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B R Stevens

Publications and source records attributed to B R Stevens.

54 records · Page 3Linked to original sources
PubMed1988

Human intestinal brush border angiotensin-converting enzyme activity and its inhibition by antihypertensive Ramipril.

Angiotensin-converting enzyme (ACE) has been identified as a prominent brush border membrane-bound enzyme of human jejunum. In this study, we purified brush border membrane vesicles enriched in ACE, and characterized the ACE with regard to (a) its stability in the membrane, (b) substrate hydrolysis kinetics compared with pulmonary endothelial ACE, and (c) pharmacologic interaction with Ramipril. These investigations resulted in the following findings. The uninhibited enzyme is stable in native membranes in vitro, with a half-life of 195 +/- 7 h. Kinetic analysis of ACE hydrolysis activity revealed the presence of a single enzyme species, which yielded a high Vmax and displayed a Km similar to purified ACE from lung endothelium. Brush border ACE was inhibited by Ramipril, one of the most specific and potent orally administered ACE inhibitors indicated for hypertension. We determined the brush border ACE value of IC50 = 3 X 10(-9) M Ramipril-diacid, which is the same value for serum and lung ACE. Brush border ACE remains 100% inhibited by 10 microM Ramipril during at least 8 days in vitro. The data indicate that ACE is a prominent jejunal brush border enzyme that behaves pharmacologically and kinetically like its peripheral circulation counterpart. This study suggests that high doses of orally administered ACE inhibitors may affect intestinal epithelial function.

Angiotensin-Converting Enzyme Inhibitors↗
PubMed1988

Intestinal brush border membrane-bound disaccharidases of the American alligator, Alligator mississippiensis.

1. The disaccharidase activities of the small intestines of American alligators (Alligator mississippiensis) were studied in epithelial scrapes and brush-border membrane preparations. 2. Maltase, isomaltase and trehalase activities were found. Activities of these enzymes were higher in the proximal small intestine and decreased distally. 3. Disaccharidase activities were enriched 12-15 times in brush-border membrane preparations, compared with mucosa/enterocyte crude homogenates and were co-enriched with the brush-border membrane marker alkaline phosphatase. 3. The pH optima were: maltase 6.5; isomaltase 5.6; and trehalase 5.8. The Q10 of maltase, the most active enzyme, was equal to 1.82. 4. In reptiles, as in mammals, disaccharidase activities may be correlated with feeding habits. The co-occurrence of sucrase and isomaltase may not be a common feature of vertebrates.

Alligators and Crocodiles↗
PubMed1988

Ramipril inhibition of rabbit (Oryctolagus cuniculus) small intestinal brush border membrane angiotensin converting enzyme.

1. Rabbit small intestinal brush border membranes possessed prominent angiotensin converting enzyme (ACE) activity. 2. Intestinal ACE was located on the lumen surface, as verified by ACE co-enrichment with brush border membrane marker enzymes. 3. Hydrolysis kinetics of rabbit intestinal ACE were comparable to the lung, utilizing the substrate (N-[3-(2-furyl)acryloyl]-L-phenylalanylglycylglycine; the Vmax = 543 +/- 51 mumol/min/g and Km = 0.62 +/- 0.09 mmol/l. 4. Intestinal brush border ACE activity was strongly inhibited by the antihypertensive drug Ramipril, which yielded an IC50 value of 5 nmol/l; the ACE activity remained completely inhibited during 15 days after a single dose of 10 mumol/l Ramipril.

Angiotensin-Converting Enzyme Inhibitors↗
PubMed1988

Angiotensin converting enzyme in brush-border membranes of avian small intestine.

Angiotensin converting enzyme activity was identified in brush-border membranes purified from the small intestinal epithelium of the common grackle, Quiscalus quiscula. Angiotensin converting enzyme was enriched 20-fold in the membrane preparation, compared with intestinal epithelial cell scrapes, and was coenriched with the brush-border markers, alkaline phosphatase and aminopeptidase N. The kinetics of hydrolysis of N-[3-(2-furyl)acryloyl]-L-phenylalanylglycylglycine (FAPGG) gave a Vmax of 907 +/- 41 units g-1 and a Km of 55 +/- 6 mumol l-1. The avian intestinal angiotensin converting enzyme was inhibited by the antihypertensive drug, Ramipril, with a median inhibitory concentration (IC50) of 1 nmol l-1. In the light of previous studies on angiotensin converting enzyme in mammalian epithelia, these results may implicate a physiological role for angiotensin converting enzyme in regulating electrolyte and fluid uptake in bird small intestines.

Animals↗
PubMed1987

Kinetics of the intestinal brush border proline (Imino) carrier.

