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Biomedical subjects

E F Johnson

Publications and source records attributed to E F Johnson.

At least 109 records · Page 6Linked to original sources
PubMed1984

Elastic fibers in the anulus fibrosus of the dog intervertebral disc.

A light microscopic investigation of the anulus fibrosus in cervical intervertebral discs of the dog was conducted to ascertain the arrangement and distribution of elastic fibers. Elastic fibers were observed in all lamellae of the anulus fibrosus. However, collagenous fibers were the predominant type of connective tissue fiber, and elastic fibers were randomly dispersed among them. Intralamellar (collagenous and elastic) fibers were vertically and obliquely oriented in both superficial and deep lamellae of the anulus fibrosus. All intralamellar fibers were densely and regularly arranged in superficial lamellae, but they were more loosely organized in deep lamellae. A narrow border of interlamellar, elastic fibers was observed between broader, contiguous lamellae in the superficial zone of the anulus fibrosus. Interlamellar elastic fibers wer vertically and obliquely arranged in superficial lamellae; however, they were radially oriented in deep lamellae. The deepest lamella of the anulus fibrosus consisted of a loose, three-dimensional network of intermeshing collagenous and elastic fibers. These observations suggest that elastic fibers are integral components of the articular and shock absorption mechanisms of the anulus fibrosus, and the cervical intervertebral disc of the dog is a suitable model for experimental investigation of the role of elastic fibers in intervertebral disc herniation.

Animals↗
PubMed1983

The effect in vivo and in vitro of allylisopropylacetamide on the content of hepatic microsomal cytochrome P-450 2 of phenobarbital treated rabbits.

Rabbits treated with phenobarbital were given a single injection of allylisopropylacetamide (AIA) s.c. and/or heme i.v. Hepatic microsomes were isolated 1, 5 and 24 hours post injection and the microsomal contents of both total cytochrome P-450 chromophore, and the protein moiety of P-450 2 were determined by spectrophotometric and immunochemical methods respectively. AIA caused the levels of total P-450 chromophore and of P-450 2 protein to decline to 30% of the control values at 5 hours post-injection. Concurrent administration of heme with AIA diminished the decrease in the total microsomal content of P-450 chromophore but not in that of P-450 2 protein. These findings suggest that the destruction of the heme prosthetic group of P-450 by suicide substrates such as AIA may lead to an enhanced degradation of the apo-P-450.

Acetamides↗
PubMed1983

Allosteric regulation of the 16 alpha-hydroxylation of progesterone as catalyzed by rabbit microsomal cytochrome P-450 3b.

A variety of compounds that can arise from the metabolism of progesterone in vivo stimulate the 16 alpha-hydroxylation of progesterone as catalyzed by highly purified, reconstituted preparations of cytochrome P-450 3b obtained from rabbit strain IIIVO/J. In general, reduction of the 20-keto moiety of progesterone or related compounds increases the extent of stimulation. Reduction of the 3-keto group also results in increased stimulatory activity in most cases. The resulting 3 beta-hydroxy derivatives are more active than the corresponding 3 alpha-isomers. In a similar fashion, 5 beta-pregnanes exhibit greater activity than the corresponding 5 alpha-pregnanes. The effect of these allosteric effectors is saturable at relatively low concentrations when compared to other positive effectors of P-450-mediated metabolism. These compounds increase the apparent ratio of Vmax/Km without altering the amount of reductase required for half-maximal activity when reconstituted with the cytochrome. In contrast, many of these compounds do not affect or inhibit the 6 beta-hydroxylation of progesterone catalyzed by a subform of P-450 3b that is expressed in New Zealand White rabbits but not in strain IIIVO/J. Many of the compounds investigated here are metabolites of progesterone and, therefore, may modulate P-450 3b-mediated metabolism during pregnancy.

Allosteric Regulation↗
PubMed1982

Functional and structural polymorphism of rabbit microsomal cytochrome P-450 form 3b.

