic water permeability (Pf) and urea permeability were measured in large unilamellar vesicles composed with egg yolk phosphatidylcholine (EYPC) +/- cholesterol (Ch) or rat liver microsomal membranes by monitoring self-quenching of entrapped carboxyfluorescein (CF). Ammonia permeability was determined utilizing the pH dependence of CF fluorescence. Submicellar bile salt concentrations did not significantly alter Pf of EYPC +/- Ch or rat liver microsomal membranes. At taurodeoxycholate (TDC) or tauroursodeoxycholate concentrations approaching those that solubilized membrane lipids, CF leakage occurred from vesicles, but Pf remained unchanged. Higher bile salt concentrations (0.5-2 mM TDC) did not alter Pf of equimolar EYPC/Ch membranes. The activation energy for transmembrane water flux was unchanged (12.1 +/- 1.2 kcal/mol for EYPC) despite the presence of bile salts in one or both membrane hemileaflets, suggesting strongly that bile salts do not form transmembrane pores that facilitate water flux. Furthermore, submicellar bile salt concentrations did not increase membrane permeability to urea or ammonia. We conclude that at submicellar concentrations, bile salts do not form nonselective convective channels that facilitate transmembrane transport of small uncharged molecules. These results suggest that bile salt-mediated transport of specific substrates, rather than nonselective enhancement of membrane permeability, underlies bile salt cytotoxicity for enterocytes and hepatocytes.

Ammonia↗

PubMed1996

Individual leaflets of a membrane bilayer can independently regulate permeability.

Water rapidly crosses most membranes, but only slowly crosses apical membranes of barrier epithelia such as bladder and kidney collecting duct, a feature essential to barrier function. How apical membrane structure reduces permeabilities remains unclear. Cell plasma membranes contain two leaflets of distinct lipid composition; the role of this bilayer asymmetry in membrane permeability is unclear. To determine how asymmetry of leaflet composition affects membrane permeability, effects on bilayer permeation of reducing single leaflet permeability were determined using two approaches: formation of asymmetric bilayers in an Ussing chamber, with only one of two leaflets containing cholesterol sulfate, and stabilization of the external leaflet of unilamellar vesicles with praeseodymium (Pr3+). In both systems, permeability measurements showed that each leaflet acts as an independent resistor of water permeation. These results show that a single bilayer leaflet can act as the barrier to permeation and provide direct evidence that segregation of lipids to create a low permeability of barrier epithelial apical membranes.

Cell Membrane Permeability↗

PubMed1996

Phosphorylation of aquaporin-2 does not alter the membrane water permeability of rat papillary water channel-containing vesicles.

Antidiuretic hormone modulates the water permeability (Pf) of epithelial cells in the rat kidney by vesicle-mediated insertion and removal of the aquaporin-2 (AQP-2) water channel. AQP-2 possesses a single consensus cAMP-dependent protein kinase A (PKA) phosphorylation site (Ser-256) hypothesized to regulate channel Pf(Kuwahara, M., Fushimi, K., Terada, Y., Bai, L., Sasaki, S., and Marumo, F. (1995) J. Biol. Chem. 270, 10384-10387). To test whether PKA phosphorylation of AQP-2 alters channel Pf, we compared the Pf values of purified AQP-2 endosomes after incubation with either PKA or alkaline phosphatase. Studies using [gamma-32P]ATP reveal that AQP-2 endosomes contain endogenous PKA and phosphatase activities that add and remove 32P label from AQP-2. However, the Pf (0.16 +/- 0.06 cm/s) of endosomes containing phosphorylated AQP-2 (0.7 +/- 0. 3 mol of PO4/mol of protein) is not significantly different from the same AQP-2 endosomes where 95 +/- 8% of the phosphate has been removed (Pf 0.14 +/- 0.06 cm/s). These data do not support a role for PKA phosphorylation in alteration of AQP-2's Pf. Instead, AQP-2 phosphorylation by PKA may modulate AQP-2's distribution between plasma membrane and intracellular vesicle compartments.

Adenosine Triphosphate↗

PubMed1996

Functional analysis of aquaporin-1 deficient red cells. The Colton-null phenotype.

The aquaporin-1 (AQP1) water transport protein contains a polymorphism corresponding to the Colton red blood cell antigens. To define the fraction of membrane water permeability mediated by AQP1, red cells were obtained from human kindreds with the rare Colton-null phenotype. Homozygosity or heterozygosity for deletion of exon I in AQP1 correlated with total or partial deficiency of AQP1 protein. Homozygote red cell morphology appeared normal, but clinical laboratory studies revealed slightly reduced red cell life span in vivo; deformability studies revealed a slight reduction in membrane surface area. Diffusional water permeability (Pd) was measured under isotonic conditions by pulsed field gradient NMR. Osmotic water permeability (Pf) was measured by change in light scattering after rapid exposure of red cells to increased extracellular osmolality. AQP1 contributes approximately 64% (Pd = 1.5 x 10(-3) cm/s) of the total diffusional water permeability pathway, and lipid permeation apparently comprises approximately 23%. In contrast, AQP1 contributes > 85% (Pf = 19 x 10(-3) cm/s) of the total osmotic water permeability pathway, and lipid permeation apparently comprises only approximately 10%. The ratio of AQP1-mediated Pf to Pd predicts the length of the aqueous pore to be 36 A.

