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Biomedical subjects

M A Dudley

Publications and source records attributed to M A Dudley.

29 records · Page 2Linked to original sources
PubMed1992

Feeding status affects in vivo prosucrase.isomaltase processing in rat jejunum.

Fed and fasted (18 h) adult male rats received a primed constant infusion of [3H]leucine for 15 to 180 min. Prosucrase-isomaltase (pro-SI) and sucrase were isolated from mixed jejunal mucosal membranes by immunoprecipitation and separated from one another by polyacrylamide gel electrophoresis under denaturing and reducing conditions. The rate at which pro-SI was processed to sucrase was calculated on the assumption that the steady-state specific radioactivity of leucine in pro-SI defined the pool of amino acids used in the formation of brush border sucrase. At isotopic steady state, pro-SI achieved a specific radioactivity that was higher than that of mucosal free leucine in both feeding groups. The relationship between the isotopic equilibrium of the free amino acid pools and pro-SI was sensitive to feeding status; the specific radioactivity of pro-SI was 25 and 55% (P less than 0.05) of the blood specific radioactivity in fed and fasted animals, respectively. The fractional rate of pro-SI processing tended to be higher (P less than 0.07) in fed (407%/d) than in fasted animals (274%/d). We conclude that the general mucosal free amino acid pool is not the amino acid pool from which pro-SI is synthesized and that the rate of pro-SI processing from the endoplasmic reticulum-Golgi membranes to the brush border membrane is sensitive to the feeding status of the animal.

Amino Acids↗
PubMed1992

Effect of oral insulin on lactase activity, mRNA, and posttranscriptional processing in the newborn pig.

Insulin, found in human and pig colostrum and mature milk, appears to influence small intestinal growth and development. Ileal lactase activity is increased when porcine insulin is added to feedings administered to newborn piglets. We studied 2-day-old miniature piglets to determine whether the increase in lactase activity is accompanied by changes in enterocyte expression of lactase activity, steady-state levels of lactase mRNA, and/or posttranscriptional changes in lactase processing. We randomized the piglets to receive bottle feedings of a swine-weaning milk formula with (group F + I) or without (group F) the addition of 85 mU/ml of regular porcine insulin. The piglets were fed for 6 days (to 8 days of age), after which they were killed and the small intestine removed for analysis. Despite large differences between groups in enterocyte expression of lactase activity in the ileum, no differences were noted in the level of ileal lactase mRNA. The relative proportions of the 207, 210, and 230 kDa precursors of the 160 kDa mature lactase protein were similar between groups. These data indicate that the insulin-induced increased expression of ileal lactase activity is not regulated at the level of its mRNA or at the level of processing of the polypeptide.

Administration, Oral↗
PubMed1992

Developmental changes in lactase-phlorizin hydrolase precursor isoforms in the rat.

The purpose of this study was to determine whether the developmental decline in lactase specific activity (mumol/min/g protein) in the rat was associated with (a) changes in the relative quantities of immunoisolated precursor and mature forms of the enzyme purified by SDS-PAGE and/or (b) immunohistologic changes in the jejunal mucosa. We studied 10- and 16-day-old suckling rat pups, 22-day-old weaned rat pups, and adult female rats (nongravid, pregnant, and lactating). Lactase activity was three- to fourfold higher in 10-day-old pups than in adult rats. Lactase activity was 27% greater in lactating compared with nongravid or pregnant rats. Three molecular forms of the enzyme that migrated identically in all animals were observed on SDS-polyacrylamide gels stained with Coomassie blue: 140-kDa (mature brush border form), 200-kDa, and 220-kDa (apparent precursor forms). There was a striking difference in the proportions of the three polypeptides at different ages that was unrelated to animal status, i.e., pregnant or lactating. As the animals aged, the relative amount of the 140-kDa band declined from 86 +/- 1.1% of the total immunoprecipitated lactase in 10-day old suckling pups to 68 +/- 0.7% in adults. Simultaneously, the relative concentration of the 200-kDa band rose from 1.7 +/- 0.4% in the 10-day-old to 19 +/- 0.6% in adults. The relative concentration of the 220-kDa polypeptide did not change as a function of age.(ABSTRACT TRUNCATED AT 250 WORDS)

Aging↗
PubMed1988

Diluting power of thick limbs of Henle. II. Bumetanide-sensitive 22Na+ influx in medullary vesicles.

