bluuebunny/crossref_metadata_2025 huggingface dataset:
REDUCTION IN SURFACE CHARGE AS AN EXPLANATION OF THE RECOGNITION BY MACROPHAGES OF NUCLEI EXPELLED FROM NORMOBLASTS
Authors: Skutelsky E. , Danon D.
Published: May 2004

The density and distribution of electric charge on the surface of rabbit bone marrow cells was visualized by electron microscopy after the cell surfaces had been stained with charged colloidal iron particles. Expulsed erythroid nuclei are less negatively charged than any other cell in the bone marrow. They carry from about one-half to one-third of the charge density on the remaining future reticulocyte. The reduction in the surface charge density is already apparent when the nucleus is partially expulsed. Practically no positive charge was found on its surface or on the surface of any other bone marrow cell. The possibility that the reduced negative charge on the surface of expelled erythroid nuclei is one of the means by which the macrophage distinguishes it from other bone marrow cells is discussed.
OSMIUM ZINC IODIDE REACTIVE SITES IN THE EPIDERMAL LANGERHANS CELL
Authors: Niebauer Gustav , Krawczyk Walter S. , Kidd Richard L. , Wilgram George F.
Published: May 2004

Fixation of epidermis with a mixture of osmium tetroxide and zinc iodide (OsO4-ZnI2) for 24 hr renders the central periodic lamella of the Langerhans cell granule (LCG), the Golgi region, and the nuclear envelope of epidermal Langerhans cells preferentially visible. The use of this technique on Langerhans cells in normal epidermis and in epidermis of patients with histiocytosis (Letterer-Siwe disease) allows a broader visualization of the LCG's than was heretofore possible with routine glutaraldehyde-osmium tetroxide fixation and uranyl acetate-lead staining. The identical staining of Golgi apparatus and LCG favors the view that there is close relation between the Golgi area and the LCG's. Different staining characteristics of the LCG's near the Golgi region and at the cell periphery, respectively, may suggest that the LCG undergoes changes on its way from the Golgi area towards the extracellular space. The hypothesis is advanced that the material which is heavily impregnated with metal after fixation with OsO4-ZnI2 might be a lipid.
PHAGOCYTOSIS OF LATEX BEADS BY ACANTHAMOEBA CASTELLANII (NEFF)
Authors: Wetzel Mary G. , Korn Edward D.
Published: May 2004

A method is described for the rapid and efficient isolation of phagocytic vesicles from large scale cultures of Acanthamoeba castellanii (Neff) that have been incubated with polystyrene latex beads. Cells were allowed to phagocytose latex beads for 30 min and then were homogenized, and the phagocytic vesicles were isolated by one centrifugation through several layers of sucrose. Identity and purity of the phagocytic vesicles were determined by electron microscopy, chemical analyses, and assays of acid phosphatase, α- and ß-glucosidase, and reduced nicotinamide adenine dinucleotide dehydrogenase. When phagocytosis was allowed to occur for longer periods the phagocytic vesicles appeared to fuse with each other and perhaps with digestive vacuoles. The resultant vesicles which contained many beads were heavier than those which consisted of only one bead or a few beads with a closely applied membrane. Ultrasonication ruptured the isolated vesicles, and the membranes could then be isolated in 30–50% yield based on phospholipid analysis. These membranes were essentially free of acid hydrolases and, presumably, other soluble proteins, as was also indicated by their low ratio of protein to phospholipid. The membranes have been prepared both as closed vesicles and as open sheets.
TEMPORAL ORDER IN MAMMALIAN CELLS
Authors: Klevecz Robert R.
Published: May 2004

Chinese hamster cells were synchronized by the Colcemid-selection system. In cells with a division cycle time of 11.5–12 hr, the activity of the enzyme lactate dehydrogenase (LDH) underwent marked oscillations with a 3.5-hr period. Precipitation of labeled LDH enzyme with specific antibody indicated that the enzyme activity changes were the result of intermittent enzyme synthesis and relatively constant degradation. Inhibition of normal DNA replication with 4 mM of thymidine, while reducing the amount of new enzyme synthesized, did not prevent oscillations from occurring. Similarly, actinomycin D (AcD) added at the time of synchronization allowed some new enzyme synthesis to proceed in an oscillatory manner. LDH synthesis went on at nearly normal rates when AcD was added in the middle of S phase. However, addition of cycloheximide to cultures at any time in the cycle caused an immediate drop in levels of activity and in enzyme protein. The half-life of LDH, calculated either from loss of enzyme activity or precipitable radioactivity in cycloheximide-treated cultures, was between 2 and 2.5 hr.
CYTOCHEMICAL LOCALIZATION OF PEROXIDATIC ACTIVITY OF CATALASE IN RAT HEPATIC MICROBODIES (PEROXISOMES)
Authors: Fahimi H. Dariush
Published: May 2004

Prominent staining of rat hepatic microbodies was obtained by incubating sections of aldehyde-fixed rat liver in a modified Graham and Karnovsky's medium for ultrastructural demonstration of peroxidase activity. The electron-opaque reaction product was deposited uniformly over the matrix of the microbodies. The microbodies were identified by their size, shape, presence of tubular nucleoids, and other morphologic characteristics, and by their relative numerical counts. The staining reaction was inhibited by the catalase inhibitor, aminotriazole, and by KCN, azide, high concentrations of H2O2, and by boiling of sections. These inhibition studies suggest that the peroxidatic activity of microbody catalase is responsible for the staining reaction. In the absence of exogenous H2O2 appreciable staining of microbodies was noted only after prolonged incubation. Addition of sodium pyruvate, which inhibits endogenous generation of H2O2 by tissue oxidases, or of crystalline catalase, which decomposes such tissue-generated H2O2, completely abolished microbody staining in the absence of H2O2. Neither diaminobenzidine nor the product of its oxidation had any affinity to bind nonenzymatically to microbody catalase and thus stain these organelles. The staining of microbodies was optimal at alkaline pH of 8.5. The biological significance of this alkaline pH in relation to the similar pH optima of several microbody oxidases is discussed. In addition to staining of microbodies, a heat-resistant peroxidase activity is seen in some of the peribiliary dense bodies. The relation of this reaction to the peroxidase activity of lipofuscin pigment granules is discussed.
FORMATION OF ARROWHEAD COMPLEXES WITH HEAVY MEROMYOSIN IN A VARIETY OF CELL TYPES
Authors: Ishikawa Harunori , Bischoff Richard , Holtzer Howard
Published: May 2004

