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Biomedical subjects

W A Olsen

Publications and source records attributed to W A Olsen.

At least 19 recordsLinked to original sources
PubMed1992

The effect of enteral carnitine administration in humans.

We previously determined that the L-carnitine uptake by human duodenal tissue occurs by both active (KT 558 mumol/L) and passive mechanisms. The effects of enteral carnitine was studied in humans. A hamburger meal (345 mumol total carnitine) induced peak jejunal fluid free (unesterified) and short-chain acylcarnitine concentrations (SCAC) of 209 and 130 mumol/L, respectively. Plasma carnitine concentrations and the percent renal reabsorption remained unchanged. By contrast, a pharmacologic dose of free carnitine (25,298 mumol) raised peak intraluminal free and SCAC to 20,660 and 4204 mumol/L. Plasma total carnitine concentrations doubled to 93 mumol/L, and the percent renal reabsorption of free and SCAC declined to 76% and 52%, respectively. In triple-lumen perfusions, 200 mumol carnitine/L was absorbed at 484 nmol.min-1.30 cm-1 jejunum, a rate sufficient for prandial but not pharmacologic assimilation. Our findings indicate that absorption of physiologic and pharmacologic amounts of carnitine occurs predominantly by active transport and passive diffusion, respectively.

Absorption↗
PubMed1990

Uptake of L-carnitine by rat jejunal brush border microvillous membrane vesicles. Evidence of passive diffusion.

We have previously described apparent active transport of carnitine into rat intestinal mucosa with intracellular accumulation against a concentration gradient in a process dependent upon the presence of sodium ions, oxygen, and energy. In the work described here, we sought to define the interaction between carnitine and the brush border membrane, which we presumed contained the transport mechanism. Using isolated rat jejunal brush border microvillous membrane vesicles, we found evidence of passive diffusion alone. We found no evidence of carrier-mediated transport--in particular no saturation over a concentration range, inhibition by structural analogs, transstimulation phenomenon, and no influence of sodium ions, potential difference or proton gradients. We conclude that a carnitine transporter does not exist in the brush border membrane of enterocytes and that other cellular mechanisms are responsible for the apparent active transport observed.

Animals↗
PubMed1987

A study of the molecular pathology of sucrase-isomaltase deficiency. A defect in the intracellular processing of the enzyme.

The intestinal brush-border enzyme sucrase-isomaltase splits sucrose into its component monosaccharides, glucose and fructose. A deficiency of the enzyme leads to sucrose intolerance. We studied the synthesis and intracellular processing of sucrase-isomaltase, using human intestinal explants in organ culture. Pulse-chase experiments with [35S]methionine followed by immunoprecipitation, sodium dodecyl sulfate-polyacrylamide-gel electrophoresis, and fluorography of labeled sucrase-isomaltase demonstrated that the molecule was initially recognized as a protein with a relative molecular weight (Mr) of 205,000. This was apparently converted to a species of 225,000 Mr within two hours. We studied the glycosylation of the protein using endo-beta-N-acetylglucosaminidase H and peptide-N4-(N-acetyl-beta-glucosaminyl)-asparagine amidase digestion of oligosaccharide side chains of the two forms of sucrase-isomaltase. The results showed that the early-appearing 205-kd (kilodalton) molecule contained high-mannose asparagine-linked oligosaccharides, and that the later-appearing, 225-kd molecule contained highly processed (mature) carbohydrate chains. Studies in a patient with primary sucrase-isomaltase deficiency demonstrated normal translation and high-mannose glycosylation of the precursor but a failure in further processing of the oligosaccharides, with subsequent intracellular degradation of the glycoprotein and undetectable enzymatic activity of intestinal sucrase. Abnormal intracellular processing of the enzyme was the probable mechanism of enzyme deficiency in this patient.

Adult↗
PubMed1987

Localization of sucrase-isomaltase in the rat enterocyte.

We used immune electron microscopy to study the intracellular localization of sucrase-isomaltase, an intrinsic glycoprotein of the brush border membrane, to provide insight regarding the sites of its synthesis and intracellular processing and the mechanisms of its transfer to the brush border membrane. We identified the protein by postembedding staining with protein A-colloidal gold and by preembedding staining with peroxidase. The protein was found not only in the brush border membrane, but also in the endoplasmic reticulum including nuclear envelope, Golgi complex, smooth apical vesicles, and to a variable extent in the multivesicular bodies. Our findings are consistent with current concepts of biosynthesis of plasma membrane proteins, with synthesis, translocation, and initial glycosylation occurring at the membrane of endoplasmic reticulum and further processing occurring in the Golgi complex. The findings suggest the possibility that some intracellular degradation of sucrase-isomaltase occurs. Finally, our results appear to indicate that at least the final step of intracellular movement, transfer to the brush border membrane, is mediated by smooth apical membrane vesicles.

