Isolation of a somatomedin from plasma of rats bearing growth hormone secreting tumors. We have purified a protein which has somatomedin-like properties from the serum of Wistar-Furth rats bearing a growth hormone producing pituitary tumor (MStT/W15). Activity was measured by a placental insulin and/or somatomedin C radioreceptor assay (SmC-RRA). The serum was initially filtered through Sephadex G-150 equilibrated with 0.1 M NH4HCO3 and 0.02% NaN3. On the G-150 column, radioreceptor insulin (RRI) and radioreceptor somatomedin C (RRSm-C) activities coincided and appeared predominantly in the 160,000 mol wt range with a minor proportion in the 50,000 mol wt range. The pooled active fractions were boiled for 30 min at pH 5.5. After removing denatured protein by centrifugation, the extract was passed through G-50 Sephadex equilibrated with 1% formic acid and 0.15 M NaCl. Sixty to 90% of the SmC-RRA activity in the effluent appeared in the 9000 mol wt range. This material has an isoelectric focusing range of 8.4--9.6, similar to that described for human somatomedin C. On SDS-urea polyacrylamide gel electrophoresis only one protein band was seen. The isolated peptide (rSm) stimulated sulfate uptake in hypophysectomized rat cartilage. The potency of two preparations was variously assayed from 14.0 to 54.7 units/mg. Rat somatomedin was iodinated and purified by absorption on and elution from placental membranes. Eight to 12% of rat [125I]Sm was specifically bound by human placental membranes. Rat [125I]Sm was displaced by hSmC and rSm and human NSILA-S, partially displaced by procine proinsulin and poorly displaced by rat insulin. In preliminary studies, rat [125I]Sm was displaced from receptors on human placental membranes by sera from pituitary tumor bearing rats greater than normal rat sera greater than hypophysectomized rat sera.


Effect of hormones on collagen metabolism and collagenase activity in the pubic symphysis ligament of the guinea pig. The ligament which forms between the pubic bones of the pregnant guinea pig offers a unique system for the study of hormonal regulation of connective tissue metabolism. During the 5 days prior to parturition the length and hydroxyproline content of the pubic symphysis increase threefold. In nonpregnant animals, ligament growth can be induced in an estrogen-primed animal only following the administration of the hormone relaxin. A further increase in length can be achieved when progesterone is also injected. Removal and culture of post-partum ligaments, which undergo a 75% reduction in length and hydroxyproline content by the fifth post-partum day, allowed the demonstration of high collagenase levels released into the media. In contrast, when ligaments from pre-partum period were cultured, low levels of collagenase were detected in the media. The rapid post-partum ligament resorption could be partially inhibited if estrogen was injected immediately following parturition, whereas progesterone or relaxin significantly impaired ligament resorption. In a corresponding fashion, when the collagenase levels of these ligaments were measured progesterone treatment was shown to inhibit collagenase activity markedly while estrogen was less effective. Relaxin however, appeared to have no direct inhibitory effect.


Development of 1,25-dihydroxycholecalciferol receptor in the duodenal cytosol of chick embryo. The development of 1,25-(OH)2D3 receptor in the duodenal cytosol of chick embryo was studied by the sucrose density gradient analysis. The binding profile for 1,25-(OH)2D3 in the cytosol of vitamin D-deficient chick duodenum on the sucrose density gradient revealed 3 binding components, and the sedimentation constant was estimated as 2.5, 3.5 and 5.5S respectively. The 3.5S binding component has high affinity and low capacity for 1,25-(OH)2D3 and is thought to be 1,25-(OH)2D3 receptor. During the development of chick embryo, the 3.5S binding component was not detected in 13-day embryonic duodenum, it appeared on 15th day of incubation and then gradually increased to the level of vitamin D-deficient chick on 19th day of incubation. The 5.5S binding component was specific for 25-OH-D3 and it was found even in 13-day embryo, but it did not show any significant change during development. On the other hand, the 2.5S component was not specific for either 1,25-(OH)2D3 or 25-OH-D3. However, it was main binding component in early stages of development and decreased during development. From these results, it is suggested that the receptor for 1,25-(OH)2D3 is available a few days before hatching and the inability to produce CaBP in the duodenum of chick embryo could not be ascribed to the absence of the receptor.


Effects of mannoheptulose and DL-glyceraldehyde on glucose induced insulin release and adenosine 3',5'-monophosphate levels in isoalted islets of rat pancreas. The effects of mannoheptulose and DL-glyceraldehyde on glucose-induced insulin release and cycli AMP levels in islets isolated from rat pancreas were investigated. Mannoheptulose inhibition on glucose-induced insulin release was observed after only 5-min incubation period, indicating an inhibitory effect on the early phase of insulin release. This inhibition on insulin release was accompanied with the simultaneous depression of cyclic AMP levels in islets. By the addition of DL-glyceraldehyde to the medium in which glucose and mannoheptulose were present, the depressed cyclic AMP levels in islets were recovered to the control level completely but the restoration of insulin release in the early phase was not complete. In the absence of glucose, DL-glyceraldehyde did not demonstrate a significant increase of insulin release during 5 min incubation, though a marked stimulation was observed after 30-min incubation. Cyclic AMP levels in islets were not affected by DL-glyceraldehyde. When DL-glyceraldehyde was added to the medium with glucose, significant inhibition of glucos-induced insulin release in its early phase was observed without the reduction of cyclic AMP levels in islets. From these findings, the following possibilities are suggested and discussed. 1. Maintenance of the cyclic AMP levels in islets is a necessary but insufficient condition for glucose-induced insulin release particularly for its early phase. 2. Glucose-induced insulin release seems to depend on both the binding of glucose with glucoreceptor and the supply of some metabolites. Mannoheptulose inhibits both mechanisms. DL-glyceraldehyde may supply metabolites but competitively inhibit the binding of glucose to the glucoreceptor.


Endocrine function in a case of beta-adrenergic hyperdynamic circulatory state. Endocrine functions were investigated in a case of "beta-adrenergic hyperdynamic circulatory state". This state was diagnosed by (1) typical symptoms of cardiac awareness, (2) physical findings (increments of pulse rate and blood pressure by changing positions or walking), (3) increase in cardiac output (5.25 l/min leads to 14.03 l/min) and decrease in circulatory time (10.8 sec leads to 5.5 sec) by isoproterenol infusion (0.02 mug/min/kg body weight), (4) rapid loss of symptoms and above findings by propranolol treatment (30 mg per os daily) and reappearance by discontinuing medication. The mechanism of insulin response to glucose has been a controversy as to whether the secretion is transmitted by beta-receptor or independent glucose receptor. And in this physiologic beta-adrenergic state, it was found that insulin responses in IVGTT and OGTT were within normal limit. When beta-adrenergic condition was corrected by propranolol treatment, insulin responses were shown lowered, though in the normal range. This could be reproduced by discontinuing medication. Insulin, glucagon and growth hormone secretions caused by arginine were also found normal, but during the period the patient was on propranolol therapy, all responses were decreased, within the normal range. These results do not positively support the idea that glucose receptor is linked to beta-receptor. They do not either agree with the contention that secretions of insulin, glucagon and growth hormone induced by arginine are mediated through beta-receptors.


