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C A Boyd

Publications and source records attributed to C A Boyd.

At least 55 records · Page 3Linked to original sources
PubMed1995

Active transport in the alveolar epithelium of the adult lung: vestigial or vital?

Active secretion by mammalian fetal pulmonary alveolar epithelium is well recognized, as is the role of the adult epithelium in the secretion of surfactant. Recent studies have demonstrated active absorption by adult epithelium involving two sodium-dependent pathways. This finding has focused attention on how poorly we understand both the disposition of alveolar liquid and the physiological role of surfactant. In this paper we review the evidence that the adult mammalian alveolar epithelium absorbs solutes by active transport, and we assess the physiological importance of the resulting liquid movements.

Animals↗
PubMed1995

Dipeptide transport characteristics of the apical membrane of rat lung type II pneumocytes.

The transport of a hydrolysis-resistant dipeptide, D-phenylalanyl-L-alanine (D-Phe-L-Ala), has been studied by high-performance liquid chromatography in rat lung epithelial cells and apical membrane vesicles. Time-dependent uptake of D-Phe-L-Ala into isolated type II pneumocytes was shown. Uptake was saturable, and Michaelis-Menten kinetics were fitted to the data and gave an apparent Michaelis constant (Km) of 3.4 mM and a maximum velocity (Vmax) of 7.0 nmol.mg protein-1.min-1. However, known peptide transport inhibitors unexpectedly increased intracellular D-Phe-L-Ala concentration when initial rates of peptide uptake were studied. Apical (brush-border) membrane vesicles prepared from rat lung also showed time- and concentration-dependent influx of D-Phe-L-Ala (apparent Km 2.0 mM, Vmax 0.53 nmol.mg protein-1.min-1). Influx of this neutral dipeptide into the vesicles was shown to be both electrogenic and stimulated by an inwardly directed proton gradient. Influx was inhibitable by mercuric chloride and by the amino acid residue modifying compounds N-acetylimidazole and diethylpyrocarbonate. These findings strongly suggest the presence of a proton-coupled peptide transport protein in the apical surface of the type II cell. This transporter may play a role in lung homeostasis.

Animals↗
PubMed1994

Membrane potential dependence of the kinetics of cationic amino acid transport systems in human placenta.

1. Mediated influx of L-lysine into human placental brush-border membrane vesicles occurs through two systems, one of lower affinity but high capacity, the other of very high affinity but low capacity. These transporters have features characteristic of systems y+ (the classical system) and y+L (recently described in the erythrocyte), respectively. 2. In solutions containing sodium the entry of lysine through the high-affinity system (y+L) is inhibited by the neutral amino acids L-leucine, L-methionine and L-glutamine with comparable high affinity. The removal of sodium reduces the affinity but not the maximal extent of this inhibition. Leucine and methionine, but apparently not glutamine, inhibit lysine entry through system y+ with a much lower affinity. 3. The influx of lysine through system y+ changes markedly in response to alterations of membrane potential. In the presence of an inwardly directed negative diffusion potential created by an inwardly directed thiocyanate (SCN-) gradient, the influx of lysine through this route is accelerated; with an inwardly directed positive potassium diffusion potential, lysine influx through this route is reduced. The influx of lysine through system y+L is much less sensitive to such alterations of potential. 4. Analysis of the kinetic constants characterizing system y+ shows that with a change of potential from zero to negative (approximately -60 mV) the maximum velocity (Vmax) is roughly doubled and the half-saturation constant (Km) halved leading to a 4-fold increase in permeability. For system y+L smaller changes are seen and Km does not change; the resulting increase in y+L permeability is 1.5-fold. 5. These findings are discussed with respect both to the mechanism of membrane transport and placental epithelial function.

Amino Acids↗
PubMed1994

Identification and properties of pathways for K+ transport in guinea-pig and rat alveolar epithelial type II cells.

