PubMed Health⌕ Search

Biomedical subjects

L J Mandel

Publications and source records attributed to L J Mandel.

At least 73 records · Page 4Linked to original sources
PubMed1986

Multiple effects of presumed glutathione depletors on rabbit proximal tubules.

The role of glutathione (GSH) in the protection of normal renal function has been investigated using rabbit proximal tubules. Compounds known to deplete GSH in various biological systems by alkylation (via GSH S-transferases, phorone; 2-cyclohexen-1-one, CHX; diethyl maleate, DEM) or by inhibiting GSH synthesis (buthionine sulfoximine, BSO) were added to suspensions of proximal tubules and incubated for 60 min. BSO (1 or 5 mM) did not decrease GSH concentrations, O2 consumption, or cause lactate dehydrogenase release (LDH). Concentrations of CHX (2 mM) and phorone (10 mM) that decreased GSH concentrations also inhibited O2 consumption and caused LDH release. DEM (10 mM) did not significantly decrease GSH concentrations but did inhibit oxygen consumption and cause slight LDH release. Time-course studies using CHX (3 mM) showed that GSH levels and O2 consumption decreased as early as 15 min while LDH release did not occur until 60 min. These results show that: there may be a relationship between O2 consumption and GSH levels; agents that have been used historically to reduce GSH concentrations have other cytotoxic effects; and rabbit renal proximal tubules appear to be resistant to GSH depletion.

Animals↗
PubMed1986

In vitro studies simultaneously examining effect of oxacillin on uptake of radiolabeled streptomycin and on associated bacterial lethality in Staphylococcus aureus.

We studied the effects of various concentrations of oxacillin on streptomycin uptake and killing for several strains of Staphylococcus aureus. When streptomycin was present in concentrations below the MIC, addition of oxacillin at concentrations greater than or equal to the MIC was associated with both significantly increased aminoglycoside uptake and killing. In contrast, when streptomycin was present in concentrations above the MIC, no increase of streptomycin uptake was noted with the addition of oxacillin, and killing was no greater than what would have been expected with a simply additive effect. Similar studies in a strain of S. aureus selected for high-level streptomycin resistance also demonstrated increased streptomycin uptake in the presence of concentrations of oxacillin above the MIC; however, killing was no greater than that seen with oxacillin alone. These studies provide data which are potentially important in designing a rational approach to clinical use of combination antibiotic therapy.

Dose-Response Relationship, Drug↗
PubMed1986

Volume regulation by Amphiuma red blood cells: cytosolic free Ca and alkali metal-H exchange.

Osmotic swelling of Amphiuma red blood cells results in activation of electroneutral K-H exchange, whereas cell shrinkage activates an electroneutral Na-H exchange. These K-H and Na-H exchangers function to restore cell volume to normal after cell swelling and shrinkage, respectively. Our previous studies have suggested that Ca plays a role in volume-dependent activation of K-H exchange. In the present studies, intracellular free Ca levels were measured employing the Ca-sensitive extracellular dye arsenazo III and a previously described null-point method. Control values for intracellular free Ca averaged 0.46 +/- 0.15 microM. Cell shrinkage caused this value to decrease to 0.16 +/- 0.11 microM, whereas either cell swelling or addition of 5 microM A23187 resulted in saturation of intracellular Ca buffers, suggesting that both treatments caused an increase in intracellular free Ca. In the presence of 7 microM A23187, the rate of K-H exchange displayed a hyperbolic relationship as a function of extracellular Ca (Cao). The apparent half-maximal concentration for Cao (in the presence of 7 microM A23187) was 0.27 mM for osmotically swollen cells and 1.9 mM for cells in isotonic medium, suggesting that the Ca affinity of a modulating site is increased in swollen cells. Inhibitors of Ca-mediated processes, such as quinidine and the phenothiazines, inhibited K-H exchange. In contrast, the phenothiazines chlorpromazine and trifluoperazine stimulated Na-H exchange by osmotically shrunken cells. These results suggest that increases in intracellular free Ca are involved in stimulating K-H exchange while repressing Na-H exchange in Amphiuma red blood cells.

Acid-Base Equilibrium↗
PubMed1986

Amiloride analogues inhibit proximal tubule metabolism.