Heavy meromyosin (HMM) forms characteristic arrowhead complexes with actin filaments in situ. These complexes are readily visualized in sectioned muscle. Following HMM treatment similar complexes appear in sectioned fibroblasts, chondrogenic cells, nerve cells, and several types of epithelial cells. Thin filaments freshly isolated from chondrogenic cells also bind HMM and form arrowhead structures in negatively stained preparations. HMM-filament complexes are prominent in the cortex of a variety of normal metaphase and Colcemid-arrested metaphase cells. There is no detectable binding of HMM with other cellular components such as microtubules, 100-A filaments, tonofilaments, membranes, nuclei, or collagen fibrils. The significance of HMM-filament binding is discussed in view of the finding that arrowhead complexes form in types of cells not usually thought to contain actin filaments.
MICROSPECTROPHOTOMETRY AND THE PHOTORECEPTOR OF PHYCOMYCES I
Authors: Wolken Jerome J.
Published: May 2004

By applying microspectrophotometry to the sporangiophore of Phycomyces blakesleeanus wild-type and the albino car-10(-) type II, absorption spectra were obtained for 1- to 5-day cultures. Spectra in the growing-zone of the wild-type during Stage IVb, taken from 0.1 to 3 mm below the base of the sporangium, show two distinctly different spectra: one is more characteristic of a carotene, the other of a flavin. Combined, these absorption spectra reproduce closely the action spectrum. For the albino car-10(-), which is deficient in carotenes, only the spectrum characteristic of lumichrome or a reduced flavin was found. A c-type cytochrome was isolated from both strains which, if coupled with a flavin, could permit a photoreversible oxidation-reduction system. Birefringent crystals were observed to be aligned in the growing zone in which the photoreceptor is believed to lie. Micro-spectrophotometry of these crystals shows absorption peaks similar to those of riboflavin crystals.
THE INFLUENCE OF THE MODE OF NUTRITION ON THE DIGESTIVE SYSTEM OF OCHROMONAS MALHAMENSIS
Authors: Stoltze H. J. , Lui N. S. T. , Anderson O. R. , Roels O. A.
Published: May 2004

The intracellular distribution and level of acid hydrolases in Ochromonas malhamensis were studied in cells grown osmotrophically in a defined medium, in a carbon-free starvation medium, and during phagotrophy in each of these media. By cytochemical techniques, little enzymic reaction product was observed in the vacuoles of osmotrophic cells grown in the defined medium. Starved cells, however, contained autophagic vacuoles and cannibalized other Ochromonas cells. Dense enzymic reaction product was observed in the digestive vacuoles and in the Golgi cisternae of these starved cells. Moreover, starved cells and cells grown in a nutritionally complete medium ingested Escherichia coli which appeared in digestive vacuoles containing enzymic reaction product. Biochemical assays for lysosomal acid phosphatase (E.C. 3.1.3.2 orthophosphoric monoester phosphohydrolase) and acid ribonuclease (E.C. 2.7.7.16 ribonucleate nucleotido-2'-transferase) were done on Ochromonas cultures in the same experimental treatments and under identical assay conditions as the cytochemical study. During starvation, the acid hydrolase specific activities were consistently twice those found in cells grown in an osmotrophic complete medium. Ochromonas fed E. coli showed no increase in acid hydrolase specific activity as compared to controls not fed E. coli. The latency of lysosomal acid hydrolases in cells fixed with glutaraldehyde was reduced, suggesting that this fixative increases lysosomal membrane permeability and may release enzymes or their reaction products into the cytoplasmic matrix during cytochemical analysis. This could explain the cytoplasmic staining artifact sometimes observed with glutaraldehyde-fixed cells when studied by the Gomori technique. This study confirms that Ochromonas malhamensis, a phytoflagellate, does produce digestive vacuoles and can ingest bacteria, thereby fulfilling its role as a heterotroph in an aquatic food chain. When Ochromonas is grown in a nutritionally complete osmotrophic medium, phagocytosis causes appearance of acid hydrolases in the digestive vacuoles, whereas the total activity of the enzymes remains unchanged. An organic carbon-free medium strongly stimulates acid hydrolaes activity and causes these enzymes to appear in the digestive vacuoles whether phagocytosis occurs or not.
THE METABOLISM OF CHYLOMICRON CHOLESTERYL ESTER IN RAT LIVER
Authors: Stein Olga , Stein Yechezkiel , Goodman Dewitt S. , Fidge Noel H.
Published: May 2004

