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Biomedical subjects

R H Allen

Publications and source records attributed to R H Allen.

At least 127 records · Page 7Linked to original sources
PubMed1982

Immunocytochemical localization of intrinsic factor--cobalamin bound to the guinea pig ileum in vivo.

The absorption of dietary cobalamin requires that a complex of intrinsic factor-cobalamin bind to specific receptors in the distal small intestine. The cellular location of these binding sites is unknown. To localize the intrinsic factor-cobalamin binding site, homogeneously purified hog intrinsic factor (7 pmol) and cobalamin (7 pmol) were instilled into ileal loops (n = 10) in anesthetized guinea pigs. Tissue controls for binding specificity were intrinsic factor-cobalamin in colon (n = 3) and jejunal (n = 3) loops, as well as intrinsic factor alone in ileal loops (n = 3). Intrinsic factor-[57Co]cobalamin was instilled in three ileal loops maintained for 90 min, and the radioactivity in the liver and kidneys was measured. Portions of each loop were washed in Krebs-Ringer buffer + Ca++, pH 7.4, or in either 1 mM Na2 ethylenediaminetetraacetic acid or in pH 4.5 acetate buffer. All tissue was processed for ultrastructural immunocytochemistry using a monospecific antisera to hog intrinsic factor as part of a well-characterized indirect immunoperoxidase method. Essentially all immunoreactive intrinsic factor-cobalamin was associated with the microvillous pits on mature ileal absorptive cells. Intrinsic factor binding was not present in colon or jejunum, or in the ileal loops instilled with intrinsic factor alone. Ethylenediaminetetraacetic acid and acid pH decreased intrinsic factor-cobalamin binding but caused severe disruption of the microvillous architecture. Absorption of [57Co]cobalamin was demonstrated, but intrinsic factor was not identified within enterocytes. The localization of specific intrinsic factor-cobalamin binding sites to the microvillous pit suggests that the intrinsic factor-cobalamin receptor may maintain a relatively fixed position within the surface membrane, and although alternative explanations are possible, our findings support the hypothesis that intrinsic factor is not internalized into the enterocyte during cobalamin absorption.

Animals↗
PubMed1981

Human intrinsic factor secretion: immunocytochemical demonstration of membrane-associated vesicular transport in parietal cells.

The human gastric parietal cell synthesizes and secretes intrinsic factor (IF) and acid. In contrast to the cellular mechanisms of acid secretion, little is known about the mechanisms of IF secretion. To elucidate these mechanisms we obtained gastric secretions and sequential fundic biopsies from three subjects before and after pentagastrin stimulation (6 microgram/Kg s.c.). IF was localized in the biopsies using an ultrastructural immunoperoxidase technique using a well-characterized, monospecific antibody to human IF. IF output was quantified using a specific radioimmunoassay in concurrently obtained gastric secretions. Before stimulation, IF was associated with tubulovesicles scattered throughout the cytoplasm and with some in rough endoplasmic reticulum (RER). The tubulovesicles associated with IF migrated to the periphery of the secretory canaliculi within 8 min of stimulation. IF was present on secretory microvilli between 8 and 30 min when IF output in gastric juice was at its maximum. The cessation of IF secretion coincided with the depletion of IF associated with tubulovesicles. IF appeared in the perinuclear space and RER as the IF associated with tubulovesicles was secreted. These observations indicate that IF secretion depends upon membrane-associated vesicular transport and provides support for a membrane translocation-fusion hypothesis to explain the morphologic changes that occur in the parietal cell during secretion.

Adult↗
PubMed1981

Nitrous oxide has multiple deleterious effects on cobalamin metabolism and causes decreases in activities of both mammalian cobalamin-dependent enzymes in rats.

