Biomedical subjects

S H Wright

Publications and source records attributed to S H Wright.

At least 73 records · Page 4Linked to original sources

PubMed1989

Integumental nutrient uptake by aquatic organisms.

Integumental nutrient transport is a widespread characteristic of soft-bodied marine invertebrates. These processes, which are qualitatively similar to the Na-dependent transporters of intestinal epithelia, have kinetic and energetic characteristics that make them particularly well suited for accumulating materials from the extremely low substrate concentrations found in seawater. Despite the low concentrations of DOM in natural waters, rates of DOM uptake are large and clearly capable of supporting a significant fraction of the metabolic needs of some species. Indeed, recent studies with larval invertebrates suggest that integumental uptake of DOM may play a pivotal role in animal nutrition. Current and future studies on the mechanism and regulation of these processes, on the metabolic fate of accumulated DOM, and on the distribution of DOM in natural waters, promise to resolve the remaining issues on the role of integumental transport in the nutrition of aquatic organisms.

Adaptation, Physiological↗

PubMed1989

Mechanisms of integumental amino acid transport in marine bivalves.

The marine bivalve integument accumulates amino acids from seawater at rates that can significantly supplement other nutritional strategies. Reviewed here are studies that used both intact bivalve gill tissue and membranes isolated from gills to examine the mechanism of integumental transport. The evidence supports a model for secondary active transport in which an inward flux of Na+ down its electrochemical gradient provides the energy to support uptake of amino acids. Indeed, the Na+-cotransport paradigm appears to be a ubiquitous strategy by which animal cells energize uphill transport of organic solutes. What appears to distinguish integumental processes from others are the conditions under which they must routinely work: exposed to ambient concentrations of substrate of less than 1 microM and to cytoplasmic concentrations that exceed 0.1 M, transport of amino acid occurs against gradients that can exceed 10(6) to 1! Somewhat paradoxically, these transporters, which are characterized by an extremely high affinity for amino acid (Kt values of congruent to 5 microM), work so well because they have a low affinity for Na+; low cytoplasmic Na+ concentrations prevent the system from acting as an avenue for efflux, whereas the very high concentrations of Na+ in seawater are adequate to fully activate the influx mode of operation. The combination of these kinetic characteristics results in a class of transporters that makes efficient use of the scarce nutritional resource represented by dissolved organic material in near-shore waters.

Alanine↗

PubMed1989

Brush-border TEA transport in intact proximal tubules and isolated membrane vesicles.

The efflux of the organic cation, tetraethylammonium (TEA), across proximal cell luminal membranes was studied using intact, perfused rabbit proximal tubules and isolated rabbit cortical brush-border membrane vesicles (BBMV). Increases of either the extravesicular H+ concentration (from pH 7.5 to pH 6.5) or the extravesicular concentration of unlabeled TEA (from 0.1 to 0.5 mM) increased the rate of efflux of radioactively labeled TEA from BBMV. Similarly, when proximal tubules were preloaded with labeled TEA and then submerged in a mineral oil bath, a rapid increase in either the H+ concentration (from pH 7.5 to pH 5.8) or the TEA concentration (from 0 to 1 mM) of the tubular perfusate produced an acute increase in efflux of the labeled TEA across the luminal brush-border membrane. These results with intact tubules and isolated membranes are consistent with previous suggestions that TEA transport across the brush border of proximal tubule cells involves a carrier-mediated countertransport process and represent the first demonstration with intact proximal tubules that the secretory flux of TEA occurs by an exchange with H+.

Animals↗

PubMed1989

Amiloride transport in rabbit renal brush-border membrane vesicles.