The effects of two amiloride analogues, ethylisopropylamiloride and methylisopropylamiloride, were tested on active Na+ transport and oxidative metabolism of a rabbit proximal renal tubule suspension. These two analogues have been recently reported to inhibit Na+-H+ antiport activity of various tissues with greater potency than amiloride. In proximal tubules, no effects were detected at concentrations less than 10(-4) M. At concentrations greater than 10(-4) M, both of these compounds inhibited the ouabain-sensitive O2 consumption (a direct measure of Na+ pump activity) with a greater potency than amiloride. Investigations into possible metabolic effects revealed that both amiloride analogues inhibited mitochondrial production of ATP at these concentrations, whereas amiloride did not directly affect metabolism. The amiloride analogues inhibited the nystatin-stimulated O2 consumption (which measures Na+-K+-ATPase activity in the intact cells) as well as the ATP content. These results suggest that the primary effects of these analogues on rabbit proximal tubules are to inhibit both the Na+-K+-ATPase and oxidative metabolism.

Adenine Nucleotides↗
PubMed1986

Effects of calcitonin on cytosolic Ca in a suspension of rabbit medullary thick ascending limb tubules.

A basal level of cytosolic free Ca (Caf) of 112 +/- 7 nM (n = 15) was measured in rabbit medullary thick ascending limb tubules that were loaded with the fluorescent Ca indicator quin 2. Calcitonin addition (10(-6) M) caused a greater than twofold increase in Caf. The KD for calcitonin stimulation of Caf was 8 X 10(-11) M. The rise in Caf had a half time of approximately 1.5 min. Neither parathyroid hormone (10(-6) M), dibutyryladenosine 3',5'-cyclic monophosphate (10(-4) M), nor forskolin (2 X 10(-5) M) had any significant effect on Caf levels, consistent with the rise in Caf being independent of an elevation in adenosine 3',5'-cyclic monophosphate. The effects of ouabain and the mitochondrial uncoupler, 1799, were also tested to obtain further information regarding the regulation of Caf. Ouabain (10(-4) M) caused no significant elevation in Caf; in contrast, 15 min of incubation with 1799 (10(-5) M) produced a twofold increase in Caf.

Aminoquinolines↗
PubMed1986

Substrate support of medullary thick ascending limb oxygen consumption.

The oxygen consumption of medullary thick ascending limb tubule suspensions was measured in the presence and absence of exogenous glucose. In the absence of exogenous glucose, the control oxygen consumption decreased 15%. Under identical conditions, the nystatin-stimulated oxygen consumption was inhibited 36%, indicating that oxidation of endogenous substrates could not meet the ATP demand of the fully stimulated Na+-K+-ATPase. Addition of inhibitors of fatty acid oxidation (bromooctanoate and tetradecylglycidic acid) further inhibited oxygen consumption, revealing that endogenous fats are oxidized in control and nystatin-stimulated states. Inhibition of endogenous carbohydrate (2-deoxy-D-glucose present) or amino acid (aminooxyacetate present) oxidation suppressed the nystatin response. In the presence of 10 mM glucose, only aminooxyacetate inhibited the nystatin-stimulated oxygen consumption, suggesting that the involvement of the malate-aspartate shuttle in redox balance during increased glycolytic flux. Addition of fatty acids or acetoacetate increased the control, nystatin-stimulated and uncoupler-stimulated oxygen consumption of tubules suspended in 10 mM glucose, indicating that these tubules oxidize exogenous fatty acids and ketones and oxidation of exogenous glucose alone may not meet all the energy demands of the tissue. Conversely, the addition of organic acids failed to enhance control oxygen consumption, possibly due to the absence of transport systems for these compounds in the medullary thick ascending limb tubules.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1986

Cellular toxicity of bromobenzene and bromobenzene metabolites to rabbit proximal tubules: the role and mechanism of 2-bromohydroquinone.

An in vitro model using a suspension of rabbit renal proximal tubules was developed to investigate the mechanism of nephrotoxicity of bromobenzene. Using oxygen consumption, glutathione concentrations and retention of lactate dehydrogenase activity as markers of toxicity, the rank order of potency was bromobenzene (5 mM) less than 2-bromophenol (2 mM) less than 3-, 4-bromophenol (1 mM) less than 2-bromohydroquinone (0.1 mM). These data support in vivo results and are consistent with the hypothesis that 2-bromohydroquinone or a metabolite thereof is responsible for bromobenzene-induced nephrotoxicity. Inhibitors of cytochrome P-450 and the cyclooxygenase and peroxidase components of prostaglandin H synthase did not protect the proximal tubules from 2-bromohydroquinone-induced toxicity, suggesting that these enzymes do not play a role in 2-bromohydroquinone bioactivation. There is a specific sequence of events in 2-bromohydroquinone-induced toxicity. Early events include decreased glutathione levels and inhibited mitochondrial respiration, whereas an increase in plasma membrane permeability is a late event.

Animals↗
PubMed1985

Intracellular compartmentation of glutathione in rabbit renal proximal tubules.

