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Biomedical subjects

W N Fishbein

Publications and source records attributed to W N Fishbein.

At least 37 records · Page 2Linked to original sources
PubMed1985

Myoadenylate deaminase deficiency and malignant hyperthermia susceptibility: is there a relationship?

Muscle biopsies from 35 patients referred for possible malignant hyperthermia were subjected to contracture testing with halothane, caffeine, and the combined agents, histopathological and fiber-type-distribution analysis, and quantitative assay of three major muscle enzymes: adenylate deaminase, adenylate kinase, and creatine kinase. Adenylate kinase and creatine kinase were in the normal range in all biopsies and each averaged 92% of expected normal value when corrected for their fiber-type distribution. Of the 14 cases with a positive halothane test, 2 had primary myoadenylate deaminase deficiency, and 5 others had low levels of this enzyme (less than one-third normal). In contrast, only 3 of 21 cases negative to halothane testing had low adenylate deaminase levels, and none were deficient. This association was significant by several statistical tests, although it would not be highly predictive for an individual case. A positive halothane test also correlated with a high type 2 fiber contribution, but this was probably secondary, since cases with low enzyme levels had significantly higher type 2 fiber areas. Caffeine contractures did not correlate with either low enzyme levels or with fiber-type distribution. Sixty percent of the biopsies were entirely normal histologically, and showed a significant correlation with a negative combined contracture test. Data on the one family included in this study suggest separate inheritance of the trait for myoadenylate deaminase deficiency and the trait for positive contracture tests. The present findings suggest that patients with myoadenylate deaminase deficiency (and the carrier state as well) may be at increased risk of malignant hyperthermia when subjected to anesthesia.

AMP Deaminase↗
PubMed1985

Some practical considerations in quantitative absorbance microspectrophotometry. Preparation techniques in DNA cytophotometry.

An experimental review of the Feulgen and gallocyanine-chrome-alum stains for quantitative cytophotometry of DNA in tissue sections yielded information on the preparation and staining of tissue for quantitative absorbance microspectrophotometry. (1) Tissues routinely fixed in formalin are suitable for either stain. Specimens fixed with glutaraldehyde-containing fixatives are not satisfactory for Feulgen staining, nor are ethanol-fixed specimens, unless they are post-fixed in formalin. (2) The pararosaniline dyes, used in the Feulgen stain, are sufficiently pure to use if a solution of the dye in ethanol shows an absorbance peak at 543 to 546 nm. (3) The Feulgen stain provides good reproducibility when the staining solution is adjusted to pH 1.5. (4) Gallocyanine is the best stain to use on Bouin-fixed or glutaraldehyde-fixed tissues. (5) Where fixation is an option, Carnoy and methanol-formalin-glacial acetic acid are excellent fixatives that can be followed by either stain. (6) Selection of the thickness of a tissue section involves a compromise. Requirements of minimum nuclear overlap and sharp focusing favor a section thickness of 4 micron to 6 micron. On the other hand, the requirement for full nuclear thickness, as judged by absorbance equivalent to that of a touch preparation, demands sections as thick as 8 micron in the case of mouse liver. Within this range, the optimum thickness, therefore, is determined by the particular tissue, the range of its nuclear sizes and its packing density. (7) The refractive index of the mounting medium should be closely matched to that of the background structure of the tissue sections. For animal tissues, we found that media with refractive indices of 1.54 to 1.56 are suitable.

Animals↗
PubMed1984

Levels of adenylate deaminase, adenylate kinase, and creatine kinase in frozen human muscle biopsy specimens relative to type 1/type 2 fiber distribution: evidence for a carrier state of myoadenylate deaminase deficiency.

Myoadenylate deaminase deficiency is believed to reflect a genetic deficiency of skeletal muscle, but its pattern of inheritance has not been established. We examined, histochemically and by quantitative biochemical assay, muscle biopsy specimens from 3 putative carriers of this disorder. Adenylate kinase and creatine kinase were assayed in parallel with adenylate deaminase in order to establish enzyme activity ratios and the variation of each enzyme with fiber-type distribution. Control tissue consisted of 34 biopsy specimens without notable abnormalities from 30 patients, and included 4 specimen pairs with disparate fiber-type contributions. By linear regression analysis, adenylate deaminase level averaged 2.8-fold higher, and adenylate kinase 4.5-fold higher, in type 2 than in type 1 fibers, whereas creatine kinase level did not differ. The slopes of the regression lines resulting from analysis of the four specimen pairs from individual patients agreed well with the overall regression line in each plot. The 3 putative carriers had adenylate deaminase levels 2.5 to 5.7 times lower than the mean control value for their fiber-type distribution, but at least 20 times higher than their enzyme-deficient kinfolk. This finding indicates that a carrier state does exist, and that the deficiency state reflects an autosomal recessive inheritance pattern. Three additional biopsy specimens were excluded from evaluation when preliminary analysis showed elevated adenylate kinase/adenylate deaminase ratios that were outliers at the 1% level. This result suggests a carrier incidence of 10% in the muscle biopsy specimen population, which would markedly bias population estimates if undetected.

