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Biomedical subjects

R Zak

Publications and source records attributed to R Zak.

At least 109 records · Page 6Linked to original sources
PubMed1981

Assessment of fractional rates of protein synthesis in cardiac muscle cultures after equilibrium labeling.

Protein synthesis, accumulation, and breakdown were examined in growing cultures of contractile embryonic chick heart cells, and the fractional synthetic rates of several individual proteins were compared. Fractional rates of protein synthesis were evaluated with a procedure that combined equilibrium and pulse labeling for determination of specific radioactivities of the precursor in the medium and in proteins isolated by electrophoresis. Kinetic analysis during equilibration of the precursor specific radioactivity with that in labeled proteins indicated that the specific radioactivity of leucine in the culture medium closely approximated that of the immediate reaction precursor for protein synthesis. Protein accumulation and breakdown were evaluated by standard growth and decay kinetics. Fractional protein synthesis rates determined during either short pulse labeling or during continuous labeling to equilibrium were found to be in agreement with independent measurements of protein accumulation and breakdown. We also determined fractional synthetic rates of proteins isolated from cultured heart cells on single and two-dimensional electrophoresis and found the following order of fractional synthesis rates: fibronectin greater than or equal to alpha-actinin greater than myosin heavy chain = tropomyosin = myosin light chains greater than 55,000-dalton proteins (desmin and tubulin) greater than or equal to actin. The half-lives of these proteins ranged from T 1/2 = 1.1 days for fibronectin and 2.0 days for myosin heavy chain to 4.7 days for actin.

Actins↗
PubMed1981

Equilibration of leucine between the plasma compartment and leucyl-tRNA in the heart, and turnover of cardiac myosin heavy chain.

By 30min continuous infusion of [3H]leucine into rats, the specific radioactivities of plasma leucine and tissue-free and tRNA-bound leucine in heart were equal. The specific radioactivity of leucyl-tRNA in heart therefore follows a time course identical with that of plasma leucine soon after the start of infusion. The half-life of cardiac myosin heavy chain (5.5 days) was the same as that reported by other investigators who used the pulse-labelling protocol.

Animals↗
PubMed1981

Molecular cloning of two fast myosin heavy chain cDNAs from chicken embryo skeletal muscle.

Recombinant DNA clones containing sequences for two different types of myosin heavy chain (HC) genes from chicken embryonic skeletal muscle were constructed and analyzed. Specificity of the clones for myosin HC was demonstrated by hybrid-arrested translation, by hybridization to a 7.0-kb mRNA, and by comparison of DNA sequences with known amino acid sequences of rabbit skeletal muscle myosin HC. Restriction enzyme and electron-microscopic heteroduplex analysis showed the presence of two distinct but homologous cDNA sequences. Hybrid melting curves indicated that both types of sequences represent fast myosin HC sequences.

Amino Acid Sequence↗
PubMed1980

Purification of messenger ribonucleic acids for fast and slow myosin heavy chains by indirect immunoprecipitation of polysomes from embryonic chick skeletal muscle.

Fast and slow myosin heavy chain mRNAs were isolated by indirect immunoprecipitation of polysomes from 14-day-old embryonic chick leg muscle. The antibodies were prepared against myosin heavy chains purified by NaDod-SO4-polyacrylamide gel electrophoresis and were shown to be specific for fast and slow myosin heavy chains. The RNA fractions directed the synthesis of myosin heavy chains in a cell-free translation system from wheat germ. Several smaller peptides were also synthesized in lower concentrations. These probably are partial products of myosin heavy chains, since they are immunoprecipitated with antibodies to myosin heavy chains. Immunoprecipitation of the translation products with the antibodies to fast and slow myosin heavy chains showed the RNA preparations to be approximately 94% enriched for fast myosin heavy chain mRNA and approximately 84% enriched for slow myosin heavy chain mRNA with respect to myosin HC type. Peptides having slightly different mobilities on NaDodSO4-polyacrylamide gels were immunoprecipitated by antibodies to fast and slow myosin heavy chains.

