Stimulation of cartilage macromolecule synthesis by adenosine 3',5'-monophosphate.

The role of cyclic AMP in the regulation of cartilage macromolecule synthesis in vitro was studied in pelvic cartilage from 10-12 day chick embryos. Incubation of cartilages in medium containing 0.5 mM cyclic AMP resulted in a 30% inhibition of 35SO4-2, [3H]leucine and [3H]uridine incorporation into proteoglycan, total protein and RNA, respectively. Higher concentrations of cyclic AMP had no greater effects. In contrast, butyrylated cyclic AMP derivatives (0.5-5.0 mM) added to the incubation medium stimulated (50-100%) the incorporation of these radiolabeled precursors into cartilage macromolecules. Theophylline, in concentrations (0.1-0.5 mM) which raise intracellular cyclic AMP, also increases the incorporation of radiolabeled precursors into macromolecules. The data indicate that exogenous cyclic AMP and butyrylated cyclic AMP derivatives have paradoxical effects on cartilage macromolecule synthesis. Butyrylated cyclic AMP derivatives, not exogenous cyclic AMP, mimic the effects of intracellular cyclic AMP. Incubation of embryonic chicken cartilage with exogenous cyclic AMP results in the extracellular degradation of the cyclic AMP to adenosine. Adenosine (0.125 mM) inhibits precursor incorporation into cartilage macromolecules. The metabolism of exogenous cyclic AMP generates sufficient adenosine to account for the observed inhibitory effects of exogenous cyclic AMP on cartilage macromolecule synthesis. Butyrylated cyclic AMP derivatives are not degraded during incubation with cartilage. The data indicate that cartilage is a tissue in which the effect of cyclic AMP is to stimulate anabolic processes.


Non-lytic, non-ionic detergent extraction of plasma membrane constituents from normal and transformed fibroblasts.

A technique has been developed for the selective extraction of plasma membrane protein constituents from normal and transformed cell employing non-ionic detergents. The extraction procedure does not damage cells as judged by cell viability, 51Cr release, and trypan blue staining. Lactoperoxidase-catalyzed iodination followed by detergent extraction permits demonstration of a 100,000 dalton protein which is found on the surface of normal but not transformed hamster and mouse fibroblasts.


Morphological and biochemical characterization of light and heavy myelin isolated from developing rat brain.

Myelin from developing rat brains was separated on a discontinuous sucrose gradient into subfractions of two different densities, i.e. light and heavy myelin. Electron photomicrographs showed that heavy myelin consisted primarily of large compacted multilamellar structures with a distinct intraperiod line characteristic of myelin in situ. Light myelin, on the other hand, was composed of small vesicles having a unilamellar structure. Similar to whole myelin, both membrane subfractions were highly enriched in 2',3'-cyclic nucleotide-3'-phosphohydrolase. The specific activity of the enzyme, however, showed no developmental trend. Both subfractions contained all the four major proteins characteristic of the whole myelin membrane. There were, however, quantitative differences in the relative distribution of these proteins between light and heavy myelin. Basic protein accounted for 55% and proteolipid protein for 46% of the total myelin proteins of light and heavy myelin, respectively. DM-20 (Agrawal, H.C., Burton, R. M., Fishman, M.A., Mitchell, R.F. and Prensky, A.L. (1972) J. Neurochem. 19, 2083-2089) exhibited a developmental "switch" between light and heavy myelin. Light myelin appeared to contain more DM-20 in 15- to 20-day-old rat brain, whereas the concentration of this protein was higher in heavy myelin at subsequent ages studied.


Altered surface topology and membrane functions of rat thymocytes eluted from nylon wool columns.

(1) Following incubation of thymocytes with nylon wool at 37 degrees C, the eluted cells showed an increase in the number of microvilli per cell and a concominant elongation of the microvilli (0.22 mum versus 1.15 mum. (2) Cyclic adenosine monophosphate (cylic AMP) levels were lowered by 30-50% in nylon wool-treated thymocytes. (3) Nylon wool-treated cells showed an impaired Na+-dependent amino acid transport system (2-aminoisobutyrate) whereas the Na+-independent amino acid transport system (1-aminocyclopentane-1-carboxylate) was unaffected.


On the use of the spin labeling technique in the study of erythrocyte membranes.

ESR spectra and scanning electron micrographs of human erythrocytes spin labeled with the conventional stearic acid nitroxide substituted at the 5-position have been obtained over a range of label-to-lipid ratios. While morphological changes as previously reported (Bieri, V. G., Wallach, D. F. H. and Lin, P. S. (1974) Proc. Natl. Acad. Sci. U.S. 71, 4797-4801) are reproduced, it is shown that at label-to-lipid ratios of 1:10 or less the basic ESR spectrum is not significantly affected. At low label concentrations the spin labeling technique is a viable one and can be used to investigate membrane properties.


Endogenous cyclic AMP in thyroid cell culture. Effect of thyroid-stimulating hormone and dibutyryl cyclic AMP.

We have studied the variations of endogenous cyclic AMP levels in thyroid cells cultured over a period of 7 days in several conditions: in the presence of thyroid-stimulating hormone or dibutyryl cyclic AMP which both promote the aggregation of isolated cells into follicles, and in their absence when cells develop as a typical monolayer. In follicle-forming cells, the cyclic AMP level was found to rise during the first day of culture, then to fall rapidly. In monolayer-forming cells, the cyclic AMP content slightly increases attaining the same level as found in other cells at the fourth day, which remains stable till the seventh day. We have investigated the response of these cells cultured in the presence of dibutyryl cyclic AMP retain the capability of increasing their cyclic AMP concentration whereas monolayer-forming cells do not preserve this quality of thyroid cells.


Sedimentation behavior of solubilized gonadotropin receptor from plasma membranes of bovine corpus luteum.

The gonadotropin receptors associated with plasma membrane fractions were solubilized by detergents, including Triton X-100, Lubrol WX, Lubrol PX and sodium deoxycholate before and after equilibration with 125I-labelled human chorionic gonadotropin. The binding activity remained in solution even after centrifugation at 300 000 X g for 3 h. The solubilized gonadotropin receptor or gonadotropin receptor complex was characterized by gel filtration and sucrose density gradient centrifugation. Sucrose density gradient centrifugation of solubilized gonadotropin-receptor complex in the presence of Triton X-100 had a sedimentation coefficient of 6.5 S whereas the solubilized uncomplexed receptor had a sedimentation coefficient of 5.1 S. In the absence of the detergent, solubilized hormone receptor complex from plasma membrane fractions I and II sedimented with an apparent sedimentation coefficient of 6.6 S and 7.4 S, respectively. Similarly, the free receptor also showed higher sedimentation profile with an apparent sedimentation coefficient of 6.7 S for fraction I and 7.2 S for fraction II. Treatment of plasma membranes with phospholipase A and C inhibited the binding of 125I-labelled human chorionic gonadotropin in a dose dependent manner, whereas phospholipase D was without any effect. Doses of 1.4 mI. U. of phospholipase A or 0.6 mI.U. of phospholipase C were required to produce 50% inhibition of the binding activity. These phospholipases had no effect on the preformed 125I-labelled human chorionic gonadotropin-receptor complex nor on the sedimentation profile of solubilized gonadotropin receptor complex.


An adenosine 3':5' monophosphate dependent protein kinase from sea urchin spermatozoa.

A cyclic AMP dependent protein kinase (EC 2.7.1.37) from sea urchin sperm as purified to near homogeneity and characterized. A 68-fold purification of the enzyme was obtained. This preparation had a specific activity of 389 000 units/mg protein with protamine as the substrate. On the basis of the purification required, it may be calculated that the protein kinase constitutes as much as 1.5% of the soluble protein in sperm. There appeared to be a single form of the enzyme in sea urchin sperm, based on the behavior of the enzyme during DEAE-cellulose and Sephadex G-200 column chromatography. Magnesium ion was required for enzyme activity. The rate of phosphorylation of protamine was stimulated 2.5-fold by an optimal concentration of 0.9 M NaCl. The Km for ATP (minus cyclic AMP) was 0.119 +/- 0.013 (S.D.) and 0.055 mM +/- 0.009 (S.D.) in the presence of cyclic AMP. The specificity of the enzyme toward protein acceptors, in decreasing order of phosphorylation, was found to be histone f1 protamine, histone f2b, histone f3 and histone f2a; casein and phosvitin were not phosphorylated. The holoenzyme was found to have an apparent molecular weight of 230 000 by Sephadex G-200 chromatography. In the presence of 5 - 10(-6) M cyclic AMP, the holoenzyme was dissociated on Sephadex G-200 to a regulatory subunit of molecular weight 165 000 and a catalytic subunit of Mr 73 000. The dissociation could also be demonstrated by disc gel electrophoresis in the presence and absence of cyclic AMP.


Nucleoside-dependent protein kinase from Trypanosoma gambiense.

A nucleoside-dependent protein kinase (EC 2.7.1.37) was partially purified from Trypanosoma gambiense, the pathogenic agent of sleeping sickness. This enzyme catalyzes the phosphorylation of histone and protamine. Various nucleosides at the concentration of 10(-4) M stimulated the histone kinase activity about two-fold, whereas cyclic AMP and cyclic GMP were without effect. The pH-optimum for histone phosphorylation was at about pH 7.0. The enzyme activity absolutely depends on Mg2+, Mn2+ or Co2+. The apparent Km-value for histone was 0.3 mg/ml and those for ATP were 2 - 10(-4) M and 6 - 10(-5) M in the absence or presence of 10(-4) M adenosine respectively. IDP and ADP complete with ATP. The inhibition constants were calculated to be 2 - 10(-4) M and 2.5 - 10(-4) M, respectively. The molecular weight of the histone kinase was found to be 95 000 by gel filtration and 88 000 by sedimentation in a sucrose gradient.


Characterization of protein phosphokinase activities in horse thyroid nuclei.

The distribution of protein phosphokinase (EC 2.7.1.37) activities has been established in horse thyroid nuclei. The presence of several enzyme activities has been demonstrated, two of which are clearly distinct. The first one acts on histone as substrate and is activated by cyclic AMP. Physico-chemical properties of this nuclear cyclic AMP-dependent histone kinase and of the cytosol histone kinase are different, demonstrating the absence of a contamination from the cytosol. The second enzyme acts on casein as substrate and is not stimulated by cyclic AMP POR CYCLIC GMP. The findings are consistent with the observation of thyrotropin stimulation of histone phosphorylation in thyroid nuclei.


Activation of rat liver pyrimidine nucleoside monophosphate kinase.