The present report describes several characteristics of rabbit microsomal cytochrome P-450 form 3b isolated from liver microsomes prepared from an incipient inbred strain, IIIVO/J. Whereas most preparations of P-450 3b isolated from outbred New Zealand White (NZW) rabbits catalyze both 6 beta- and 16 alpha-hydroxylation of progesterone, form 3b prepared from strain IIIVO/J exhibits a greatly diminished 6 beta-hydroxylase activity. This difference is attributable to a large difference in Km for progesterone, and similar differences are seen when the metabolism of progesterone by microsomes prepared from each rabbit source is examined. The dependence of the rate of 16 alpha-hydroxylation of progesterone and similar differences are seen when the metabolism of progesterone by microsomes prepared from each rabbit source is examined. The dependence of the rate of 16 alpha-hydroxylation of progesterone on the initial substrate concentration also differs in that most preparations from NZW rabbits contain two enzymatically distinct forms and that one of these forms is apparently absent from the preparations of P-450 3b obtained from strain IIIVO/J. The two enzymatic subforms are differentially inhibited by 16 alpha-methylprogesterone. In contrast, the rates of 4-hydroxylation of biphenyl and of the 7-hydroxylation of 2-acetylaminofluorene catalyzed by form 3b prepared from strain IIIVO/J are similar to those catalyzed by the cytochrome obtained from NZW rabbits. When the two types of P-450 3b were compared by peptide mapping, they displayed similar yet distinguishable peptide closely related yet distinct. The selective loss of 6 beta-hydroxylase activity in the preparations of P-450 3b is suggestive of the genetically determined absence of a subform of the cytochrome that catalyzes this reaction and that is normally expressed in New Zealand White rabbits.

Animals↗
PubMed1982

Rabbit hepatic progesterone 21-hydroxylase exhibits a bimodal distribution of activity.

Progesterone 21-hydroxylase activity varies extensively among liver microsomes prepared from individual New Zealand White (NZW) rabbits. The 21-hydroxylase activities are distributed between two groupings that differ by more than tenfold in mean activity. Both male and female animals are represented in the two groupings. However, females exhibited the higher activity more frequently than males. The 21-hydroxylation of progesterone is catalyzed by one of the liver microsomal cytochrome P-450 isozymes, form 1, and these differences in activity are suggestive of differences in the occurrence of this isozyme among NZW rabbits.

Animals↗
PubMed1982

The role of cytosolic proteins in the intracellular transport of haem in rat liver. A dual-label approach.

After consecutive injections of delta-amino[3H]- and -[14C]-laevulinic acid, the incorporation of the two labels into haem associated with different subfractions of the liver was determined. Marked differences in the 14C/3H ratios were observed between haem associated loosely and tightly with microsomes and mitochondria and haem associated with three subfractions of the cytosol obtained by gel filtration. The effect of changing the amounts of delta-aminolaevulinic acid injected and of changing the interval between injections and killing of the animal on the ratios of labels in the haem of each subfraction was studied. The results are discussed in terms of the flow of haem from the mitochondria to other parts of the cell via putative cytosolic carrier proteins.

Aminolevulinic Acid↗
PubMed1982

Effect of 2,3,7,8-tetrachlorodibenzo-p-dioxin and phenobarbital on the occurrence and distribution of four cytochrome P-450 isozymes in rabbit kidney, lung, and liver.

The effect of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) and of phenobarbital (PB) on the distribution and occurrence of four cytochrome P-450 isozymes, Forms 2, 3, 4, and 6, in the kidney, lung, and liver of adult male rabbits was investigated using immunofluorescence. In the kidney, Forms 2 and 3 were localized in the proximal tubules of both untreated and PB-treated animals, while antibodies to Forms 4 and 6 showed weak to negative staining. In TCDD-pretreated animals, Forms 4 and 6 appeared in the renal endothelium, in addition to staining the proximal tubular epithelium intensely. Form 2 was the only isoenzyme of those studied found to be present in the lungs of normal and PB-pretreated rabbits; it was also present in lungs of TCDD-pretreated rabbits. Form 3 was not detected in any of the rabbit lungs examined. Forms 4 and 6, while not apparent in the lungs of normal or PB-treated animals, were found in the lungs of TCDD-treated animals and also appeared in the endothelium of the pulmonary arteries and veins. All forms tested were present in control liver. The staining for Form 2 was intense in the livers of PB-pretreated animals, as was the staining for Forms 4 and 6 in the livers of TCDD-pretreated animals. Our results indicate that, while PB altered the intensity of staining for Form 2 in the liver and kidney, TCDD altered both the staining intensity and distribution of the isozymes in kidney, lung, and liver, producing, for example, a localization of Forms 4 and 6 in the endothelium of both the kidney and lung which was not seen in either untreated or PB-pretreated rabbits.