The kinetics of L-proline transport across intestinal brush borders via the Imino carrier were studied using membrane vesicles. The Imino carrier is defined as the agent responsible for L-alanine insensitive. Na+-dependent uptake of L-proline. Initial rate measurements were made under voltage clamped conditions (pD = 0) to investigate L-proline transport as a function of cis and trans Na+ and proline concentrations. Under zero-trans conditions, increasing cis Na+ activated proline uptake with a Hill coefficient of 1.7 and decreased the apparent Kt with no change in Jimax. The Jimax was approximately 60 pmol mg-1 s-1 and the apparent Kt ranged from 0.25 mM at cis Na = 100 to 1.0 mM at cis Na+ = 30 mM. Trans Na inhibited proline uptake via a reduction in Jimax. Trans proline had no significant effect in the absence of trans Na+, but it relieved the trans Na+ inhibition. Under equilibrium exchange conditions, the Jimax was twice that observed under zero-trans conditions. These kinetics of L-proline transport suggest a model in which uptake occurs by a rapid equilibrium iso-ordered ter ter system. Two Na+ ions bind first to the carrier on the cis face of the membrane to increase the affinity of the carrier for proline. The fully loaded complex then isomerizes to release the substrates to the trans side. The partially loaded Na+-only forms are unable to translocate across the membrane. A rate-limiting step appears to be the isomerization of unloaded carrier from the trans to the cis side of the membrane.

Amino Acid Transport Systems, Neutral↗
PubMed1987

Dipeptidyl peptidase-II activity in cultured astroglial cells from neonatal rat brain.

Astrocytic glial cells in primary culture from neonatal rat brain possess prominent dipeptidyl peptidase-II activity. This enzyme has been previously isolated and purified from whole brain tissue. The glial enzyme characteristically hydrolyzed glycyl-proline-p-nitroanilide (GPN) substrate to release glycyl-proline dipeptide plus p-nitroaniline products. At the enzyme's optimal pH of 5.4, the activities of other tested amino exopeptidases were virtually zero. At pH greater than 8, activity was less than 2% of the activity at pH 5.4, which suggested a paucity of the related enzyme, dipeptidyl peptidase-IV. No competitive inhibition was observed for glycine, proline nor their permuted dipeptides. Glial dipeptidyl peptidase-II activity was strongly inhibited by Hg2+, while other redox sulfhydryl agents were ineffective. Tested cations did not affect activity, except K+ which was mildly inhibitory. Chelating agents were not inhibitory. Of the peptidase inhibitors tested, only phenylmethylsulfonyl fluoride and puromycin were partially inhibitory. We suggest that dipeptidyl peptidase-II may play a role in glial processing of brain peptides which possess an N-terminal penultimate proline residue.

Animals↗
PubMed1987

Ubiquitin-directed antibodies inhibit neuronal transporters in rat brain synaptosomes.

Affinity-purified antibodies specific for ubiquitin were found to inhibit the sodium-dependent uptake of [3H]choline, gamma-[3H]aminobutyric acid [( 3H]GABA), [3H]glutamate, [3H]norepinephrine, [3H]aspartate, and [3H]serotonin in rat cerebral cortical synaptosomes at a low concentration (10 micrograms/ml). These antibodies (termed anti-Ub) had no effect on the sodium-independent uptake of these substances or their calcium-dependent efflux. Synaptosomal [3H]deoxyglucose uptake was not affected in normal Krebs Ringer buffer containing 10 mM glucose, but was inhibited in glucose-free medium. Other nonneuronal sodium-dependent transport processes were found to be unaffected by 10 micrograms/ml anti-Ub, suggesting that anti-Ub does not bind indiscriminantly to sodium-binding sites on sodium-dependent organic solute transporters. Finally, anti-Ub inhibited sodium-dependent [3H]GABA and [3H]glutamate uptake in plasma membrane ghosts, devoid of membrane potential, which were derived from rat cerebral cortical synaptosomes. These results suggest that neuronal transporters or sites proximal to them may be ubiquitinylated on the plasma membrane surface.