Lack of a direct effect of homocysteic acid on growth hormone and prolactin receptors. The effects of homoystine derivatives upon GH and prolactin receptors which are reported to stimulate growth in the rat were investigated. Several concentrations of homocysteic acid, homocysteine, homocysteinethiolactone, homoystine as well as other amino-acids were used in GH and prolactin receptor assay systems to determine their growth promoting or the lactogenic activities. None of the aminoacids used displaced labelled hGH or ovine PRL. It os concluded that derivatives of homoystine stimulate growth by an indirect mechanism and exhibit a lack of a direct stimulation of GH and prolactin receptors.


Effect of hypophysectomy on responsiveness of rat adrenal slices to ACTH--pregnenolone formation from endogenous cholesterol. Responsiveness of rat adrenal slices to ACTH was studied by measuring pregnenolone formation from endogenous cholesterol. The yield of pregnenolone by adrenal slices of hypophysectomized for 48 hrs rats was markedly higher as compared to intact animals.


7-O-(2-Amino-2-deoxy-alpha-D-glucopyranosyl)-L-glycero-D-manno-heptose. A constituent of the endotoxin of Bordetella pertussis. Hydrolysis of the Bordetella pertussis endotoxin, extracted from both "phase I" and "phase IV" bacteria, with 4 M HCl for 1 h at 100 degrees C, released the disaccharide named in the title; it was isolated by paper electrophoresis or by ion-exchange chromatography in about 1% yield (w/w). The structure of the heptose could be rigorously established by chemical degradation; the facts that the glucosaminidic linkage was hydrolysed by an enzyme preparation containing both, alpha and beta-N-acetylglucosaminidase activities, whereas it was resistant to cleavage by pure beta-N-acetylglucosaminidase strongly support the assumption that the disaccharide contains an alpha-D-glucosaminide linkage.


Characterization of the amino-terminal segment in type III procollagen. Native type III collagen and procollagen were prepared from fetal bovine skin. Examination of the cleavage products produced by digestion with tadpole collagenase demonstrated that the three palpha1(III) chains of type III procollagen were linked together by disulfide bonds occurring at both the amino-terminal and carboxy-terminal portions of the molecule. Type III collagen contained interchain disulfide bonds only in the carboxy-terminal region of the molecule. After digestion of procollagen with bacterial collagenase an amino-terminal, triple-stranded peptide fragment was isolated. The reduced and alkylated chain constituents of this fragment had molecular weights of about 21 000. After digestion of procollagen with cyanogen bromide a related triple-stranded fragment was isolated. The chains of the cyanogen bromide fragment had a molecular weight of about 27 000. When the collagenase-derived peptide was fully reduced and alkylated, it became susceptible to further digestion with bacterial collagenase. This treatment released a fragment of about 97 amino acid residues which contained 12 cystein residues and had an amino acid composition typical for globular proteins. A second, non-helical fragment of about 48 amino acid residues contained three cysteines. This latter fragment is formed from sequences that overlap the amino-terminal region in the collagen alpha1(III) chain by 20 amino acids and possesses an antigenic determinant specific for the alpha1(III) chain. The collagenase-sensitive region exposed by reduction comprised about 33 amino acid residues. It was recovered as a mixture of small peptides. These results indicate that the amino-terminal region of type III procollagen has the same type of structure as the homologous region of type I procollagen. It consists of a globular, a collagen-like and a non-helical domain. Interchain disulfide bonding and the occurrence of cysteines in the non-helical domain are, however, unique for type III procollagen.


Interaction of the apoproteins of very low density and high density lipoproteins with synthetic phospholipids. The interaction of synthetic dimyristoyl phosphatidylcholine (lecithin) liposomes with isolated apoC-I and apoC-III proteins from very low density lipoproteins has been studied by microcalorimetry. Complex formation is a highly exothermal process characterized by a maximal enthalpy of -130 kcal/mol (-544 kJ) apoC-III-1 and -65 kcal/mol apoC-I proteins (-272 kJ). The complex composition determined after its isolation by ultracentrifugal flotation agrees with the value derived from the enthalpy binding curves. The binding of a constant amount of dimyristoyl lecithin to apoprotein mixtures containing various proportions of apoA-I and apoC-III failed to demonstrate the existence of any preferential association between the two apoproteins, in contrast with results obtained previously with apoA-I/apoA-II protein mixtures. Finally the various contributions to the enthalpy of binding such as that arising from an increase in apoprotein helicity have been evaluated. A classification of the apolipoproteins according to their lipid-binding affinity is proposed as: apoA-II congruent to apoC-III greater than apoC-I greater than apoA-I proteins.


Uridine-diphosphate-glucose 4-epimerase from Saccharomyces fragilis. Inactivation by heat and reconstitution of the inactive enzyme. UDP-glucose 4-epimerase from Saccharomyces fragilis is rapidly inactivated by heating at 42 degrees C for 7 min and at 45 degrees C for 4 min. The effector site, specific for sugar phosphates, is destroyed still earlier. The enzyme is inactivated by the dissocation of NAD from it leaving the dimeric structure unaffected. It can be reactivated by mercaptoethanol and NAD, both of which are essential for reactivation, and NAD becomes associated with the dimeric protein moiety.


Protein degradation during yeast sporulation. Enzyme and cytochrome patterns. The levels of several enzymes have been studied during sporulation of Saccharomyces cerevisia. The specific activities of ribonuclease and aminopeptidase I raised several-fold after transfer of the cells to sporulation medium, whereas the specific activities of phosphofructokinase, glucose-6-phosphate dehydrogenase, tryptophan synthase and pyruvate decarboxylase were not significantly altered. The specific activities of NAD-dependent glutamate dehydrogenase, isocitrate lyase, malate dehydrogenase and fructose bisphosphatase all decreased from the onset of sporulation. The inactivation of these latter enzymes was inhibited by cycloheximide and by inhibitors of energy metabolism. Hexokinase, alcohol dehydrogenase and glutamate oxaloacetate transaminase were partially lost from the cells during the period of ascus maturation. None of the enzyme changes observed proved to be 'sporulation-specific' in that it occurred exclusively in sporulating diploid yeast cells. Therefore it is postulated that the meiotic events and the metabolic changes required for ascospore formation are under separate genetic control in this organism. During sporulation, the cellular content of cytochromes b, c, and aa3 was reduced to 20% or less of that present in vegetative derepressed cells. Since the relative percentage of total to cycloheximide-insensitive mitochondrial protein synthesis was not significantly altered throughout sporulation, and the pattern of mitochondrially synthesized polypeptides was rather similar both in vegetative and in sporulating cells, it appeared that not only degradation but also synthesis and therefore turnover of the mitochondrially coded polypeptides of cytochromes b and aa3 took place during sporulation. The activity ratio of cytochrome c oxidase to F1-ATPase in submitochondrial particles isolated from vegetative cells and from purified asci was almost identical. This indicates that the loss of membrane-bound mitochondrial cytochromes during sporulation is probably due to a nonselective degradation of inner mitochondrial membrane proteins.