AMP Deaminase↗
PubMed1984

Myoadenylate deaminase deficiency. Functional and metabolic abnormalities associated with disruption of the purine nucleotide cycle.

To assess the role of the purine nucleotide cycle in human skeletal muscle function, we evaluated 10 patients with AMP deaminase deficiency (myoadenylate deaminase deficiency; MDD). 4 MDD and 19 non-MDD controls participated in an exercise protocol. The latter group was composed of a patient cohort (n = 8) exhibiting a constellation of symptoms similar to those of the MDD patients, i.e., postexertional aches, cramps, and pains; as well as a cohort of normal, unconditioned volunteers (n = 11). The individuals with MDD fatigued after performing only 28% as much work as their non-MDD counterparts. Muscle biopsies were obtained from the four MDD patients and the eight non-MDD patients at rest and following exercise to the point of fatigue. Creatine phosphate content fell to a comparable extent in the MDD (69%) and non-MDD (52%) patients at the onset of fatigue. Following exercise the 34% decrease in ATP content of muscle from the non-MDD subjects was significantly greater than the 6% decrease in ATP noted in muscle from the MDD patients (P = 0.048). Only one of four MDD patients had a measurable drop in ATP compared with seven of eight non-MDD patients. At end-exercise the muscle content of inosine 5'-monophosphate (IMP), a product of AMP deaminase, was 13-fold greater in the non-MDD patients than that observed in the MDD group (P = 0.008). Adenosine content of muscle from the MDD patients increased 16-fold following exercise, while there was only a twofold increase in adenosine content of muscle from the non-MDD patients (P = 0.028). Those non-MDD patients in whom the decrease in ATP content following exercise was measurable exhibited a stoichiometric increase in IMP, and total purine content of the muscle did not change significantly. The one MDD patient in whom the decrease in ATP was measurable, did not exhibit a stoichiometric increase in IMP. Although the adenosine content increased 13-fold in this patient, only 48% of the ATP catabolized could be accounted for by the combined increases of adenosine, inosine, hypoxanthine, and IMP. Studies performed in vitro with muscle samples from seven MDD and seven non-MDD subjects demonstrated that ATP catabolism was associated with a fivefold greater increase in IMP in non-MDD muscle. There were significant increases in AMP and ADP content of the muscle from MDD patients following ATP catabolism in vitro, while there was no detectable increase in AMP or ADP in non-MDD muscle. Adenosine content of MDD muscle increased following ATP catabolism, but there was no detectable increase in adenosine content of non-MDD muscle following ATP catabolism in vitro. These studies demonstrate that AMP deaminase deficiency leads to reduced entry of adenine nucleotides into the purine nucleotide cycle during exercise. We postulate that the resultant disruption of the purine nucleotide cycle accounts for the muscle dysfunction observed in these patients.

AMP Deaminase↗
PubMed1984

Interstitial 5'-nucleotidase stain for frozen biopsy specimens of skeletal muscle. A useful adjunct in the diagnosis of polymyositis.

We applied a simple lead salt-based stain for interstitial and vascular 5'-nucleotidase to 150 muscle biopsy specimens. No reaction was obtained with 2'- or 3'-adenosine monophosphate, indicating that the stain was specific, and distinct from phosphatases. Staining was not inhibited by alpha, beta-methylene adenosine 5'-diphosphate, but was prevented by formaldehyde fixation or by brief immersion in octoxynol 9 (Triton X-100). Nucleotidase stains the following specific histologic sites that distinguish it from alkaline phosphatase: the intima and adventitia of medium-sized and large arteries, perineural and muscle spindle sheaths, and tendon insertions. Aside from these structures, normal muscle shows little reaction, as the sarcoplasm and sarcolemma do not stain. Neither of these enzymes shows a compensatory increase, histochemically, in myo-adenylate deaminase deficiency. In Duchenne's muscular dystrophy, however, and particularly in inflammatory myopathy, interstitial staining of 5'-nucleotidase is increased, leading to investment of most muscle fibers in the affected area. The stain rarely identifies regenerating fibers. Although alkaline phosphatase commonly shows a corresponding increase in interstitial staining, we encountered six cases of inflammatory myopathy in which this was absent, despite pronounced endomysial staining in the 5'-nucleotidase reaction. 5'-Nucleotidase thus appears to provide a valuable adjunct in the diagnosis of inflammatory myopathy.