Animals↗
PubMed1986

Radiation inactivation probe of membrane-bound enzymes: gamma-glutamyltranspeptidase, aminopeptidase N, and sucrase.

gamma-Glutamyltranspeptidase (GGT), aminopeptidase N (AP-N), and sucrase in purified rabbit intestinal brush border membrane vesicles were irradiated in situ at -135 degrees C using high energy electrons. Surviving activities of the enzymes were measured as a function of radiation dose, and the functional unit target sizes (corresponding to carbohydrate-free polypeptides) were determined using target analysis. The in situ functional unit sizes were GGT 59 kDa, AP-N 59 kDa, and sucrase 63 kDa. Together with biochemical data determined previously, it is concluded that the noncovalently attached large (approximately 40 kDa) and small (approximately 25 kDa) subunits of GGT are both required for catalytic activity. Furthermore, these data suggest that (i) the membrane-bound form of AP-N consists of one or more noncovalently attached subunits of 59 kDa, each of which is enzymatically active; and (ii) in situ sucrase activity is associated with a subunit of 63 kDa which is noncovalently attached within the sucrase-isomaltase complex.

Aminopeptidases↗
PubMed1985

Specificity of intestinal brush-border proline transport: cyanine dye studies.

The ability of rabbit jejunal brush borders to transport inhibitors of the imino carrier was investigated in membrane vesicles by measuring their ability to depolarize the membrane potential. Membrane potentials were monitored using a voltage-sensitive cyanine dye. Piperidine and pyrrolidine carboxylic acids, which are potent inhibitors of Na+-dependent proline transport (Ki less than 0.5 mM) depolarize the potential in a Na+-dependent, saturable manner indicating transport. On the other hand, N-methylated amino acids, which are fair inhibitors (Ki 2-10 mM), do not depolarize the membrane to any significant extent, but they competitively inhibit the L-proline transport signal. This indicates that these analogs are nontransported inhibitors of the imino carrier. The poor inhibitors niacin and pipolinic acid (Ki greater than 60 mM) depolarize the membrane about twice as much as proline and with low Kf values. This suggests separate carriers for these substrates.

Amino Acids↗
PubMed1985

Substrate specificity of the intestinal brush-border proline/sodium (IMINO) transporter.

L-proline uptake via the intestinal brush-border IMINO carrier was tested for inhibition by 41 compounds which included sugars, N-methylated, alpha-, beta-, gamma- and epsilon- amino and imino acids, and heterocyclic analogs of pyrrolidine, piperidine and pyridine. Based on competitive inhibitor constants (apparent Ki' 's) we find that the IMINO carrier binding site interacts with molecules which possess a well-defined set of structural prerequisites. The ideal inhibitor must 1) be a heterocyclic nitrogen ring, 2) have a hydrophobic region, 3) be the L-stereoisomer of 4) an electronegative carbonyl group which is 5) separated by a one-carbon atom spacer from 6) an electropositive tetrahedral imino nitrogen with two H atoms. Finally, 7) the inhibitor conformation determined by dynamic ring puckering must position all these features within a critical domain. The two best inhibitors are L-pipecolate (apparent Ki' 0.2 mM) and L-proline (apparent Ki' 0.3 mM).

Amino Acid Transport Systems, Neutral↗
PubMed1984

Intestinal transport of amino acids and sugars: advances using membrane vesicles.

Glucose crosses the intestinal brush border via one diffusive and two saturable systems. Under zero-trans conditions, the major saturable system has the characteristics of an Iso Ordered Bireactant system with one Na binding and, additionally, shows the properties of trans inhibition by Na and sensitivity to Em. An additional high-affinity saturable system exists with a Kt in the micromolar range and has the capacity for binding multiple Na+ ions. Biochemical and transport studies indicate that the brush-border Na-glucose cotransporter may be asymmetric with respect to the inner and outer membrane face. The Na-independent glucose carrier found in enterocyte basolateral membranes strongly resembles the facilitated glucose carrier found in other membranes.

Amino Acids↗
PubMed1983

Kinetics of sodium-dependent solute transport by rabbit renal and jejunal brush-border vesicles using a fluorescent dye.

The kinetics of Na-coupled solute transport by renal and jejunal brush-border vesicles in the rabbit were examined using the potential-sensitive fluorescent dye diS-C3-(5). All organic solutes known to be transported across these membranes by Na-coupled mechanisms increase the fluorescence of the dye in the presence of Na, but not K. An increase in fluorescence (delta F) corresponds to a depolarization of the electrical potential difference (5-60 mV) across the brush-border membrane in the intact cell. delta F was independent of the valency of the transported solute. The fluorescence response was saturable, and for twelve solutes the Kf, i.e. the concentration of the substrate generating 50% of the maximal response, agreed quite closely with the Kt values reported from tracer studies. For six solutes increasing the Na concentration decreased Kf, and this agrees with the effect of Na on the kinetics of succinate transport in renal vesicles. We conclude that D-glucose, neutral amino acids and imino acids are co-transported with Na across both renal and jejunal brush-border membranes, and that carboxylic acids, beta-amino acids, and dibasic amino acids are co-transported with Na across the renal, but not jejunal, membranes.