The cytochrome bc) complex of yeast mitochondria. Site of translation of the polypeptides in vivo. 1. Yeast cells were labelled with radioactive amino acids in the presence of cycloheximide and the cytochrome bc1 complex was isolated from them as described in the preceding paper (Katan, M.B.., Pool, L. & Groot, G.S.P. (1976)Eur. J. Biochem, 65, 95-105). After analysis of this preparation by sodium dodecylsulphate polyacrylamide gel electrophoresis only one band, with an apparent Mr of 32000, was found to have incorporated radioactivity. The amount of label in the band was low, but could be increased approximately 5-fold by preincubating the cells in erythromycin before the labelling period. 2. Cells were labelled in the presence of chloramphenicol and the cytochrome bc1 complex was isolated by (NH4)2SO4 fractionation. Upon electrophoresis in the presence of sodium dodecylsulphate only four of the six bands that belong to the complex were found to have incorporated radioactivity; no radioactivity was found in the bands with an Mr of 40000 and 17000. The same result was obtained after labelling in the presence of acriflavin. If, however, the cytochrome bc1 complex was isolated by immunoprecipitation, all bands were found to have incorporated radioactivity in the presence of chloramphenicol. The amount of radioactivity in the Mr 32000 band was now clearly depressed. 3. It is concluded that of the seven polypeptides of the cytochrome bc1 complex of yeast only one is made on mitochondrial ribosomes. This polypeptide has an Mr of 32000 and is probably associated with cytochrome b.


Separation and characterization of two phosphorylase phosphatase inhibitors from rabbit skeletal muscle. Two heat-stable and trypsin-labile inhibitors of phosphorylase phosphatase, designated inhibitor-1 and inhibitor-2, were partially purified from extracts of rabbit skeletal muscle by heating and coloumn chromatography using DEAE-dellulose and Bio-gel P-60. Inhibitor-1 exists in an active phosphorylated form and an inactive dephosphorylated form. The interconversion of phosphorylated inhibitor-1 and dephosphorylated inhibitor-1 is mediated by protein kinase dependent on adenosine 3':5'-monophosphate (cyclic AMP) and a Mn2+-stimulated phosphoprotein phosphatase. Inhibitory activity of inhibitor-2 is not influenced by treatment with either the kinase or the Mn2+-stimulated phosphatase. The molecular weights of inhibitor-1 and inhibitor-2 estimated by sodium dodecylsulfate-polyacrylamide gel electrophoresis are 26000 and 33000 respectively. Both inhibitor-1 and inhibitor-2 inhibit phosphorylase phosphatase by a mechanism which appears to be non-competitive with respect to the substrate phosphorylase a. Inhibitor fractions at early stages of purification also inhibit cyclic-AMP-dependent histone phosphorylation, but this kinase inhibitory activity resides with a protein moiety which is separable from inhibitor-1 and inhibitor-2.


Metabolic regulation of steroidogenesis in isolated adrenal cells of rat. Relationship of adrenocorticotropin-, adenosine 3':5'-monophosphate-and guanosine 3':5'-monophosphate-stimulated steroidogenesis with the activation of protein kinase. The data presented with the isolated adrenal cells, in the present study, show that adrenocorticotropin in the physiological concentration range stimulates the synthesis of guanosine 3':5'-monophosphate(cyclic GMP), protein kinase activity, and steroidogenesis in a concentration-dependent manner without detectable rise in the levels of adenosine 3':5'-monophosphate (cyclic AMP). Millimolar concentrations of cyclic AMP and cyclic GMP, which stimulate corticosterone synthesis, also activate kinase activity and steroidogenesis in a sigmoid concentration-response manner. The process of phosphorylation activated by corticotropin, cyclic AMP and cyclic GMP is not inhibited by cycloheximide or actinomyin D. It is therefore proposed that the hormonal responses mediated by cyclic GMP and cyclic AMP are via the protein kinase enzymatic steps, and the inhibitory effect of cycloheximide and actinomycin D in corticotropin-stimulated steroidogenesis follows this step. In conjuction with our previous observations that the biosynthetic steps from (20S)-20-hydroxycholesterol to corticosterone are neither inhibited by cycloheximide nor affected by cyclic GMP, it is inferred that the rate-limiting step of adrenal steroidogenesis is the transformation of cholesterol to (20S)-20hydroxycholesterol and this very step is regulated by cyclic GMP and cyclic AMP. Of further significance are the findings that micromolar cincentrations of cyclic AMP and cyclic GMP, which do not stimulate steroidogenesis, effectively stimulate protein kinase activity in a concentration-dependent manner. It is therefore concluded that all cyclic-nucleotide-dependent protein kinase activities of the cell are not necessarily related to steroidogenesis.


Purification of poly(ADP-ribose) polymerase from Ehrlich ascites tumor cells by chromatography on DNA-agarose. Poly(ADP-ribose) polymerase with a high specific activity was obtained from Ehrlich ascites tumor cells by extraction of nuclei with 175 mM potassium phosphate, followed by chromatography on DNA-agarose. Electrophoretic analysis indicated that the preparation contained two proteins, one of which was shown to catalyze the synthesis of poly(ADP-ribose). As expected from results obtained by other workers, the synthesis was inhibited by nicotinamide and thymidine, and stimulated by DNA. Addition of histones gave inhibition of the synthesis, unless DNA was present in the reaction mixture.


Subunit structure of simian-virus-40 minichromosome. Electron microscopic evidence indicates that Simian virus 40 (SV40) minichromosomes extracted from infected cells consist of 20 +/- 2 nucleosomes, each containing 190 -- 200 base pairs of DNA. About 50% of the nucleosomes are not close together, but connected by segments of DNA of irregular lengths which correspond to about 15% of the viral genome, irrespective of the ionic strength. Micrococcal nuclease digestion studies show that there is about 200 base pairs of DNA in the biochemical unit of SV40 chromatin. Therefore, the visible internucleosomal DNA of the SV40 minichromosome does not arise from an unfolding of a fraction of the 190 - 200 base pairs of DNA initially wound in the nucleosome. These results support the chromatin model which proposes that the same DNA length is contained in the nucleosome and the biochemical unit. Results from extensive micrococcal nuclease digestion suggest that an SV40 nucleosome consists of a 'core' containing a DNA segment of about 135 base pairs associated to a DNA fragment more susceptible to nuclease attack. The addition of histone H1 results in a striking condensation of the SV40 minichromosome, which supports the assumption that histone H1 is involved in the folding of chromatin fibers.


