Biomedical subjects

K G Dickman

Publications and source records attributed to K G Dickman.

13 recordsLinked to original sources

PubMed2006

Clues to VIP function from knockout mice.

We have taken advantage of the availability of vasoactive intestinal polypeptide (VIP) knockout (KO) mice to examine the possible influence of deletion of the VIP gene on: (a) airway reactivity and airway inflammation, as indicators of bronchial asthma; (b) mortality from endotoxemia, a model of septic shock; and (c) the pulmonary circulation. VIP KO mice showed: (a) airway hyperresponsiveness to the cholinergic agonist methacholine, as well as peribronchial and perivascular inflammation; (b) a greater susceptibility to death from endotoxemia; and (c) evidence suggestive of pulmonary hypertension.

Animals↗

PubMed2000

Pathways of inflammation and cell death in the lung: modulation by vasoactive intestinal peptide.

The pathogenesis of tissue injury in disease is a complex process that is only partially understood. We have investigated different models of acute lung injury, representing the clinical entity known as the acute respiratory distress syndrome, and tested their possible modulation by the neuropeptide vasoactive intestinal peptide (VIP). Three major mechanisms of injury appear to be involved in many of these models as common denominators: (1) activation of nuclear transcriptions factor NFkappaB; (2) apoptotic cell death; and (3) excitotoxic phenomena, due to activation of N-methyl-D-aspartate glutamate receptors. These pathogenetic mechanisms and pathways are logical targets of therapeutic intervention. Protection by VIP against lung injury, and against related forms of injury/cell death of neuronal cells and heart muscle, is attributable, in large measure, to the ability of VIP to suppress these mechanisms, and to additional anti-inflammatory and anti-oxidant actions. Finally, a hypothesis is presented for survival-promoting pathways that can be augmented by VIP and the related pituitary adenylyl cyclase-activating peptide.

Animals↗

PubMed2000

Rotavirus alters paracellular permeability and energy metabolism in Caco-2 cells.

Rotaviruses infect epithelial cells of the small intestine, but the pathophysiology of the resulting severe diarrhea is incompletely understood. Histological damage to intestinal epithelium is not a consistent feature, and in vitro studies showed that intestinal cells did not undergo rapid death and lysis during viral replication. We show that rotavirus infection of Caco-2 cells caused disruption of tight junctions and loss of transepithelial resistance (TER) in the absence of cell death. TER declined from 300 to 22 Omega. cm(2) between 8 and 24 h after infection and was accompanied by increased transepithelial permeability to macromolecules of 478 and 4,000 Da. Distribution of tight junction proteins claudin-1, occludin, and ZO-1 was significantly altered during infection. Claudin-1 redistribution was notably apparent at the onset of the decline in TER. Infection was associated with increased production of lactate, decreased mitochondrial oxygen consumption, and reduced cellular ATP (60% of control at 24 h after infection), conditions known to reduce the integrity of epithelial tight junctions. In conclusion, these data show that rotavirus infection of Caco-2 intestinal cells altered tight junction structure and function, which may be a response to metabolic dysfunction.

Animals↗

PubMed1993

Enhanced Na+ transport in an air-liquid interface culture system.

Use of the air-liquid interface culture technique has produced improved morphological differentiation of rodent, canine, and human tracheal epithelia. We have investigated the effect of this culture technique on ion transport activities of cultured canine bronchial epithelia. These cells were isolated from excised airways by enzymatic digestion and plated on permeable collagen membrane substrates. All cultures were maintained utilizing standard culture techniques, by bathing both apical and basolateral sides with hormone supplemented, serum-free media until confluent (days 4-6). Half of the cultures were converted to air-liquid interface cultures (ALIC) by gentle aspiration of the apical medium and half were continued under standard technique culture (STC) conditions. After three additional days, preparations cultured under both conditions were mounted in modified Ussing chambers where bioelectric properties were measured under short-circuit conditions. Mean short-circuit current (Isc) was significantly greater in ALIC (-91.3 +/- 7.84 microA/cm2) than in STC (-54.8 +/- 5.03 microA/cm2). The sodium channel blocker, amiloride, reduced Isc by 68.4 +/- 5.0% in STC and by 84.8 +/- 3.0% in ALIC. 22Na and 36Cl fluxes confirmed the presence of enhanced sodium absorption in ALIC when compared with STC. The depth of the apical fluid, measured by microelectrodes during ALIC, was approximately 15 microns. Studies of cellular metabolism demonstrated a shift in metabolism from an anaerobic to an oxidative pattern in ALIC. This change in the pattern of metabolism suggests that the ALIC technique enhanced sodium transport in canine bronchial epithelia by increasing oxygen delivery to the epithelium.