Chylomicrons containing labeled cholesterol, mainly (70%) present as cholesteryl ester, were injected intravenously into intact rats, and samples of liver were obtained 27–210 min later. Most (58–75%) of the injected label was recovered in the liver after 27–75 min. Hepatic uptake occurred without hydrolysis of the labeled cholesteryl ester. In separate experiments, in vitro perfusion of livers of similarly treated rats for 30–35 min washed out only 3–9% of the labeled sterol. Samples of liver and small intestine were prepared for electron microscopy with Aquon as the dehydrating agent. Good retention (70% or more) of labeled cholesterol and satisfactory preservation of ultrastructure were obtained. After 30 min, the radioautographic reaction was localized mainly over the region of the cell boundary of the parenchymal liver cells, with fewer grains being present over intracellular organelles. At later time intervals, when considerable hydrolysis of the labeled cholesteryl ester had occurred, the radioautographic reaction was more evenly distributed. Phagocytosed labeled lipid was seen in Kupffer cells after the larger lipid load; phagocytosis by parenchymal cells was not seen. In other experiments, cholesteryl ester hydrolase activity was found in all subcellular fractions, the microsome and plasma membrane fractions showing the highest activity per mg protein. The mechanism of cholesteryl ester transport into the liver cell may involve: (1) hydrolysis at the cell surface; or (2) slow entry of intact molecules followed by intracellular hydrolysis of the ester bond.
CONTROL OF GROWTH HORMONE PRODUCTION BY A CLONAL STRAIN OF RAT PITUITARY CELLS
Authors: Bancroft Frank C. , Levine Lawrence , Tashjian Armen H.
Published: May 2004

Addition of hydrocortisone to the medium of a clonal strain of rat pituitary cells (GH3) stimulated the rate of production of growth hormone. The stimulation had a lag period of about 24 hr, reached a maximum at 70–100 hr, and was observed at a hydrocortisone concentration as low as 5 x 10-8 M. Cells maximally stimulated with 3 x 10-6 M hydrocortisone produced 50–160 µg growth hormone/mg cell protein/24 hr. These rates were four to eight times those observed in control cells. At maximum stimulation, intracellular levels of growth hormone in both stimulated and control cells were equal to the amount secreted into the medium in about 15 min. Removal of hydrocortisone from the medium of GH3 cells caused a return of the rate of growth hormone production to that in control cells. Addition of hydrocortisone to the medium of cells growing exponentially with a population-doubling time of 60 hr caused both an increase in the doubling time to 90 hr and a stimulation of growth hormone production. Cycloheximide (3.6 x 10-5 M) and puromycin (3.7 x 10-4 M) suppressed incorporation of labeled amino acids into protein by 93 and 98%, respectively, and suppressed growth hormone production by stimulated and control cells by at least 94%.
OPTICAL DIFFRACTION STUDIES OF CRYSTALLINE STRUCTURES IN ELECTRON MICROGRAPHS
Authors: Berger Jacob E.
Published: May 2004

Determination of the unit cell of crystalline particles by optical diffraction analysis of electron micrographs may establish the identity and help in approximating the molecular weight of the substances contained in the crystal. This technique may be particularly helpful when isolation and purification of the crystalline material cannot be accomplished.
CONFIGURATION OF A FILAMENTOUS NETWORK IN THE AXOPLASM OF THE SQUID (LOLIGO PEALII L.) GIANT NERVE FIBER
Authors: Metuzals J.
Published: May 2004

High-resolution electron microscopy is integrated with physicochemical methods in order to investigate the following preparations of the giant nerve fibers of the squid (Loligo pealii L.): (1) Thin sections of fibers fixed in four different fixatives; (2) fresh axoplasm stained negatively in solutions of different pH and composition; (3) chemically isolated threadlike elements of the axoplasm. A continuous, three-dimensional network can be identified in all these preparations of the axoplasm. The network is composed of coiled or looped unit-filaments ∼30 A wide. The unit-filaments are intercoiled in strands ∼ 70–250 A wide. The strands are oriented longitudinally in the axoplasm, often having a sinuous course and cross-associations. Microtubules are surrounded by intercoiled unit-filaments and filamentous strands. Calcium ions cause loosening and disintegration of the network configuration. UO2++ ions of a 1% uranyl acetate solution at pH 4.4 display a specific affinity for filamentous protein structures of the squid giant nerve fiber axoplasm, segregating the filamentous elements of the axoplasm in a coiled, threadlike preparation. The uranyl ions combine probably with the carboxyl groups of the main amino acids of the protein—glutamic and aspartic acids. It is proposed that by coiling/decoiling and folding/unfolding of the unit-filaments, shifts in physicochemical properties of the axoplasm are maintained.
HIGH-YIELD PREPARATION OF ISOLATED RAT LIVER PARENCHYMAL CELLS
Authors: Berry M. N. , Friend D. S.
Published: May 2004

A new technique employing continuous recirculating perfusion of the rat liver in situ, shaking of the liver in buffer in vitro, and filtration of the tissue through nylon mesh, results in the conversion of about 50% of the liver into intact, isolated parenchymal cells. The perfusion media consist of: (a) calcium-free Hanks' solution containing 0.05% collagenase and 0.10% hyaluronidase, and (b) magnesium and calcium-free Hanks' solution containing 2 mM ethylenediaminetetraacetate. Biochemical and morphologic studies indicate that the isolated cells are viable. They respire in a medium containing calcium ions, synthesize glucose from lactate, are impermeable to inulin, do not stain with trypan blue, and retain their structural integrity. Electron microscopy of biopsies taken during and after perfusion reveals that desmosomes are quickly cleaved. Hemidesmosome-containing areas of the cell membrane invaginate and appear to pinch off and migrate centrally. Tight and gap junctions, however, persist on the intact, isolated cells, retaining small segments of cytoplasm from formerly apposing parenchymal cells. Cells which do not retain tight and gap junctions display swelling of Golgi vacuoles and vacuoles in the peripheral cytoplasm. Cytoplasmic vacuolization in a small percentage of cells and potassium loss are the only indications of cell injury detected. By other parameters measured, the isolated cells are comparable to normal hepatic parenchymal cells in situ in appearance and function.
FINE STRUCTURE OF PACINIAN CORPUSCLES IN THE MESENTERY OF THE CAT
Authors: Nishi Katsuhide , Oura Chikayoshi , Pallie Wazir
Published: May 2004