A new plant-type ferredoxin from halobacteria. A stable, 2Fe-type ferredoxin has been prepared from Halobacterium halobium and purified by chromatography. A similar ferredoxin was also found in three other Halobacteria. The ferredoxin is present in large amounts-about 1 percent of the total soluble protein. From amino acid composition a molecular weight of 14800 +/- 200 was calculated. The ferredoxin was found to contain two atoms each of iron and sulphide. The midpoint redox potential of the protein is about -345 mV. The electron paramagnetic resonance spectrum of the reduced form shows much similarity to plant and algal ferredoxins with gx = 1.90, gy = 1.97 and gz = 2.07. The same similarity is observed in the optical absorption, optical rotatory dispersion and circular dichroism spectra. However it does not seem to mediate electron transport in the NADP-photoreduction system of chloroplasts. Extracts of the bacterial cells catalyze the reduction of the ferredoxin by NADH.


A spin-label study of the chromaffin granule membrane. The structure of the chromaffin granule membrane has been probed using a number of different spin labels. Both the effect of temperature and high levels of calcium have been studied. 1. The results from three positional isomers of the stearic acid spin label demonstrate that a substantial part of the membrane lipid (that is sensed by the probe) is in a bilayer structure which undergoes a structural transition at 32-36 degrees C, characterized by an increase in the population of gauche isomers in the lipid chains. A possible mechanism for this transition would be the preferential segregation of cholesterol. 2. The covalently bound iodoacetamide spin label reveals a transition within the protein component of the membrane or its immediate lipid environment at 32 degrees C. This transition corresponds to an increased degree of motional freedom of the spin label above the transition temperature. 3. The lipid-soluble spin label 2,2,6,6-tetramethyl-piperidine-1-oxyl exhibits a break at 34 degrees C in the temperature-dependence of its partitioning into the membrane. This could correspond to the onset of a lateral separation in the membrane lipid, again possible involving a re-distribution of cholesterol. 4. Calcium abolishes, diminishes or shifts the transition observed by the spin label and decreases the amplitude of motion of the stearic acid spin labels, again possibly involving a redistribution of cholesterol and also lysolecithin. The temperatures of the structural transition agree well with the changes in the enzymic activity of the membrane ATPase and NADH oxidase functions and also with the results from fluorescent probes [Bashford et al., Eur. J. Biochem. 67, 105-114(1976)]. It is possible that triggering of the transition either by calcium or some other stimulus may play a role in catecholamine release and membrane fusion.


Isolation and characterization of hydrophobic proteins (H proteins) in the membrane fraction of Bacillus subtilis. Involvement in membrane biosynthesis and the formation of biochemically active membrane vesicles by combining H proteins with lipid. Cytoplasmic membranes of Bacillus subtilis, grown in complex medium containing glucose, were fractionated into three membrane subfractions [light band (1.155 - 1.158 g/cm3); medium band (1.181 - 1.183 g/cm3); heavy band (1.21 - 1.25 g/cm3)] by sucrose density gradient centrifugation. Among these subfractions, the light and medium bands consisted mainly of membranes but the heavy band consisted of an irregular arrangement or aggregate of small globular protein components of 5 - 8 nm in diameter. We named this H-protein. H-protein formed trilamellar unit membrane structure when combined with lipid. In pulse-labeling and pulse-chase experiments with radioactive leucine, it was found that H-protein consisted of the newest membrane protein synthesized in the cells and the label incorporated into H-protein was shifted into light and medium band of the membranes during the chase. Cytochromes were not found in H-protein. However, when H-protein was incubated with haem alpha and protohaem, these compounds were incorporated into the apoproteins of the cytochromes present in H-protein and form cytochromes a and b. Cytochromes were also formed in H-protein which were isolated from the cells grown in the presence of haemin (haemin-grown H protein). Succinate dehydrogenase activity was increased about 4-fold by combining H-protein or haemin-grown H protein with lipid. H-protein had no cytochrome oxidase activity; however, haemin-grown H protein was found to have some of the activity and this was increased about 4-fold by combining the protein with lipid. Haemin-grown H protein was also found to form succinate: cytochrome c oxidoreductase when combined with lipid and vitamin K2. On the other hand, succinate oxidase was required for the addition of lipid, vitamin K2 and cytochrome c. NADH oxidase was also found in haemin-grown H protein and was activated about 9-fold in constituted reaction systems. Vesicles formed by haemin-grown H protein and lipid, could accumulate alanine and proline by addition of NADH or reduced phenazine methosulfate. Alanine and proline was also accumulated into the vesicles when transport energy was supplied as a membrane potential introduced by K+-diffusion via valinomycin. These results would indicate that H-protein contains the apoprotein of cytochromes, and a carrier involved in the active transport of alanine and proline.


Difficulties in determining accurate molecular motion parameters from proton relaxation enhancement measurements as illustrated by the immunoglobulin G-Gd(III) system. Longitudinal and traverse proton magnetic relaxation rates for water in the hydration sphere of Gd(III) bound to non-immune rabbit IgG (immunoglobulin G) have been determined over a wide range of frequencies (4-84 MHz) at constant temperature (19 degrees C) using pulsed nuclear magnetic resonance spectrometry. The rates have also been determined at temperatures between 0 and 40 degrees C for two frequencies (61 and 84 MHz). The rates were fitted to existing theory using a computer least-squares procedure. Further computer analysis was then carried out to determine the sensitivity of the best-fit error to variation in the variable parameters in the theoretical expressions used. These include the water co-ordination number (q) for which it was found large variations could occur (between approximately 2 and 8) for only small changes in the error of best-fit. It was concluded that a slow exchange contribution to the relaxation rates was important in deciding which parameters are poorly determined. A rotational correlation time (tau r) was obtained which suggested there might be considerable internal motion of the Fc region (C-terminal half of heavy-chain dimer) of the IgG molecule. However the possibility of large errors in this value prevented unequivocal conclusions being drawn.


Kinetic studies on a 4-methoxybenzoate O-demethylase from Pseudomonas putida. A direct, sensitive and reliable photometric assay procedure for monitoring the activity of non-specific 4-methoxybenzoate O-demethylases of microorganisms is described. The assay is based on the O-demethylation of 3-nitro-4-methoxybenzoate to the yellow-coloured product 3-nitro-4-hydroxybenzoate. Using this assay and by monitoring the oxidation rate of reduced pyridine nucleotides, the kinetic properties of a purified, reconstituted enzyme system composed of 4-methoxybenzoate monooxygenase (O-demethylating) and a reductase from Pseudomonas putida have been investigated. It has been found that the KM value of the monoxygenase of this enzyme system towards different substrates (i.e. tight couplers, uncouplers and partial uncouplers) rises from the extremely low value of 0.07 muM for the tight couplers to about 55 muM for the uncouplers. The effect of possible inhibitors and metal ions on the reconstituted enzyme system was investigated. The inhibition pattern was almost identical to that found for the purified reductase, only batho-phenanthrolinedisulfonate showing a greater inhibition of the reconstituted enzyme system. The affinity of the reductase towards NADH was found to be approximately 200-fold greater than that towards NADPH. Futhermore, the affinity of this reductase to NADH depended on the nature of the electron acceptor. The affinity to NADH was more than 10 times higher when the monooxygenase-substrate complex was used as the electron acceptor, than when cytochrome c or 2,6-dichloroindophenol was used. These differences are discussed on the basis of enzyme-enzyme interactions between the reductase and the monooxygenase.