The activity of the pyrimidine nucleoside monophosphate kinase (ATP:dCMP phosphotransferase, EC 2.7.4.14) from rat liver is dependent upon the presence of sulfhydryl-reducing agents. Addition to the inactive enzyme of 2-mercaptoethanol (5 mM), a reagent specific for cleavage of disulfide bonds, effects a reduction in molecular weight from approx. 53 000 to 17 000, measured by molecular sieve chromatography. This low molecular weight form is partially active in the presence of 2-mercaptoethanol (f mM). In absence of 2-mercaptoethanol, the low molecular weight form is inactive. Higher concentrations of 2-mercaptoethanol (50 mM) fully reactivate the CMP(ATP) kinase activity followed by dCMP(ATP) and CMP(dCTP) kinase activities in a sequential manner, without further change in moelcular weight. Alkylation by iodoacetamide of the enzyme at different stages of reactivation in dithiothreitol suggests an ordered appearance of the various enzyme activities. Furthermore, iodoacetamide inactivates the fully active enzyme. Thioredoxin was found to activate the enzyme in a manner similar to 2-mercaptoethanol and dithiothreitol. These results are consistent with the interpretation that the mechanism of activation of the enzyme involves cleavage of inter- and intramolecular disulfide bonds.


Procollagenase from bovine gingiva.

1. Collagenase (EC 3.4.24.3) is released from bovine gingival explants in vitro as a zymogen. The zymogen does not hydrolyze collagen and does not form a complex with alpha2-macroglobulin (alpha2-M). It elutes in gel filtration with an apparent molecular weight of approx. 80 000. 2. Incubation of the zymogen with trypsin results in a 15 000-20 000 dalton decrease in molecular weight and imparts to the enzyme the ability to hydrolyze collagen and to form a complex with alppha2-M. 3. The zymogen can be completely separated from the active enzyme to alpha2-M. Likewise, the zymogen can be harvested from cultures supplemented with serum.


Chemical characterization and study of the autodigestion of pure collagenase from Achromobacter iophagus.

Only one collagenase (EC 3.4.24.3) is produced by the non-pathogenic Achromobacter iophagus strain. The chromatography of the crude enzyme on DE-32 cellulose followed by gel filtration on Sephadex G-100 in the presence of 1 M sodium chloride led to the isolation of a homogeneous enzyme. Its specific activity (1.642 mukat/mg) represents the highest value ever obtained for a bacterial collagenase. The amino acid composition of A. iophagus collagenase differs from that of Clostridium histolyticum mainly in the sulfur-containing amino acids. 1 mol of zinc was found for 1 mol of enzyme of molecular weight 104 000. The autodegradation of the A. iophagus collagenase results in the formation of at least three active fractions which can be separated by preparative polyacrylamide gel electrophoresis as well as rechromatography on DE-32 cellulose. They are active towards the synthetic substrate as well as towards the native collagen. The results of ORD have shown that the digestion of the last one occurs in the helical parts of the substrate.


Inhibition of hydroxyproline synthesis by palladium ions.

Palladium ions, administered as PdSO4, markedly affect the incorporation of L-[3,4-3H2] proline into non-dialyzable fractions in 10-day chick embryo cartilage explants with a 55-65% reduction in the concentration range 0.06-0.6 mM. Under these conditions the synthesis of [3H]hydroxyproline was nearly completely inhibited. Experiments with prolyl hydroxylase (EC 1.14.11.2) indicated a strong irreversible inhibition of the enzyme with a competition between Fe2+ and Pd2+. The Ki for the inhibition was 0.02 mM. Pd2+-treated enzyme remained inactive after extensive dialysis. These studies suggest that Pd2+ may inhibit collagen synthesis by replacing Fe2+ in the active site of prolyl hydroxylase and forming strong complexes with the enzyme. These studies also point to a potential mechanism of Pd2+ toxicity.


[Research on subcellular localization of procollagen proline hydroxylase from chick embryo liver (author's transl)].

Subcellular fractionation by differential centrifugation was used to study protocollagen proline hydroxylase (EC 1.14.11.2) localization from chick embryo liver. The fractions have been characterized by marker enzymes and electron microscopy. By these methods, it was observed that procollagen-proline-hydroxylase is concentrated in the microsomal fraction which is sedimented at 145 000 X g in 250 mM sucrose.


The effect of several diphosphonates on acid phosphohydrolases and other lysosomal enzymes.

Diphosphonates are known to inhibit bone resorption in tissue culture and in experimental animals. This effect may be due to their ability to inhibit the dissolution of hydroxyapatite crystals, but other mechanisms may be important. Since lysosomal enzymes have implicated in the process of bone resorption, we have examined the effect of several phosphonates and of a polyphosphate (P20,2) on lysosomal hydrolases derived from rat liver and rat bone. Dichloromethylene diphosphonate strongly inhibited acid beta-glycerophosphatase (EC 3.1.3.2) and acid p-nitrophenyl phosphatase (EC 3.1.3.2) and to a lesser degree (in descending order) acid pyrophosphatase (EC 3.1.3.-), arylsulfatase A (EC 3.1.6.1), deoxyribonuclease II(EC 3.1.4.6) and phosphoprotein phosphatase (EC 3.1.3.16) of rat liver. Inhibition of acid p-nitrophenyl phosphatase and arylsulfatase A was competitive. Ethane-1-hydroxy-1, 1-diphosphonate did not inhibit any of these enzymes, except at high concentrations. Neither dichloromethylene diphosphonate nor ethane-1-hydroxy-1, 1-diphosphonate had any effect on beta-glucuronidase (EC 3.2.1.31), arylesterase (EC 3.1.1.2) and cathepsin D (EC 3.4.23.5). Of several other phosphonates tested only undec-10-ene-1-hydroxy-1, 1-diphosphonic acid inhibited acid p-nitrophenyl phosphatase strongly, the polyphosphate (P20, I) had little effect. Acid p-nitrophenyl phosphatase in rat calvaria extract behaved in the same way as the liver enzyme and was also strongly inhibited by dichloromethylene diphosphonate, but not by ethane-1-hydroxy-1, 1-diphosphonate. It is suggested that the inhibition of bone resorption by dichloromethylene diphosphonate might be due in part to a direct effect of this diphosphonate on lysosomal hydrolases.


Catalytic and regulatory properties of two forms of bovine heart cyclic nucleotide phosphodiesterase.

The soluble supernatant fraction of bovine heart homogenates may be fractionated on a DEAE cellulose column into two cyclic nucleotide phosphodiesterases (EC 3.1.4.-):PI and PII phosphodiesterases, in the order of emergence from the column. In the presence of free Ca2+, the PI enzyme may be activated several fold by the protein activator which was discovered by Cheung((1971) J. Biol. Chem. 246, 2859-2869). The PII enzyme is refractory to this activator, and is not inhibited by the Ca2+ chelating agent, ethylene glycol bis (beta-aminoethyl ether)-N, N'-tetraacetate (EGTA). The activated activity of PI phosphodiesterase may be further stimulated by imidazole or NH+4, and inhibited by high concentrations of Mg2+. These reagents have no significant effect on either the PII enzyme or the basal activity of PI phosphodiesterase. Although both forms of phosphodiesterase can hydrolyze either cyclic AMP or cyclic GMP, they exhibit different relative affinities towards these two cyclic nucleotides. The PI enzyme appears to have much higher affinities toward cyclic GMP than cyclic AMP. Km values for cyclic AMP and cyclic GMP are respectively 1.7 and 0.33 mM for the non-activated PI phosphodiesterase; and 0.2 and 0.007 mM for the activated enzyme. Each cyclic nucleotide acts as a competitive inhibitor for the other with Ki values similar to the respective Km values. In contrast with PI phosphodiesterase, PII phosphodiesterase exhibits similar affinity toward cyclic AMP and cyclic GMP. The apparent Km values of cyclic AMP and cyclic GMP for the PII enzyme are approx. 0.05 and 0.03 mM, respectively. The kinetic plot with respect to cyclic GMP shows positive cooperativity. Each cyclic nucleotide acts as a non-competitive inhibitor for the other nucleotide. These kinetic properties of PI and PII phosphodiesterase of bovine heart are very similar to those of rat liver cyclic GMP and high Km cyclic AMP phosphodiesterases, respectively (Russel, Terasaki and Appleman, (1973) J. Biol. Chem. 248, 1334).


Protein inhibitor of cyclic adenosine 3':5'-monophosphate phosphodiesterase in retina.

A protein acting as inhibitor of cyclic 3':5'-nucleotide phosphodiesterase (EC 3.1.4.1.) activity was found in the ox retina tissue. An inhibitor from one tissue (ox retina) effectively cross-inhibited a phosphodiesterase from another tissue (rat brain), indicating a lack of tissue specificity. Kinetic analysis showed that inhibition was independent of the time of preliminary incubation of the inhibitor with enzyme but dependent on its concentration in the reaction mixture. An inhibitor decreased the V of the enzyme and had no effect on its Km for cyclic adenosine-3':5'-monophosphate. The inhibitory effect was more pronounced with cyclic adenosine-3':5'-monophosphate than with cyclic guanosine-3':5'-monophosphate used as substrates of the reaction. The extractable form of the phosphodiesterase of the retina rod outer segments was much more sensitive to the inhibitory action than the membrane-bound one. The binding of labeled cyclic adenosine-3':5'-monophosphate to the inhibitory protein was shown not to occur. The inhibitor was sensitive to trypsin treatment, indicating that it was a proten attempt was mode to purify the inhibitory factor. Gel filtration indicated that the inhibitor had a molecular weight of 38 000.


Human blood platelet 3': 5'-cyclic nucleotide phosphodiesterase. Isolation of low-Km and high-Km phosphodiesterase.

Human blood platelet contained at least three kinetically distinct forms of 3': 5'-cyclic nucleotide phosphodiesterase (3': 5'-cyclic-AMP 5'-nucleotidohydrolase, EC 3.1.4.17) (F I, F II, and F III) which were clearly separated by DEAE-cellulose column chromatography. Although a few properties of the platelet phosphodiesterases such as their substrate affinities and DEAE-cellulose profile resembled somewhat those of the three 3': 5'-cyclic nucleotide phosphodiesterase in rat liver reported by Russell et al. [10], there were pronounced differences in some properties between the platelet and the liver enzymes: (1) the platelet enzymes hydrolyzed both cyclic nucleotides and lacked a highly specific cyclic guanosine 3': 5'-monophosphate (cyclic GMP) phosphodiesterase and (2) kinetic data of the platelet enzymes indicated that cyclic adenosine 3': 5'-monophosphate (cyclic AMP) and cyclic GMP interact with a single catalytic site on the enzyme. F I was a cyclic nucleotide phosphodiesterase with a high Km for cyclic AMP and a negatively cooperative low Km for cyclic GMP. F II hydrolyzed cyclic AMP and cyclic GMP about equally with a high Km for both substrates. F III was low Km phosphodiesterase which hydrolyzed cyclic AMP faster than cyclic GMP. Each cyclic nucleotide acted as a competitive inhibitor of the hydrolysis of the other nucleotide by these three fractions with Ki values similar to the Km values for each nucleotide suggesting that the hydrolysis of both cyclic AMP and cyclic GMP was catalyzed by a single catalytic site on the enzyme. However, cyclic GMP at low concentration (below 10 muM) was an activator of cyclic AMP hydrolysis by F I. Papaverine and EG 626 acted as competitive inhibitors of each fraction with virtually the same Ki value in both assays using either cyclic AMP or cyclic GMP as the substrate. The ratio of cyclic AMP hydrolysis to cyclic GMP hydrolysis by each fraction did not vary significantly after freezing/thawing or heat treatment. These facts also suggest that both nucleotides were hydrolyzed by the same catalytic site on the enzyme. The differences in apparent Ki values for inhibitors such as cyclic nucleotides, papaverine and EG 626 would indicate that three enzymes were different from each other. Centrifugation in a continuous sucrose gradient revealed sedimentation coefficients F I and II had 8.9 S and F III 4.6 S. The molecular weight of these forms, determined by gel filtration on a Sepharose 6B column, were approx. 240 000 (F I and II) and 180 000 (F III). F III was purified extensively (70-fold) from homogenate, with a recovery of approximately 7%.