Pacinian corpuscles in the mesentery of adult cats were fixed with either glutaraldehyde, osmium tetroxide or permanganate solutions by close intra-arterial injection through the mesenteric artery, and were processed, after electron staining and Epon embedding, for electron microscopy. Better resolution of the corpuscle's ultrastructure was obtained than available heretofore. The myelinated segment of the corpuscle contains blood vessels separated from the axon by collagen fibers and 3 to 4 layers of lamellae. No blood vessels are found in the central core, though access from the vessels is afforded by diffusion through the "cleft" of the inner core. Two cell types are discernible in the inner core hemilamellae; the "clear cells" in which pinocytotic vesicles and organelles abound and reflect the greater metabolic activity of these cells, in contrast to the "dark cells." The ultraterminal is ellipsoidal in form with projections into the "cleft" which give this portion an irregular appearance in section. The terminal and ultraterminal are packed with mitochondria, and "synaptic" vesicles are seen in the ultraterminal. The innermost laminae of the inner core cells are in close apposition to the terminal and break their regular pattern of hemilamellation to surround the small ultraterminal projections at the apical part of the corpuscle.
OBSERVATIONS ON THE ASSOCIATION OF HELICAL POLYRIBOSOMES AND FILAMENTS WITH VINCRISTINE-INDUCED CRYSTALS IN EARLE'S L-CELL FIBROBLASTS
Authors: Krishan Awtar , Hsu Dora
Published: May 2004

Earle's L-929 fibroblasts from cultures treated with 5–10 µg/ml of vincristine sulfate have a large number of eosinophilic, proteinaceous crystals in their cytoplasm. In electron micrographs, large arrays of helical polyribosomes, stacks of Golgi lamellae, and membranes of granular endoplasmic reticulum are seen in the cytoplasm of these cells. "Stalks" of fine granular material, approximately 300 A wide, are often seen in association with the arrays of the helical polyribosomes. In many sections rows of helical polyribosomes and filaments emerging from individual polyribosomes are seen in intimate contact with the crystals. A gradual reduction in the number of crystals and crystal-bearing cells is seen in cultures removed from the drug-containing medium and reincubated in fresh medium. In electron micrographs, these reincubated cells show large aggregates of filamentous material in the cytoplasm, and in many sections filaments are seen in continuity with the crystals.
A STUDY OF THE T SYSTEM IN RAT HEART
Authors: Forssmann W. G. , Girardier L.
Published: May 2004

The technique of extracellular space tracing with horseradish peroxidase is adapted for labeling the transverse tubular system (T system) in rat heart. In rat ventricular muscle the T system shows extensive branching and remarkable tortuosity. The T system can only be defined operationally, since it does not display specific morphological features throughout its entire structure. Owing to branching of the T system, a sizable proportion of the apposition between the T system and L system (or closed system) occurs at the level of longitudinal branches of the T system and is not restricted to the Z line region. The regions of apposition between the T system and L system are analyzed in rat ventricular muscle and skeletal muscle (diaphragm) and compared with the intercellular tight junctions (nexuses) of heart muscle by the use of a photometric method. The over-all thickness of the nexus is significantly smaller than that of T-L junctions in both cardiac and skeletal muscles. The thickness of the membranes of the T and L systems are not significantly different in the two muscles, but the gap between both membranes is larger in the heart. In atrial muscle the following two types of cells are found: (a) those cells with a well-developed T system in which the tubular diameter is quite uniform and the orientation predominantly longitudinal and, (b) cells with no T system, but with a well-developed L system. Atrial cells possessing a T system are richly provided with specific granules and show little micropinocytotic activity, whereas cells devoid of T system show intense micropinocytotic activity and few specific granules. The possible functional implications of these findings are discussed.
THE OUTWARD TRANSPORT OF CORTISOL BY MAMMALIAN CELLS IN VITRO
Authors: Gross Stephen R. , Aronow Lewis , Pratt William B.
Published: May 2004

It has been determined that cortisol and a few other steroids are transported outward from certain mammalian cells growing in vitro. The extrusion process is temperature dependent, glucose dependent, saturable, and operates for only a few selected steroids. Many, but not all, steroids are able to block the extrusion process but are not themselves transported. The outward transport process for steroids has been found in mouse fibroblasts, mouse lymphoma cells, and functional mouse adrenal gland tumor cells growing in vitro. The transport process is not present in two varieties of cells cultured from human sources—HeLa or diploid fibroblasts, WI-38.
ORIGIN, DEVELOPMENT, AND NATURE OF INTRANUCLEAR RODLETS AND ASSOCIATED BODIES IN CHICKEN SYMPATHETIC NEURONS
Authors: Masurovsky Edmund B. , Benitez Helena H. , Kim Seung U. , Murray Margaret R.
Published: May 2004