Glucose effect in tgl mutant of Escherichia col K12 defective in methyl-alpha-D-glucoside transport. 1. The dependence of the rate of accumulation of methyl-alpha-D-glucoside on its extracellular concentration was studied in the tgl mutant of Escherichia coli K12, isolated earlier. It has been shown that the kinetics of methyl-alpha-D-glucoside transport differ sharply from those in wild-type bacteria. 2. The beta-galactosidase synthesis in tgl strain is much less sensitive both to permanent and transient glucose catabolite repression. The level of cyclic AMP in mutant cells under the conditions of glucose catabolite repression is several times higher than in the parent strain. 3. The tgl mutation does not affect the manifestation of catabolite inhibition and inducer exclusion with glucose. 4. The data obtained are discussed in the light of a hypothesis concerning the existence of two sites, binding and pecific enzyme II of the phosphoenolpyruvate-dependent phosphotransferase system. The tgl mutation alters the first site, and the second one is damaged by the pgt mutation. 5. It is suggested that the products of the tgl and gpt genes are necessary for the manifestation of the phenomena of glucose permanent and transient repression. The effects of catabolite inhibition and inducer exclusion are realized irrespective of the existence or absence of the tgl product.


Binding studies of a spin-labelled oxidized coenzyme to bovine-liver glutamate dehydrogenase. NAD+ with a nitroxide piperidine ring linked to the NH2 group of the adenine possesses full coenzymatic activity with glutamate dehydrogenase. Electron spin resonance spectra in the presence of glutamate dehydrogenase show mixtures of free and strongly immobilized spin-label. Binding studies in phosphate buffer demonstrate: (a) weak binary binding to the enzyme with a dissociation constant in the order of 2mM;(b) an indication for negative cooperativity or different sites for binding to enzyme-2-oxoglutarate, with dissociation constants in the order of 20--250muM; (c) similar but much weaker binding to enzyme-2-oxoglutarate-ADP; (d) a strong positive cooperative binding to enzyme-2-oxoglutarate-GTP, dependent on the enzyme concentration. Binding of phosphate to the enzyme with a Kd of about 20 mM or binding of pyrophosphate or tripolyphosphate with a Dd of about 2.5 mM enhances the binding of spin-labelled NAD+ in the presence of 2-oxoglutarate. There is evidence that the binding sites for these phosphates coincide with phosphate binding subsites of GTP.


Urinary cyclic AMP and renal concentrating capacity in infants. In 15 infants between 1 and 31 weeks the effect of antidiuretic hormone (ADH) on the renal concentrating capacity and urinary cyclic AMP (cAMP) was tested. A significant decrease of urine flow and a significant increase of osmolality, urea and cAMP was observed indicating that the distal nephron of the infant kidney is responsive to exogenous ADH and that its effect is mediated by cAMP. The results of a second series with 52 normally hydrated infants demonstrate that the nonlinear age-related increase of osmolality and urea in urine is accompanied by a similar pattern of cAMP excretion, pointing out that the maturation of the concentrating capacity seems to be related to an increasing responsiveness of the cAMP system to ADH. Furthermore the results raise the possibility that increasing concentrations of urea and solutes in the medulla and papilla of the infant kidney may have--in the presence of very low ADH secretion--an additional stimulating effect on cAMP formation.


Extracerebrospinal metastases in glioblastoma. Case report and review of the literature. A malignant glioblastoma adherent to the dura mater was removed from the parieto-occipital lobe in a 12-year-old boy. The site of the tumor was subsequently irridiated by 4000 rads of Cobalt-60. Five months later the boy was readmitted complaining  of pains in the pelvis an in both thighs. X-ray examination of the pelvis demonstrated multiple metastases. Investigation of bone marrow revealed replacement of normal haematopoiesis by a tumor cell population histologically identical to that of the brain tumor. Reviewing the literature 58 reports on glioblastomas with extracerebrospinal metastases could be found. Metastases were preferably localized in cervical or mediastinal lymph nodes, lungs, bones, liver, dura mater, and operative flap. It is suggested that extracerebrospinal metastases occur most frequently after the tumor has infiltrated the cranium and extracranial soft tissues. In the case reported here it is speculated that the tumor spread to extraneural tissues after invading the dural veins. The possible occurrence of extracerebrospinal metastases in glioblastoma emphasizes the necessity of additional chemotherapy.


Cyclic 3',5'-adenosine monophosphate and bronchial tone. The present studies demonstrate that adenylate cyclase and cyclic 3',5'-adenosine monophosphate (cAMP)-phosphodiesterase activities in dog bronchus are comparable to those found in other smooth muscle preparations. Catecholamines, in the order isoproterenol greater than epinephrine greater than norepinephrine, increase the rate of cAMP formation. This effect can be competitively inhibited by propranolol and potentiated by a cAMP-phosphodiesterase inhibitor. The kinetic study of bronchial cAMP-phosphodiesterase showed two different rates of cAMP hydrolysis, with apparent Km values of 1.4 and 48.0 muM. The high affinity cAMP-phosphodiesterase was inhibited competitively by theophylline and papaverine, the latter being about 20 times more potent than the former. The potency of each compound to inhibit the enzyme and to relax the bronchial strip was comparable. These results, the similar order of potency of the catecholamines to relax the bronchus and to increase the rate of cAMP formation, the competitive inhibition of both effects by propranolol, and the relaxing effect of dibutyryl cAMP on bronchial strip, are compatible with the assumption that the cAMP system is one of the biochemical mechanisms mediating bronchial smooth muscle relaxation.


Strain differences in responsiveness of norepinephrine-sensitive adenosine 3',5'-monophosphate-generating systems in rat brain slices after intraventricular administration of 6-hydroxydopamine. The development of hyperresponsiveness in cyclic AMP-generating systems has been investigated in brain slices of Sprague-Dawley and F-344 rats following intraventricular administration of 6-hydroxydopamine. Hyperresponsiveness to adrenergic agonists in cerebral cortical slices of Sprague-Dawley rats pertained during the period 5-25 days after treatment with 6-hydroxydopamine. In contrast, hyperresponsiveness did not develop in cerebral cortical slices of F-344 rats. Reductions in norepinephrine levels of the cerebral cortex and hypothalamus following 6-hydroxydopamine treatment were comparable in the two rat strains. A hyperresponsiveness to norepinephrine and isoproterenol failed to develop in mesencephalic slices of either strain 12-14 days after treatment with 6-hydroxydopamine. The accumulation of cyclic AMP elicited by norepinephrine in cortical slices of F-344 rat is normally about 50% greater than the accumulation elicited in slices from Sprague-Dawley rats. However, after 6-hydroxydopamine treatment, there was no significant difference in the accumulations of cyclic AMP elicited in cortical slices from the two rat strains. These data indicate there may be a limit to the responsiveness of catecholamine-sensitive cyclic AMP generating systems which can develop following alterations in synaptic input.