The intracellular compartmentation of glutathione in rabbit renal proximal tubules was determined using digitonin, the non-ionic detergent Lubrol PX, and rapid centrifugation. Glutathione was distributed between two pools within the proximal tubules. The mitochondrial pool was the largest, containing 72 percent of the cellular glutathione while the cytoplasmic pool contained the remaining 28 percent. These results are in marked contrast to rat hepatocytes in which 85 percent of the cellular glutathione is cytoplasmic and only 15 percent is mitochondrial (1).

Animals↗
PubMed1985

Phenamil: an irreversible inhibitor of sodium channels in the toad urinary bladder.

Several new amiloride analogues and two reported photoaffinity analogues were tested for irreversible inhibition of short-circuit current, Isc, in toad bladder. Bromoamiloride, a photoaffinity analogue, induced 40% irreversible inhibition at 500 microM after irradiation with ultraviolet light greater than or equal to 320 nm. Iodoamiloride caused no irreversible inhibition. Of the new analogues tested, only 3,5-diamino-6-chloro-N-[(phenylamino) amino-methylene] pyrazinecarboxamide, phenamil, irreversibly inhibited Isc at concentrations of 0.05 to 5 microM when added to the mucosal solution. Irreversible inhibition of Isc by phenamil may be attributed to specific blockage of the mucosal sodium channels, which depended on: 1) time of exposure; 2) mucosal pH; 3) mucosal sodium concentration. For example, 5 microM phenamil irreversibly inhibited Isc by 38% in 103 mM Na at pH 8.6 and nearly 75% in 30 mM Na at pH 6.4 after a 40-min exposure. Irreversible inhibition occurred in two phases with time constants of less than or equal to 10 min and approximately 140 min. Due to its irreversible nature, phenamil may be used to measure channel density.

Amiloride↗
PubMed1985

Role of glucose in corneal metabolism.

Glucose catabolism by glycolysis and the Krebs cycle was examined in the isolated rabbit cornea incubated with [6-14C]glucose. The production of [14C]lactate and 14CO2 from this substrate provided minimal values for the fluxes through these pathways since the tissue was in metabolic steady state but not isotopic steady state during the 40-min incubation. The specific activity of lactate under these conditions was one-third of that for [6-14C]glucose, and label dilution by exchange with unlabeled alanine was minimal, suggesting that glycogen degradation was primarily responsible for this dilution of label in the Embden-Meyerhof pathway. In addition, considerable label accumulation was found in glutamate and aspartate. Calculations revealed that these endogenous amino acid pools were not isotopically equilibrated after the incubation period, suggesting that they were responsible for the isotopic nonsteady state by exchange dilution through transaminase reactions with labeled intermediates. An estimate of glucose oxidation by the Krebs cycle, which was corrected for label dilution by exchange, indicated that glucose could account for most of the measured corneal oxygen consumption that was coupled to oxidative phosphorylation. A minor component of this respiration could not be accounted for by glucose or glycogen oxidation. Additional experiments suggested that endogenous fatty acid oxidation was probably also active under these conditions. Finally, reciprocal changes in plasma membrane Na+-K+-ATPase activity induced by ouabain and nystatin were found to concomitantly alter oxygen consumption rates and [14C]lactate production from [6-14C]glucose. These results demonstrated the capacity for regulating glycolysis and the Krebs cycle in response to changing energy demands in the cornea.

Amino Acids↗
PubMed1985

Intracellular respiratory dysfunction and cell injury in short-term anoxia of rabbit renal proximal tubules.

The effects of short-term anoxia and hypoxia were studied in a rabbit proximal renal tubule suspension in order to avoid the hemodynamic consequences of clamp-induced ischemia. The suspension was subjected to anoxia for 10-40 min and the effects on a number of cellular transport and respiratory parameters were monitored. Cellular respiration was measured upon addition of nystatin (Nys) to maximally stimulate Na pump activity. Mitochondrial respiration was measured in the tubules by addition of digitonin and ADP to obtain the state 3 respiratory rate. The release of lactate dehydrogenase (LDH) was measured as an index of plasma membrane damage. The cellular contents of K and Ca were also measured. Results show that 10 and 20 min of anoxia partially inhibited Nys-stimulated and mitochondrial respiration, and partially decreased the K contents, but all these effects were largely reversible after 20 min of reoxygenation. After 40 min of anoxia and 20 min of reoxygenation, all these variables remained irreversibly inhibited: Nys-stimulated respiration by 54%, mitochondrial respiration by 50%, K content by 42%, and LDH release was 40% of total. Ca content decreased slightly during anoxia, but increased up to fourfold during severe hypoxia; the excess Ca was released during the first 10 min of reoxygenation. The degree of respiratory impairment was identical during anoxia or hypoxia, suggesting that Ca accumulation was not associated with the impairment. Decreasing the extracellular Ca to 2.5 microM decreased LDH release significantly during anoxia, suggesting that plasma membrane damage during anoxia may be associated with increased intracellular free Ca. Addition of Mg-adenosine triphosphate during anoxia dramatically improved recovery of all the measured parameters after the anoxic period.