Animals↗
PubMed1982

Effect of microsomal cytochrome P-450 isozyme induction on the mutagenic activation of 2-aminoanthracene.

Two rabbit microsomal cytochrome P-450 isozymes. Forms 4 and 6, which are differentially induced by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) in the liver in an age-dependent fashion, catalyze the activation of 2-aminoanthracene to a mutagen in the Ames Salmonella mutagenesis assay. We have shown previously that in the presence of saturating concentrations of 2-aminoanthracene, Form 4 is 15-fold more active than Form 6 in the activation of this mutagen. Similar differences in the activation of 2-aminoanthracene are observed between liver microsomes isolated from TCDD-treated adult rabbits, in which Form 4 predominates, and microsomes from rabbit neonates exposed transplacentally to TCDD prior to birth, in which Form 6 predominates. However, when the extent of mutagenesis is limited by the amount of 2-aminoanthracene and is independent of the rate of activation, the number of revertants produced is similar for microsomes isolated from either newborn or adult TCDD-treated rabbits. Under these conditions, the extent of mutagenesis obtained for a given amount of 2-aminoanthracene will depend on the balance between activation and competing reaction pathways leading to detoxication. Thus, differences in the rate of activation between adult and newborn microsomes are probably offset by similar differences in the rates of competing pathways of metabolism. This is consistent with the finding that the overall rate of 2-aminoanthracene metabolism by the adult microsomes is greater than that of the neonate. In order for the extent of mutagenesis to be independent of rate, the half-life of 2-aminoanthracene was seen to be less than approximately 12 min.

Animals↗
PubMed1982

The distribution and arrangement of elastic fibres in the intervertebral disc of the adult human.

Selected cervical and thoracic intervertebral discs were removed from twelve human adults at autopsy. The discs were prepared by the paraffin method, stained with orcein or by Verhoff's technique and examined with the light microscope for the presence of elastic fibres. Elastic fibres were located only in the lamellae of the anulus fibrosus and the superficial zones of the nucleus pulposus from regions of intervertebral disc that connected or interfaced with osseous vertebrae. Elastic fibres were longitudinally, circularly and obliquely arranged in lamellae of the anulus fibrosus and penetrated bony vertebrae as Sharpey's fibres. Elastic fibres assumed a three dimensional lattice work in the superficial zone of the nucleus pulposus and connected it to the hyaline cartilage end plate as Sharpey's fibres. Irregularly arranged elastic fibres interconnected the superficial zone of the nucleus pulposus and lamellae of the anulus fibrosus. Elastic fibres were a major histological component of the ligamenta flava and anterior longitudinal ligaments.

Aged↗
PubMed1981

Amino acid sequence of an analogous peptide from two forms of cytochrome P-450.

Two cytochrome P-450 preparations, a constitutive isozyme, form 3b, and a phenobarbital-induced isozyme, form 2, were isolated from rabbit liver microsomes and compared by peptide mapping following digestion with trypsin and by partial sequence analysis. The NH2-terminal sequence of form 3b differed from form 2 in 15 out of 18 amino acids, but both forms have an NH2-terminal methionine residue followed by an acidic residue. Comparisons of many of the tryptic peptides of the two forms by means of high pressure liquid chromatography, as well as amino acid composition and sequence analysis, indicated that peptides from these forms, with one exception, are different. A tridecapeptide, differing only in a methionine (form 3b)/isoleucine (form 2) replacement was isolated from both forms. The amino acid sequence of this peptide is as follows: Met-Pro-Tyr-Thr-Asp-Ala-Val-Ile/Met-His-Glu-Ile-Gln-Arg. Taken together, these data indicate that forms 2 and 3 represent dissimilar gene products. The observation that these two cytochromes share an analogous peptide suggests that this tridecapeptide may contribute structural information necessary for common functional properties.