Rabbit renal brush-border membrane vesicles (BBMV) were used to study amiloride transport across the luminal membrane of proximal tubular cells. An outwardly directed H+ gradient (pHi 6.0; pHo 7.5) stimulated 8 microM [14C]-amiloride uptake into BBMV and supported a transient "active" accumulation of substrate consistent with the presence of an amiloride-H+ exchange process. Uptake was inhibited, in the presence or absence of a pH gradient, by 1 mM unlabeled amiloride or 20 mM tetraethylammonium (TEA). Amiloride transport was not directly affected by the presence of 100 mM Na+ in the extravesicular medium, suggesting that Na-H exchange did not mediate amiloride flux. Amiloride transport was a saturable process with a maximal flux (under pH gradient conditions) of 3 nmol.mg-1.min-1 and an apparent Kt of 8 microM. TEA acted as a competitive inhibitor of this process with an apparent Ki of approximately 80 microM, similar to the Kt of TEA transport via the TEA-H+ exchanger. Likewise, amiloride acted as a competitive inhibitor of TEA uptake with an apparent Ki of approximately 11 microM. Preloading BBMV with 1-2 mM TEA stimulated the rate of amiloride uptake and supported a transient active accumulation of amiloride. We conclude that amiloride and TEA are transported by a common pathway in BBMV, which involves a carrier-mediated exchange with H+ and which may play a role in the tubular secretion of these compounds.

Amiloride↗

PubMed1989

Intracellular Na+ and the control of amino acid fluxes in the integumental epithelium of a marine bivalve.

The accumulation of amino acids from sea water into the integumental epithelium of the bivalve gill can occur against chemical gradients in excess of 10(6) to 1. The energy to drive this transport has been proposed to come from the inwardly directed Na+ electrochemical gradient. The present study examined the influence of intracellular and extracellular [Na+] on influx and efflux of amino acids in gill tissue from the mussel, Mytilus californianus. Influx of alanine was inhibited by more than 90% when external [Na+] was reduced from 425 to 2 mmol l-1, and by 85% when intracellular [Na+] was increased from approximately 11 to approximately 100 mmol l-1 (by means of a 30-min exposure to the ionophore, nigericin). Efflux of taurine and alanine from gill tissue into normal-Na+ sea water was very low (less than 5% of the Jmax of the carrier-mediated influx pathways). Reducing the external Na+ from 425 to 2 mmol l-1 increased taurine efflux by only 20%. Raising cell [Na+] to approximately 100 mmol l-1 increased taurine efflux 2.7-fold; further increases in cell [Na+] increased taurine efflux another 7.5-fold. These data, in conjunction with results from earlier studies, suggest that activation of integumental amino acid transporters requires an interaction of multiple sodium ions with binding sites of low affinity for this ion. This set of characteristics results in transport systems that are well-adapted for the net accumulation of amino acids from sea water.

Alanine↗

PubMed1988

Mechanism of cis- and trans-substrate interactions at the tetraethylammonium/H+ exchanger of rabbit renal brush-border membrane vesicles.

The kinetic basis for trans-effects of intravesicular substrates on the uptake of the organic cation, tetraethylammonium (TEA), into rabbit renal brush-border membrane vesicles (BBMV) was studied. Preloading BBMV with 1, 2, or 4 mM TEA stimulated the initial rate of uptake and the total net accumulation of 0.1 mM [3H]TEA. The stimulatory effect of intravesicular TEA on the initial rate of uptake was a saturable function of the trans-TEA concentration, with a half-maximal effect noted at an intravesicular concentration of 0.28 mM. A 1 mM trans-concentration of TEA increased the Jmax of [3H]TEA uptake (from 4.3 to 6.8 nmol.mg-1.min-1) without affecting the apparent Kt. An outwardly directed H+ gradient also increased Jmax (to 10.7 nmol.mg-1.min-1), although the addition of an outwardly directed TEA gradient did not produce further increases in the rate of TEA uptake. External H+ acted as a competitive inhibitor of TEA uptake, and an increase in external [H+] (from 32 nM to 100 nM) produced an increase in the apparent Kt for TEA transport (from 0.12 to 0.26 mM) without affecting the Jmax. The results suggested that TEA and H+ compete for a common site or set of mutually exclusive sites on the cytoplasmic and luminal aspects of TEA/H+ exchanger in the renal brush border, and that these sites have a similar affinity for TEA.