Dopamine-sensitive adenylate cyclase in the rat kidney particulate preparation. Dopamine, apomorphine, isoproterenol and norepinephrine all increased the concentration of adenosine 3',5'-monophosphate in the rat kidney particulate preparation, which was composed of tubules, glomeruli and blood vessels. The concentrations of dopamine, apomorphine, isoproterenol and norepinephrine causing a half-maximal increase were 50, 83, 0.1 and 10 muM, respectively. The alpha-blocker, phentolamine, at a concentration as high as 1 mM, did not significantly reduce the effect of these drugs on the adenosine 3',5'-monophosphate concentration. The beta-blocker, propranolol (50 muM), blocked the effect of isoproterenol and norepinephrine, but not that of dopamine. The effect of dopamine was selectively blocked by spiroperidol (50 muM), a dopamine receptor antagonist, whereas the effects of isoproterenol and norepinephrine was not blocked by spiroperidol. These results suggest that in the rat kidney particulate preparation there is a specific dopamine receptor which can lead to the increase in the concentration of adenosine 3',5'-monophosphate.


Effects of adenosine and related compounds on adenylate cyclase and cyclic AMP levels in smooth muscle. The hypotheses were tested that the relaxant effect of adenosine and related compounds in the longitudinal muscle of the rabbit small intestine involves interaction with adenylate cyclase and/or the elevation of tissue cAMP levels. Adenylate cyclase was prepared by gentle homogenization of an isolated smooth muscle cell fraction obtained after collagenase digestion of longitudinal muscle strips. A number of analogs and derivatives of adenosine possessing a primary or secondary 6-amino group were found to inhibit the enzyme similarly to adenosine; however, there was no correlation between compounds known to relax the intact tissue and the existence, or the degree of, cyclase inhibition. Isolated muscle strips were exposed to adrenaline, isoprenaline, adenosine or ATP, at doses causing 30-60% relaxation, for 60 sec prior to sampling and analysis of cAMP content. While small increments in cAMP levels were found after administering adrenaline or isoprenaline, no change was found with adenosine in the absence or presence of theophylline of 1-methyl-3-isobutylxanthine. Neither adenylate cyclase inhibition nor changes in cAMP levels appear to be part of the mechanism of the smooth muscle relaxant action of adenosine or ATP.


Hypoglycemia induced by alpha-adrenergic stimulation during alkalosis. Hypoglycemia developed during respiratory alkalosis in fasted rats. This hypoglycemia was markedly potentiated by the simultaneous injection of inhibitors of hepatic gluconeogenesis or a beta-adrenergic blocking agent; was not influenced by anti-insulin serum; was attenuated by hexamethonium; and was abolished by an alpha-adrenergic blocking agent. The rate of removal of injected glucose by peripheral tissues increased during alkalosis in insulin-deficient rats. The uptake of [14C]-glucose by the adipose tissue in vivo, which is stimulated by a very minute amount of insulin, was also stimulated during alkalosis whether or not the circulating insulin was neutralized with anti-insulin serum. It was concluded that, in alkalotic rats, blood glucose is rapidly utilized by peripheral tissues dependent on alpha-adrenergic stimulation, but without mediation of insulin and that this leads to development of hypoglycemia.


Erythroid precursor fusion induced by Sendai virus. The ultrastructural events of the interaction of Sendai virus (SV) with fetal mouse erythroid precursors, and SV-induced fusion of erythroid precursors at different maturation stages are described. SV was shown to affect the erythroid nuclei causing interruption of the nuclear membrane, enlargement of the nucleolus and nuclear fusion. SV induced fusion also between dividing and non-dividing cells.


Integration in descending motor pathways controlling the forelimb in the cat. 2. Convergence on neurones mediating disynaptic cortico-motoneuronal excitation. With intracellular recording from forelimb motoneurones the spatial facilitation technique has been used to investigate interaction between descending pathways and forelimb afferents. As previously shown for the hindlimb, pyramidal volleys effectively facilitate interneuronal transmission in reflex pathways from different primary afferents. Evidence is presented suggesting disynaptic excitation from corticospinal fibres of interneurones in the reciprocal Ia inhibitory pathway. Interneurones of other reflex pathways from group I muscle afferents recieve monosynaptic pyramidal excitation. During pyramidal facilitation volleys in cutaneous afferents may evoke PSPs in motoneurones after a central delay of 1.3 ms suggesting that the minimal linkage is disynaptic. Information regarding convergence on the neurones intercalated in the disynaptic cortico-motoneuronal pathway was obtained by investigating the effect from primary afferents and from other descending pathways on the disynaptic pyramidal EPSPs. Volleys in cutaneous and group I muscle affferents facilitate transmission in the disynaptic cortico-motoneuronal transmission with a time course showing oligosynaptic (probably monosynaptic) action the intercalated neurone. Rubrospinal volleys likewise effectively facilitate disynaptic cortico-motoneuronal pathway with a time course showing oligosynaptic (probably monosynaptic) action on the intercalated neurone. Rubrospinal volleys likewise effectively facilitate disynaptic cortico-motoneuronal transmission with a time course showing monosynaptic action on the intercalated neurone. Spatial facilitation experiments involving three tests revealed that those intercalated neurones which receive convergent monosynaptic excitation from corticospinal and rubrospinal fibres are excited also from cutaneous forelimb afferents. Disynaptic cortico-motoneuronal transmission was also monosynaptically facilitated by stimuli in the dorsal mesencephalic tegmentum probably activating tectospinal fibres. Disynaptic, presumed tectospinal EPSPs were facilitated from cutaneous forelimb afferents. The convergence onto the neurones intercalated in the disynaptic excitatory cortico-motoneuronal pathway suggests that these neurones integrate the activity in different descending pathways and primary forelimb afferents.


Timing of mitosis in Physarum polycephalum: effects of agents affecting cyclic AMP concentrations. Cultures of Physarum polycephalum incubated with caffeine or theophylline for over 100 min prior to mitosis exhibited mitotic delay proportional to the time of treatment before 100 min. Starved cultures exhibited mitotic delay at times of starvation longer than 180 min and slight stimulation from 100-180 min. Dibutyryl cAMP appeared to accelerate reconstruction of the nucleus following mitosis.


Inhibition in the rat of gastric acid secretion and cyclic AMP analogs accumulation in vitro by somatostatin. Various somatostatin (S) analogs exhibited similar degree and similar, or shorter, duration of inhibition of basal gastric acid secretion as S in the unanesthetized rat and similar, or less, inhibition of the cyclic AMP accumulation induced by prostaglandin E2 in the rat anterior pituitary in vitro. With the analogs examined, the gastrointestinal and pituitary receptors appear to exhibit generally similar recognition specificity with the differences within the gastro-intestinal activities reflecting duration of availability rather than receptor affinity.