Amino Acid Sequence↗
PubMed1981

Specificity in the activation and inhibition by flavonoids of benzo[a]pyrene hydroxylation by cytochrome P-450 isozymes from rabbit liver microsomes.

The effect of flavone and 7,8-benzoflavone on the metabolism of benzo[a]pyrene to fluorescent phenols by five cytochrome P-450 isozymes obtained from rabbit liver microsomes was determined. Benzo[a]pyrene metabolism was stimulated more than 5-fold by the addition of 600 microM flavone to a reconstituted monooxygenase system consisting of NADPH, cytochrome P-450 reductase, dilauroylphosphatidylcholine, and cytochrome P-450LM3c or cytochrome P-450LM4. In contrast, an inhibitory effect of flavone on benzo[a]pyrene metabolism was observed when cytochrome P-450LM2, cytochrome P-450LM3b, or cytochrome P-450LM6 was used in the reconstituted system. 7,8-Benzoflavone (50-100 microM) stimulated benzo[a]pyrene metabolism by the reconstituted monooxygenase system about 10-fold when cytochrome P-450LM3c was used, but benzo[a]pyrene hydroxylation was strongly inhibited when 7,8-benzoflavone was added to the cytochrome P-450LM6-dependent system. Smaller effects of 7,8-benzoflavone were observed on the metabolism of benzo[a]pyrene by the cytochrome P-450LM2-, cytochrome P-450LM3b-, and cytochrome P-450LM4-dependent monooxygenase systems. These results demonstrate that the activating and inhibiting effects of flavone and 7,8-benzoflavone on benzo[a]pyrene metabolism depend on the type of cytochrome P-450 used in the reconstituted monooxygenase system.

Animals↗
PubMed1980

Isolation and characterization of a constitutive form of rabbit liver microsomal cytochrome P-450.

A heretofore unrecognized form of cytochrome P-450 was purified from rabbit liver microsomes with an average yield and purity similar to that of other highly purified forms of cytochrome P-450. Several properties of this cytochrome are contrasted with those of form 2, the major phenobarbital-inducible cytochrome P-450, form 4, the major 2,3,7,8-tetrachlorodibenzo-p-dioxin-inducible cytochrome, and form 6, a cytochrome that is selectively induced in liver microsomes by 2,3,7,8-tetrachlorodibenzo-p-dioxin during the perinatal period. Thes four forms can be distinguished by virtue of their molecular weights as determined using polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate, by their respective peptide fingerprints, and by the monospecificity of their antisera. Since the enumerated properties are thought to reflect the primary structure of the cytochromes and since the observed differences are extensive, we suggest that these four forms are not derived from a common protein precursor.

Animals↗
PubMed1980

Catalysis of divergent pathways of 2-acetylaminofluorene metabolism by multiple forms of cytochrome P-450.

Four highly purified forms of rabbit hepatic, microsomal cytochrome P-450 catalyze the N- and ring-hydroxylation of 2-acetylaminofluorene (AAF) at different rates. Form 4, the major form of the cytochrome induced by 2,3,7,8-tetrachlorodibenzo-p-dioxin in adult rabbit liver, catalyzed the N-hydroxylation of AAF more rapidly than did the other three forms. N-Hydroxy-2-acetylaminofluorene accounted for 70% of the metabolites formed by the action of this cytochrome. Form 6, the major form of the cytochrome induced by 2,3,7,8-tetrachlorodibenzo-p-dioxin in neonate rabbit liver, and Form 3, a constitutive form of the cytochrome, both metabolized AAF at one-half the rate observed for Form 5. Phenols accounted for more than 90% of the metabolites produced by these two cytochromes. The major phenobarbital-inducible cytochrome P-450, Form 2, exhibited practically no catalytic activity (< 1% of the other forms) with AAF as a substrate. Since N- and ring-hydroxylation are thought to represent divergent pathways of carcinogen metabolism (activation versus detoxification), the differential occurrence of the various cytochrome forms should affect the balance between these two reaction pathways. In this sense, cytochrome P-450 induction by 2,3,7,8-tetrachlorodibenzo-p-dioxin differentially affects the magnitude and direction of in vitro microsomal metabolism of AAF as a function of age.

2-Acetylaminofluorene↗