Absorption↗

PubMed1992

Relationship between HCO3- transport and oxidative metabolism in rabbit proximal tubule.

HCO3-/CO2 can affect proximal tubule energy metabolism directly by serving as a substrate for metabolic reactions and indirectly through ATP utilization by HCO3(-)-coupled Na+ reabsorption and proton secretion. In this study, metabolic and transport roles of HCO3-/CO2 were examined by measuring the effects of HCO3-/CO2 removal and transport inhibitors on oxygen consumption (QO2) in suspensions of rabbit proximal tubules. Removal of medium HCO3-/CO2 inhibited ouabain-sensitive, ouabain-insensitive, and uncoupled QO2. Consistent with metabolic inhibition, the absence of HCO3-/CO2 also reduced tubule ATP content and stimulated lactate production. Analysis of the dependence of mitochondrial state 3 respiration on HCO3-/CO2 in digitonin-permeabilized tubules traced the metabolic inhibition to limitations in tricarboxylic acid cycle intermediate supply. Energy requirements for HCO3- transport were examined by measuring QO2 in response to acetazolamide, 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS) and 4-acetamido-4'-isothiocyanostilbene-2,2'-disulfonic acid (SITS), and the H(+)-adenosinetriphosphatase (H(+)-ATPase) inhibitor bafilomycin A. Acetazolamide had no effect on QO2, whereas DIDS-SITS and bafilomycin A reduced ouabain-insensitive QO2, consistent with inhibition of active proton secretion. DIDS-SITS did not affect ouabain-sensitive respiration, suggesting that HCO3(-)-dependent Na+ reabsorption may not be mediated through the Na(+)-K(+)-ATPase in this preparation.

4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid↗

PubMed1990

Streptomycin toxicity in primary cultures of flounder renal proximal tubule cells.

The aminoglycoside antibiotic streptomycin is a known nephrotoxin in vivo and a common component of cell culture media. The effects of streptomycin (100 micrograms/ml) on transepithelial electrical properties, glucose transport, glycolytic metabolism, and morphology were examined in primary proximal tubule cell cultures from winter flounder (Pseudopleuronectes americanus) kidney. Streptomycin treatment on either Days 2 to 12 or Days 8 to 13 abolished the transepithelial potential difference and short-circuit current across the monolayer but had no effect on transepithelial resistance in confluent 12 to 13-d cultures, suggesting the loss of active transepithelial transport. Consistent with these findings, mucosal-to-serosal glucose fluxes were greatly reduced in streptomycin-treated cultures and insensitive to the transport inhibitor phlorizin, indicating the absence of the apical Na-dependent glucose transport system associated with net glucose reabsorption. In addition to transport processes, antibiotic treatment also interfered with cellular energy metabolism as judged by the rapid reduction in glycolytic lactate production observed in the presence of streptomycin. Scanning and transmission electron microscopy revealed that streptomycin-treated cultures were composed of cuboidal-to-columnar shaped cells which maintained intact tight junctions similar to control cultures. However, apical microvilli, the presumed sites of mucosal transport systems, were severely reduced in number in streptomycin-treated cultures. We concluded that streptomycin, at a dose commonly used in cell culture, inhibited the expression of differentiated function by flounder proximal tubule cell cultures. These cell cultures may provide a suitable model system for examination of the mechanisms of aminoglycoside nephrotoxicity.

Animals↗

PubMed1990

Cell volume regulation by skate erythrocytes: role of potassium.