Aquaporin 1↗

PubMed1996

Chronic regulation of transepithelial Na+ transport by the rate of apical Na+ entry.

In several settings in vivo, prolonged inhibition of apical Na+ entry reduces and prolonged stimulation of apical entry enhances the ability of renal epithelial cells to reabsorb Na+, an important feature of the load-dependent regulation of renal tubular Na+ transport. To model this load dependency, apical Na+ entry was inhibited or stimulated for 18 h in A6 cells and vectorial transport was measured as short-circuit current (Isc) across monolayers on filter-bottom structures. Basal amiloride-sensitive Isc represents the activity of apical Na+ channels, whereas Isc after permeabilization of the apical membrane to cations with nystatin represents maximal activity of the basolateral Na(+)-K(+)-ATPase. Chronic inhibition of apical Na+ entry by 18-h apical exposure to amiloride or replacement of apical Na+ with tetramethylammonium (TMA+), followed by washing and restoration of normal apical medium, revealed a persistent decrease in Isc that remained despite exposure to nystatin. Both basal and nystatin-stimulated Isc recovered progressively after restoration of normal apical medium. In contrast, chronic stimulation of apical Na+ entry by short circuiting the epithelium increased Isc in the absence and presence of nystatin, indicating upregulation of both apical Na+ channels and basolateral Na(+)-K(+)-ATPase. Basolateral equilibrium [3H]ouabain binding was reduced to 67 +/- 5% in TMA+ vs. control cells, whereas values in 18-h short-circuited cells increased by 42 +/- 19%. The results demonstrate that load dependency of tubular Na+ transport can be modeled in vitro and indicate that the regulation of Na(+)-K(+)-ATPase observed in these studies occurs in part by changes in the density of functional transporter proteins within the basolateral membrane.

Amiloride↗

PubMed1996

Phosphodiesterases (PDEs) hydrolyze the 3' phosphoester bond of the purine 3',5'-cyclic monophosphates, cAMP and cGMP.

Resistance to the natriuretic action of atrial natriuretic peptide (ANP) is a hallmark of states of pathological sodium retention including congestive heart failure, cirrhosis of the liver, and nephrotic syndrome. A variety of mechanisms including reduced delivery of filtrate to ANP-sensitive sites in the inner medullary collecting duct and diminished receptor density in this tubular segment have been offered to account for this resistance. Recent studies in experimental nephrotic syndrome and in liver disease produced by ligation of the common bile duct in rats suggest that increased activity of cyclic guanosine 3',5'-monophosphate (cGMP) phosphodiesterase may be an important mediator of renal resistance to ANP. Such increased enzyme activity rapidly catabolizes the second messenger cGMP, normally formed when ANP interacts with its biologically active natriuretic peptide. A receptors, thereby leading to blunted ANP responsiveness. This increased phosphodiesterase activity offers a novel approach to the management of clinical conditions associated with sodium retention and edema formation.

Animals↗

PubMed1996

Low permeabilities of apical membranes of barrier epithelia: what makes watertight membranes watertight?

Several epithelia, including those of collecting duct (in the absence of antidiuretic hormone), thick ascending limb of Henle, and mammalian bladder exhibit extremely low permeabilities for water, small nonelectrolytes like urea, and protons. This brief review describes how apical membrane structure might account for these low permeabilities by incorporating recent biophysical and molecular modeling results.

Animals↗

PubMed1996

Permeability properties of the intact mammalian bladder epithelium.

Because the mammalian bladder must store urine of composition which differs markedly from that of plasma for prolonged periods, the bladder permeability barrier must maintain extremely low permeabilities to substances which normally cross membranes relatively rapidly, such as water, protons, and small nonelectrolytes like urea and ammonia. In the present studies, permeabilities of the apical membrane of dissected rabbit bladder epithelium to water, urea, ammonia, and protons were measured in Ussing chambers and averaged (in cm/s) for water, 5.15 +/- 0.43 x 10(-5); for urea, 4.51 +/- 0.67 x 10(-6); for ammonia, 5.14 +/- 0.62 x 10(-4); and for protons, 2.98 +/- 1.87 x 10(-3), respectively. These permeability values are exceptionally low and are expected to result in minimal to no leakage of these normally permeable substances across the epithelium. Water permeabilities in intact whole rabbit bladders were indistinguishable from those obtained in the dissected epithelial preparation. Moreover, addition of nystatin to the apical solution of dissected epithelia rapidly increased water permeability in conjunction with loss of epithelial resistance. These results confirm that the apical membrane of the bladder epithelial cells represents the bladder permeability barrier. In addition, they establish a model system that will permit examination of how membrane structure reduces permeability and how epithelial injury compromises barrier function.