Cytochrome c dependent, antimycin-A resistant respiration in mitochondria from potato tuber (Solanum tuberosum L.). Influence of wounding and storage time on outer membrane NADH-cytochrome-c-reductase.

Cytochrome c has two stimulatory effects on respiration of mitochondria especially those from wounded potato tuber. In the first place a stimulation of succinate- and NADH-consuming, antimycin-A-sensitive respiration, which reaches a maximal value at low cytochrome c concentrations, has been found. In the second place, at higher concentrations of cytochrome c a stimulation of NADH-consuming respiration occurs, which is antimycin-A-resistant, but KCN-sensitive. This antimycin-A-resistant, NADH-consuming respiration is absent, when no cytochrome c is added to the reaction medium. It is insensitive to metal chelators, to which the antimycin-A-and KCN-resistant plant mitochondrial alternative oxidase is sensitive. By measurements of NADH-cytochrome c reductase activities a corresponding antimycin-A-resistant NADH-cytochrome c reductase has been found, which is insensitive to osmotic shock treatment. A localization of this antimycin-A-resistant electron transport with NADH as the electron donor in the outer mitochondrial membrane is likely. In the mitochondrial preparations cytochrome c might stimulate by acting as an electron-carrier between the outer membrane reductase and the inner membrane cytochrome oxidase. A big increase of the outer membrane mediated electron transport in the mitochondria has been observed after wounding of potato tuber tissue. The ability of the tissue to produce this electron transport pathway after wounding disappeared after prolonged storage of the tubers. A possible function of this electron transport pathway in fatty acid desaturation during the wound-reaction is suggested.


Catalytic activity of cytochromes c and c1 in mitochondria and submitochondrial particles.

1. Beef heart mitochondria have a cytochrome c1:c:aa3 ratio of 0.65:1.0:1.0 as isolated; Keilin-Hartree submitochondrial particles ahve a ratio of 0.65:0.4:1.0. More than 50% of the submitochondrial particle membrane is in the 'inverted' configuration, shielding the catalytically active cytochrome c. The 'endogenous' cytochrome c of particles turns over at a maximal rate between 450 and 550 s-1 during the oxidation of succinate or ascorbate plus TMPD; the maximal turnover rate for cytochrome c in mitochondria is 300-400 s-1, at 28 degrees-30 degrees C, pH 7.4. 2. Ascorbate plus N,N,N',N'-tetramethyl-p-phenylene diamine added to antimycin-treated particles induces anomalous absorption increases between 555 and 565 nm during the aerobic steady state, which disappear upon anaerobiosis; succinate addition abolishes this cycle and permits the partial resolution of cytochrome c1 and cytochrome c steady states at 552.5-547 nm and 550-556.5 nm, respectively. 3. Cytochrome c1 is rather more reduced than cytochrome c during the oxidation of succinate and of ascorbate + N,N,N',N'-tetramethyl-p-phenylene diamine in both mitochondria and submitochondrial particles; a near equilibrium condition exists between cytochromes c1 and c in the aerobic steady state, with a rate constant for the c1 leads to c reduction step greater than 10(3) s-1. 4. The greater apparent response of the c/aa3 electron transfer step to salts, the hyperbolic inhibition of succinate oxidation by azide and cyanide, and the kinetic behaviour of the succinate-cytochrome c reductase system, are all explicable in terms of a near-equilibrium condition prevailing at the c1/c step. Endogenous cytochrome c of mitochondria and submitochondrial particles is apparently largely bound to cytochrome aa3 units in situ. Cytochrome c1 can either reduce the cytochrome c-cytochrome aa3 complex directly, or requires only a small extra amount of cytochrome c to carry the full electron transfer flux.


The reduced nicotinamide adenine dinucleotide "oxidase" of Acholeplasma laidlawii membranes.

An NADH dehydrogenase possessing a specific activity 3-5 times that of membrane-bound enzyme was obtained by extraction of Acholeplasma laidlawii membranes with 9.0% ethanol at 43 degrees C. This dehydrogenase contained only trace amounts of iron (suggesting an uncoupled respiration), a flavin ratio of 1:2 FAD to FMN and 30-40% lipid. Its resistance to sedimentation is probably due to the high flotation density of the lipids. It efficiently utilized ferricyanide, menadione and dichlorophenol indophenol as electron acceptors, but not O2, ubiquinone Q10 or cytochrome c. Lineweaver-Burk plots of the dehydrogenase were altered to linear functions upon extraction with 9.0% ethanol. A secondary site of ferricyanide reduction could not be explained by the presence of cytochromes, which these membranes lack. In comparison to other respiratory chain-linked NADH dehydrogenases in cytochrome-containing respiratory chains, this dehydrogenase was characterized by similar Km's with ferricyanide, dichlorophenol indophenol, menadione as electron acceptors, but considerably smaller V's with ferricyanide, dichlorophenol indophenol, menadione as electron acceptors, and smaller specific activities. It was not stimulated or reactivated by the addition of FAD, FMN, Mg2+, cysteine or membrane lipids, and was less sensitive to respiratory inhibitors than unextracted enzyme. The ineffectiveness of ADP stimulation on O2 uptake, the insensitivity to oligomycin and the very low iron content of A. laidlawii membranes were considered in relation to conservation of energy by these cells. Some kinetic properties of the dehydrogenation, the uniquely high glycolipid content and apparently uncoupled respiration at Site I were noteworthy characteristics of this NADH dehydrogenase from the truncated respiratory chain of A. laidlawii.


RNA polymerases from a rat hepatoma. Partial purification and comparison of properties with corresponding liver enzymes.

DNA-dependent RNA polymerases were extracted from the nuclei of poorly differentiated tumor, Morris hepatoma 3924A, and purified by an initial chromatography on a DEAE-Sephadex column followed by fractionation on phosphocellulose and finally on a second DEAE-Sephadex column. Three major forms of RNA polymerase (IA, IB and II) were resolved chromatographically. Enzymes IA, IB and II eluted from DEAE-Sephadex at 75, 150 and 210 mM (NH4)2SO4, respectively. The specific activities (nmol UMP incorporated mg protein per 15 min) of polymerases IA, IB and II were 40, 43 and 182, respectively. Concurrently, DNA-dependent RNA polymerases were extracted from normal liver and subjected to similar chromatographic procedure. Upon the final DEAE-Sephadex chromatography, enzymes IA, IB and II eluted at 110, 180 and 210 mM (NH4)2SO4, respectively. The recovery of polymerases IA, IB and II after purification was 0.21, 0,28 and 0.42 unit/mg DNA, respectively, for hepatoma enzymes and 0.07, 0.05 and 0.42 unit/mg DNA for the corresponding liver enzymes.


Biosynthesis of plasma membrane components by SV40-virus-transformed 3T3 mouse cells temperature sensitive for expression of some transformed cell properties.

We have studied the plasma membranes of an SV40-transformed 3T3 cell line temperature sensitive for the transformed growth phenotype (ts H6-15 cells), and have found that they vary little as a function of temperature of cultivation. Analysis by polyacrylamide gel electrophoresis was performed on plasma membranes prepared from ts H6-15 cells cultured at the permissive (32 degrees C) and non-permissive (39 degrees C) temperatures and radioactively-labelled in several ways. No significant differences were seen when the electrophoretic patterns of polypeptides of the plasma membranes of ts H6-15 cells, grown through 3-4 generations in medium containing radioactive leucine (32 degrees C and 39 degrees C temperatures) were compared. Plasma membranes derived from cells similarly grown in medium with radioactive glucosamine indicated that extensive alterations in the intrinsic glycopeptides occurred in association with alteration in growth phenotype. A shift towards decreased synthesis of large molecular weight (congruent to 100 000-160 000) glycopeptides occurred in cells grown at the temperature of non-transformed growth (39 degrees C). A decrease in amount of a 120 000 molecular weight glycopeptide at 39 degrees C was the most prominent of these alterations. We have studied the surface exposure of polypeptides and glycopeptides of intact cells grown at 32 and 39 degrees C, using lactoperoxidase-catalyzed iodination, NaBH4 reduction of galactose oxidase-treated cells, and metabolic-labelling with glucosamine of trypsin-sensitive molecules. We found no major qualitative differences between whole cell extracts or between plasma membrane preparations of cells cultivated at the permissive and non-permissive temperatures. Of special interest was the observation that the formation and surface exposure of a trypsin-sensitive, 240 000 molecular weight polypeptide appeared not to be ts in ts H6-15 cells. The significance of these observations will be discussed.


Liver plasma membranes from essential fatty acid-deficient rats. Isolation, fatty acid composition, and activities of 5'-nucleotidase, ATPase and adenylate cyclase.

Tod determine whether changes in unsaturation of fatty acids in rat liver plasma membranes might alter activities of membrane-associated enzymes, liver plasma membranes were prepared from rats fed purified diets lacking or supplemented with essential fatty acids. Two methods of membrane purification were used. A similar degree of purification was obtained with both methods for both depleted and control membranes, as indicated by marker enzyme purification. The proportion of essential fatty acids of the linoleate series was significantly lower in phospholipids from depleted rats. The specific activity of 5'-nucleotidase was lower, and the activity, V and apparent Km for total (Na+ +K+ +Mg2+)-ATPase were higher in the depleted liver plasma membranes. Arrhenius plots of total ATPase activity showed a discontinuity at the same temperature for both the depleted and control membranes. Activity with the depleted membranes was higher at all temperatures tested. Supplementation of deficient rats with a source of essential fatty acids (corn oil) restored V and apparent Km values to normal. Adenylate cyclase activity in the presence of fluoride, glucagon or glucagon plus GTP was significantly lower in the depleted plasma membranes.


Measurement of the translational mobility of concanavalin A in glycerol-saline solutions and on the cell surface by fluorescence recovery after photobleaching.