Animals↗
PubMed1980

Zinc and osteoporosis in patients with spinal cord injury.

Thirty-eight patients (8 women and 30 men) with spinal cord injury were investigated. All had been immobilised after the traumatic event. The time elapsed since their accidents varied from 2 to 74 weeks. Blood and urine samples were collected to investigated calcium, zinc, magnesium, sodium, alkaline phosphate, phosphore, haemoglobin, creatinine, uric acid and proteins in blood, and the urinary excretion of phosphore, hydroxyproline, creatinine, amino acids, calcium, calcium, magnesium and zinc. The methods were estimately by atomic absorption spectrophotometry. The serum zinc levels did not differ statistically from normal and the calcium and magnesium levels in the serum were lower among the patients than in normal controls. The urinary excretion of zinc, calcium, phosphore and hydroxyproline was higher among the patients without correlation to the patients' age. The zinc excretion is negatively correlated to the time elapsed since the injury, but it is still high 3 months after trauma. The highly significant correlation between urinary zinc and hydroxyproline excretion, together with increased calcium and phosphore excretion, suggests that zinc may be involved in the process of osteoporosis in patients with spinal cord injury.

Adolescent↗
PubMed1979

Cardiac hypertrophy: its characteristics as a growth process.

The causal relation between cardiac function and growth is analyzed in this review article. Three different levels of development are discussed: cytodifferentiation, embryogenesis and postnatal development. The earliest stage of cardiac morphogenesis, that is, the appearance of cell-specific proteins and of spontaneous contractions, appears to be independent of hemodynamic forces. Also, the first major morphologic transformation of the primitive heart, looping, is the intrinsic property of the heart itself. However, at any later stage of life, hemodynamic function in both health and disease is closely coupled to cardiac growth.

Animals↗
PubMed1979

Molecular aspects of cardiac hypertrophy.

Despite continuous interest in cardiac hypertrophy, our knowledge of its molecular aspects is still elementary. Recently, however, several advancements of particular interest have been made: (a) Nuclei of muscle and nonmuscle cells have been separated, allowing for the first time the study of nuclear activity in specified cells (18). (b) Cardiac growth induced by pressure-overload (72) or by hormone treatment (26) has been shown to lead to myosin of altered ATPase, and strong evidence suggests that new species of myosin molecules thus appear. (c) The basis for assessment of protein synthesis and degradation has been established (46, 48). (d) Methods are being developed to supplement radioautography in evaluating cell proliferation (42, 59, 69). (e) In spontaneously hypertensive rats it has been shown that blood pressure might not be the sole factor responsible for cardiac enlargement, but that hypertrophy can be the result of genetic cardiovascular abnormality (19, 66). (f) A hypothesis relating the extent of energy utilization to the nuclear activity via NAD+ metabolism has been proposed, which allows for experimental verification (43).

Adenosine Triphosphatases↗
PubMed1979

Changes in mitochondrial DNA in cardiac hypertrophy in the rat.

We studied DNA (mtDNA) replication in adult female rat hearts undergoing hypertrophy secondary to constriction of the ascending aorta. MtDNA was measured in isolated mitochondria by a fluorometric method adapted for that purpose. The conditions for removal of contaminating nuclear DNA were developed, and the purity of the mtDNA was assessed from its molecular conformation (open and closed circles) and by renaturation-kinetic analysis. The mtDNA concentration in mitochondria, expressed as micrograms of DNA per milligram of mitochondrial protein, increased 2, 4, and 7 days postoperatively by 21, 73, and 98%, respectively. Similar results were obtained when mtDNA was expressed per nonomole of cytochrome a. The population of replicative intermediates of mtDNA was analyzed by electron microscopy. In normal hearts, we observed molecular forms characteristic of animal mtDNA, such as circular monomers and dimers, catenated molecules, D-loops, expanded D-loops, and gapped molecules. D-loop frequency, which was near 50% in the mtDNA of control hearts, was markedly reduced to 5-7% in hypertrophying hearts. This result indicates that the increase in replicative flux of mtDNA is associated with the removal of a block in the conversion of D-loops to other intermediates.