Morphometric study of the aortic body type I cell. The subclavian glomus (aortic body) of New Zealand white rabbits was examined ultrastructurally using stereological morphometric analysis. The Type I cells of the glomus possess numerous electron-opaque vesicles which occupy approximately 12% of the cytoplasmic volume of the cells. The amine-containing vesicles comprise a heterogeneous population of vesicles with a mean caliper diameter of 113.5 nm. Differences in vesicle diameters may indicate the storage of different biogenic amines, different secretion or maturation states between glomera and/or additional physiological functions for the glomera.


The effect of distamycin A on some biochemical functions of tissue cultures infected with Herpes simplex virus. The antibiotic distamycin A displays no inhibition of protein synthesis in HeLa cell cultures. In HSV infected HeLa cell monolayers the drug exerts a partial inhibition of virus induced thymidine kinase synthesis but is devoid of inhibitory effect on the increase of activity of DNA polymerase and on total DNA synthesis. The antiviral activity of distamycin A is based on a mechanism which appears to be different from that displayed by other drugs such as actinomycin and IUD.


Micropuncture studies of the motility of rete testis and epididymal spermatozoa. The effect of cyclic adenosine 3':5'-monophosphate (cAMP) and caffeine on the motility of spermatozoa obtained in vivo by micropuncture from the rat rete testis, caput epidiymidis, and cauda epididymidis was studied. Spermatozoa from all sites were immobile in their native fluid. Rete testis spermatozoa were not motile under any experimental conditions. After dilution in salt solution, some caput sperm exhibited circular motion, whereas most cauda sperm swam progressively. Dextrose enhanced the motility of sperm from both epidiymal sites. Caffeine further increased the motility of epididymal sperm. Dibutyryl cAMP and cAMP stimulated caput spermatozoa but had no effect on cauda spermatozoa.


[Electrical activity of layers of isolated cortex during drowsiness and different sleep stages]. In cats with isolated cerebral cortex, with the aid of electordes indwelled into different cortical layers, first electrographic changes during onset of sleep were shown to occur, as with intact cortex, in the lower layers of the isolated cortex. 3-4 months after the cortex isolation, changes of electrical activity during sleep occured simultaneously in the cortex of both hemispheres. Awakening produced desynchronization of electrical activity in all cortical layers of both hemispheres. Origin of primary electrical activity changes in the lower cortical layers, is discussed.


[Myocardial cell adenyl cyclase activity and the effect of thyrocalcitonin]. The thyrocalcitonin (TCT) effect under influence of some factors affecting the activity of membrane adenylcyclase (AC) of the ventricle myocardial cells in cold-blooded animals, was studied. The TCT positive inotropic effect is intensified by decrease in the potassium concentration in the medium, by blockade of alpha-adrenoreceptors with phentolamine, and inhibited by increase in the potassium concentration and blockade of beta-adrenoreceptors. The TCT action is preserved in presence of adrenaline.


[Participation of histamine in activating the hypothalamo-hypophyseo-adrenal system during stress]. In 289 male rats, accumulation of histamine in diencephalon, augmentation of activity in the hypothalamus, and increase of the ACTH content in the hypophysis and 11-OCS in adrenal and plasma were revealed under a functional stress for the hypothalamo-hypophyseal-adrenal system (HHAS). Further accumulation of histamine was associated with increased secretion of CRF, ACTH, and 11-OCS. Blocking of histamine by diphenylhydramine hydrochloride (10.0 mg/kg i. p. -- 30 min before the experiment) depressed the rate and intensity of the initial reaction of HHAS activation.


[Effect of a periodic growth-retarding diet on changes with age in the hypothalamo-hypophyseo-adrenal system]. Two pacemakers localized in sinoatrial and atrioventricular valves were revealed in the tench heart. In their absence, the ventricular and sometimes the atrial myocardium are capable of initialting the automaticity. Pacemaker's cells of the sinoatrial valve are surrounded by specific area similar to sinoauricular ring in amphibia or sino-caval region in mammals. Characteristics of the action potential in different parts of the tench heart are presented.


Somatostatin-induced inhibition of insulin secretion by isolated pancreatic rat islets prepared by micro-dissection or collagenase digestion. The effect of somatostatin on insulin secretion in response to a variety of stimuli was investigated in micro-dissected as well as collagenase isolated pancreatic rat islets. Somatostatin inhibited insulin secretion in both islet preparations in the presence of different initiators, but failed to affect the hormone output induced by theophylline, 1-methyl-3-isobutylxanthin (IBMX) or p-chloromercuriphenylsulfonic acid (CMBS). The inhibitory action was associated with decreased glucose metabolism by islets (measured as conversion of U-14C-glucose to 14CO2).


Hypolipidemic effects of metformin in hyperprebetalipoproteinemia. Metformin's hypolipidemic effects (2.55 g/day for 3 months) have been studied in 19 subjects with Fredrickson's Type IV hyperprebetalipoproteinemia. The majority of patients were above ideal body weight (relative body weight = 118 +/- 2.7 %). Eleven of the subjects presented chemical diabetes, 5 fasting hyperglycemia, and 3 normal glucose tolerance. After treatment with metformin, body weight showed a slight, but significant reduction (--2.4 +/- 0.3 kg). Glucose tolerence was not substantially altered while basal glucose was significantly reduced in the 5 subjects with fasting hyperglycemia. Basal plasma insulin was significantly reduced in all the patients following metformin treatment. Insulin response to OGTT was slightly reduced in the subjects with fasting hyperglycemia. Independent of the patients' glucose tolerance, metformin treatment induced a marked decrease in plasma triglycerides (-- 40 %) and a reduction in plasma cholesterol (-- 12 %). No correlation was found between triglyceride and cholesterol reduction and body weight, glucose, and plasma insulin variations. Like phenformin, metformin acts not only on glucose metabolism and insulin secretion but on lipid metabolism as well.


The interaction of hCG with rat adipose tissue: apparent lack of hCG--LH receptors. The interaction of human chorionic gonadotropin (hCG) with rat adipose tissue was investigated by both metabolic and binding studies. Highly purified preparations of hCG did not affect the adenylate cyclase activity nor the lipolysis of rat adipocytes in the presence or in the absence of GTP. However, it was demonstrated that (a) the hCGs used were biologically active since they stimulated cAMP and testosterone production by rat Leydig cells, and (b) there are receptor sites on the rat ovary that bind [125I]hCG and recognize rat luteinizing hormone (LH). The lack of response cannot then be attributed to a loss of activity of the hormone preparation tested nor to a failure of the rat tissues to recognize an hormone of human origin, but rather to an absence of hCG--LH receptors on the fat cell membrane surface. It is suggested that results previously reported in other laboratories could be explained by the presence of contaminating amounts of lipolytic hormones in their preparations.