We evaluated the effects of osmotic gradients on 22Na+ influx in vesicles prepared from rat outer renal medulla. 22Na+ influx driven in a coflow mode by an inwardly directed 100 mM KCl gradient was measured at 20 and 60 s; 1 mM bumetanide inhibited approximately 30% of 22Na+ influx. The bumetanide-sensitive 22Na+ influx was reduced by approximately 65% when either K+ or Cl- was omitted from the aqueous phases. We found that an osmotic gradient for vesicle shrinkage, that is, 600 mM urea in the extravesicular medium, enhanced the bumetanide-sensitive 22Na+ influx twofold. Conversely, an osmotic gradient for vesicle swelling, that is, with vesicles but not extravesicular media loaded with 600 mM urea, produced a 50% suppression of bumetanide-sensitive 22Na+ influx. Moreover, 600 mM extravesicular urea, an osmotic gradient for vesicle shrinkage, also reduced uptake of the nonspecific marker [14C]mannitol. These effects of osmotic gradients were not due to alterations in ionic driving forces, since bumetanide-sensitive 22Na+ influx driven in a counterflow mode by loading the vesicles with 100 mM NaCl also was activated or suppressed by osmotic gradients for vesicle shrinkage or swelling, respectively. We conclude that osmotic gradients, and/or vesicle volume changes, modulate bumetanide-sensitive Na+:K+:2Cl- activity.

Alkaline Phosphatase↗
PubMed1986

Rapid vesicle formation and aggregation in abnormal human biles. A time-lapse video-enhanced contrast microscopy study.

Rapid nucleation of cholesterol crystals has previously been shown to provide a sharp discrimination between abnormal (cholesterol gallstone-associated) and normal human gallbladder bile. In the present study, we sought to further clarify the crystal nucleation process by time-lapse microscopy using a novel high-resolution video-enhanced microscopy technique. Using a previously described method for removal of particles from abnormal biles, we found a strikingly rapid rate of de novo formation of unilamellar vesicles, soon followed by massive vesicular aggregation, culminating in crystal formation. In normal biles, by contrast, this rapid aggregation process was not observed and the isolated unilamellar vesicles showed prolonged stability. Morphometric analysis of interval particle counts showed statistically significant differences. The process of cholesterol monohydrate crystal nucleation in supersaturated human bile is characterized by a sequential combination of vesicle formation, vesicle aggregation, and subsequent crystal formation. The primary distinction between abnormal and normal biles resides only in the consistent rapidity of onset and completion of these events in the abnormal biles.

Bile↗
PubMed1986

Vesicle aggregation in model systems of supersaturated bile: relation to crystal nucleation and lipid composition of the vesicular phase.

The presence of small vesicles composed of phospholipid and cholesterol has recently been demonstrated in super-saturated model and in dilute native human biles by several groups using differing methods. Among compositional factors shown to favor spontaneous vesicle formation and prolong the cholesterol monohydrate nucleation time in model bile systems are dilution, a raised cholesterol saturation index (CSI), and a low bile salt/phospholipid ratio. Time-lapse video-enhanced microscopy of a series of model bile systems representing systematically designed variations in the above factors revealed strong evidence for an essential linkage between antecedent vesicle aggregation and subsequent crystal nucleation. Stability of vesicles was inversely related to their degree of cholesterol saturation, i.e., the greater the degree of vesicular cholesterol saturation, the less their stability (metastability). Instability of vesicles was reflected by their early aggregation followed by rapid cholesterol crystal nucleation. The lowest degree of vesicular cholesterol saturation was found in dilute systems which also exhibited the greatest metastability despite a high degree of cholesterol solubility (raised CSI). Conversely, the more concentrated and least metastable systems exhibited both rapid vesicle aggregation and rapid onset of crystal nucleation. These systems, while influenced by the other compositional factors, were found to have a high degree of vesicular cholesterol saturation, i.e., cholesterol/phospholipid molar ratio = 2.0. An additional finding was the extreme variability in the proportionate distribution of total solution cholesterol distributed to the vesicular phase, i.e., from zero to as high as 37%. Higher solute concentration, raised bile salt/lecithin ratio, and raised CSI were interactive and almost equally capable of increasing the proportionate amount of cholesterol in the vesicular phase. In conclusion, lipid compositional differences in model bile systems drastically affect the cholesterol saturation of spontaneously formed phospholipid-cholesterol vesicles. This effect, in turn, exerts a potent influence upon the metastability of vesicles, subsequently affecting the cholesterol crystal nucleation time.