The fluorescence recovery kinetics of succinyl-fluorescein Concanavalin A (S-F-ConA) in glycerol-physiological saline solutions of high viscosity and when bound to the surface of mouse fibroblasts were measured following brief photobleaching using a laser excited fluorescence microscope. In the high viscosity solutions, the recovery kinetics, interpreted on the basis of a simple diffusion model, yielded a diffusion coefficient in close agreement with the values predicted by the Stokes-Einstein equation. Recovery kinetics for S-F-ConA bound to the surface of mouse 3T3 and SV3T3 cells cultured in vitro yielded diffusion coefficients in the range of 5-10-10(-11) cm2/s, values considerably lower than those reported previously for membrane proteins. These measurements indicated that a considerable fraction of the S-F-ConA molecules bound to the cell surface are immobilized. These results are discussed in relation to current concepts of lateral motion of protein components within natural membranes.


Localization in the synaptic junction of the cyclic AMP stimulated intrinsic protein kinase activity of synaptosomal plasma membranes.

Synaptosomal plasma membrane fragments contain a tightly bound protein kinase which can catalyse the phosphorylation of endogenous protein the reaction bein stimulated by cyclic AMP. A fraction enriched in synaptic junctions, which can be isolated from Triton X-100-treated synaptosomal plasma membranes, is also enriched in the cyclic AMP stimulated intrinsic protein kinase. The location of the enzyme in the synaptic junction suggests that cyclic AMP-stimulated phosphorylation may have some role in synaptic transmission.


Permeability properties of the membrane of vesicular stomatitis virions.

Observations of the light-scattering properties of several enveloped viruses indicate that virions (vesicular stomatitis, SV5 and influenza), in common with other membrane systems, are osmotically active, responding to NaCl gradients by swelling in hypo-osmolar solutions and shrinking in hyperosmolar solutions. The permeability barrier responsible for this osmotic response in vesicular stomatitis virions was modified both by protease treatment to remove the viral glycoprotein and by treatment with the polyene antibiotic filipin, an agent known to interact with cholesterol in liposomes and membranes. Filipin altered the kinetic and equilibrium permeability behavior of virions but the extent of leakage of osmotic shocking agent was less than that in lecithin/cholesterol and lecithin/ergosterol liposomes and in ergosterol-containing ciliary membranes. Negative-staining electron microscopy revealed that filipin treatment caused structural changes in the viral membrane. Intact virions exhibited appreciably larger responses to osmotic change than did protease-treated virus particles. Thus, the osmotic barrier in intact vesicular stomatitis virions may not be exclusively lipid in nature.


[Two-dimensional immunoelectrophoresis of spontaneous dissociation of low density lipoproteins].

Plasma-lipoproteins isolated between d 1.020 and d 1.055 g/ml were partially delipidated with ethyl ether at 4 degrees C. This treatment induces a transformation of the lipoproteins which is evolutive during several days. Bidimensional immunoelectrophoresis reveals the lipoproteins dissociation and the appearance of 4 immunologically different fractions. The time dependent formation of these subunits is slowed down by EDTA and less efficiently by antioxydants. Once started, the dissociation can be accelerated by heating at 37 degrees C or by UV exposition. Another lipopeptide is more easily revealed by anti VLDL antiserum. It can be shown in the native and in the partially or completely delipidated LDL. Its presence does not depend on lipoprotein dissociation.


[Phospholipid composition and turnover in normal and SV40 transformed fibroblasts. Effect of cell density].

The phospholipid composition and turnover in normal and in SV 40 transformed hamster fibroblasts were studied. The amount of phospholipid phosphorus relative to protein is lower in transformed hamster fibroblasts than in normal fibroblasts. This amount decreases with increasing cell density until stationary growth is reached. The decrease is largest for the normal fibroblasts. In transformed cells, less sphingomyelin and more diphosphatidyl glycerol are found than in normal cells. The turnover of 32P in sphingomyelin is slower in transformed cells than in normal cells ; the contrary is observed with diphosphatidyl glycerol. On the other hand, in transformed cells, phosphatidyl ethanolamine has a faster turnover than phosphatidyl choline, whereas the contrary is observed in normal cells. Finally, the change to stationary growth slows down the turnover of 32P of all phospholipids, this decrease being more important in transformed cells.


Effect of substrate - binding on the immunologic reactivity of lobster - muscle arginine kinase : a comparison with rabbit - muscle creatine kinase.

The effects of substrate-binding upon the immunologic reactivity of rabbit creatine kinase and lobster arginine kinase have been investigated. The separate binding of the guanidine or the nucleotide substrate to creatine kinase yields no alteration of antigenicity and a substantial effect is only observed when all the loci at the active center of the enzyme, including that for the transferable phosphoryl group, are occupied. In contrast, the antigenic reactivity of arginine kinase is affected by the separate binding of either the guanidine or the nucleotide substrate, and the simultaneous binding of the two substrates results in a cumulative effect, which is irrespective of the phosphorylated or non-phosphorylated form of the complex. These results support the existence of substrate-induced conformational changes demonstrated by other methods, and they reveal appreciable differences in their effect on the antigenic reactivity of the two enzymes.


[Isolation of peptide containing essential tyrosine from lobster arginine kinase].

The essential tyrosine residue of Lobster muscle arginine kinase, which is part of an antigenic determinant, has been modified by tetranitromethane. Cleavage of the S-carboxymethylated nitrated enzyme with cyanogen bromide gives rise to eight peptides, one of which containing the labelled essential tyrosyl group. Ion exchange chromatography on sulfoethyl-Sephadex C-25 in urea medium has been used with success for isolation and purification of the nitrated peptide. From its amino acid composition and end groups structure this peptide is the N-terminal fragment of the protein.


[Adenylate cyclase/Lipase. Hormone receptor induction].

An adenylate cyclase activity (AC) was found in guinea pig brown adipose tissue (BAT), since the tissue's apparition. This enzymatic activity increased during the development and showed high values at the end of gestation. An increase of AC units per cell was observed, in addition to the cell multiplication. A norepinephrine stimulation of AC activity was observed at the end of gestation : this regulating action disappeared in the first days of extrauterine life. Neither glucagon nor ACTH had any regulating role upon AC activity during fetal and newborn life. The basal lipolytic activity which was observed in BAT of fetuses (61rst day) and neonate dramatically around the 15th day. A potent lipolysis activation by norepinephrine was observed, but only after birth. The correlation observed between these enzymatic activities in presence of norepinephrine seems to indicate that the AC/lipase system was involved in the neonatal thermogenesis of guinea pigs.


Evidence for three enzymatic activities in one electrophoretic band of 3-phosphoglycerate mutase from red cells.

Electrophoresis of 3-phosphoglycerate mutase from erythrocytes of man and several animal species has been performed on cellulose acetate strips. In most cases the electrophoretic pattern of this enzymatic activity shows three bands. 2,3-diphosphoglycerate phosphatase and diphosphoglycerate mutase from erythrocytes of the same species have been revealed after migration during the same electrophoresis. We found that the band of 2,3-diphosphoglycerate phosphatase and the band of diphosphoglycerate mutase activities migrate at the same level as one of the bands corresponding to 3-phosphoglycerate mutase. Here, we discuss the possible existence of a single molecule carrying three enzymatic activities.


Involvement of macrophages in genetic resistance to bone marrow grafts. Studies with two specific antimacrophage agents, carrageenan and silica.

Carrageenans and silica, agents toxic for macrophages, were used in this study to examine the role of macrophages in resistance of irradiated mice to inbred parental and rat bone marrow grafts. Administration of 2.5 mg of carrageenans or 2.5-5 mg of silica particles intravenously to prospective graft recipients resulted in a prompt abrogation of hybrid and xenogeneic resistance. The macrophage stabilizer poly-2-vinylpyridine N-oxide (PVNO) injected subcutaneously in the dose of 150 mg/kg, 24 hr before silica prevented or reduced the suppression of resistance. PVNO, however, did not antagonize the suppression of resistance by carrageenen, horse anti-mouse thymocyte serum and cyclophosphamide. These results suggest that a) a subpopulation is involved in marrow graft rejection by irradiated mice; b) carrageenan and silica apparently act on macrophages by different mechanisms c) horse anti-mouse thymocyte serum and cyclophosphamide may act on cells other than macrophages or they act on macrophages by a different mechanism than silica, to resistance to bone marrow transplantation.


Effect of cyproheptadine on the spontaneous diurnal variations of plasma ACTH-cortisol and ACTH-GH secretion induced by l-dopa.

Cyproheptadine administration resulted in marked reduction and complete inhibition of the GH and ACTH-cortisol responses to L-dopa in a group of healthy subjects. If this drug acts by antagonizing serotonin, as assumed, this study suggests that serotonin is also involved in L-dopa induced GH and ACTH release.


Interferon induction by a tumoral DNA in an homologous culture system.

The interferon induction by Rous sarcoma DNA in an homologous culture system and a further insight of its kinetics and antimetabolites action were the principal aim of the present study. There was a direct relation between the dose of the inducer and the protection against the cytoplathogenic effect of the challenging virus (VSV) reaching the highest activity (68-75% CPE inhibition) with 100 mug RS-DNA. The kinetics of the induction revealed a peak inhibition by 18 hours after the inducer. Treatment with Actinomycin D evidenced that both interferon production and activity are modified. Its early addition resulted in a poor protection; but an accentuated interferon release was observed when antimetabolite was added 18 hours after the inducer. Similar results were obtained when its effect was studied on the activity of exogenous interferon. Interferon induction by Rous sarcoma DNA in an homologous system, allows the detection of a difference between tumoral and normal DNA at a biological level.


An inhibitor of hepatoma cells multiplication in the efferent fluid from isolated perfused rat liver.

Efferent fluid from isolated rat livers perfused with a synthetic medium, when concentrated, inhibits the multiplication of hepatoma LF 20 cells "in vitro". The inhibitory factor is thermolabile, and is inactivated at a slow rate by proteolytic enzymes. Sephadex and Acrylamide-agarose chromatographic fractionation leads us to assume a molecular weight around 100,000 for the inhibitory factor. The release of the factor by the perfused liver depends on the temperature of the perfusing fluid. Potential role of this factor in the control of liver cell proliferation is discussed.


Activity of human adenylate cyclase from human fat cell membranes.

Purified adipocytes plasma membranes have been prepared from human adipose tissue. The presence of an adenylate cyclase sensitive to epinephrine and fluoride has been demonstrated. Activation of the adenylate cyclase was usually 2 to 4 fold in the presence of epinephrine 5.10-5M and 8 to 10 fold in the presence of fluoride 10 mM. The adenylate cyclase from human adipose tissue was insensitive to glucagon and ACTH; these results are in support of previous studies of lipolysis in isolated fact cells or tissue fragments from human adipose tissue.


Characterization of Epstein-Barr virus (EBV) antigens II. Detection of early antigen (S) using anticomplement immunofluorescence (ACIF) et complement fixation (CF) tests.

Using human sera having antibodies against Epstein-Barr virus (EBV) early antigens (EA)  but lacking antibodies directed against the EB nuclear antigen (EBNA), the detection of EBV/EA was possible both by the anticomplement immunofluorescent test (ACIF) and by the classical complement fixation test (CF). The latter is particularly suitable for biochemical studies on EA.


Letter: In vitro induction of EBV early antigens in NPC epithelial tumour cells by IUDR or superinfection.