In man, use of the general anesthetic nitrous oxide, N2O, is associated with hematologic and neurologic abnormalities that mimic those seen in cobalamin (Cbl, vitamin B12) deficiency. We have measured a number of aspects of Cbl metabolism in rts exposed to various concentrations of N2O for various periods of time. As little as 2% N2O given for 15 h resulted in 30% inhibition of methionine synthetase (MS) in rat liver. With 50% N2O, inhibition of 70% occurred with 1 h and did not change during the next 48 h. Under these conditions, no inhibition of methylmalonyl-CoA mutase (MMCoAM) was observed. The recovery of MS activity was slow and was only 80% of control values 72 h after N2O was stopped. Studies employing rats previously injected with [57Co]Cbl showed that N2O displaced [57Co]Cbl from MS in a manner that temporally and quantitatively paralleled the loss of MS activity. Recovery of MS activity paralleled the reappearance of [57Co]Cbl on MS. N2O also caused the hepatic content of CH3-[57Co]Cbl to decrease by 20-60%. When [57Co]-Cbl was extracted from liver and analyzed by paper chromatography, [57Co]Cbl analogues were present (10-40% of total [57Co]Cbl) in rats exposed to N2O, but not in control animals. When rats were exposed to 50% N2O for 33 d, the total of endogenous Cbl and Cbl analogues in liver decreased to 35% of control values and endogenous Cbl decreased to 10% of control values. At this time, MS activity was 15% of control values and MMCoAM was only 26% of control values. We conclude that N2O causes multiple defects in Cbl metabolism that include the following: (a) rapid inhibition of MS activity with a slow recovery when N2O is stopped; (b) displacement of Cbl from MS; (c) decreased CH3-Cbl; (d) conversion of Cbl to Cbl analogues; (e) the gradual development of Cbl deficiency and (f) an eventual decrease in MMCoAM activity with a further decrease in MS activity.

5-Methyltetrahydrofolate-Homocysteine S-Methyltran↗
PubMed1980

Isolation and characterization of methylmalonyl-CoA mutase from human placenta.

Methylmalonyl-CoA mutase, one of two known cobalamin-dependent enzymes present in mammalian tissues, has been isolated from 2.5 kg of human placenta utilizing affinity chromatography on 5'-deoxyadenosylcobalamin-Sepharose as the major purification step. The enzyme gives a single band on polyacrylamide disc gel electrophoresis. The Mr of the enzyme is 145,000 and it has two subunits of Mr = 72,000. Amino acid analysis reveals major differences from other human cobalamin-binding proteins. Based on x-ray fluorescence, the enzyme has 2 mol of cobalamin bound/mol of enzyme. In contrast to purified cobalamin transport proteins, most of the cobalamin bound to the enzyme is not released by boiling at low pH in the presence of KCN, or dialysis against 7.5 M guanidine containing 0.2 M dithiothreitol, or both, suggesting the possibility that cobalamin may be covalently attached to the purified enzyme. Both precipitating antibodies and antibodies that inhibit enzyme activity have been raised in a chicken.

Amino Acids↗
PubMed1980

Presence of cobalamin analogues in animal tissues.

Cobalamin (Cbl, vitamin B-12) has been extracted and isolated from a number of animal tissues by using (i) reverse-affinity chromatography on R protein-Sepharose followed by adsorption to and elution from charcoal-coated agarose and (ii) paper chromatography. Radioisotope dilution assays showed that only 75-97% of the Cbl chromatographed in the position of crystalline Cbl. The remaining 3-25% was present in a number of slower and faster moving fractions. This suggested that Cbl analogues are present in animal tissues because appropriate controls ruled out the possibility that this material was artifactually derived from Cbl during the extraction and purification procedures. With a large-scale isolation from rabbit kidney, the material in five such fractions contained cobalt and had absorption spectra that were similar to but different from the spectrum of Cbl, indicating that they were Cbl analogues. Compared to Cbl, these Cbl analogues had decreased but definite affinities for Cbl-binding proteins with the following order of strength of binding: R protein > transcobalamin II > intrinsic factor. Compared to Cbl, they also had decreased but definite growth-promoting activity for two microorganisms, Euglena gracilis and Lactobacillus leichmannii, which require Cbl for growth. These Cbl analogues differed from each other and from 18 synthetic Cbl analogues, including the most common Cbl analogues synthesized by microorganisms, in at least one of the above features. These studies indicate that animal tissues contain a number of Cbl analogues whose origins, structures, and biologic activities remain to be determined.