Adenosine Triphosphate↗
PubMed1984

Regulation of cytosolic free calcium in rabbit proximal renal tubules.

The relative role of various Ca2+ transport systems in the regulation of Ca2+ cytosolic free Ca2+ concentration was evaluated using rabbit renal proximal tubules. Intracellular compartmentation was evaluated through Ca2+ releases induced by carbonyl cyanide p-trifluoromethoxyphenylhydrazone (FCCP), A23187, and ruthenium red (RR) alone and in combination. In a Ca2+-free solution after 1 h of incubation, FCCP released 43 +/- 4%, A23187 54 +/- 3%, and RR 29 +/- 5% of total cellular Ca2+; in addition, RR inhibited the rate of FCCP-induced release, confirming its mitochondrial origin. In 1 mM Ca2+, the releases were 57 +/- 9%, 70 +/- 5%, and 34 +/- 10%, respectively. Therefore, the mitochondrial Ca2+ content is 20-60 nmol/mg of mitochondrial protein, sufficiently large to effectively buffer cell Ca2+. To evaluate the role of the plasma membrane Na:Ca exchanger, 10(-4) M ouabain was added and caused a slight decline in total cell Ca2+ content and no change in ionized Ca2+ measured by the null-point method, suggesting that the plasmalemmal Na+:Ca2+ exchanger does not play an important role in Ca2+ extrusion. Cytosolic free Ca2+ increased when 100 mM sodium was replaced with equimolar choline or tetramethylammonium. However, tetramethylammonium replacement released 55% of the mitochondrial Ca2+ content by increasing mitochondrial Ca2+ efflux without affecting the Ca2+ influx pathway. These results suggest that Na+ replacements in this tissue increase ionized Ca2+ by increasing mitochondrial Ca2+ efflux and not by inhibition of Na+:Ca2+ exchange at the plasma membrane.

Adenosine Triphosphate↗
PubMed1984

Glucose-dependent respiration in suspensions of rabbit cortical tubules.

The effects of glucose on cellular respiration were examined in suspensions of rabbit cortical tubules. When glucose was removed from the bathing fluid, oxygen consumption (QO2) decreased from 18.6 +/- 0.8 to 15.7 +/- 0.5 nmol O2/mg protein X min (P less than 0.01). The transported but nonmetabolized analogue of glucose, alpha-methyl-D-glucoside (alpha MG), was found to support QO2 to the same extent as glucose. These observations were also evident in the presence of butyrate, a readily oxidized substrate of the renal cortex. Additional studies with nystatin and ouabain indicated that glucose-related changes in QO2 were the result of changes in Na, K-ATPase associated respiration. The effect of glucose was localized to the luminal membrane since phlorizin (10(-5) M), a specific inhibitor of luminal glucose-sodium cotransport, also significantly reduced QO2 by 10 +/- 1%. Phlorizin inhibition of QO2 was also evident in the presence of alpha MG but was abolished when glucose was removed from the bathing medium. Finally, measurement of NADH fluorescence showed that addition of glucose (5 mM) to a tubule suspension causes an oxidation of NAD. These data are all consistent with glucose acting to increase respiration by stimulating sodium entry at the luminal membrane (via glucose-sodium cotransport) followed by increased sodium pump activity and its associated increase in mitochondrial respiration.

Animals↗
PubMed1984

Active ion transport in the renal proximal tubule. I. Transport and metabolic studies.

Various aspects of the interrelationship between ion transport and cellular metabolism were investigated using a suspension of rabbit cortical tubules that were mainly proximal in nature. Using the intact tubules, the compartmentation of K within the renal cell was studied by performing 42K uptake studies. The oxygen consumption (QO2) of the tubules was measured under similar conditions, as well as when the Na pump was stimulated by increasing Na+ entry with nystatin. In addition, the state 3 rate of respiration was measured when the mitochondria of digitonin-permeabilized tubules were stimulated by ADP. At 37 and 25 degrees C, a single-compartmental uptake of 42K was observed, which suggests that extracellular K+ communicates with a single compartment within the renal cell. Between 37 and 15 degrees C, the ouabain-sensitive QO2 and the initial 42K uptake rate were parallel in an Arrhenius-type plot, which indicated that active ion transport and oxidative phosphorylation remain tightly coupled within this temperature range. At all temperatures between 37 and 15 degrees C, nystatin stimulated the QO2, which demonstrates that the entry of Na+ into the renal cells was rate limiting for active Na+ transport throughout this temperature range. Between 37 and 20 degrees C, the nystatin-stimulated QO2 was nearly equal to the state 3 rate of respiration, which suggests that active ion transport may be limited by ATP availability under these conditions. At 15 degrees C, nystatin addition stimulated the QO2 well below the state 3 respiratory rate.