Animals↗
PubMed1986

Carnitine transport in human intestinal biopsy specimens. Demonstration of an active transport system.

Although carnitine is present in a variety of foods, the mechanism of its absorption has not been previously studied in humans. We investigated the absorption of carnitine by studying uptake into human intestinal mucosal biopsy specimens. We found evidence of active transport in the duodenum and ileum, but not in the colon. We demonstrated that intracellular concentrations exceeded concentrations in the incubation media at steady states and that uptake against a concentration gradient was abolished by anoxia and by replacement of sodium ion with potassium. Studies of initial rate of uptake over a range of concentrations revealed a curve consistent with a two-component system: a saturable system with a KT of 558 microM and a linear component probably representing passive diffusion. Addition of D-carnitine and L-acetylcarnitine resulted in diminished uptake of L-carnitine, suggesting that these substrates utilize the same transport mechanism. These studies demonstrate the presence of an active intestinal transport system for L-carnitine in human intestinal mucosa.

Adult↗
PubMed1986

Intestinal mucosa in diabetes: synthesis of total proteins and sucrase-isomaltase.

The effects of insulin deficiency on nitrogen metabolism in muscle and liver have been extensively studied with recent in vivo demonstration of impaired protein synthesis in rats with streptozotocin-induced diabetes. Despite the significant contribution of small intestinal mucosa to overall protein metabolism, the effects of insulin deficiency on intestinal protein synthesis have not been completely defined. We studied the effects of streptozotocin-induced diabetes on total protein synthesis by small intestinal mucosa and on synthesis of a single enzyme protein of the enterocyte brush-border membrane sucrase-isomaltase. We used the flooding-dose technique of McNurlan, Tomkins, and Garlick (Biochem. J. 178: 373-379, 1979) to minimize the difficulties of measuring specific radioactivity of precursor phenylalanine and determined incorporation into mucosal proteins and sucrase-isomaltase 20 min after injection of the labeled amino acid. Diabetes did not alter mucosal mass as determined by weight and content of protein and DNA during the 5 days after injection of streptozotocin. Increased rates of sucrase-isomaltase synthesis developed beginning on day 3, and those of total protein developed on day 5. Thus intestinal mucosal protein synthesis is not an insulin-sensitive process.

Animals↗
PubMed1985

In vivo studies of intestinal carnitine absorption in rats.

We have studied small intestinal absorption of carnitine in vivo using a combination of segmental perfusion techniques and bolus intraluminal injection. We found evidence of a partially saturable absorption process (with Km values of 1035 and 1267 microM for jejunum and ileum calculated for the saturable component) that appeared to be separate from the imino acid transport system. Absorption was characterized by slow mucosal uptake, prolonged mucosal retention, and a very slow mucosal exit process with blood levels of [3H] carnitine still rising 8 h after intraluminal administration. We have also demonstrated the presence of carnitine acetyltransferase in intestinal mucosa and have shown that the intestine forms significant amounts of acetylcarnitine from exogenous carnitine.

Animals↗
PubMed1985

Studies of carnitine metabolism in relation to intestinal absorption.

We studied the postabsorptive fate of L-[3H]carnitine after intraluminal injection into the proximal intestine of anesthetized rats. Carnitine absorption was characterized by slow appearance in the circulation with blood levels still rising 2 h after administration. Absorption via the portal vein was followed by hepatic extraction and appearance in bile with reabsorption of a fraction, thus establishing an enterohepatic circulation. About half of the [3H]carnitine in blood obtained 4 h after administration was free, with the rest largely acetylcarnitine. In contrast the increase in blood carnitine content after intraluminal administration of unlabeled carnitine was almost exclusively limited to the esterified fraction. We hypothesize that release of esterified endogenous or stored carnitine from some other site accounted for the increase in esterified carnitine. The liver may be that site: although about 50% of hepatic [3H]carnitine was in ester form after administration of labeled carnitine, the increase after unlabeled carnitine was primarily in the free fraction, suggesting that a large amount of esterified carnitine had been released. Thus the liver appears to be an important storage and excretory site for exogenous as well as endogenous carnitine, which may be released with an appropriate signal from the intestine.