Animals↗
PubMed1979

Dissociation of adult mammalian heart into single cell suspension: an ultrastructural study.

Adult rat heart was dissociated into a single cell suspension by a perfusion technique which used 0.05% collagenase and 0.1% hyaluronidase in Krebs-Ringer phosphate buffer (KRP). The non-muscle cells of the suspension were separated from the myocytes by centrifugation through 3% Ficoll solution in KRP with 0.01 mM Ca2+. An approximately 90% pure suspension of isolated single muscle cells was obtained with this method. The effects of the successive steps in the dissociation procedure on the ultrastructure of the heart were studied by scanning and transmission electron microscopy. After 30 minutes of enzyme digestion, dissociation of the inner endothelial lining of the ventricle into single cells or small groups of cells became apparent. In addition, the underlying cardiac skeleton began to disintegrate and linear arrays of cardiac muscle cells were observed. After 45 minutes of enzyme digestion the number of released single cells was higher because of the separation of intercalated discs. The majority of non-muscle cells were by now dissociated from the surfaces of muscle cells. Widening of the lateral intercellular spaces between the myocardial cells was associated with separation of desmosomes. In some regions of the heart, intact desmosomes, fasciae adherentes and gap junctions were observed even though lateral intercellular spaces had widened greatly. The majority of myocardial cells had become separated from one another after 60 minutes of enzyme digestion. Separation of gap junctional sites took place in two ways: (1) by 'unzipping' them through enzyme action; (2) by tearing them mechanically. Gap junction remnants were sometimes observed in a vesiculated state within the cell. The dissociation of the heart was ineffective when perfused with media containing 1.0 or 2 mM Ca2+. Alcian blue treatment after 60 minutes of enzyme digestion revealed that the basement membrane, and its accompanying collagen fibrils, was still present on the plasma membrane of dissociated single cells. The isolated myocardial cells retained their normal morphological characteristics. This study has enabled us to understand in detail how dismantlement of highly ordered adult cardiac tissue into a single cell suspension takes place. Cell suspensions of this type should be invaluable in the study of metabolic and synthetic activities in adult myocardial cells.

Animals↗
PubMed1979

Cellular growth of cardiac muscle after birth.

Present concepts concerning DNA synthesis in the heart have been reviewed, and a speculative theory presented involving control of DNA synthesis in the developing heart. Research describing DNA synthesis in myocytes of the weanling rat heart undergoing hypertrophy has been presented. Its possible use for testing the control theory is suggested.

Aging↗
PubMed1977

Measurements of half-life of rat cardiac myosin heavy chain with leucyl-tRNA used as precursor pool.

The kinetics of labeling of myosin heavy chain, following a single intravenous injection of L-[4,5-3H]leucine, were analyzed with the help of a computer, in conjunction with the labeling kinetics of the specific radioactivities of the precursor amino acid pool. As precursor we used leucyl-tRNA which, as we show here, differs significantly from the intracellular free leucine pool. The half-life of myosin heavy chain was determined from the initial period (0 to 60 min) of incorporation of label into protein after a single injection of tritiated leucine, and also from the period (7 to 14 days) when there is exponential decay of the labeled protein. Myosin heavy chain was separated from other myofibrillar proteins by polyacrylamide gel electrophoresis before measurement of leucine specific radioactivity. The specific radioactivity was measured in both protein and precursor pools by a sensitive isotope dilution procedure (range, 100 to 1500 pmol). The values for the half-life of myosin heavy chain determined at both intervals were similar (5.4 and 5.9 days). Substitution of the specific radioactivity of the intracellular free leucine pool decreased the half-life to 2.7 dyas. Similar values were obtained when the half-life was calculated by simple graphical integration of the experimental curves.

Amino Acids↗