Correlative data are presented here on the developmental history, dynamics, histochemistry, and fine structure of intranuclear rodlets in chicken sympathetic neurons from in vivo material and long-term organized tissue cultures. The rodlets consist of bundles of ∼70 ± 10 A proteinaceous filaments closely associated with ∼0.4–0.8 µ spheroidal, granulofibrillar (gf) bodies of a related nature. These bodies are already present in the developing embryo a week or more in advance of the rodlets. In early formative stages rodlets consist of small clusters of aligned filaments contiguous with the gf-bodies. As neuronal differentiation progresses these filaments increase in number and become organized into well-ordered polyhedral arrays. Time-lapse cinemicrography reveals transient changes in rodlet contour associated with intrinsic factors, changes in form and position of the nucleolus with respect to the rodlet, and activity of the gf-bodies. With the electron microscope filaments may be seen extending between the nucleolus, gf-bodies, and rodlets; nucleoli display circumscribed regions with fine structural features and staining reactions reminiscent of those of gf-bodies, We suggest that the latter may be derivatives of the nucleolus and that the two may act together in the assemblage and functional dynamics of the rodlet. The egress of rodlet filaments into the cytoplasm raises the possibility that these might represent a source of the cell's filamentous constituents.
LOCALISATION MITOCHONDRIALE D'ENZYMES LIEES AU METABOLISME DU GLYCOGENE DANS LE SPERMATOZOIDE DE L'ESCARGOT
Authors: Personne Paul , Anderson Winston
Published: May 2004

The mitochondrial derivative of the sperm cell of the Gastropod Helix is endowed with a compartment loaded with glycogen (Personne et André, 1964). The aim of this work is to establish whether this mitochondrial glycogen is synthesized in the mitochondrial derivative itself or elsewhere in the cell. For this purpose, living sperm were first incubated in a medium containing glucose 1-phosphate, then fixed, sectioned, and stained specifically for polysaccharides by the phosphotungstic acid technique or the periodic acid-thiosemicarbazide-silver proteinate technique. Comparison with controls shows that a synthesis of glycogen occurred during the time of incubation. It can be inferred from this result that an amylophosphorylase system controlling the metabolism of polysaccharides is present in the mitochondrial derivative itself. Results obtained with the iodine technique indicate that the original glycogen molecules are elongated during the experiment. It seems probable that the amylophosphorylase system demonstrated here accounts for at least a part of the proteinaceous coat morphologically detectable around each individual glycogen particle.
PREPARATION OF ISOLATED RAT LIVER MITOCHONDRIA FOR ELECTRON MICROSCOPY
Authors: Butler W. H. , Judah J. D.
Published: May 2004

A comparative study of the fixation of isolated rat liver mitochondria was undertaken. If the criterion is adopted that after processing, the mitochondria should resemble as closely as possible rat liver mitochondria in situ, the procedure found to produce such preservation was that of fixation in suspension in veronal-buffered 2% potassium permanganate. Fixation in osmium tetroxide produced variable results, while mitochondria fixed in glutaraldehyde were contracted. We suggest that in cases where fixation procedures modify the morphological appearance of mitochondria, the significance of such changes must be treated with caution.
LYSOZYME IN EPIPHYSEAL CARTILAGE
Authors: Kuettner K. E. , Soble L. W. , Ray R. D. , Croxen R. L. , Passovoy M. , Eisenstein R.
Published: May 2004

Embryonic mouse femoral cartilage, like the epiphyseal cartilage of the calf scapula, contains large amounts of lysozyme. The addition of egg white lysozyme to organ cultures of embryonic mouse femurs induces unique alterations in the gross and microscopic morphology of the femurs. The sites of these alterations are precisely related to the natural distribution of lysozyme in calf scapula. If the exogenous lysozyme is withdrawn from the culture, the morphological changes disappear, accompanied by a resumption or derepression of growth. The effect on growth is evident only in 17-day embryos. These observations support the idea that lysozyme has a physiological role in cartilage, perhaps related to a regulatory mechanism in bone formation.
CELLULAR ENERGY METABOLISM DURING FETAL DEVELOPMENT
Authors: Warshaw Joseph B. , Terry Mary L.
Published: May 2004

In view of the importance of fatty acids as substrates for the mature heart, fatty acid oxidation by fetal and calf heart mitochondria has been investigated. Free fatty acids of 10 carbon units or less which exhibit carnitine-independent transport into mitochondria were effective substrates for oxidative phosphorylation in both fetal and calf heart mitochondria. Efficient oxidative phosphorylation with these substrates was dependent upon the presence of bovine serum albumin in the assay medium to reverse the uncoupling effects of the fatty acids. In the presence of bovine serum albumin, ADP/0 ratios were in the range of 3 when short-chain fatty acids and carnitine esters of short- and long-chain fatty acids were substrates. Compared with calf heart mitochondria, fetal heart mitochondria showed decreased carnitine-dependent oxidation of palmityl-CoA. However, the oxidation of palmitylcarnitine was identical in both. These data suggest that the formation of palmitylcarnitine is rate limiting for palmityl-CoA oxidation by the fetal heart mitochondria and that long-chain fatty acids are not readily oxidized by the fetal heart.
INTRANEURAL DISTRIBUTION OF EXOGENOUS NOREPINEPHRINE IN THE CENTRAL NERVOUS SYSTEM OF THE RAT
Authors: Descarries Laurent , Droz Bernard
Published: May 2004

Catecholaminergic neurons, which take up and retain exogenous norepinephrine labeled with tritium, were studied by means of high resolution radioautography, in the substantia nigra, the substantia grisea periventricularis, and the locus coeruleus of the rat. Under the conditions required for the radioautographic detection of exogenous norepinephrine-3H, it was established that (1) glutaraldehyde was the most suitable fixative for preserving the labeled amine in situ; (2) norepinephrine-3H itself, rather than metabolites, accounted for most of the reactions detected in catecholaminergic neurons. At various time intervals after an intraventricular injection of norepinephrine-3H, the tracer reached a concentration 15–100 times higher, and disappeared at a slower rate, in presynaptic axons (t½:4 hr) than in nerve cell bodies (t½:0.8–1.3 hr). After pretreatment with a monoamine oxidase inhibitor, the radioautographic reactions increased and persisted longer, especially in the preterminal axons. Within neurons, the labeled amine was ubiquitously distributed in the nerve cell body and concentrated in presynaptic axons and synaptic terminals of various morphological types. Although large granular vesicles were usually present in the labeled axonal bulbs, no structural characteristic could be specifically ascribed to catecholaminergic neurons. It is suggested that exogenous norepinephrine bound to macromolecular complexes is present in all parts of catecholaminergic neurons and mainly concentrated within presynaptic axons.
PREPARATION AND CHARACTERIZATION OF A PLASMA MEMBRANE FRACTION FROM ISOLATED FAT CELLS
Authors: McKeel Daniel W. , Jarett Leonard
Published: May 2004