The role of K transport during cell volume regulation in response to extracellular osmolality, protein kinase C activation, and cellular Ca was examined in skate (Raja erinacea) red blood cells (RBC). Reduction of medium osmolality from 960 to 660 mosmol/kgH2O had no effect on K uptake or efflux despite a 25% increase in cell volume. Further reduction to 460 mosmol/kgH2O caused K uptake to double and K efflux to triple resulting in net K loss. Net K efflux in 460 mosmol/kgH2O medium was correlated with the presence of a regulatory volume decrease, which was sensitive to the anion transport inhibitor 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS) and insensitive to chloride replacement. K-K exchange was absent in both isotonic and hypotonic media. Treatment with the Ca ionophore A23187 in the presence of Ca had no effect on either cell volume or K efflux in isotonic medium, indicating the absence of Ca-activated K transport. In contrast, phorbol ester treatment caused cell volume, Na content, and proton and K efflux to increase. Consistent with activation of Na-H exchange, phorbol ester effects were inhibited by dimethylamiloride. This study constitutes the first demonstration of volume-sensitive K transport in RBC from the most primitive vertebrate studied to date.

4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid↗

PubMed1990

Differential effects of respiratory inhibitors on glycolysis in proximal tubules.

The effects of inhibition of mitochondrial energy production at various points along the respiratory chain on glycolytic lactate production and transport function were examined in a suspension of purified rabbit renal proximal tubules. Paradoxically, partial blockage at site 3 by hypoxia (1% O2) induced lactate production, whereas total site 3 blockage by anoxia (0% O2) failed to stimulate glycolysis. Compared with anoxia, hypoxic tubules exhibited greater preservation of ATP and K+ contents during O2 deprivation and more fully recovered oxidative metabolism and transport function during reoxygenation. The mitochondrial site 1 inhibitor rotenone and the uncoupler carbonyl cyanide-p-trifluorome-thoxyphenylhydrazone (FCCP) were equipotent stimuli for lactate production, whereas the site 2 inhibitor antimycin A failed to stimulate glycolysis despite a 90% inhibition of O2 consumption. Compared with antimycin A, treatment with rotenone or FCCP resulted in less cell injury [measured by lactate dehydrogenase (LDH) release] and greater preservation of cell K+ and ATP contents. 2-Deoxyglucose blocked lactate production by 50% in the presence of rotenone and increased LDH release, suggesting that glycolytic ATP is partially protective. Addition of ouabain during rotenone treatment reduced lactate production by 50%, indicating that glycolytic ATP can be used to fuel the Na pump when mitochondrial ATP production is inhibited. We conclude that 1) proximal tubules can generate lactate during inhibition of oxidative metabolism by hypoxia, rotenone, or FCCP; 2) mitochondrial inhibition is not obligatorily linked to activation of glycolysis, since neither anoxia nor antimycin A stimulate lactate production; 3) when ATP can be produced through anaerobic glycolysis it serves to protect cell viability and transport function during respiratory inhibition.

Adenosine Triphosphate↗

PubMed1989

Glycolytic and oxidative metabolism in primary renal proximal tubule cultures.

Cultured cells often exhibit alterations in energy metabolism (increased glycolytic activity and decreased oxidative metabolism) during adaptation to the culture environment. The role of hypoxia as a mediator of these effects was examined by comparison of metabolism in primary rabbit renal proximal tubule (RPT) cultures maintained in stationary culture dishes (DISH), shaking Erlenmeyer flasks (SHAKE), and DISH cultures transferred back to SHAKE conditions (RESHAKE). Both oxidative metabolism and transport capacity were fully preserved in SHAKE cultures over a 24-h period. In contrast, within 6 h, DISH cultures exhibited a continuous decline in transport-dependent and -independent oxygen consumption, respiratory capacity, and ATP and K+ contents. The loss of oxidative metabolism in DISH cultures was accompanied by stimulation of lactate production, detectable within 1 h after plating. Comparison of metabolic properties of DISH cultures to those of RPT exposed to graded levels of hypoxia suggested that medium oxygen tensions may be as low as 1-3% in DISH cultures. RESHAKE cultures exhibited metabolic properties comparable to those of SHAKE cultures, indicating reversibility of DISH culture metabolism on reoxygenation. We concluded that DISH cultures rapidly become hypoxic as a consequence of static culture conditions. Shaking suspension cultures may provide a more metabolically appropriate model for long-term in vitro studies.