Acetylcarnitine↗
PubMed1984

Magnesium sulfate-induced water secretion in hamster small intestine.

We studied possible mechanisms of magnesium sulfate (MgSO4)-induced diarrhea. In vivo perfusion of hamster small intestine with an isotonic electrolyte solution containing 50 mM MgSO4 produced nearly three times as much fluid secretion as did a solution containing an equiosmotic amount of mannitol. We found that magnesium was absorbed at a faster rate than mannitol under these conditions, suggesting that differences in solute permeability do not explain the differences in secretory rates. Magnesium ion rather than sulfate appeared largely responsible for the effect as replacement of sulfate with chloride did not diminish the response. MgSO4 perfusion of a proximal intestinal segment did not affect water transport in an isolated distal segment suggesting that release of cholecystokinin or alterations in serum levels of other hormones were not responsible. Intestinal permeability, morphology, and cyclic nucleotide levels were normal after MgSO4 perfusion. Thus, MgSO4-induced diarrhea cannot be explained by the usual mechanisms, and additional processes responsible for intestinal secretion must exist.

Animals↗
PubMed1983

A bean alpha-amylase inhibitor formulation (starch blocker) is ineffective in man.

A commercial alpha-amylase inhibitor with potent inhibitory activity in vitro was used in a randomized double-blind, cross-over clinical trial in six nonobese, healthy adult males. In these subjects, this inhibitor had no effect on the response of blood glucose, insulin, or breath hydrogen to a standardized starch meal. It is concluded that this formulation has no effect on starch digestion in humans.

Amylases↗
PubMed1983

Intestinal mucosa in diabetic rats: studies of microvillus membrane composition and microviscosity.

In experimental diabetes, a number of intestinal brush-border hydrolases and transport systems are stimulated. In this study, we assessed possible effects of diabetes on the composition and membrane fluidity of rat intestinal brush-border membranes that might correlate with these functional changes. We found similar proportions of lipid and protein in the diabetic and control preparations, although there was a considerable increase in total membrane from the diabetic rats, presumably reflecting mucosal hyperplasia. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of membrane protein revealed an increase in the bands corresponding to sucrase-isomaltase, consistent with an increased enzyme activity of sucrase. Membrane lipid analysis revealed only a decrease in fatty acids of the neutral lipid fraction of diabetics--a change that may well have occurred during membrane preparation. 1-6-Diphenyl-1,3,5-hexatriene fluorescence polarization data, obtained as a function of temperature, was similar for the diabetic and control rats, with a three-phase linear model superior to one- and two-phase linear or quadratic models. The overall composition of the intestinal brush-border membrane, unlike other plasma membranes, appears little affected by experimental diabetes.

Animals↗
PubMed1983

Carnitine transport in rat small intestine.

Although L-carnitine has been given orally to patients with systemic carnitine deficiency with successful control of the disease and is present in a variety of dietary sources, there is little available information on the physiology of its absorption. We therefore studied intestinal carnitine absorption in the rat by measuring the uptake of radioactive L-carnitine by everted intestinal rings and sacs. Active transport was demonstrated in duodenum and jejunum, but not ileum, with intracellular concentrations higher than medium concentrations at steady state and by the prevention of concentration gradients with anoxia, metabolic inhibitors, and replacement of sodium ion. Studies of the relationship of uptake to carnitine concentration demonstrated the presence of two components of transport: a saturable component (with a Km of between 206 and 316 microM) that could be inhibited by the metabolically inactive D-isomer and by acetylcarnitine and a linear component that we presume represents diffusion.

Anaerobiosis↗
PubMed1982

Sucrase metabolism in germfree rats.

We studied degradation of the intestinal brush-border protein sucrase-isomaltase in germfree animals as well as in ex-germfree animals and conventional controls to determine the mechanism by which intestinal bacteria alter disaccharidase levels. Our results indicate that sucrase-isomaltase turnover is as rapid in germfree animals as in the other groups and are consistent with surface removal of disaccharidases by pancreatic proteases under physiological conditions. Our results also suggest that elevated disaccharidase levels in germfree animals are in large part the consequence of an increased number of mature enterocytes, which persists for at least 2 wk after exposure of animals to a conventional microbial flora.

Animals↗
PubMed1982

In vivo responses of rat intestinal epithelium to intraluminal dietary lectins.