5'-Nucleotidase↗
PubMed1983

Kinetic and immunologic evidence for a complete gene block in myoadenylate deaminase deficiency.

Muscle biopsies from patients with myoadenylate deaminase deficiency (mADD) have been evaluated kinetically and immunologically to ascertain the origin of residual enzyme activity. Kinetic evaluation employed 5 mM AMPS/1 mM AMP ratios, which were 0.7-0.8 for the human muscle isozyme, but 0-0.25 for the isozyme(s) of all other human blood cells and tissues examined. Of 14 control biopsies, 13 showed a ratio greater than 0.60 (one gave 0.47) regardless of the enzyme specific activity, while all 14 mADD biopsies showed a ratio less than 0.24, suggesting that a fetal muscle isozyme and/or blood cell isozyme were responsible for the residual activity. Confirmation was provided by rabbit antisera to purified human muscle AMP deaminase. These antisera fully precipitate the isozyme from crude human muscle biopsy homogenates, regardless of fiber-type composition, and cross-react effectively with the muscle isozyme of Rhesus monkeys and thoroughbred horses, but are inactive toward the isozymes of all other human blood cells and tissues examined. Of 18 mADD homogenates tested, 14 showed less than 20% reactivity with the antisera, at levels that precipitated 10 x more enzyme in control specimens. The residual activity in most cases of mADD must therefore arise from some source other than normal AMP deaminase. To evaluate the possibility of a single common determinant, 9 mADD homogenates were tested for soluble immune complexes. Seven of the 9 then showed 20-42% reactivity, suggesting that part of their residual activity may be due to an isozyme sharing one antigenic determinant with the normal muscle isozyme. Competitive antigen binding was used to assess whether catalytically inactive AMP deaminase was present in mADD. The method was demonstrated effective in identifying spontaneously inactivated purified enzyme and alkaline-inactivated crude enzyme. Nevertheless, homogenates from 17 mADD cases failed to produce more than 14% activation, under conditions in which 63-99% activation was expected. Triton X-100 extracts of homogenate residues of 11 mADD cases were also tested, in a search for insoluble antigen; none produced significant competition. The evidence thus indicates that most cases of mADD are due to a complete gene block, with total absence of all normal muscle AMP deaminase protein.

AMP Deaminase↗
PubMed1980

Stain for skeletal muscle adenylate deaminase. An effective tetrazolium stain for frozen biopsy specimens.

Procedures for staining frozen sections of human muscle biopsy specimens for adenylate deaminase are outlined. The preeminent role of this procedure is as a survey stain for myoadenylate deaminase deficiency, a new and common muscle enzyme deficiency, an example of which is illustrated. Discrimination of type I and II muscle fibers with this stain is based on color differences rather than density differences. Color photomicrographs illustrate the variations in staining that may be encountered, and experimental controls to verify the validity of the stain are summarized.

AMP Deaminase↗
PubMed1978

Myoadenylate deaminase deficiency: a new disease of muscle.

Five cases of a new disease presented with muscular weakness or cramping after exercise; three of the cases also had an elevated serum creatine phosphokinase. Muscle biopsies were histologically normal but lacked adenylate deaminase by stain and solution assay, while the erythrocyte isozyme was normal. A clinical diagnostic test has been developed, and the human enzyme was separated by acrylamide-gel electrophoresis.

AMP Deaminase↗
PubMed1977

Electron microscopy of negatively stained jackbean urease at three levels of quaternary structure, and comparison with hydrodynamic studies.

Electron microscopy, with sodium phosphotungstate as negative stain, has been carried out on purified jackbean urease prepared at three levels of quaternary structure: (a) A1 urease, Mr = 240 000, S20,W = 11.5 S (b) alpha urease, Mr = 480 000, S20,w = 18.3 S (c) polymers of alpha urease above the tetramer stage. The compatibility of the images from level to level leaves no doubt that the enzyme itself is being visualized, and the following geometry is suggested by electron microscopy: A1 molecules are cyclic trimers, which pair up in eclipsed position across a 1-nm cleft to form the hexameric alpha, which displays D3 (or 32) symmetry of a trigonal prism. Polymers consist of alpha molecules aligned with their clefts coplanar and an angle of 120 degrees between each triplet of 3-fold axes. These features correspond reasonably well with sedimentation and electrophoretic studies of the solvated enzyme, which have indicated a hemispherical A1, a spherical alpha, and string-of-beads polymers. Sedimentation constants of the urease polymers up through the pentamer level were found to be compatible with the rosette, straight-chain, and zig-zag forms seen in the electron microscope, and with the suggested protomer arrangement in A1 and alpha urease.

Macromolecular Substances↗