Induction of early antigens in the epithelial tumour cells of NPC was achieved both by IUDR treatment and EBV superinfection.


Magnetic resonance distinction between site bound and atmospherically bound paramagnetic counterions in polyelectrolyte solutions.

A combination of the water protons NMR chemical shifts, longitudinal and transversal relaxation rates and of the paramagnetic counterion EPR signal is shown to provide a clear distinction between site binding, atmospheric trapping and free counterions in solutions of polyelectrolyte TMA salts with increasing concentrations of the divalent counterions Co++ and Mn++. Site binding is defined by the loss of water in the counterion first hydration shell while atmospheric binding results in a change in the counterion correlation time as compared to a free ion.


[Morphofunctional state of the digestive tract in the early stages of atherosclerosis].

Cholesterol, alpha- and beta-lipoproteids, serotonin were determined in the blood serum of dogs which were on atherogenic diet for 2 months. The serotonin content was examined in various parts of the gastrointestinal tract (GIT). Parallel studies of the structural changes were carried out in the vascular system (VS) and various parts of the GIT. There was found a direct correlation between a rise in the cholesterol and serotonin level in the blood and serotonin in the GIT tissues. The initial stages of atherosclerotic changes were revealed in the vascular system. Along with compensatory-adaptive changes detected in the duodenum and the upper protions of the small intestine, initial stages of dystrophic-atrophic processes were observed in the lower portions. Comparative analysis of biochemical and morphological data indicated that disturbances of the morphofunctional state in the GIT played an important role in the genesis of the early stages of atherosclerosis.


[Comparative characteristics of the fatty acid composition of lipoproteins in human blood plasma and aortic wall].

A study was made of a fatty acid composition of individual lipid fractions in the lipoproteids of the blood serum of a very low, low and high density, and the wall of the aorta affected with atherosclerosis. There was a close similarity between a fatty acid composition of phospholipids, triglycerides, and cholesterol esters of all the lipoproteid classes of the vascular wall and blood plasma. The fatty acid compostion of individual fractions proved to be similar in all the lipoproteid fractions. The lipids of the vascular wall not included into the lipoproteid composition differed considerably by their fatty acid composition from the lipids isolated from lipoproteids; lipids of the wall of the aorta contained much less unsaturated fatty acids than lipoproteids.


[Effect on drugs changing the intracellular level of adenosine 3',5'-cyclic monophosphate on interferon formation in chick embryo cells of different ages].

The effect of theophylline and adrenaline on the synthesis of interferon induced by the influenza B virus, strain Lee, in a chick embryo tissue culture was studied. Both preparation were found to decrease interferon synthesis when 5-day-old cultures were used; the inhibitory effect was increased when the two drugs were used together. The degree of inhibition of interferon production depended on a dose of the preparation; the inhibition was still present even when the drugs ere introduced several hours after the cells were infected with interferonogen. The treatment of one-day-old cultures with theophylline resulted in increase of interferon synthesis, whereas administration of adrenaline alone or together with theophylline did not affect the level of interferon synthesis. The drugs used produced no effect on the reproduction of the test-virus of vesicular stomatitis, Newcastle disease and Chickungunya viruses in chick embryo cells and influenza B virus in the developing chick embryos. The results obtained are discussed from the point of view of a possible influence of the intracellular adenosine 3',5-cyclic monophosphate level on the synthesis of virus-induced interferon.


[Effect of carbidine on the process of adrenergic neurotransmission].

Methods of physiological analysis were applied to a study of the effect of an original psychotropic agent--carbidine--on the process of adrenergic neurotransmission on a model of an isolated rat Vas deferens. The noradrenaline (NA) content was determined in the same ductus spectrofluorimetrically; the capacity of the neuroleptic to block the consumption of the exogenous NA by the tissue was also investigated. Carbidine proved to possess an adrenomimetic effect caused by it capacity to release NA, leading to a reduction of endogenous neuromediator stores.


[Autoradiographic study of human scirrhous gastric carcinoma during cultivation in diffusion chambers].

The growth of the epithelial and connective tissue cells was noted in cultivation of human schirrous carcinoma of the stomach in diffuse chambers. In difference to the initial tissue of the tumour in vitro in which no incorporation of thymidine-H3 into the connective tissue cells was noted, these cells proved to be labeled under conditions of growth in the chambers. One hour after the administration of thymidine-H3 the percentage of cells with labeled nuclei averaged 25.1% this considerably exceeding the value of the label index determined under conditions of incubation in vitro of the initial tumour tissue (6.6%). Under conditions of cultivation there was revealed a rapidly proliferating subpopulation of cells with the mitotic cycle duration of 14.8 hours.


[Comparative study of the immune response of newborn and adult animals to the administration of tumor extracts].

Immunological reactivity to hepatoma 22a was studied in newborn and adult mice pretreated with tumour antigens. Treatment of newborn mice enhanced the growth rate of hepatoma 22a. Analogous treatment of adult mice resulted in the immunization effect. Evidently this difference was connected with a possible change of immunological reactivity to the tumour in newborn mice. This is supported by the results of cell dynamics--a mediated immunity assessed by the macrophage migration inhibition test. Splenic cells of the pretreated adult mice possessed the capacity to depress the macrophage migration more intensively in comparison with those of the pretreated newborn mice.


[Methods of obtaining antisera against antigens of the mammary cancer virus].

The authors used various methods of producing virus-containing preparations from mouse mammary carcinoma to obtain rabbit antisera against the mammary carcinoma antigens. The best results were achieved by means of the virus precipitation from the cultural fluids of carcinomas growing in vitro after preliminary hormonal stimulation of the virus yield followed by its purification in the sucrose density gradient.


[Cyclic adenosine-3',5'-monophosphate level in donor plasma].

The method of competitive protein binding was applied to the study of content of cyclic adenosine-3',5'-monophosphate (cAMP) in donor plasma one day before and immediately after the donation of 200 ml of blood. The work was performed by Gilman's radiochemical method. Two types of reaction of donors to donorship were revealed: without any changes and with the changes of the cAMP level. In accordance with these reactions the donors were divided into two subgroups--the stable and the reactive ones. In repeated donors the cAMP level was higher than in the primary ones, and the reactive ones. In repeated donors the cAMP level was higher than in the primary ones, and at the time of blood recovery it increased even more particularly among persons of "reactive" type. In primary donors of reactive type the cAMP content before the blood donation was either below or over the mean value and either increased or decreased to the mean level after the blood loss.


[Changes in the noradrenaline level and synthesis of nuclear RNA in the brain of rats with disturbances of the emotions and conditioned-defense behavior].

Brain noradrenaline (Na) exhaustion in reserpine-, disulfiram-or diethyldithio carbamate-treated rats was followed by a fall of the nRNA synthesis, emotional and conditioned behaviour disturbances. In accumulation of cerebral NA (ipraside and imipramine injections) there is an increase in the rate of synthesis, of the nuclear RNA. Prevention of the exhausting effect of reserpine by a preliminary injection of ipraside also prevented disturbances of the nuclear RNA synthesis.


[Metabolism of liver polyphosphoisositides in rats during stimulating and depressing influences on the organism].

Under normal conditions the rate of the 32P-orthophosphate incorporation into the polyphosphoinositides was 30--40 times greater than into the other phospholipid fraction of the normal rat liver. There was a rapid postmortem alteration of the polyphosphoinositide content in the liver. Under conditions studied the changes in the phospholipid content were revealed in the polyphosphoinositide fractions only. The changes in the content and in the metabolic intensity of rat polyphosphoinositide in the liver were in many respects similar to those in the brain. Polyphosphoinositide fractions were found to be the most labile of all the phospholipid fractions in the rat liver.


[Phosphoinositides of the brain and liver of rats in ethanol poisoning].

Alcohol intoxication induced in the rat brain significant decrease in the monophosphoinositides level, but the di- and triphosphoinositides quantity did not change; the content of all the three studied fractions of the inositol-containing phospholipids in the liver decreased. The tendency to retain the normal level of di- and triphosphoinositides in the brain following alcohol intoxication may be considered as an important factor providing physiological condition of the brain metabolism.


[Influence of nonakhlazin on adrenergic neurotransmission in the vas deferens of rats].

The effect of a new antianginal drug--nonachlazine on the adrenergic neurotransmission in the isolated rat vas deferens was studied by examining the vas deferens contractions in response to the transmural electric stimulation of the postganglionic sympathetic nerves and addition of noradrenaline (NA) or BaCl2. After the nonachlazine treatment the NA content in the vas deferens was also studied by the spectrofluorometric method. Besides, the effect of the drug on the uptake of the exogenous NA was investigated. Nonachlazine was found to possess some sympatholytic and spasmolytic effect and could block the uptake of the exogenous NA greatly.


In vitro colony-forming cells in the marrow of leukemic and preleukemic mice.

In vitro colony-forming cells (CFUc) were evaluated in preleukemic and leukemic AK mice. Increased concentrations of CFUc were found in normal appearing marrow of superinfected animals 2-3 wk prior to the onset of lymphoma. CFUc were present in marrows of mice with virus-accelerated, spontaneous, and transplanted lymphoma. CFUc concentration was often increased in mice with advanced disease and marrow replacement by lymphoblasts. When calculated on the basis of CFUc per normal marrow cells, this increase was marked. The colonies developed only in the presence of added colony-stimulating activity (CSA) and had the morphologic features of colonies from normal marrow. Leukemic cells did not form colonies. Lymphoma cells, from virus accelerated, spontaneous and transplanted lymphoma, did not produce CSA in feeder layers or conditioned medium. Leukemic and nonleukemic AK bone were found to produce similar small amounts of CSA. These studies showed that the preleukemic state as well as marrow replacement by lymphoblasts resulted in increased marrow CFUc's. No evidence for increased local production of CSA by lymphoblasts or the marrow microenvironment was found to account for this.


Abnormal parachloromercuriphenylsulfonate-sensitive cation channel in the erythrocytes of hereditary spherocytosis.

The erythrocytes of hereditary spherocytosis (HS) demonstrate an increased inward movement of sodium ions, an alteration which has been proposed as the primary defect leading to cell destruction. Parachloromercuriphenylsulfonate (PCMBS), an agent reacting with sulfhydryl groups of the membrane, increases the cation permeability of normal red cells, but does so to a much lesser extent in the HS red cells. On the other hand, pronase that is specific for amino groups of the membrane increases cation permeability and decreases anion permeability equally in normal and HS red cells. It may be postulated that a decreased number of sulfhydryl sites or a mutation of proteins in the PCMBS-sensitive cation channels of the HS cell membrane result in this hyposensitivity to PCMBS.


[Immunologic status in Hodgkin patients: correlation with Epstein-Barr virus titers].