Animals↗
PubMed1980

A multi-tiered screening system for the least restrictive setting.

The authors evaluated the placement process in a large psychiatric facility with both inpatient and outpatient services. They examined 71 inpatient and 90 outpatient new admissions using standardized tests of psychopathology and sociodemographic status. After correcting for response bias, the authors found, in contrast to other studies, that the level of morbidity in the inpatient service was significantly different from that in the outpatient unit. The measures of sociodemographic data indicate that inpatients were more dependent on social support systems and that this finding was not influenced by response bias.

Commitment of Persons with Psychiatric Disorders↗
PubMed1980

Immunocytochemical localization of human intrinsic factor: the nonstimulated stomach.

Human intrinsic factor (IF) is found on the surface of parietal cells at the light microscopic level by immunofluorescence, but there is no direct evidence that IF is synthesized in and secreted from this cell. In order to provide this evidence, IF was localized at the ultrastructural level in human stomach, using an immunochemically, monospecific antibody against purified human IF. We defined optimal immunocytochemical conditions and identified the subcellular location of IF in fundic biopsies from 4 fasting subjects. Intrinsic factor was found on the perinuclear membrane, on rough endoplasmic reticulum, Golgi apparatus, tubulovesicles, and membranes of the multivesicular body in parietal cells. Intrinsic factor was absent from nuclei, mitochondria, basolateral surface membranes, blunt microvilli, and the cytosol. Thus, we have defined the parietal cell as the site of IF synthesis. The observed distribution of intracellular IF suggested that an interrelationship existed between the stained membranous structures, and that IF secretion may depend upon membrane translocation. Further immunocytochemical and physiologic studies should increase our understanding of the cellular physiology of IF secretion as well as the parietal cell.

Adult↗
PubMed1979

Isolation and characterization of the transferrin receptor from human placenta.

The transferrin receptor has been isolated from human placenta using immunochromatography and affinity chromatography. The receptor is a glycoprotein and has a Mr = 90,000 on sodium dodecyl sulfate-gel electrophoresis in the presence of 2-mercaptoethanol. The isolated receptor is immunologically related to the transferrin receptor on the reticulocyte cell surface.

Amino Acids↗
PubMed1979

The effects of response bias on sex differences in a psychiatric population.

A sample of 58 male and 84 female new admissions to a psychiatric facility was interviewed to identify sex differences in personal and social functioning. Female subjects were found to score significantly higher on intrapsychic distress scales while male subjects scored higher on scales measuring antisocial behavior. Using analysis of covariance, keeping the L- or Lie scale of the MMPI constant, all measures which had previously shown significant sex differences now became nonsignificant. This finding suggests that reported sex differences may be accounted for by response bias.

Adult↗
PubMed1978

Cobalamin analogues are present in human plasma and can mask cobalamin deficiency because current radioisotope dilution assays are not specific for true cobalamin.

Since R protein binds cobalamin (vitamin B12) and cobalamin analogues, whereas intrinsic factor is highly specific for true cobalamin, we compared the serum cobalamin values obtained with these proteins in radioisotope dilution assays. With R protein, eight of 21 patients with cobalamin deficiency had serum cobalamin levels (mean, 204, range, 85 to 355 pg per milliliter) that overlapped with values for 74 normal subjects (mean, 576, range, 220 to 1230). With intrinsic factor, no patient values (mean, 36, range, less than 10 to 78 pg per milliliter) overlapped with the normal values (mean, 322, range, 130 to 785). Paper chromatography showed that these differences were due to the presence of cobalamin analogues. R protein constituted 51 to 85 per cent of the cobalamin-binding protein in 10 commercial serum cobalamin assay kits, which were said to contain "intrinsic factor". Human plasma contains cobalamin analogues that can mask cobalamin deficiency with current radioisotope dilution assays.

Adult↗