Ammonia↗

PubMed1995

The relationship between membrane fluidity and permeabilities to water, solutes, ammonia, and protons.

Several barrier epithelia such as renal collecting duct, urinary bladder, and gastric mucosa maintain high osmotic pH and solute gradients between body compartments and the blood by means of apical membranes of exceptionally low permeabilities. Although the mechanisms underlying these low permeabilities have been only poorly defined, low fluidity of the apical membrane has been postulated. The solubility diffusion model predicts that lower membrane fluidity will reduce permeability by reducing the ability of permeant molecules to diffuse through the lipid bilayer. However, little data compare membrane fluidity with permeability properties, and it is unclear whether fluidity determines permeability to all, or only some substances. We therefore studied the permeabilities of a series of artificial large unilamellar vesicles (LUV) of eight different compositions, exhibiting a range of fluidities encountered in biological membranes. Cholesterol and sphingomyelin content and acyl chain saturation were varied to create a range of fluidities. LUV anisotropy was measured as steady state fluorescence polarization of the lipophilic probe DPH. LUV permeabilities were determined by monitoring concentration-dependent or pH-sensitive quenching of entrapped carboxyfluorescein on a stopped-flow fluorimeter. The relation between DPH anisotropy and permeability to water, urea, acetamide, and NH3 was well fit in each instance by single exponential functions (r > 0.96), with lower fluidity corresponding to lower permeability. By contrast, proton permeability correlated only weakly with fluidity. We conclude that membrane fluidity determines permeability to most nonionic substances and that transmembrane proton flux occurs in a manner distinct from flux of other substances.

Acetamides↗

PubMed1995

Regulation of collecting duct Na+ reabsorption by ANP 31-67.

1. The bulk of studies of the actions of atrial natriuretic peptides (ANP) have focussed on the carboxyterminal derivative (ANP 99-126) of the prohormone (ANP 1-126), but recent evidence indicates that an additional peptide derived from ANP 1-126, namely, ANP 31-67 also circulates, and has natriuretic actions. 2. The effects of ANP 31-67 on inner medullary collecting duct (IMCD) Na+ transport have been examined in freshly prepared suspensions of rabbit IMCD cells. Like ANP 99-126, ANP 31-67 reduces Na+ transport in these cells. 3. However, unlike ANP 99-126, this effect is not mediated by cGMP, and does not result from inhibition of apical Na+ channels. Rather, ANP 31-67 inhibits basolateral Na/K-ATPase, probably via the stimulation of PGE2 synthesis. 4. These results are discussed in the context of other natriuretic substances (interleukin 1 and endothelin), which also inhibit Na+ reabsorption by PGE2-mediated inhibition of Na/K-ATPase.

Animals↗

PubMed1994

Mutations in aquaporin-1 in phenotypically normal humans without functional CHIP water channels.

The gene aquaporin-1 encodes channel-forming integral protein (CHIP), a member of a large family of water transporters found throughout nature. Three rare individuals were identified who do not express CHIP-associated Colton blood group antigens and whose red cells exhibit low osmotic water permeabilities. Genomic DNA analyses demonstrated that two individuals were homozygous for different nonsense mutations (exon deletion or frameshift), and the third had a missense mutation encoding a nonfunctioning CHIP molecule. Surprisingly, none of the three suffers any apparent clinical consequence, which raises questions about the physiological importance of CHIP and implies that other mechanisms may compensate for its absence.

Animals↗

PubMed1994

Membrane lipid composition and vesicle size modulate bilirubin intermembrane transfer. Evidence for membrane-directed trafficking of bilirubin in the hepatocyte.

To characterize the mechanisms underlying intracellular bilirubin transport, stopped-flow fluorometry was utilized to study the effects of membrane vesicle size and lipid composition on the kinetics of unconjugated bilirubin movement between model and native hepatocyte membranes. Bilirubin transfer rates declined asymptotically with increasing donor vesicle diameter, due primarily to a 1.4 kcal.mol-1 decrease in the entropy of activation for the larger vesicles. The incorporation of phosphatidylethanolamine and phosphatidylserine significantly enhanced the dissociation of bilirubin from phosphatidylcholine vesicles. Cholesterol induced a biphasic effect on the transfer rate constant; an initial decrease in rate from 248 to 217 s-1 associated with cholesterol:phospholipid ratios up to 20% was followed by a dramatic rise to 312 s-1 as the cholesterol concentration was increased to 70 mol %. The bilirubin dissociation rate from isolated rat liver endoplasmic reticulum (9.1 s-1) was significantly slower than for both basolateral and canalicular plasma membranes, which exhibited rate constants of 11.7 and 25.8 s-1, respectively. Collectively, these data suggest that the cholesterol: phospholipid ratio is the principal determinant of bilirubin dissociation from membranes. We postulate that the inherent cellular membrane cholesterol gradient in the hepatocyte creates a directed flux of bilirubin from the plasma membrane to teh endoplasmic reticulum and represents a potential driving force for intracellular bilirubin transport.

Animals↗