Animals↗
PubMed1984

Active ion transport in the renal proximal tubule. II. Ionic dependence of the Na pump.

The dependence of Na pump activity on intracellular and extracellular Na+ and K+ was investigated using a suspension of rabbit cortical tubules that contained mostly (86%) proximal tubules. The ouabain-sensitive rate of respiration (QO2) was used to measure the Na pump activity of intact tubules, and the Na,K-ATPase hydrolytic activity was measured using lysed proximal tubule membranes. The dependence (K0.5) of the Na pump on intracellular Na+ was affected by the relative intracellular concentration of K+, ranging from approximately 10 to 15 mM at low K+ and increasing to approximately 30 mM as the intracellular K+ was increased. The Na pump had a K0.5 for extracellular K+ of 1.3 mM in the presence of saturating concentrations of intracellular Na+. Measurements of the Na,K-ATPase activity under comparable conditions rendered similar values for the K0.5 of Na+ and K+. The Na pump activity in the intact tubules saturated as a function of extracellular Na at approximately 80 mM Na, with a K0.5 of 30 mM. Since Na pump activity under these conditions could be further stimulated by increasing Na+ entry with the cationophore nystatin, these values pertain to the Na+ entry step and not to the Na+ dependence of the intracellular Na+ site. When tubules were exposed to different extracellular K+ concentrations and the intracellular Na+ concentration was subsaturating, the Na pump had an apparent K0.5 of 0.4 mM for extracellular K. Under normal physiological conditions, the Na pump is unsaturated with respect to intracellular Na+, and indirect analysis suggests that the proximal cell may have an intracellular Na+ concentration of approximately 35 mM.

Animals↗
PubMed1984

Active ion transport in the renal proximal tubule. III. The ATP dependence of the Na pump.

The dependence of the Na pump activity of intact renal tubules on the ATP concentration was investigated using a suspension of rabbit cortical tubules. Rotenone (an inhibitor of mitochondrial oxidative phosphorylation) was used in graded fashion to alter the cellular ATP, and the Na pump activity was measured when the pump was stimulated by adding KCl to tubules suspended in a K+-free medium. The K+ uptake into the tubule was measured using an extracellular K+ electrode, and the oxygen consumption (QO2) was measured using a Clark-type oxygen electrode. The Na pump activity was found to have a linear, nonsaturating dependence on the ATP concentration. However, the Na,K-ATPase hydrolytic activity (assayed biochemically) of lysed proximal tubule membranes demonstrated saturation and had a K0.5 value of 0.4 mM ATP. Presumably, unknown cytosolic factors present in the intact renal cell but not normally present in the biochemical assay accounted for the differences between the two measurements. The data suggest that an alteration in the intracellular ATP will result in a proportional change in active ion transport activity. Moreover, additional findings also suggest that the basal (non-transport-related) QO2 may be redirected to support the proximal Na pump activity when transport activity is stressed. Thus, basal respiration is not invariant under all conditions, and ion transport activity appears to be maintained foremost among cellular ATP-dependent processes.

Adenosine Triphosphate↗
PubMed1984

Soft tissue infections in parenteral drug abusers.

Thirty-four parenteral drug abusers admitted with soft tissue infections underwent bacteriologic and immunologic evaluation. Staphylococcus aureus and beta hemolytic streptococci were the most common organisms recovered. Enteric gram negative aerobes and oral flora were common and enteric anaerobes rare. Absolute lymphopenia and elevations in the IgA, IgG and IgM fractions of the immunoglobulins were common as were false positive VDRL examinations. Cutaneous anergy was found in 83% of the group and 70% of a simultaneously noninfected addict group. Staphylococcal carriage was frequent. Because of variation in the flora between this and other reported groups, ongoing bacteriologic surveillance could be a useful guide to initial antibiotic therapy. Differences in the pattern of immune reaction in this group when compared to different addict groups suggest a difference in antigenic stimulation, possibly as a result of differences in bacteriologic exposure.

Abscess↗