Adenosine Triphosphate↗

PubMed1987

Renal metabolism and acute renal failure.

We briefly review what appear to be the most important elements responsible for renal cell injury during and after oxygen deprivation. Recent studies in numerous laboratories have vastly improved our understanding of the changes in cell function that occur during ischemia and yet, the underlying mechanisms by which tubule damage and cell death occur remain elusive. We attempt to separate the effects that occur during ischemia or anoxia from those occurring during reperfusion (reoxygenation). These are not always separable, especially because it appears that ischemia initiates a series of complex events that may only become manifested during reperfusion. Ischemia-induced renal dysfunctions are clearly multifactorial events that will require major efforts to unravel.

Acute Kidney Injury↗

PubMed1986

Primary culture of flounder renal tubule cells: transepithelial transport.

Renal proximal tubule cells from the winter flounder (Pseudopleuronectes americanus) were maintained in a functionally differentiated state for up to 16 days in primary culture on floating collagen gels. The cells were confluent after 7-8 days in culture, contracted the collagen gels, and exhibited ciliary activity. Electron microscopy indicated that the cultures were composed of continuous sheets of columnar epithelial cells that had established structural polarity. When mounted in Ussing chambers, the cultures exhibited a small mucosa-negative potential difference (0.6 +/- 0.10 mV) and a low transepithelial resistance (23 +/- 2.3 omega X cm2). Short-circuit current averaged 24 microA/cm2. The cultured epithelium was four times more permeable to Na than to Cl and actively secreted sulfate and p-aminohippuric acid and reabsorbed hexoses. Glucose reabsorption was rheogenic and occurred via a high-affinity (Km = 0.16 mM), low-capacity (Vmax = 5 microA/cm2), phlorizin-sensitive transport system. We concluded that the cultured cells express many of the differentiated properties of the intact flounder proximal tubule and thus provide a suitable model system for studying renal transport processes.

Acids↗

PubMed1982

Effect of Na+ and ATP on peritubular Ca transport by the marine teleost renal tubule.

Ca uptake rates and efflux rate coefficients were determined in isolated renal tubules of the winter flounder, Pseudopleuronectes americanus. Na-free medium and 10(-4) M ouabain depressed while 0.2 mM dinitrophenol stimulated Ca efflux. The effects on Ca movement appeared to be associated with the adenosine triphosphate (ATP) concentration of the tubular fluid space (TFS), rather than the bath-to-tissue Na gradient. Elevation of the ATP concentration of TFS by incubation of tissue in external ATP more than doubled Ca uptake rate. Inhibition of Ca uptake by lanthanum and stimulation by Ca ionophore A23187 suggested that ATP altered plasma membrane Ca transport. Incubation in external phosphate had no effect. Plasma membrane vesicles (PMV) prepared from flounder tubules showed saturable Ca uptake inhibited by Mg, unaffected by Na gradients, and stimulated by intravesicular but not extravesicular ATP. ATP appeared to stimulate PMV Ca uptake by increasing membrane Ca binding. We concluded that Ca uptake across the peritubular membranes was not directly linked to the serosal Na gradient but was greatly influenced by the intracellular ATP concentration.

Adenosine Triphosphate↗

PubMed1980

Sulfate transport across the peritubular surface of the marine teleost renal tubule.

The uptake of sulfate across the peritubular surface of isolated renal tubules of seawater-acclimated winter flounder, Pseudopleuronectes americanus, consisted of two phases. The fast exchanging component appeared to be a small sulfate compartment with a saturable uptake rate; however, the Km was quite large (14.5 mM). The fast phase was partially inhibited by the anion transport inhibitor 4-acetamido-4'-isothiocyano-2,2'-disulfonic stilbene (SITS), but was unaffected by antimycin A, which indicated lack of ATP-dependence. The slowly exchanging compartment was fourfold larger than the fast, saturable with a low Km (0.65 mM), and inhibited by antimycin A, SITS, ouabain, and Na-free incubation medium. Phosphate appeared to be a noncompetitive inhibitor of this phase. These observations support the idea that the slow phase of sulfate uptake may be driven by the peritubular membrane Na gradient. This mediated, energy-dependent uptake may be part of the active sulfate secretory pump of the marine teleost renal tubule.

4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfo↗