Animals↗

PubMed1988

Amino acid transport in the gill epithelium of a marine bivalve.

1. Amino acid transport across the intestine of bivalve molluscs is reviewed. 2. Transport of alanine or taurine is dependent on the sodium gradient across the intestine. 3. The time course of the uptake of alanine into the brush border membrane vesicles is also sodium dependent.

Amino Acids↗

PubMed1987

L-alanine uptake in brush border membrane vesicles from the gill of a marine bivalve.

Brush border membrane vesicles were prepared from mussel gills using differential and sucrose density gradient centrifugation. These vesicles contained both the maximal Na+-dependent alanine transport activity found in the gradient and the maximal activities of gamma-glutamyl transpeptidase and alkaline phosphatase. Electron micrographs showed closed vesicles of approximately 0.1-0.5 micron diameter. Transport experiments using these vesicles demonstrated a transient 18-fold overshoot in intravesicular alanine concentration in the presence of an inwardly directed Na+ gradient, but not under Na+ equilibrium conditions. A reduced overshoot (10-fold) was seen with an inwardly directed K+ gradient. Further studies revealed a broad cation selectivity, with preference for Na+, which was characteristic of alanine transport but not glucose transport in these membranes. The apparent amino acid specificity of the uptake pathway(s) was similar to that of intact gills and supported the idea of at least four separate pathways for amino acid transport in mussel gill brush border membranes. The apparent Michaelis constant for alanine uptake was approximately 7 microM, consistent with values for Kt determined with intact tissue.

Alanine↗

PubMed1987

Succinate and citrate transport in renal basolateral and brush-border membranes.

Properties of the Na-dependent transport of succinate and citrate were determined in brush-border and basolateral membrane vesicles (BBMV and BLMV, respectively) isolated from the rabbit renal cortex. As has been shown in previous studies, the initial rate of uptake of these substrates into BBMV was markedly stimulated (50- to 150-fold) by an inwardly directed Na gradient. In BLMV, uptake of 5 microM succinate was also stimulated (150-fold) by a Na gradient, and the profile of the time course of accumulation was qualitatively and quantitatively different from that in BBMV. In contrast, basolateral uptake of 2 microM citrate was stimulated only one- to fivefold by a Na gradient. The external pH had little effect on uptake of succinate into BBMV or BLMV, or on uptake of citrate into BLMV. Uptake of citrate into BBMV, however, was stimulated 10-fold by reducing external pH from 8 to 5.5. Valinomycin-induced K diffusion potentials stimulated succinate uptake into BBMV and BLMV, suggesting that Na-coupled succinate transport across both membranes is electrogenic. The Jmax and apparent Kt for uptake into BBMV were much greater than those in BLMV (90 vs. 5 nmol X mg-1 X min-1 and 0.61 vs. 0.01 mM, respectively). Inhibition of succinate uptake by several polycarboxylates suggested that the BLMV transporter had a higher apparent affinity for dicarboxylates than for tricarboxylates, compared with the BBMV transporter. We conclude that BLMV possess a low-capacity, high-affinity Na-dependent transport pathway showing a high degree of specificity for dicarboxylic, rather than tricarboxylic, acids.

Animals↗

PubMed1987

Transport of tetraethylammonium by rabbit renal brush-border and basolateral membrane vesicles.