Although a variety of plant lectins are consumed as part of the normal human diet and are capable of binding to intestinal cell surfaces in vitro, little information exists on their effects on intact intestine. We have studied the acute effects of intraluminal administration of wheat germ agglutinin and concanavalin A in normal rats. Both lectins caused increased shedding of brush border membrane and, at higher concentrations, reduction in surface area, acceleration of cell loss, and shortening of villi. These changes were prevented by simultaneous administration of the appropriate sugar to inhibit binding, indicating that the effects were related to binding to carbohydrate residues of intestinal cells. Similar changes of brush borders were found after intraluminal administration of antiserum to sucrase-isomaltase, a surface protein of the brush border membrane, suggesting that the lectin effects resulted from cell surface receptor-lectin interaction rather than a primary intracellular effect. Our results suggest that dietary lectins may be in part responsible for normal turnover of brush border membrane, and support, in addition, the possibility that certain intestinal diseases such as celiac sprue may be the consequence of increased levels of lectin receptor allowing a dietary lectin to exert a toxic effect.

Animals↗
PubMed1981

The effect of bilirubin on the function of hamster small intestine.

Jaundice phototherapy is associated with a significant incidence of watery diarrhea. We have postulated that acute intestinal secretion, rather than malabsorption of dietary carbohydrate, is an effect of a photoproduct of bilirubin upon the intestinal mucosa. Because of major effect of phototherapy is the hepatic excretion of nonconjugated bilirubin, we investigated the effect of bilirubin on small intestinal function in the hamster in vivo. The entire small intestine was luminally perfused in vivo with solutions containing bilirubin (0.125 to 0.75 mmole/liter) and net water and sodium fluxes were measured. Control animals absorbed both water (J H2O(net) = 58.9 microliter/min/g) and sodium (J Na(net) = 4.55 microEq/min/g), but animals perfused with bilirubin (greater than or equal to 0.25 mmole/liter) exhibited secretion of water (J H2O(net) = -39.0--85.9) and sodium (J Na(net)=-9.91--18.24). The rate of water secretion was positively related to the concentration of bilirubin in the infusate (r=0.749; p less than 0.001). The concentration of bilirubin in ultrafiltrates of perfusate was likewise positively related to its concentration in the infusate (r = 0.844; p less than 0.001), indicating the potential importance of soluble forms of bilirubin in inducing secretion. Possible epithelial injury was studied by measuring the concentration of DNA in the perfusate and the activity of disaccharidases in postperfusion mucosa, and the possible role of cyclic adenosine monophosphate as a mediator of the secretory process was investigated by determining its concentration in postperfusion mucosa. Perfusion with 0.5 mM bilirubin, which produced significant secretion, did not cause loss of DNA (0.284 versus 0.244 mg/liter) or mucosal lactase activity (56 versus 53 units/g) or enhancement of cyclic adenosine monophosphate concentration (14.9 versus 14.12 pmoles/mg protein).

Animals↗
PubMed1980

Circadian rhythm of intestinal sucrase activity in rats. Mechanism of enzyme change.

Past investigation has revealed that the circadian rhythm of intestinal sucrase activity in rats is primarily cued by the time of feeding. We examined the mechanism of the circadian rhythm by methods involving quantitative immunoprecipitation of sucrase-isomaltase protein and study of decay of radioactively labeled protein. Rats were placed on a controlled feeding regimen (1000-1500 h) and then sacrificed at 3-h intervals over a 24-h period. Immunotitration experiments indicated that the circadian rhythm was the result of changes in the absolute amount of sucrase-isomaltase protein present and not of changes in the enzyme's catalytic efficiency. To study the mechanism of this circadian variation in sucrase-isomaltase mass, [(14)C]sodium carbonate was injected and, after maximum incorporation into brush border protein, the rats were sacrified at 3-h intervals. Sucrase-isomaltase protein was isolated by immunoprecipitation, and the decrease in total disintegrations per minute over time was used to study degradation of the protein. Enzyme degradation was not constant but exhibited a clear circadian rhythm. The period of increasing enzyme mass was characterized by virtual cessation of enzyme degradation (t((1/2)) of 38 h), and the period of declining enzyme mass by rapid degradation (t((1/2)) of 6 h or less). We found similar changes in enzyme degradation in fasted animals, demonstrating that the changes were not the result of decreased isotope reutilization during feeding. We found no evidence of a circadian rhythm in [(14)C]leucine incorporation into the protein, suggesting that enzyme synthesis was constant. These results indicate that the circadian rhythm of sucrase activity represents changes in the total amount of enzyme protein that are, at least in large part, secondary to changes in the enzyme's degradation rate.

Animals↗