86Rb+ was used to study potassium uptake and efflux in type II pneumocytes freshly isolated from adult guinea-pig and rat lung. Both species exhibited a substantial ouabain-sensitive component of potassium influx. In rats, most of the ouabain-resistant influx was abolished by bumetanide and removal of extracellular chloride elicited no further effect. In contrast, only a proportion of the ouabain-insensitive uptake was inhibitable by bumetanide in guinea-pigs and this species showed an additional component of influx, which was chloride dependent and which was reduced by either the K(+)-H(+)-ATPase inhibitor, omeprazole, or by the stilbene derivative, 4,4'-diisothiocyanostilbene-2,2'-disulphonate (DIDS). The chloride-dependent component was also apparent in efflux experiments in guinea-pigs, but was absent in rats. Ouabain-insensitive ATPase activity was assayed in highly purified apical membranes from guinea-pig type II pneumocytes. This activity was inhibitable by omeprazole (apparent inhibition constant, Ki, was approximately 40 microM), was potassium dependent (apparent activation constant, Ka, was approximately 200 microM) and was doubled by the addition of nigericin. While potassium transport in rat type II cells is adequately accounted for by Na(+)-K(+)-ATPase and Na(+)-K(+)-2Cl- cotransport, our data suggest the additional presence of K(+)-Cl- cotransport and K(+)-H(+)-ATPase in guinea-pig type II pneumocytes. A model of how alveolar subphase acidification may occur is proposed.

4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid↗
PubMed1994

Tripeptide transport in rat lung.

Transport of L-alanyl-D-phenylalanyl-L-alanine was investigated with an in situ vascular perfusion preparation of rat lung and brush border membrane vesicles prepared from type II pneumocytes. In the perfused lung 1 mM tripeptide was transported intact from the alveolar lumen to the vascular perfusate at a mean rate of 25.1 +/- 1.29 (3) nmol/min per g dry weight. D-Phenylalanine also appeared in the vascular perfusate at a rate of 21.9 +/- 1.74 (3) nmol/min per g dry weight indicating that 47% of the absorbed tripeptide was split during passage across the epithelial layer. No dipeptide could be detected in the vascular effluent during perfusions with tripeptide. Rapid L-alanyl-D-phenylalanyl-L-alanine uptake occurred with fresh apical membrane vesicles prepared from type II pneumocytes and this was abolished by treatment with 0.1% triton. The related tripeptide, D-alanyl-L-phenylalanyl-D-alanine, was taken up significantly more slowly by the vesicles. D-phenylalanyl-L-alanine and D-phenylalanyl-D-alanine, were also studied with the vascularly perfused preparation; the mixed dipeptide appeared in the vascular perfusate significantly faster than L-alanyl-D-phenylalanyl-L-alanine whereas D-phenylalanyl-D-alanine appeared more slowly and was not hydrolysed.

Amino Acid Sequence↗
PubMed1994

Cell surface receptor for ecotropic host-range mouse retroviruses: a cationic amino acid transporter.

The cell surface receptor for ecotropic host-range murine leukemia viruses is a sodium-independent transporter for essential cationic acids. Our evidence strongly identifies this receptor as the transporter system y+, which was previously characterized by transport assays. Mutational analysis indicates that transporter activity is not necessary for viral reception. Infection of cells with ecotropic retroviruses causes only a partial down-modulation of receptor expression on cell surfaces.

Amino Acid Sequence↗
PubMed1994

Effect of lung inflation on active and passive liquid clearance from in vivo rabbit lung.

Active sodium transport contributes to liquid clearance from the alveoli. We hypothesized that the magnitude of active transport of alveolar liquid depends on the extent to which the alveolar epithelium is stretched and, consequently, on the degree of alveolar inflation. In a study on 38 adult rabbits, the left lung was filled in vivo with a solution of glucose (10 mmol/l) made isosmotic with plasma, using sodium chloride, and held at a constant airway pressure of 3, 6, or 9 cmH2O for 6 h. Alveolar liquid clearance was measured directly as a flow into a left main bronchial catheter. Control animals were compared with animals in which active epithelial sodium transport was inhibited by adding amiloride and phloridzin (both 1 mmol/l) to the instillate. At low inflation, active sodium transport reversed a secretion of liquid into the alveoli; at high inflation, active sodium transport made little or no contribution to transepithelial flow. Hydraulic conductance of the left lung was 1.57 microliters.min-1.cmH2O-1.kg body wt-1. The experiments suggest that pulmonary inflation renders active liquid clearance ineffective.

Air↗
PubMed1994

Regulation of conductive chloride transport in human placental brush border membrane vesicles.