Brush-border and basolateral membrane vesicles (BBMV and BLMV, respectively) from rabbit renal cortex were used to study transport of the organic cation, tetraethylammonium (TEA). Outwardly directed proton gradients stimulated uptake of TEA into BBMV and supported concentrative accumulation. Furthermore, an inwardly directed H+ gradient accelerated TEA efflux from BBMV. These data suggest that TEA transport in BBMV involved exchange with H+. The Jmax and Kt for TEA transport into BBMV under pH equilibrium conditions (pH 7.5) were 2.1 nmol.mg-1.min-1 and 0.15 mM, respectively. Under pH gradient conditions (6.0in:7.5out), Jmax increased by 270% with no effect on Kt. Uptake of TEA into BBMV was stimulated by an inside-positive electrical potential difference (PD), although exchange of TEA for H+ appeared to be one for one. In BLMV, H+ gradients had little effect on TEA uptake and were incapable of supporting concentrative transport. The Jmax and Kt for TEA transport in BLMV were 0.33 nmol.mg-1.min-1 and 0.37 mM, respectively. Inside-negative PDs stimulated this uptake, suggesting that it involved an electrically conductive pathway. TEA transport in BBMV and BLMV was inhibited by amiloride and cimetidine, although p-aminohippuric acid was without effect. Thus, secretion of TEA involves carrier-mediated transport steps at both the luminal and peritubular membranes, although an active step is not evident in isolated BLMV.

Animals↗

PubMed1986

Efficient homologous recombination of linear DNA substrates after injection into Xenopus laevis oocytes.

When DNA molecules are injected into Xenopus oocyte nuclei, they can recombine with each other. With bacteriophage lambda DNAs, it was shown that this recombination is stimulated greatly by introduction of double-strand breaks into the substrates and is dependent on homologous overlaps in the recombination interval. With plasmid DNAs it was shown that little or no recombination occurs between circular molecules but both intra- and intermolecular events take place very efficiently with linear molecules. As with the lambda substrates, homology was required to support recombination; no simple joining of ends was observed. Blockage of DNA ends with nonhomologous sequences interfered with recombination, indicating that ends are used directly to initiate homologous interactions. These observations are combined to evaluate possible models of recombination in the oocytes. Because each oocyte is capable of recombining nanogram quantities of linear DNA, this system offers exceptional opportunities for detailed molecular analysis of the recombination process in a higher organism.

Animals↗

PubMed1985

Transport of N1-methylnicotinamide across brush border membrane vesicles from rabbit kidney.

A preparation of isolated brush border membranes from the rabbit renal cortex was used to examine the characteristics of N1-methylnicotinamide (NMN) transport in the kidney. Transport was independent of the presence of Na+ under Na+ equilibrium conditions. However, outwardly directed Na+ gradients stimulated NMN uptake, whereas inwardly directed Na+ gradients inhibited NMN uptake. Transport appeared to involve two parallel processes: one saturable with a Jmax of 5 nmol X mg protein-1 X min-1 and an apparent Kt of 0.6 mM, and a second that behaved like passive diffusion. Countertransport of NMN was observed when vesicles were preloaded with either NMN or another organic cation, tetraethylammonium (TEA). TEA and several structural analogues, as well as a wide variety of other organic cations and bases, were effective inhibitors of NMN uptake, though nicotinamide and p-aminohippuric acid did not interact with the uptake process. Outwardly directed proton gradients (pH 6.0 in, 7.6 out) stimulated transport, suggesting that NMN uptake may involve a countertransport of H+. The electrical potential difference across the vesicle membrane was manipulated using gradients of the permeant organic ion thiocyanate (SCN-); an outwardly directed gradient of SCN- (i.e., a depolarizing condition) stimulated uptake and produced a transient accumulation of NMN above that noted at equilibrium, whereas an inwardly directed SCN- gradient inhibited uptake of NMN. The data can be explained by postulating the presence of an electrogenic NMN+-H+ antiporter in the rabbit luminal membrane that could play a role in organic cation secretion.

Animals↗

PubMed1985

Interactions between tricarboxylic acid cycle intermediates and phosphate uptake by proximal renal cells and renal brush border membranes.