1. In Hodgkin's disease patient's immunological in vitro and in vivo parameters are of prognostic importance. 2. Skin test reactivity correlates to peripheral T-lymphocyte counts and Con A induced lymphoblastogenesis. 3. Con A is the most sensitive in vitro indicator for detecting latent immunodeficiency. 4. Hodgkin patients in long term remission after tumor reductive therapy exhibit a qualitative and quantitative lymphocyte defect. 5. In Hodgkin patients Herpes virus related antibody titers are elevated against Epstein Barr virus (EBV). The elevation coincides with a decreased T-cell number and function. Antibodies against other Herpes viruses (HSV, CMV, VZV) are in the normal range, when tested by the complement fixation method.


[Biochemical study of the human palmar aponeurosis under normal and pathological conditions].

Study of oxyproline content of the palmar aponeurosis connective tissue. The knowledge of the quantity of the above mentioned aminoacid, which is specific of collagen, is considered very important to define the nature of tissue present in Dupuytren's disease and to understand the metabolic defect. After a short bibliographic analysis of the matter, methods used for the examination of total oxyproline content (Kivirikko, 1967) are discussed. The result shows a marked increase of oxyproline proportionally to the pathological aspect of aponeurotical tissue of the hand. Two hypotheses are formulated: a) an increase of the proline-hydroxilase activity. b) a slow breakdown of collagen fibers.


[Immunocytochemical study of corticotropic cells in the adenohypophysis of the chick embryo].

Corticotrophin cells of the pituitary have been studied immunocytologically with I.S. anti-A.C.T.H. (1-24) and I.S. anti-A.C.T.H. (17-39). These sera show differentiated cells as soon as the 8th of incubation. These cells remain located within the anterior part of the adenohypophysis. They are not "revelated" by I.S. anti-beta-M.S.H. and I.S. anti-beta-LPH.


[Mechanism of the appearance of nuclear inclusions with microfilaments and microtubules in sympathetic neurons. Influence of dibutyryl-adenosine 3'-5' monophosphate and theophylline].

As we have shown previously the nucleus of sympathetic neurons contains microtubules and microfilaments giving rise to highly organized inclusions. Present in vivo experiments on cat stellate ganglia have shown that dibutyryl cyclic AMP and theophyllin induce an increase of their frequency (up to 8.5 fold). These and earlier observations lead us to conclude that cyclic AMP mediates the process of nuclear microtubules and microfilaments assembly from preformed protein subunits.


The effect of premarin on blood loss attending open heart surgery.

Premarin was administered prophylactically to every other patient in a series of 148 consecutive cases of open intracardiac surgery. Of these, 132 patients were  matched to analyse the results of Premarin administration and postoperative blood loss. There were 67 patients who were given Premarin and 65 who did not receive this drug; both groups were identical in respect of age and sex. Matching was undertaken according to history of previous cardiac operations and anticoagulation, type of valve surgery or repair of congenital anomaly, duration of perfusion, platelet count and the use of fresh blood and epsilon-aminocaproic acid in the postoperative period. The prophylactic use of Premarin was not associated with a reduction in postoperative bleeding.


Malignant acanthosis nigricans and its unusual association with carcinoma of the colon and carcinoma of the cervix.

Malignant acanthosis nigricans is rarely associated with carcinoma of the bowel or cervix. However, this dermatosis is described in a case of adenocarcinoma of the ascending colon, and in another with carcinoma of the cervix.


The effects of long term feeding of Solanum glaucophyllum to growing rats on Ca, Mg, P and bone metabolism.

The effects of the long term ingestion of Solanum Glaucophyllum leaves (SG) by the rat was investigated in two series of experiments; the animals were fed a normal (for 8 weeks) or a low Ca diet (for 5 weeks). With both diets, the intestinal absorption of Ca was increased and the endogenous fecal Ca excretion was decreased by SG treatment. Added to a normal Ca diet, SG increased the urinary excretion of Ca, Mg and P and reduced the excretion of hydroxyproline and pyrophosphate. At the histological level, SG induced a higher rate of bone tissue synthesis on trabecular and endosteal surfaces. The bone content of hydroxyproline and citrate increased significantly. The total alkaline phosphatase activity of plasma decreased as a function of SG intake due to a decreased in the activity of the intestinal isoenzyme, which was not compensated by the increase in the bone isoenzyme activity. The Mg absorption was decreased by SG inducing lower Mg balances and lower plasma Mg levels. Added to a low Ca diet, SG increased the severity of the secondary hyperparathyroidism induced by the diet. The urinary excretion of hydroxyproline and the plasma alkaline phosphatase activity (both isoenzymes) were significantly increased. The Na and K content of bone decreased as a function of SG intake. 45Ca kinetic experiments revealed that SG increased the rate of Ca resorption and the rate constant of the fast exchangeable Ca pool, in both diets.


Osteoclast ultrastructure in Paget's disease.

The ultrastructural study of osteoclasts in biopsies from 12 patients with Paget's disease reveals several cytological anomalies. In Particular, nuclear inclusions, consisting essentially of striated filaments about 150 A in diameter, often organized in bundles and sometimes in paracrystalline arrays, appear to be specific to the disease. Morphologically, the inclusions are remarkably similar to those observed in various cases of viral attack, and the hypothesis of the action of a possible external agent in Paget's disease cannot be ruled out.


Early effects of synthetic bovine parathyroid hormone and synthetic salmon calcitonin on urinary excretion of cyclic AMP, phosphate and calcium in man.

Bovine synthetic parathyroid hormone infused intravenously in man increased both the urinary excretion of cyclic AMP and the urinary excretion of phosphate whereas a Salmon synthetic calcitonin infusion increased the urinary excretion of phosphate without change in urinary excretion of cyclic AMP. These data are consistent with the hypothesis that different renal mechanisms are involved in the response to each hormone.


Perturbation of lecithin bilayer structure by globoside.

The ultrastructure of aggregates formed by mixtures of pig erythrocyte lecithin, cholesterol and globoside in aqueous systems was studied by electron microscopy and X-ray diffraction. Globoside and lecithin in up to equimolar amounts formed a lamellar mesophase, although the structure of the lamellae was perturbed. Mixtures containing excess globoside formed complex tubular or reticular aggregates. Cholesterol appeared to promote mixing of lecithin and globoside. The flexibility gradient of the hydrocarbon (hc) region of the lipid bilayers was studied using electron spin resonance (esr) spectroscopy of various nitroxide-labelled stearic acid probes. Globoside in equimolar amounts greatly perturbed the order parameters of lecithin bilayers, reducing the fluidity of the hc region and flattening the flexibility gradient near the polar (p) surface. The effect of globoside on lecithin-cholesterol bilayers was not so pronounced, since the latter was already more ordered than lecithin bilayers. A phase transition of pure globoside at 55 degrees C, involving 'melting' of the hc chains was also detected using X-ray and esr spectroscopic techniques. The interbilayer spacing, dw, of equimolar lecithin-globoside lamellar phase increased by 42% from that of lecithin bilayers, indicating that the glycolipid p group may increase the net repulsive force between bilayers, as was previously predicted theoretically.


Low-affinity interaction of ethanol with the fluorescent complex between horse liver alcohol dehydrogenase and auramine O.

Quenching of the fluorescence of the complex between horse liver alcohol dehydrogenase (alcohol:NAD+ oxidoreductase (EC 1.1.1.1) and auramine O complex is inconsistent with a simple competitive displacement of auramine O by ethanol. Instead, the action of ethanol requires an explanation in terms of a solvent effect, or the formation of an enzyme-auramine O-ethanol ternary complex. The latter complex would have to be the low-affinity variety similar to the enzyme-NADH-ethanol ternary complex encountered in the kinetic system.


Effect of various sterols on the 22Na permeability and fluidity of phospholipid bilayer membranes.

Sodium-22 efflux was measured in multilamellar liposomes composed of egg lecithin, dicetylphosphate, and various sterols. In a parallel series of experiments a spin labelled fatty acid ester was incorporated into similar vesicles and the molecular motion of the spin label monitored by electron spin resonance spectroscopy. Spin lable mobility was used as a measure of phospholipid hydrocarbon chain motion. There was a poor correlation between the effects of these sterols on sodium permeability and their effects on the motion of the lipid chains. It is postulated that sterols alter sodium transport not only through a reduction in the motional freedom of membrane lipids, but also through changes in the partitioning of sodium between membrane and aqueous phases.


Inhibition of choline transport into human erythrocytes by choline mustard aziridinium ion.

Acetylcholine mustard aziridinium ion inhibited the transport of [3H]choline into human erythrocytes. Treatment of the erythrocytes with 1 X 10(-4) M tetraethylpyrophosphate prevented the inhibition of [3H]choline transport by acetylcholine mustard aziridinium ion. Hydrolyzed acetylcholine mustard aziridinium ion inhibited choline transport both in the presence and absence of 1 X 10(-4) M tetraethylpyrophosphate. The product of hydrolysis was equipotent with acetylcholine mustard in its ability to inhibit choline transport; incubation of this product with sodium thiosulfate prevented inhibition of choline transport thereby indicating the presence of an aziridinium ion. The hydrolysis product is likely to be choline mustard aziridinium ion. Results on the efflux of [3H]choline from erythrocytes in the presence of the proposed choline mustard aziridinium ion showed that the mustard moiety was transported into the red cells on the choline carrier. The rate of efflux of [3H]choline produced by choline mustard aziridinium ion was 55% of that produced by the same concentration of choline. It is concluded that acetylcholinesterase (EC 3.1.1.7) of red cells rapidly hydrolyzes acetylcholine mustard aziridinium ion to acetate and choline mustard aziridinium and the latter compound can act as a potent inhibitor of choline transport. This finding would indicate that the hemicholinium-like toxicity of acetylcholine mustard in the mouse is due to the formation of choline mustard aziridinium ion.


Monolayer cultures of dispersed cells from bovine anterior pituitary: responses to synthetic hypophysiotropic peptides.

Cells were dispersed from bovine anterior pituitary glands, by digestion with collagenase, and cultured. After 4 days the cell monolayers were incubated with fresh medium containing synthetic hypophysiotropic peptides for 2, 6, or 20 h, and hormone released into the medium was estimated by radioimmunoassay. After 2 h, thyroid releasing hormone (TRH) stimulated the release of thyroid-stimulating hormone (TSH) up to eightfold, and of prolactin (PRL) and follicle-stimulating hormone (FSH) about twofold at a minimal effective concentration of 1 ng/ml; enhanced growth hormone (GH) release was not apparent until 20 h, and release of luteinizing hormone (LH) and adrenocorticotrophic hormone (ACTH) was unaffected. Luteinizing hormone releasing hormone (LH-RH) enhanced release of LH maximally (three- to fourfold) during a 2 h incubation and was effective at 0.1 ng/ml; FSH release was significantly enhanced by about 50% above control level. Growth hormone release inhibiting hormone (GH-RIH)(somatostatin) showed significant effects only in the 20 h incubation; GH release was inhibited by 50% and release of PRL was slightly, but significantly, enhanced. Pituitary cell monolayers apparently permit maximal expression of releasing activities inherent in the hypothalamic hormones.


Effects of potassium chloride and smooth muscle relaxants on tension and cyclic nucleotide levels in rat myometrium.