Amino Acids↗
PubMed1982

Multiple transport pathways for neutral amino acids in rabbit jejunal brush border vesicles.

Amino acids enter rabbit jejunal brush border membrane vesicles via three major transport systems: (1) simple passive diffusion; (2) Na-independent carriers; and (3) Na-dependent carriers. The passive permeability sequence of amino acids is very similar to that observed in other studies involving natural and artificial membranes. Based on uptake kinetics and cross-inhibition profiles, at least two Na-independent and three Na-dependent carrier-mediated pathways exist. One Na-independent pathway, similar to the classical L system, favors neutral amino acids, while the other pathway favors dibasic amino acids such as lysine. One Na-dependent pathway primarily serves neutral L-amino acids including 2-amino-2-norbornanecarboxylic acid hemihydrate (BCH), but not beta-alanine or alpha-methylaminoisobutyric acid (MeAIB). Another Na-dependent route favors phenylalanine and methionine, while the third pathway is selective for imino acids and MeAIB. Li is unable to substitute for Na in these systems. Cross-inhibition profiles indicated that none of the Na-dependent systems conform to classical A or ACS paradigms. Other notable features of jejunal brush border vesicles include (1) no beta-alanine carrier, and (2) no major proline/glycine interactions.

Alanine↗
PubMed1982

Organic and inorganic solute transport in renal and intestinal membrane vesicles preserved in liquid nitrogen.

The transport of organic solutes (sugars, amino acids, and metabolic intermediates) and inorganic solutes (Na+/H+ exchange and Na+-SO = 4 cotransport) in renal brush border and in intestinal brush border and basal lateral membrane vesicles is preserved when the vesicles are stored in liquid nitrogen. The preservation allows comparisons among transport systems of renal and intestinal cells obtained from the same animal.

Amino Acids↗
PubMedDate not supplied

Process of care and outcomes for elderly patients hospitalized with peptic ulcer disease: results from a quality improvement project.

CONTEXT: Since publication in 1994 of guidelines for management of peptic ulcer disease (PUD), trends in physician practice and outcomes related to guideline application have not been evaluated. OBJECTIVES: To describe changes in process of care that occurred in a quality improvement program for patients hospitalized with PUD and to evaluate associations between in-hospital treatment of PUD and 1-year rehospitalization for PUD and mortality in a subset of these patients. DESIGN, SETTING, AND PATIENTS: Cohort study of 4292 sequential Medicare beneficiaries hospitalized at acute care hospitals with a principal diagnosis of PUD in 5 states (Colorado, Georgia, Connecticut, Oklahoma, and Virginia) in 1995 (baseline) and 1997 (remeasurement); outcomes were evaluated for 752 patients in Colorado. MAIN OUTCOME MEASURES: Changes in rates of screening for Helicobacter pylori infection, treatment for H pylori infection, screening for nonsteroidal anti-inflammatory drug (NSAID) use, counseling about NSAID use; outcomes included rehospitalization for PUD and all-cause mortality within 1 year of discharge in Colorado. RESULTS: Screening for H pylori infection increased significantly (12%-19% increase; P<.001) in each of the 5 states. Treatment of H pylori infection increased in each state and was significantly increased for the entire group of hospitalizations examined (8% increase overall; P =.001). Despite increased screening, detection of H pylori infection was less frequent than expected in every state, (13%-24%) and did not increase in any state. Screening for and counseling about NSAIDs did not significantly increase overall or in any state. In the Colorado cohort, the proportion of patients rehospitalized was unchanged in 1995 (8.9%) and 1997 (6.8%), and 124 patients (16%) in the combined 1995 and 1997 cohorts died within 1 year. Treatment for H pylori was not associated with a reduction in rehospitalization within 1 year (adjusted odds ratio [OR], 1.24; 95% confidence interval [CI], 0.65-2.36) or with a reduction in mortality (adjusted OR, 1.08; 95% CI, 0.68-1.71). Counseling about NSAID use was associated with a decrease in risk of 1-year rehospitalization for PUD (adjusted OR, 0.47; 95% CI, 0.22-0.99) and risk of all-cause mortality (adjusted OR, 0.44; 95% CI, 0.26-0.75). CONCLUSIONS: This quality improvement program for elderly patients with PUD resulted in increased screening for H pylori and increased treatment of H pylori infection but no change in counseling about NSAID use. However, with the low prevalence of H pylori detected, treatment of H pylori infection was not associated with a reduction in repeat hospitalization for PUD or subsequent mortality, whereas counseling about the risks of using NSAIDs was associated with a reduction in the risk of both outcomes.

Aged↗