Glucose and other hexoses as well as amino acids have been shown to inhibit the renal transport of phosphate (Pi). Although studies with renal brush border membrane vesicles showed that such an inhibitory effect on Pi transport is due to the dissipation of the Na electrochemical gradient, the mechanism(s) responsible for such an action in the intact cell is not clear. The present study examined the effects of tricarboxylic acid (TCA) cycle intermediates (succinate and fumarate) and acetate on the uptake of Pi and alpha-methylglucoside (AMG) at 37 degrees C by intact rabbit renal cells. These TCA cycle compounds significantly (p less than 0.05) inhibited the uptake of both Pi and AMG. In the presence of 5-10 mM succinate the ATP content of the renal cells increased by 40% (p less than 0.02). Inhibition of succinate-induced gluconeogenesis by 3-mercaptopicolinic acid did not modify the inhibition of Pi uptake. Studies with renal brush border membrane vesicles showed that succinate inhibited Pi uptake at 15 and 60 s but not at 1 s and only under conditions of Na gradient (outside greater than inside). Succinate did not inhibit Pi uptake during Na equilibrium conditions. The data demonstrate that the succinate-induced inhibition of the Pi uptake by intact proximal renal cells is not due to competition for metabolic energy, is not related to stimulation of gluconeogenesis nor due to allosteric interaction between Pi carrier and succinate transporter. The results support the notion that the inhibition of Pi uptake by succinate in the intact renal cell is due to dissipation of the Na chemical gradient.

Animals↗

PubMed1984

Genetic recombination of Xenopus laevis 5 S DNA in bacteria.

The behavior in genetic recombination of Xenopus laevis 5 S DNA has been examined, with particular emphasis on the role of 15-base-pair tandem repeats in the A + T-rich spacer. Fragments of 5 S DNA were introduced into Escherichia coli cells as inserts in the recombination vectors, lambda rva and lambda rvb. Intermolecular recombinants were selected in which, because of properties of the phage vectors, the crossover event must have occurred within the 5 S DNA inserts. Inserts from individual recombinants have been characterized in detail. The effects of varying the number (n) of 15-base-pair repeats and the recombination capabilities of the phage and host have been investigated. In these crosses, unequal crossovers can occur, yielding inserts different in size from the parental inserts. When the number of 15-mers is large (n = 12 or 20), most of the unequal crossovers have occurred within the 15-mers, resulting in an altered n value, although other homologies within the 5 S DNA sequence can also support unequal events. Increasing n in the parental inserts modestly increases the overall frequency of recombination and the percentage of altered inserts. We conclude that, in a bacterial setting, the 15-base-pair repeats stimulate recombination only slightly by allowing alternative registers for heteroduplex formation. The degree of stimulation observed is less than predicted by one simple model.

Animals↗

PubMed1984

Does early diagnosis and delivery in acute fatty liver of pregnancy lead to improvement in maternal and infant survival?

Two cases of acute fatty liver of pregnancy resulting in maternal and infant survival are described. There have only been six such cases reported previously. The two described here are unique because the diagnosis was made prepartum by an oil red O stain of a frozen section of a liver biopsy, and the patients were promptly delivered by cesarean section under spinal anesthesia. The role of early diagnosis and delivery in this disease is discussed.

Acute Disease↗

PubMed1984

T-cell abnormalities in inflammatory bowel disease are mediated by interleukin 2.