A rapid method of preparing plasma membranes from isolated fat cells is described. After homogenization of the cells, various fractions were isolated by differential centrifugation and linear gradients. Ficoll gradients were preferred because total preparation time was under 3 hr. The density of the plasma membranes was 1.14 in sucrose. The plasma membrane fraction was virtually uncontaminated by nuclei but contained 10% of the mitochondrial succinic dehydrogenase activity and 25–30% of the RNA and reduced nicotinamide adenine dinucleotide cytochrome c reductase activity of the microsomal fraction. Part of the RNA and NADH-cytochrome c reductase activity was believed to be native to the plasma membrane or to the attached endoplasmic reticulum membranes demonstrated by electron microscopy. The adenyl cyclase activity of the plasma membrane fraction was five times that of Rodbell's "ghost" preparation and retained sensitivity to epinephrine. The plasma membrane ATPase activity was five times that of the homogenate and microsomal fractions. Electron microscopic evidence suggested contamination of the plasma membrane fraction by other subcellular components to be less than the biochemical data indicated.
A COMPARISON BETWEEN HETEROGENEOUS NUCLEAR RNA AND POLYSOMAL MESSENGER RNA IN HELA CELLS BY RNA-DNA HYBRIDIZATION
Authors: Soeiro Ruy , Darnell James E.
Published: May 2004

Heterogeneous nuclear RNA (HnRNA) and mRNA from cytoplasmic polyribosomes of HeLa cells have been compared by RNA-DNA hybridization tests. 1 µg of HeLa cell DNA binds 0.05–0.10 µg of either HnRNA or mRNA. In addition, HeLa DNA that is preexposed to unlabeled HnRNA was found to have a reduced capacity to bind either HnRNA or mRNA. The results are compatible with considerable sequence similarity in the two types of RNA but, as is discussed, firm conclusions are precluded by imperfections of the hybridization reaction as presently employed.
A COMPARISON OF THE HETEROGENEOUS NUCLEAR RNA OF HELA CELLS IN DIFFERENT PERIODS OF THE CELL GROWTH CYCLE
Authors: Pagoulatos Gerassimos N. , Darnell James E.
Published: May 2004

HeLa cells synthesize heterogeneous nuclear RNA (HnRNA) in the G1, S, and G2 portions of the cell cycle. HnRNA prepared from these various periods was compared by RNA-DNA hybridization experiments. The results indicated that some of the HnRNA molecules were equivalent at all times in the cell cycle, but limitations in the sensitivity of the hydridization reactions, as well as in the spectrum of hybridizing molecules, restrict the conclusions that can be drawn from these comparisons.
ISOLATION OF A GOLGI APPARATUS-RICH FRACTION FROM RAT LIVER
Authors: Morré D. James , Hamilton R. L. , Mollenhauer H. H. , Mahley R. W. , Cunningham W. P. , Cheetham R. D. , Lequire V. S.
Published: May 2004

Golgi apparatus were released without fixatives from rat hepatocytes by gentle homogenization, concentrated by differential centrifugation, and purified by sucrose gradient centrifugation. Examination of sections of purified fractions by electron microscopy showed fields of morphologically intact units of Golgi apparatus consisting of stacks of parallel flattened cisternae, secretory vesicles, and small vesicular profiles. Negative staining of unfixed pellets revealed a complex network of anastomotic tubules continuous with platelike structures and secretory vesicles. These structures corresponded, respectively, to the small vesicular profiles and parallel flattened cisternae with attached secretory vesicles of sectioned material. Small fragments of granular endoplasmic reticulum were often closely associated with the peripheral tubules, suggesting sites of continuity in intact hepatocytes.
HEMOGLOBIN UPTAKE BY RAT HEPATOCYTES AND ITS BREAKDOWN WITHIN LYSOSOMES
Authors: Goldfischer Sidney , Novikoff Alex B. , Albala Arline , Biempica Luis
Published: May 2004

The peroxidatic activity of hemoglobin permitted visualization of its uptake by rat hepatocytes by means of the Graham-Karnovsky 3,3'-diaminobenzidine (DAB) procedure. Lysosomes were visualized by their acid phosphatase, ß-glucuronidase, and glucosaminidase activities. When large doses of rat, cow, or human hemoglobin are intravenously injected, or when hemoglobinemia is induced by injection of distilled water, DAB-positive hemoglobin is engulfed by pinocytosis. Pinocytotic vacuoles become digestive vacuoles ("phagolysosomes") by fusion with lysosomes of the dense body type that have moved from their pericanalicular position. By 16–24 hr after even massive amounts of hemoglobin (400 mg/100 g), the protein is barely demonstrable in hepatocytes. At the lowest doses of injected hemoglobin (15 mg/100 g body weight), DAB-positive vacuoles are demonstrable only in the Kupffer cells.
CHLOROPLAST STRUCTURE AND FUNCTION IN ac-20, A MUTANT STRAIN OF CHLAMYDOMONAS REINHARDI
Authors: Togasaki R. K. , Levine R. P.
Published: May 2004