Bile↗
PubMed1985

Factors affecting cholesterol monohydrate crystal nucleation time in model systems of supersaturated bile.

We explored the influence of several compositional factors considered capable of influencing the nucleation time of model biles supersaturated in cholesterol. In addition to the classical techniques, e.g., electron microscopy and quasielastic light scattering, employed for size measurement and structural assessment, we employed a novel technique, i.e., video-enhanced microscopy, for particle evaluation in these polydisperse systems which often may simultaneously contain isolated small vesicles, their complex aggregates, and small cholesterol monohydrate crystals. The factors we studied included dilution, degree of cholesterol supersaturation, bile salt/lecithin molar ratio, and Ca2+ concentration. Dilution markedly raised the degree of cholesterol saturation, prolonged nucleation time for cholesterol monohydrate crystals, and favored formation of metastable small unilamellar vesicles. Increasing the degree of cholesterol supersaturation as an independent variable in more concentrated systems both shortened the nucleation time and favored spontaneous formation of a relatively small number of isolated vesicles. A decrease in bile salt/lecithin molar ratio within the physiologically relevant range was accompanied by a prolonged nucleation time and favored spontaneous vesicle formation. Large numbers of small unilamellar vesicles were observed even in concentrated model bile solutions (total lipids: 20 g/dl) when the bile salt/lecithin molar ratio was 1.9 or less. At physiological concentrations, Ca2+ promoted nucleation of cholesterol monohydrate crystals only in vesicle-containing solutions. Taken together, the following conclusions can be drawn. First, spontaneous vesicle formation in dilute systems prolongs solid cholesterol crystal nucleation. It can thus provide a supplementary non-micellar mode of cholesterol transport in micellar systems of supersaturated human bile. Second, dilution, degree of cholesterol supersaturation, and a decrease in bile salt/lecithin ratio prolong cholesterol crystal nucleation time and favor spontaneous vesicle formation. With increasing calcium concentrations, opposite effects are observed. Third, the presence of vesicles may help to account for the frequently observed and otherwise unexplained remarkable degree of metastable supersaturation and prolonged metastability (delayed nucleation time) for cholesterol in human bile.

Bile↗
PubMed1970

Kinetics of neutralization of bacteriophage f2 by rabbit gamma-G-antibodies.

The neutralization of bacteriophage f2 by intact gammaG-immunoglobulin or fragments is first order with respect to both bacteriophage and antibody. Minimum values for the rate constants are of the order of 10(7)M(-1) sec(-1). The temperature dependence of the rates corresponds to the activation parameters: DeltaHdouble dagger = 6.7 kcal mole(-1) and DeltaSdouble dagger = -4 cal deg(-1) mole(-1) (gammaG-immunoglobulin); DeltaHdouble dagger = 8.0 kcal mole(-1) and DeltaSdouble dagger = -0.9 cal deg(-1) mole(-1) (5S pepsin fragment); and DeltaHdouble dagger = 13.3 kcal mole(-1) and DeltaSdouble dagger = 12 cal deg(-1) mole(-1) (3.5S fragment). The kinetic observations are consistent with the view that the binding of a single antibody molecule can bring about phage neutralization. There are two ways in which a single antibody molecule can affect neutralization: (1) binding at or near some critical site on the phage may block its function, (2) binding may disturb the general architectural design of the protein shell of the phage. Although the rate of neutralization varied directly with antibody size, consideration of the activation parameters speaks against the dependence of neutralization on simple steric factors. Addition of antibodies directed against rabbit gammaG-immunoglobulin or the 5S pepsin fragment caused approximately a threefold neutralization enhancement. This enhancement may result from the detection of a class of infectious bacteriophage antibody complexes.

Animals↗