Inflammatory bowel disease (IBD) may be an immunologically mediated disorder in which T cells are unable to respond appropriately to cell surface-associated antigens. To test this possibility, 37 patients with IBD, 24 with Crohn's disease and 13 with ulcerative colitis who were not being treated with immunosuppressive therapy were studied. The ability of T cells to proliferate in response to autologous or allogeneic cells, i.e., the autologous or allogeneic mixed-lymphocyte reaction (MLR) was tested. The autologous MLR was depressed using patient cells compared to control cells, regardless of disease type or activity (1564 +/- 223 cpm versus 3300 +/- 381 cpm, P less than 0.05) while the allogeneic MLR was depressed in patients with active disease only (29,833 +/- 2871 cpm versus 46,799 +/- 3340 cpm, P less than 0.01). The ability of T cells to recognize and lyse allogeneic cells, allogeneic cell-mediated lympholysis (CML), was also low in patients with active disease (24 +/- 4% versus 37 +/- 3%, P less than 0.05). Since T-cell proliferation and cytotoxicity depend upon adequate production of and response to a T-cell growth factor, interleukin 2 (IL-2), IL-2 production and responsiveness in IBD were studied. IL-2 production by patient T cells in response to phytohemagglutinin was only 39% of control values, P less than 0.05. The response to IL-2 was measured by the increase in T-cell proliferation in the autologous MLR in medium alone or medium supplemented with IL-2. Control T-cell proliferation rose from 3300 +/- 381 cpm to 10,761 +/- 428 cpm with exogenous IL-2 (P less than 0.001). Patient T-cell proliferation rose from 1564 +/- 223 cpm to 6817 +/- 771 cpm with IL-2 (P less than 0.001) but did not reach the level of the IL-2-supplemented control autologous MLR (P less than 0.05). In addition, the percentage of activated patient T cells having Tac antigen (IL-2 receptor) was depressed (P less than 0.05). These findings did not vary with disease type or activity. It is concluded from these data that peripheral blood T lymphocytes from patients with IBD have a diminished response to cell surface antigens which is associated with a decrease in IL-2 production and receptor generation. These defects may be responsible for the depressed T-cell proliferation and cytotoxicity that accompany IBD.

Adolescent↗

PubMed1984

Epidermal taurine transport in marine mussels.

We examined the structural specificity and kinetic characteristics of epidermal taurine transport in the marine mussels Mytilus edulis and M. californianus. The gill was the primary site of taurine uptake in both species, responsible for 70% of the accumulation of radioactively labeled substrate. Taurine transport was inhibited by short-chain beta-amino acids, including beta-alanine and beta-aminobutyric acid (beta-ABA), and by gamma-ABA; alpha-neutral, acidic, and basic amino acids had no effect on this transport. The uptake process was clearly a saturable phenomenon and was adequately described by Michaelis-Menten-type kinetics in both intact animals and isolated preparations of gill. The maximal rates of taurine uptake, expressed per gram of gill tissue, were 8.4 and 3.2 mumol X g-1 X h-1 for M. edulis and M. californianus, respectively. In intact mussels the half-saturation constant for both species ranged from 5 to 8 microM. A mathematical model was developed that describes the epidermal transport of taurine in the gill of the actively pumping mussel. The model takes into account the geometry and pattern of water flow in the gill. Calculations based on this model indicate that because of the convective movement of water through the gill, the actual Michaelis constant of the taurine transporter must be much lower (i.e., 0.5-2.5 microM) than the half-saturation constants determined with intact animals and isolated gills. The model also predicts that the presence of the transporter in the gill can result in a significant reaccumulation of taurine lost from the gill by passive diffusion. We suggest that epidermal taurine transport is involved with maintenance of the high concentrations of this compound commonly associated with gill tissue in mussels.

Amino Acids↗

PubMed1983

Kinetics of sodium succinate cotransport across renal brush-border membranes.

The kinetics of Na/succinate cotransport across renal brush-borders was studied using membrane vesicles. Initial rates of succinate uptake (Js) were estimated from 1-s uptakes measured under voltage-clamped conditions. Lowering the external (cis) sodium concentration reduced the succinate Kt without affecting the Jsmax. Increasing the intravesicular (trans) sodium concentration reduced both Jsmax and Kt. This trans inhibition by Na was relieved by trans-succinate. Varying the membrane potential from -60 to +60 reduced succinate transport in an exponential manner, which was reflected by a Kt effect, i.e. an increase in the Kt with no change in Jsmax. The membrane potential results suggest that the succinate/sodium-carrier complex bears a net positive charge. On the basis of these and earlier observations, we propose that Na/succinate cotransport across renal brush-borders is described by an Iso Ordered Quad Quad reaction kinetics in which three Na ions first bind to the carrier to increase its affinity for succinate.

Animals↗