Conductive chloride transport in the brush border of the human placental epithelium has been determined by measurement of radioactive chloride flux (influx and efflux) and anion diffusion potentials. This conductance is inhibited specifically by low micromolar concentrations of arachidonic acid whereas the K+ permeability of this membrane is increased by arachidonic acid. The conductive pathway for anions is inhibited by 9-AC and has the permeability sequence SCN > I > NO3 > Br > Cl > F > gluconate; it is also permeant to HCO3-. Functionally the control of these anion and cationic conductances by arachidonic acid will be important in regulating epithelial transport and nutrient delivery to the fetus since they will set the value of the resting potential across the apical surface of the syncytiotrophoblast.

Arachidonic Acid↗
PubMed1994

Cationic amino acid transport in human T lymphocytes is markedly increased in the CD45RA, CD8+ population after activation.

Membrane transport of cationic amino acids is essential for cells which are actively metabolizing L-arginine or L-lysine. In human cells most of this transport occurs through y+, a transport system which is only now being characterized at the molecular level. We have previously shown that phytohaemagglutinin (PHA) stimulation of peripheral blood E rosette positive (T) lymphocytes specifically activated lysine transport through system y+, whereas Staphlyococcus aureus Cowan A (SAC) stimulation of the E rosette negative fraction did not. We have now analysed this effect in PHA-activated CD4, CD8, CD45RO and CD45RA T-cell subsets. Both PHA-activated CD4+ and CD8+ T cells have increased lysine transport through y+, and in seven out of eight experiments, more activity was seen in the CD8+ fraction. In contrast, marked differences in y+ activity were seen between the PHA-activated CD45RO and CD45RA subsets. Thus in six experiments y+ activity was markedly increased in the CD45RA (naive T cell) population but not in the CD45RO (memory) cells. In one further experiment the activated CD45RO, CD4- population (enriched for CD45RA+, CD8+) was studied and y+ activity was shown to be maximal in this cell subset. Transport of arginine is essential for nitric oxide synthesis. Our findings therefore suggest that activated CD45RA, CD8+ T cells are capable of nitric oxide production.

Adult↗
PubMed1993

Anion exchange in type II pneumocytes freshly isolated from adult guinea-pig lung.

We have studied chloride influx and efflux in a highly purified preparation of type II cells freshly isolated from adult guinea-pig lung using 36Cl-. Chloride uptake was time-dependent, saturable (Km < 10 mM) and was inhibited by 4,4'-diisothiocyanatostilbene-2,2'-disulphonic acid (DIDS; Ki approximately 80 microM). In the absence of external chloride (substituted by gluconate), 36Cl- uptake exhibited an overshoot above equilibrium. The rate of 36Cl- entry was strongly inhibited by addition of external nitrate; sulphate was a weaker inhibitor. 36Cl- efflux was stimulated by external bromide > bicarbonate > or = chloride > or = citrate; and was inhibited by propionate > acetate > oxalate. Although the "chloride channel blocker" 4-nitro-2-(3-phenylpropylamino)benzoate (0.14 mM) caused an inhibition, 36Cl- influx did not appear to be electrogenic. These data are compatible with the existence of a substantial electroneutral anion-exchange pathway for chloride transport in freshly isolated adult type II pneumocytes.

4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid↗
PubMed1992

Substrate-specific differences in the rate of bile acid carrier reorientation: studies on human placental basal vesicles.

The initial rate of transport of the bile acid glycocholic acid (GCA) has been measured in influx and efflux across placental basal membrane vesicles, and the mechanism of inhibition of its transport by the analogue taurochenodeoxycholic acid (TCDCA) analysed kinetically. This analogue, although trans-stimulating GCA efflux, inhibits influx in a way which does not depend upon substrate concentration; moreover, its potency as an inhibitor is markedly influenced by whether it is placed on one or on both sides of the vesicles membrane. These findings can be accounted for by postulating that both GCA and TCDCA are translocated through the carrier, but that the rate of loaded carrier reorientation is higher than that of the free carrier only when loaded with TCDCA and not with GCA.

Bile Acids and Salts↗
PubMed1992

Activation of cationic amino acid transport through system y+ correlates with expression of the T-cell early antigen gene in human lymphocytes.