A mutant strain of the green alga Chlamydomonas reinhardi, ac-20, is described in which both the rate of CO2 fixation by whole cells and the rate of carboxylation of ribulose-1,5-diphosphate in cell-free extracts are reduced, particularly when sodium acetate is present in the growth medium. Of the enzymes of the reductive pentose phosphate cycle tested, only ribulose-1,5-diphosphate carboxylase activity is reduced in the mutant strain, and it appears that the low carboxylase activity limits the strain's rate of photosynthetic carbon metabolism. Evidence is presented to show that the fluctuation in the level of the enzyme activity in the presence or absence of acetate results from the fluctuation in the level of some factor(s) limiting the rate of synthesis of the protein.
BIOGENESIS OF CHLOROPLAST MEMBRANES
Authors: Goldberg I. , Ohad I.
Published: May 2004

The glycolipid, phospholipid, pigment, and fatty acid content in whole y-1 cells during the greening process have been investigated. The time course of their changes indicates that phosphatidyl glycerol and glycolipids are the main lipids synthesized specifically during illumination of dark-grown cells, concomitant with an increase in the polyunsaturated C18:2 and C18:3 fatty acids. The pigment complex of light-grown cells consists mainly of chlorophylls a and b, lutein, ß-carotene, violaxanthin, and neoxanthin. During the greening process, chlorophylls a and b are synthesized in constant proportions (ratio a/b equals 2.6), ß-carotene and violaxanthin do not change significantly, and lutein and neoxanthin increase. The molar ratios of the different lipids and pigment to total chlorophyll during greening has been calculated. It was found that during the initial phase of greening when chlorophyll is synthesized at increasing rates, the molar ratios of various lipids and pigments to chlorophyll decrease and tend to become constant when chlorophyll and membrane synthesis proceed at constant rates. The implication of these findings with respect to the concept of membrane assembly through a spontaneous single step process is discussed
WOUND HEALING AND COLLAGEN FORMATION
Authors: Ross Russell , Everett Newton B. , Tyler Ruth
Published: May 2004

Healing skin wounds were studied in a series of parabiotic rats. The femurs of one parabiont of each pair were shielded whilst both animals were given 800 r from a Co60 source. The animals were wounded 3 days after irradiation. Each animal with partially shielded marrow was then given tritiated thymidine intraperitoneally daily while the cross-circulation was arrested by clamping. After the thymidine-3H had cleared the blood, the clamp was released. Animals were sacrificed, and wounds were prepared for radioautography 1, 2, and 6 days after wounding. In the wounds of the shielded animals thymidine-3H was observed in epidermis, endothelium, leukocytes, fibroblasts, and mast cells. Only neutrophilic leukocytes, monocytes, and lymphocytes were labeled, as determined by light and electron microscope radioautography, in the wounds of each nonshielded parabiont. None of the many fibroblasts present were found to contain label in the wounds of the nonshielded parabionts through the 6 day period. These observations provide further evidence that wound fibroblasts do not arise from hematogenous precursors and, therefore, must arise from adjacent connective tissue cells.
CYTOCHEMICAL LOCALIZATION OF ENDOGENOUS PEROXIDASE IN THYROID FOLLICULAR CELLS
Authors: Strum Judy M. , Karnovsky Morris J.
Published: May 2004

Endogenous peroxidase activity in rat thyroid follicular cells is demonstrated cytochemically. Following perfusion fixation of the thyroid gland, small blocks of tissue are incubated in a medium containing substrate for peroxidase, before being postfixed in osmium tetroxide, and processed for electron microscopy. Peroxidase activity is found in thyroid follicular cells in the following sites: (a) the perinuclear cisternae, (b) the cisternae of the endoplasmic reticulum, (c) the inner few lamellae of the Golgi complex, (d) within vesicles, particularly those found apically, and (e) associated with the external surfaces of the microvilli that project apically from the cell into the colloid. In keeping with the radioautographic evidence of others and the postulated role of thyroid peroxidase in iodination, it is suggested that the microvillous apical cell border is the major site where iodination occurs. However, that apical vesicles also play a role in iodination cannot be excluded. The in vitro effect of cyanide, aminotriazole, and thiourea is also discussed.
CYTOPLASMIC INCLUSIONS RESEMBLING NUCLEOLI IN SYMPATHETIC NEURONS OF ADULT RATS
Authors: Grillo Mary A.
Published: May 2004

A distinctive cytoplasmic inclusion consisting of a convoluted network of electron-opaque strands embedded in a less dense matrix was identified in the neurons, but not in the supporting cells, of rat sympathetic ganglia. This ball-like structure, designated "nematosome," measures ∼ 0.9 µ and lacks a limiting membrane. Its strands (diameter = 400–600 A) appear to be made of an entanglement of tightly packed filaments and particles ∼ 25–50 A thick. Cytochemical studies carried out with the light microscope suggest the presence of nonhistone proteins and some RNA. Usually only one such structure is present in a cell, and it appears to occur in most ganglion cells. Although they can be seen anywhere in the cell body, nematosomes are typically located in the perinuclear cytoplasm, where they are often associated with smooth-surfaced and coated vesicles. In fine structure and stainability, they bear a resemblance to the fibrous component of the nucleolus. Subsynaptic formations, which are a special feature of some of the synapses in sympathetic ganglia, appear similar to the threadlike elements in the nematosomes. The possibility that these three structures—nucleolus, nematosome, and subsynaptic formation—may be interrelated in origin and function is discussed.
CONTROLLED PROTEOLYSIS OF NASCENT POLYPEPTIDES IN RAT LIVER CELL FRACTIONS
Authors: Blobel G. , Sabatini D. D.
Published: May 2004