Ten minutes after KCl-depolarization of rat myometrial strips, at which time the muscles were in a state of sustained contracture, tissue levels of adenosine 3',5'-cyclic monophosphate (cyclic AMP) were increased by approximately 40% over relaxed controls, and levels of guanosine 3',5'-cyclic monophosphate (cyclic GMP) were decreased by 40%. At this point both nitroglycerin (4 X 10(-4) M) and papaverine (2 X 10(-5) M) were capable of relaxing the depolarized muscles without significantly increasing cyclic AMP levels. Isoproterenol, in concentrations from 5 X 10(-9) M to 10(-6) M, relaxed the depolarized muscles and significantly increased tissue levels of cyclic AMP. However, the magnitudes of the cyclic AMP increases seen after the lower concentrations of isoproterenol were small relative to the increases observed during KCl-contracture alone. For example, the 40% elevation of cyclic AMP seen 10 min after KCl-depolarization did not cause the muscles to relax, whereas 5 X 10(-9) M isoproterenol caused relaxation with an increase in cyclic AMP levels of only 16% over depolarized controls. It was concluded that changes in total tissue levels of cyclic AMP were not responsible for the uterine relaxation caused by nitroglycerin, papaverine or isoproterenol in these experiments. Cyclic GMP levels in the depolarized muscles were not significantly changed by isoproterenol or papaverine but were increased approximately 80% by nitroglycerin. The above results are not consistent with the previously suggested roles for cyclic GMP and cyclic AMP as mediators of smooth muscle contraction and relaxation, respectively.


On the specificity of histamine H2-receptor antagonists in the rat cerebral cortex.

The effects of iontophoretically applied histamine H2-receptor antagonists and their antagonism of various amines, acetylcholine (ACh), and adenosine 5'-monophosphate (5'-AMP) were studied on spontaneously active rat cerebral cortical neurons. Metiamide selectively blocked the depressant actions of histamine. Burimamide, in amounts necessary for histamine antagonism, also antagonized the depressant effects of noradrenaline, dopamine, and 5-hydroxytryptamine. Neither antagonist affected 5'-AMP-induced depressions, but both reduced or blocked the excitatory actions of ACh. It is concluded that metiamide may be useful as a reliable antagonist of H2 receptors on cerebral cortical neurons.


Quantitative aspects of cyclic AMP and relaxation in the rabbit anterior mesenteric-portal vein.

Rabbit anterior mesenteric-portal vein was shown to exhibit a positive correlation between relaxation and cyclic AMP levels using different concentrations of isoproterenol for varying exposure times. No consistent relationship was found between cyclic AMP and relaxation using a series of phosphodiesterase inhibitors (papaverine, SC2964, or RA233). In sodium-free (Tris-substituted) Kreb's solution, the addition of isoproterenol still increased cyclic AMP levels, but there was no relaxation. Dibutyryl cyclic AMP was also ineffective in sodium-free solution. Under identical conditions, both relaxation and increases in cyclic AMP produced by papaverine and SC2964 were found to be independent of extracellular sodium. We conclude, therefore, that increased cyclic AMP may be involved in isoproterenol-induced relaxation possibly acting via increased Na+-Ca2+ exchange, but plays only a small role, if any, in smooth muscle relaxation produced by papaverine, RA233, or SC2964.


Comparative ultrastructure of selected oral streptococci: thin-sectioning and freeze-etching studies.

The ultrastructure of Streptococcus mutans, serotypes a-e, S. sanguis, S. mitis, and S. salivarius HHT, were examined by the techniques of thin-sectioning and freeze-etching. The cell walls varied in width between 15 and 46 nm and were covered with an electron-dense fibrillar or fuzz layer. The cytoplasmic membrane was in close association with numerous mesosomes which were, in turn, either closely associated or in contact with the bacterial chromosome. In freeze-etch replicas, the outermost layer of the cell wall was fibrous, and the cytoplasmic membrane was covered with particles composed of several subunits. Both particle-clusters and particle-free areas occurred on the surfaces of the cytoplasmic membrane, as well as a crystalline array in the ground plasm of the cell.


Anaplastic carcinoma of the esophagus. Report of three cases and their histogenetic consideration.

The autopsies of three cases of esophageal anaplastic carcinoma with an oat cell pattern are presented. Grossly, each case revealed a fungating growth and showed extensive metastases all over the body. The histology bears a striking resemblance to that of oat cell carcinoma of the lung, occasionally showing rosette formation, mucin secretion, and intracytoplasmic argyrophil granules in each case. These peculiar carcinomas most probably were derived from the esophageal submucosal glands. They were composed of both a cell group of argyrophil variety and a group of mucus-secreting variety which may have originated from the more primitive cells capable of differentiation into either variety.


Pulmonary tumorlet. A form of peripheral carcinoid.

Twenty cases of pulmonary tumorlet are presented. Approximately one-third of the tumors occurred in lungs severely scarred by bronchiectasis or other inflammatory processes; the other two-thirds were found in lungs with little to no scarring. In the former instances, tumorlets were found in large scars in which identifying architectural features had been destroyed. In the latter, the tumorlets were seen in minute fibrous nodules, surrounding or obliterating small bronchi or bronchioles. Argyrophilic granules were demonstrated in 14 of 15 cases; in the electron microscope these were consistent with neurosecretory granules. Their location, silver staining properties, and ultrastructure suggest that pulmonary tumorlets are minute peripheral carcinoid tumors.


Primary intracranial Burkitt's lymphoma in an infant.

A case of intracranial Burkitt's lymphoma is reported in a child whose symptoms began at 3 months of age with a definite histologic diagnosis established at 18 months. Serologic studies demonstrated high antibody titers to Epstein-Barr virus (EBV) in the patient and in four out of five members of the immediate family. The patient also demonstrated immunity to antigens derived from African Burkitt's lymphoma cell lines. The autopsy findings strongly support the case for the primary intracranial origin of the neoplasm and a perinatal infection with EBV is probable in this case.


Wilms' tumor in adolescence.

Two cases of Wilms' tumor in adolescent males are presented. The clinical and radiographic findings were unusual and both presented atypical gross and microscopic features that could be correlated with the radiographic findings. Histologic examination of both tumors showed evidence of tubular and glomerular maturation, a feature usually associated with Wilms' tumors of early infancy. One of the tumors contained a large amount of cartilage and bone. The other was grossly cystic and incorporated some features of multilocular cystadenoma, a benign metanephric tumor.


Glioblastoma multiforme with extraneural metastases in the absence of previous surgery.

A 63-year-old man was found to have an intracerebral glioblastoma multiforme and preoperative roentgenographic evidence of a mass in the middle lobe of the right lung. Because of the rarity of extraneural metastases from glioblastoma, especially in the absence of prior surgery, the lesions were considered to be separate neoplasms until death. The histologic appearance of the lung tumor obtained at autopsy was identical to the cerebral tumor. Additional metastases were found to bronchial lymph nodes and a lumbar vertebra. This case demonstrates that a glioblastoma can spontaneously metastasize extraneurally. Invasion of the glioblastoma into the lumen of a blood vessel was demonstrated within the primary tumor. Embolization of cells to the lung and beyond is the suspected mode of spread.


Inflammatory fibrous histiocytoma. An aggressive and lethal lesion.

During an investigation of soft tissue fibrohistiocytic tumors, seven cases were found which we believe represent a new and specific diagnostic subgroup of fibrous histiocytomas. All patients were adults (mean 52.6 years), only one of whom was younger than age 40. There were four females and three males; all were Caucasian. The tumors which occurred in the retroperitoneum, anterior chest wall, anterior abdominal wall, femoral area, and oral cavity, were large, averaging 8.5 cm., and although appearing encapsulated, were microscopically infiltrative. The common histologic feature of all cases was a diffuse and at times intense neutrophilic infiltrate unassociated with tissue necrosis, in combination with bland and anaplastic appearing histiocytes. The presence of this acute exudative reaction is a unique feature of these lesions, and was present not only in the original tumors, but in recurrences and metastatic foci as well. Other cell types including foam cells, lymphocytes, plasma cells, eosinophils, Reed-Sternberg-like and ganglion-like cells were commonly present. A storiform fibrous pattern, a hallmark of the fibrous histiocytomas was seen at some time in all but one case. The treacherous aspect of these lesions was their bland initial histologic appearance. The foci of foam cells associated with the inflammatory elements often led to a diagnosis of a benign inflammatory reaction or xanthogranuloma. However, follow-up has confirmed the aggressive and neoplastic nature of these lesions. The clinical course was usually protracted with multiple recurrences and eventual metastases. All patients died of their tumor, the average survival being 53 months; four patients survived over 5 years. At this time, we propose the term inflammatory fibrous histiocytoma to designate these lesions.


Morphologic study of virus-like particles in a case of acute leukemia.

Virus-like particles, grouped in clusters not bound by a membraine, were seen in electron micrographs of fresh leucoblasts of an acute leukemic patient. They consisted of large (100-nm diameter), round particles apparently composed of subunits. Tubular structures (30-50-nm diameter) were also seen in leucoblasts of the same patient. The two types of structures were never seen simultaneously in the same cell. The authors review the literature pertaining to the morphology of virus-like particles associated with malignant hematologic disease.


Müllerian adenosarcoma of the uterine cervix with heterologous elements: a light and electron microscopic study.

A müllerian adenosarcoma with heterologous elements having the gross appearance of a sarcoma botryoides occurred in the uterine cervix of a 14-year-old girl. Histologically, the tumor was composed of an admixture of benign-appearing glands and a sarcomatous stroma, the latter containing areas of undifferentiated sarcoma and endometrial stromal sarcoma, as well as heterologous elements consisting of rhabdomyosarcoma and hyaline cartilage. No malignant epithelial component was observed. The distribution of benign müllerian-type glands throughout the tumor simulated a malignant mixed müllerian tumor in appearance. Ultrastructural studies of the stromal elements of the neoplasm suggest origin from primitive müllerian stroma, and the glands have features of primitive müllerian epithelium. This study supports the concept that the malignant mixed müllerian tumor and related müllerian sarcomas originate from a common undifferentiated multipotential müllerian stem cell.


Metastatic cystosarcoma phyllodes: a light and electron microscopic study.

Two cases of metastatic cystosarcoma phyllodes, one in lung and the other in axillary lymph nodes, were studied with the light and the electron microscopes. The malignant element of these tumors appears to be a poorly differentiated mesenchymal cell. No evidence supporting the presence of epithelial cells was found. The tumor metastatic to the axilla exhibited intracellular virus-like particles similar to those described in adenocarcinoma of breast and other sarcomas.


So-called "minute pulmonary chemodectoma": a tumor not related to paragangliomas.