Lysine influx (2 microM) into activated human B and T lymphocytes through transport systems y+ and y+L was measured. In activated T cells very substantial activation of system y+ was detected; system y+L was also activated in these cells but with a slower time course and to a smaller extent. No stimulation of either system was found in activated B cells. The time course of activation of system y+ precisely matched the expression of the T cell early antigen gene described by MacLeod et al, 1990. The functional significance of these observations with respect to L-arginine transport and nitric oxide synthesis in activated T lymphocytes is discussed.

Amino Acids↗
PubMed1992

Identification of a new transport system (y+L) in human erythrocytes that recognizes lysine and leucine with high affinity.

1. The effect of neutral amino acids on the transport of L-lysine across the human erythrocyte membrane was studied. 2. All neutral amino acids tested (range 0.3-5 mM) inhibit the influx of L-[14C]lysine (1 microM). The inhibition pattern is biphasic, and tends to reach a maximum at approximately 50% of the original flux. The concentrations that give 25% inhibition are (mM): L-cysteine (2.7), L-alanine (1.3), L-serine (0.9), L-isoleucine (0.6), L-phenylalanine (0.35), L-methionine (< 0.3), L-leucine (< 0.3). L-lysine and L-arginine completely inhibit the rate at the highest concentration. 3. These results can be explained by assuming that L-lysine transport occurs through two independent transporters that differ in their affinity for neutral amino acids. A detailed kinetic analysis of the effect of L-leucine on L-lysine entry is consistent with this hypothesis. 4. Using a new experimental strategy, the substrate and inhibitor transport parameters for the two systems were determined. The half-saturation constants for lysine (+/- S.E.M.) are found to be: KmA, 0.014 +/- 0.002 mM and KmB, 0.112 +/- 0.017 mM. The maximum rates differ by a factor of 8.2 (VmaxB/VmaxA). The leucine inhibition constants are: KiA, 0.022 +/- 0.003 mM and KiB, 30.36 +/- 7.9 mM. If the sodium in the incubation medium is replaced by potassium, the apparent affinity for leucine (1/KiA) is reduced approximately 30-fold. 5. The maximum inhibition caused by leucine decreases as the lysine concentration is raised, showing that leucine acts upon the higher affinity system. 6. When added to the trans side, L-leucine, L-phenylalanine and L-isoleucine do not cause inhibition, but stimulate the flux by approximately 30%. This indicates that these analogues are also transported. 7. In conclusion, in the concentration range 1-100 microM, lysine crosses the red cell membrane through two distinct transport systems, one of which recognizes both neutral and cationic amino acids with high affinity.

Amino Acids↗
PubMed1992

Pathways for glucose transport in type II pneumocytes freshly isolated from adult guinea pig lung.

Previous in vivo studies of sugar transport across the mature pulmonary epithelium have provided evidence for the existence of a specific phlorizin-inhibitable, sodium-dependent transport process for D-glucose, although no direct evidence for the cellular location of this transport system in fresh cells has been shown to date. With the use of elastase digestion and lectin agglutination, a pure preparation of type II alveolar epithelial cells was isolated from adult guinea pig lung. This preparation always contained >90% type II cells and typically showed approximately 85% cell viability 2-3 h after the isolation procedure had begun. At 37 degrees C, cells showed specific [3H]phlorizin binding that was attenuated by D-glucose and completely abolished by sodium replacement. Substantial accumulation of the hexose [14C]methyl alpha-D-glucopyranoside (14C-labeled AMG), a substrate specific for the sodium-dependent glucose cotransporter was found in the presence of extracellular sodium; this accumulation above equilibrium was abolished on removal of sodium, addition of phlorizin, or in the presence of a saturating concentration (69 mM) of D-glucose. The apparent inhibition constant (Ki) for glucose inhibition of AMG uptake was 0.4 mM and for phlorizin, 0.5 microM. The Hill plot of sodium activation of AMG uptake gave a coefficient of 2.8, suggesting cooperativeness between sodium and AMG transport. 3-O-[14C]methyl-D-glucose (3-OMG) transport was also blocked by phlorizin. Phloretin, in the presence of phlorizin, slowed the initial rate of entry but did not affect the equilibrium that was attained in the presence of phlorizin alone.(ABSTRACT TRUNCATED AT 250 WORDS)

3-O-Methylglucose↗