Free ribosomes containing nascent polypeptide chains labeled in vitro were submitted to proteolysis at 0° by a mixture of trypsin and chymotrypsin. Sucrose gradient analysis showed that polysome patterns are retained even after 24 hr of proteolysis in the cold, while messenger RNA-free ribosomes (generated progressively during in vitro incorporation) are, within 2 hr, completely dissociated into subunits by trypsin. Although ribosomes and subunits are not extensively degraded into smaller fragments during low temperature proteolysis, changes in the acrylamide gel electrophoresis pattern showed that most ribosomal proteins are accessible to and are partially degraded by the proteases. Ribosome-bound nascent polypeptides are partially resistant to proteolysis at 0°, although they are totally digested at 37° or when the ribosomal subunit structure is disrupted by other means. Radioactivity incorporated into nascent chains during incubation times shorter than 3 min was mostly resistant to digestion at 0°. A larger fraction of the initial radioactivity became degraded in ribosomes which incorporated for longer times. In these ribosomes, the amount of radioactivity which was resistant to proteolysis was constant and independent of the initial value, which reflects the labeled length of the nascent chains. These results suggest that the growing end of the nascent polypeptide is resistant to digestion and is protected from proteolytic attack by the ribosomal structure. A pulse and chase experiment confirmed this suggestion, showing that the protected segment is located at the carboxy-terminal end of the nascent chain. The protected segment was contained in the large ribosomal subunit and had a length of ∼39 amino acid residues, as estimated by chromatography on Sephadex G-50.
THE FORMATION OF MULTIVESICULAR BODIES FROM THE NUCLEAR ENVELOPE
Authors: Kilarski Wincenty , Jasiński Andrzej
Published: May 2004

Cells of the gas gland of the perch Perca fluviatilis L., stimulated to increased generation of gas by the repeated emptying of the swim-bladder, were examined in the electron microscope. Intense activity of the nuclear envelope was demonstrated. Simple vesicles originating from the external nuclear membrane and the so-called multivesicular bodies derived from the outpocketings of both membranes of the nuclear envelope were observed. The multivesicular bodies were filled with numerous fine vesiculae arising from the active proliferation of their internal membrane. The authors offer two alternative mechanisms of formation of fine vesiculae inside the multivesicular bodies and the mechanism of the tearing away of these bodies from the nuclear envelope.
THE ASSOCIATION OF A CLASS OF SALTATORY MOVEMENTS WITH MICROTUBULES IN CULTURED CELLS
Authors: Freed Jerome J. , Lebowitz Marcia M.
Published: May 2004

Particulate structures in the cytoplasm of HeLa and other cultured cells in interphase undergo rapid individual linear displacements (long saltatory movements, LSM). By the use of time-lapse microscopy to locate saltating particles prior to fixation and histochemical examination of the cells, structures of several kinds have been shown to move in this manner. Elements that show LSM include lysosomes, pinosomes, ingested carbon particles, lipoidal granules, and unidentified particles that appear as bright objects in positive phase contrast. The pattern of movement of the particles suggests the presence of linear guiding elements radially disposed from the cytocenter (centriole region). The participation of microtubules in these movements is inferred from the observation that LSM cease after treatment with drugs which depolymerize microtubules, i.e., colchicine, Vinblastine, and podophyllin. The directions of the microtubules in the cytoplasm of HeLa cells found by electron microscopy are consistent with the aster-like configuration predicted from study of LSM. Further support for this arrangement of cytoplasmic microtubules is provided by light microscope observations of colchicine-sensitive radial arrays of acid phosphatase granules in the cytoplasm of some cell lines.
A STUDY OF SIEVE ELEMENT STARCH USING SEQUENTIAL ENZYMATIC DIGESTION AND ELECTRON MICROSCOPY
Authors: Palevitz Barry A. , Newcomb Eldon H.
Published: May 2004

The fine structure of plastids and their starch deposits in differentiating sieve elements was studied in bean (Phaseolus vulgaris L.). Ultrastructural cytochemistry employing two carbohydrases specific for different linkages was then used to compare the chemical nature of "sieve tube starch" (the starch deposited in sieve elements) with that of the ordinary starch of other cell types. Hypocotyl tissue from seedlings was fixed in glutaraldehyde, postfixed in osmium tetroxide, and embedded in Epon-Araldite. Treatment of thin sections on uncoated copper grids with α-amylase or diastase at pH 6.8 to cleave α-(1 → 4) bonds resulted in digestion of ordinary starch grains but not sieve element grains, as determined by electron microscopy. Since α-(1 → 6) branch points in amylopectin-type starches make the adjacent α-(1 → 4) linkages somewhat resistant to hydrolysis by α-amylase, other sections mounted on bare copper or gold grids were treated with pullulanase (a bacterial α-[1 → 6] glucosidase) prior to digestion with diastase. Pullulanase did not digest sieve element starch, but rendered the starch digestible subsequently by α-amylase. Diastase followed by pullulanase did not result in digestion. The results provide evidence that sieve element starch is composed of highly branched molecules with numerous α-(1 → 6) linkages.
THE SYNTHESIS AND TURNOVER OF RAT LIVER PEROXISOMES
Authors: Poole Brian , Higashi Tokuhiko , de Duve Christian
Published: May 2004

Rat liver peroxisomes have been separated according to size by zonal sedimentation. A method is described for calculating the size of the particles from their final position in the gradient. Peroxisomes seem biochemically homogeneous throughout their size distribution. 3 hr after injection of tritiated leucine, the specific radioactivity of catalase is the same in peroxisomes of different sizes, and it remains so for up to 1 wk after administration of the precursor. This observation rules out the possibility that peroxisomes have an extended period of independent growth. If individual particles maintain an independent existence, they must be formed very rapidly. The other possible explanation is that peroxisomes exchange material within the liver cell.