Twenty-six cases of so-called "minute pulmonary chemodectoma" are presented. The patient population showed a marked female preponderance, and there appeared to be an association of the lesion with pulmonary injury from a variety of causes including cardiac failure, chronic bronchitis and emphysema, and thromboemboli. Half the cases had multiple tumors. Microscopically, the tumors consisted of nests of cells in the interstitial tissue near small veins. Argentaffin and argyrophil stains failed to demonstrate cytoplasmic granules in any case. By electron microscopy, the nests were composed of large cells with broadly interdigitating processes connected by many well-formed desmosomes. The cytoplasm was filled with numerous 60-A filaments. The Golgi apparatus was prominent, while other organelles were sparse. No secretory granules were identified. It is concluded that the fine structure and lack of silver-positive granules are inconsistent with the morphology of previously reported paragangliomas, but that there is a resemblance at the light and electron microscopic level to meningeal arachnoed cells and the cells of meningiomas.


Multiple molecular forms of lactate dehydrogenase and glucose 6-phosphate dehydrogenase in normal and abnormal human breast tissues.

Normal and abnormal breast samples of women were analyzed for multiple molecular forms of lactate dehydrogenase and glucose 6-phosphate dehydrogenase, using acrylamide disc gel electrophoresis and specific enzyme staining techniques for separation and quantitation. Infiltrating ductal carcinomas demonstrated a significant increase (three to six-fold) in the proportion of LDH-5 compared to samples of normal breast, fibrocystic disease and fibroadenoma, indicative of a shift toward the muscle-type lactate dehydrogenase in neoplasia. For glucose 6-phosphate dehydrogenase, carcinomas were found to contain increased proportions of the fastest migrating species, G6PD-I. Total enzyme activity/mg DNA was elevated in meoplastic tissues. Little or no alteration in isoenzyme profiles could be related to menopausal status of the patient.


Small cell undifferentiated carcinoma of salivary gland origin: an ultrastructural study.

A case of small cell carcinoma of salivary gland was studied by light and electron microscopy. Light microscopy showed sheets of small uniform cells with scanty cytoplasm and pyknotic nuclei. Electron microscopy showed two distinct cell types; an electron-lucent epithelial-appearing cell, and a denser cell with processes containing bundles of filaments and other features suggesting myoepithelial differentiation. Neurosecretory granules were absent. These findings support a salivary duct origin, and are evidence against a neurendocrine derivation for this tumor.


Estrogen receptor protein in lesions of the male breast: a preliminary report.

Eleven specimens of breast lesions obtained from 10 male patients were analyzed for estrogen receptor protein (ERP). Three patients (ages 49, 77, 82 years) had infiltrating duct carcinomas with no axillary metastases. ERP  in each of these was positive. Eight specimens with gynecomastia, one of which was obtained from the 77-year-old patient with carcinoma in the same breast, were also analyzed. Of these ERP was positive in a 59-year-old man who had cirrhosis of the liver; two patients with borderline ERP had hepatitis and testicular seminoma, respectively. No relationship between histopathologic features of the lesions and ERP results was found and it is too early to relate these ERP studies to prognosis in these patients. Review of available literature, including our cases, reveals that six of eight male breast carcinomas were ERP-positive.


Mucinous carcinoma of the colon and rectum.

The clinicopathologic significance of mucus production by adenocarcinoma of the colon and rectum was analyzed in retrospective study with stage matched non-mucus producing control carcinomas. Mucinous carcinoma of the colon and rectum comprised 132 (15%) of 893 cases of colorectal carcinoma. The rectum was the most common site (33% of cases). While 120 mucinous cancers had a poorer five-year survival than non-mucinous tumors (34% vs. 53%, p less than .005), these had a particularly bad prognosis in the rectum (18% 5 year survival vs. 49% for the non-mucinous tumor controls, p less than .00k). The theoretical basis for this location-dependent behavior is considered. From this study, distinctive clinico-pathologic features emerge. There were seven documented cases of ulcerative colitis and 8 additional patients gave a history of "colitis". An additional five patients had received prior pelvic irradiation. Of particular note was the fact that 31% of mucinous carcinomas were associated with villous adenomas, implying a histogenetic relationship. Moreover, this finding again emphasizes the neoplastic potential of the villous adenoma, especially in the rectum where the development of mucinous carcinoma is particularly ominous.


Strumal carcinoid of the ovary.

A 45-year-old woman presented with an apparently nonfunctioning strumal carcinoid of the ovary. On ultrastructural examination the cells in the solid parts of the tumor as well as in the "follicular" areas contained dense-core secretory granules, indicating that this neoplasm is a pure carcinoid tumor. The thyroid-like appearance in areas is due to the accumulation of secretory material within unusually dilated acinar spaces. This study raises the question whether any strumal carcinoid contains thyroid tissue since the cells forming the follicle-like spaces in this tumor contain dense-core granules.


Localized submassive liver cell necrosis as a terminal event of liver carcinoma.

Six cases of hepatocellular carcinoma (HCC) and one case of metastatic liver carcinoma in which SGOT, SGPT, and SLDH were suddenly and markedly elevated immediately before death are described. All had a large blood loss and systemic hypotension in the preterminal period; autopsy disclosed irregularly shaped, patchy necrotic areas or infarcts, often clearly demarcated by hemorrhagic rims, in the noncancerous liver parenchyma. Tumor growths in the intrahepatic portal branches were extensive in all six cases with HCC; in the metastatic case, invasion and narrowing of the portal branches were extensive. The incidence of this terminal catastrophe was 3.3% (6 of 184 cases) for HCC and 1.15% (1 of 87) for metastatic carcinoma. The terminal liver necrosis was probably a result of sudden reduction in portal perfusion which had been inadequate because of tumor thrombosis, combined with hypotension of hepatic arteries.


Mediastinoscopy: a diagnostic aid in metastatic carcinoma of the breast.

Six patients with metastatic carcinoma of the breast underwent mediastinoscopy to obtain tissue for estrogen receptor analysis and pathologic confirmation of metastatic tumor. The indication for mediastioxcopy was an abnormal mediastinal accumulation of gallium in five patients, only two of whom had an abnormality noted on tomography. All six patients had tumor recovered by mediastinoscopy as demonstrated by pathologic examination and/or estrogen receptor assay, Therefore, the diagnosis of mediastinal metastases in breast cancer may be suggested by either chest roentgenograms, mediastinal tomography, or gallium scintigraphy. Mediastinoscopy is a safe, effective procedure capable of establishing the diagnosis of metastatic carcinoma of the breast and of procuring sufficient tissue for estrogen receptor analysis in patients without more readily accessible sites of metastases.


Peripheral pulmonary carcinoid tumors.

The light microscopic and ultrastructural features of five asymptomatic peripheral carcinoids presented as distinct pulmonary solitary nodules are described. By conventional microscopy the tumors displayed a variety of histologic patterns, the most unusual one showing tumor cells embedded in a richly vascular hyalinized stroma and forming papillary structures or cystic spaces lined by low cuboidal cells which ultrastructurally bore a strong resemblance to intermediate or transitional forms between types I and II pneumocytes. A striking feature of these tumors was their rich vasculature associated with a marked perivascular sclerosis composed of basement membrane-like material and collagen fibrils most likely produced by the increased numbers of pericytes surrounding these sclerotic vessels. The clinical implications, biologic behavior, and association of these tumors with other pulmonary neoplasms are also discussed.


Biology of the human malignant lymphomas. III. Intracranial heterotransplantation in the nude, athymic mouse.

Intracranial heterotransplantation in the nude, athymic mouse has been found to be an effective method for the experimental growth of human malignant lymphomas. Transplants of 11 primary lymphomas and six derived cell lines yielded a high take rate (90%) and a low mean latent period (36 days). Relatively small inocula produced extensive intracerebral infiltrates which could be identified as human in origin by immunofluorescence. Although confined to the central nervous system and meninges, the tumors were highly invasive and displayed morphologic features strikingly similar to those of the original primary tumors. Heterotransplantation of the lymphomas to extracranial sites was only rarely successful. Nude mice previously grafted with isologous neonatal thymuses failed to develop intracerebral tumors. Secondary cell cultures successfully established from several of the intracranial heterotransplants were found to be infected with NIH type-C xenotropic virus. The distinctive growth patterns and other neuropathologic features of the heterotransplants are described, and the relevance of these observations to the development of intracerebral lymphomas in immunosuppressed organ transplant recipients is noted. This method of studying human malignant lymphomas in vivo may permit better histopathologic characterization of the tumors, and may serve as a basis for further experimental lines of investigation, including viral, immunologic, and therapeutic studies.


Cellular whorls in brain tumors other than meningiomas.

Cellular whorls with or without secondary calcification are generally regarded as reliable diagnostic criteria in the differential diagnosis of meningiomas. They may however occasionally occur in other primary and metastatic brain tumors. Five cases (metastatic laryngeal carcinoma, metastatic mammary carcinoma, metastatic melanoma, medulloblastoma, and giant cell glioblastoma) are presented to illustrate this phenomenon occurring in non-meningothelial brain tumors.


Hepatocellular carcinoma in the Malaysian Orang Asli.

Necropsies were performed on 285 consecutively unclaimed Orang Asli bodies from Gombak Orang Asli Hospital during an eight-year period from May 1967 to April 1975. Of the 25 malignant neoplasms, hepatocellular carcinoma was by far the commonest (36%). The nine patients with this neoplasm had coexistant macronodular cirrhosis. There were 20 cases of cirrhosis; 45% of these had coexistant hepatocellular carcinoma. The 53,000 Orang Aslis living in West Malaysia comprise three tribes, the Negrito, Senoi, and Melayu Asli (Proto Malays). The Sinoi appear to have a high predilection for liver cancer, all our nine cases occurring in this group. These aboriginal people live in the jungles where they practice shifting cultivation and maintain their own dietary and social customs. Detailed studies of their dietary habits may provide a clue to the etiology of liver cancer in these people.


An ultrastructural and histochemical study of the mucous membrane adjacent to and remote from carcinoma of the colon.

The material was taken from surgical specimens of large intestine resected for carcinoma. A parallel study by electron microscopy and mucin histochemistry was made on fragments from "transitional" (TR) mucosa (adjacent to carcinoma) and "normal" (N) mucosa (remote from the tumor). These were compared with similar studies on the mucosa from control individuals. Histochemically, the "TR" mucosa shows an increase in sialomucins as compared with the "N" and control mucosae, where sulphomucins normally predominate. At the ultrastructural level, the "TR" mucosa is characterized by the following changes which may precede the histochemical variations: A) alteration in the relative proportions of the different cell types, along the crypt, with a persistence of immature and intermediate cells at higher levels of the crypt than in the control. Mature absorptive cells are fewer and bear an inverse relationship to intermediate cells. Goblet cells are increased in number and size. B) Appearance of electron-dense bodies 0.15-0.3mum in in diameter and membrane limited. C) An elaborated and enlarged Golgi zone showing increased secretory activity. It is suggested that mucin and ultrastructural changes described in the "TR" zone may indicate a failure in the normal process of cell differentiation along the crypt. Previous work7,8 further suggests that the mucin changes may be primary, reflecting a cellular response to unknown stimuli (i.e., carcinogens) rather than a local secondary effect of tumor growth. As for the ultrastructural features in the "TR" mucosa, the secondary effect cannot be excluded at the present.