Adenosine 3',5'-monophosphate-dependent protein kinase from normal human adrenal. The protein kinase of normal human adrenal cytosol has been resolved by DEAE-cellulose chromatography into two major components, the protein kinases I and II, which are both adenosine 3',5'-monophosphate (cAMP) dependent. Both enzymes have similar substrate specificities, cAMP-dependency, and sensitivity to the stimulation by this nucleotide, but differ in their states of activation after preincubation with histone. The DEAE--cellulose charomatography of dissociated cytosol protein kinase reveals only one peak of kinase activity and two peaks of cAMP binding activity (A and B). Both binding proteins are able to inhibit the kinase activity of the catalytic subunit. Recombination experiments suggest that the regulatory subunit A originated from protein kinase I and subunit B from protein kinase II. The phosphorylation of histone by adrenal protein kinases is inhibited by a heat-stable protein inhibitor isolated from human fetal brain and human adult adrenal.


Apparent positive cooperativity of ACTH action on adrenocortical cells: the effect of hormone degradation. The ability of adrenocortical cells to degrade ACTH1--39 and [125I]ACTH has been assessed under various conditions. Under conditions leading to increased hormone degradation there was an elevation of both the ED50 and the value of the Hill coefficient derived from concentration-effect curves for ACTH-stimulated steroidogenesis. Such degradative mechanisms offer a simple explanation for tha apparent positive cooperativity proposed by others for ACTH-receptor-adenylate cyclase interactions.


Insulin receptor sites as membrane markers during embryonic development. I. Data obtained with unfertilized and fertilized sea urchin eggs. Binding of insulin to sea urchin egg plasma membrane has been studied by biochemical and immunocytochemical methods. Unfertilized and fertilized eggs as well as embryos during the first cell division have been used. 1. Competition experiments between 125I-insulin (1 nM) and an excess of native insulin (30 muM) indicate a specific hormone fixation to membrane crude extracts from unfertilized and fertilized eggs. The magnitude of "specific binding" is comparable to values recorded for mammalian cells. 2. Inhibition of insulin fixation by concanavalin A (100 mug/ml) suggests the glycoprotein composition of plasma membrane receptors. 3. An 30-min incubation of unfertilized and fertilized eggs in the presence of insulin leads to a significant increase in cyclic AMP content. 4. An immunocytochemical method demonstrates that insulin is selectively and specifically bound to the plasma membrane of eggs incubated in the presence of insulin before fixation. It can be concluded that insulin receptor sites are components of sea urchin eggs plasma membrane. Insulin binding which leads to cyclic AMP accumulation is not deeply modified by fertilization and does not include visible morphological changes in the eggs.


Squamous cell differentiation in a clonal teratocarcinoma cell line. Several clonal teratocarcinoma cell lines restricted to squamous cell differentiation have been established from a subculture of totipotential murine teratocarcinoma stem cells. These lines contain a continuum of cell types from less differentiated precursor cells to differentiated squamous cells. In contrast to their highly malignant progenitors, these cells are nontumorigenic. Chromosomally, the cells are near-tetraploid, and their DNA distributions are tetraploid. These cells lines may prove useful in the study of normal squamous cell differentiation and also will be important in studies concerning the conversion of malignant cells to non-malignant progeny.


Abnormal plasma lipoproteins and lecithin-cholesterol acyltransferase deficiency in alcoholic liver disease. Serial studies of plasma lipids and lipoproteins were performed in 4 patients with acute alcoholic liver disease characterized by a massive fatty liver and laboratory evidence of intrahepatic cholestasis. There were striking alterations in the plasma lipoprotein electrophoretic patterns characterized by the absence of alpha- and pre-beta-lipoprotein bands and the presence of a single band of abnormal mobility. These changes were associated with an extreme decrease in plasma lecithin-cholesterol acyltransferase activity, resulting in greatly reduced levels of plasma cholesteryl esters and increased levels of unesterified cholesterol. In 2 patients hypertriglyceridemia and hypercholesterolemia were present, the latter because of an increase in unesterified cholesterol. Lipoproteins were isolated from the plasma by sequential ultracentrifugation at the densities used for separation of normal very low density, low density, and high density lipoproteins; however, the patients' lipoproteins were different from normal in lipid composition and ultrastructure. All of the lipoprotein fractions were decreased in cholesteryl esters and the major lipoprotein was a triglyceride-rich low density lipoprotein. Electron microscopic studies of the low and high density lipoprotein fractions revealed the presence of bilamellar vesicles and stacked discs. All of the changes in lipoprotein composition and ultrastructure gradually returned to normal with clinical improvement. These observations indicate that alcoholic liver injury is associated with profound alterations in lipoprotein composition and metabolism which may be related in part to lecithin-cholesterol acyltransferase deficiency.


Cytomegalovirus-associated gastric ulcer. A patients with a cytomegalovirus (CMV) post-transfusion syndrome developed upper gastrointestinal tract bleeding; subsequently, a gastric ulcer was found. CMV was searched for in the gastroscopic biopsy material because the gastric ulcer had occurred in a setting of CMV mononucleosis. CMV cells were found in gastroscopic biopsy sections and CMV was also cultured from biopsy material. This study illustrates the feasibility of antemortem diagnosis of CMV-associated disease of the upper gastrointestinal tract.


Cytological studies of the effect of hepatocarcinogens on liver cells of rats. Morphological and cytological changes of liver cells were studied under sequential feeding of p-dimethylaminoazobenzend (DAB) and CCl4 injection. It was found that in hepatic cells of experimental animals definite macroscopical alterations occurred manifested in a change in the colour and the external texture of the liver, the appearance of nodules of different sizes, segmentation, and the abdominal cavity became filled with ascitic fluid, associated with spleenomegaly. The histopathological changes include a disturbance of the normal regularity of hepatic cells, nuclear atypia, hyperchromatism, vaculation of the cytoplasm and decreased mitochondrial and Golgi elements.


[The subcapsular blastema of the adrenal cortex of the swine after continuous long-term infusion of exogenous ACTH]. After long time application of homologous ACTH the morphokinesis of the adrenal cortex of the pig was investigated experimentally. Following results were obtained: 1. In view of the controls the absolute and relative weight of the adrenals is raised considerably. 2. The progressive transformation is followed by the disappearance of the zonal structure of the adrenal cortex, and the parenchyma get the picture of fasciculata cells generally. 3. Nearly exclusive the zona fasciculata consists of great, pale activated spongiocytes with 2 nucleoli frequently. Topochemically glycogen and the lipids are inconstant, however the histochemical activity of succinodehydrogenase, lactate dehydrogenase, acid and alkaline phosphatase are considerable raised in regard of the controls. 4. The zona fasciculata contains degenerated cells isolated only. Signs of extensive regressive changes are not present. 5. The zona glomerulosa is dissolving or eliminating respectively. The consequences for the synthesis of the adrenal steroid hormones are discussed. 6. A large, spongy subcapsular blastema with several cell layers and a rich capillary network develop between the fibrous capsula of the adrenal and the zona fasiculata. The fasciculata cells are the direct continuation of the subcapsular blastema. The blastema contains neither glycogen nor lipids and histochemical activities of the enzymes are absent, too. The significance of the subcapsular blastema for the morphological and functional adaptation of the adrenal cortex in stress are discussed. Under the conditions of the closed hypothalamo-hypophyseal-adrenal control system the new origin of cells (hyperplasia) is not significant for the morphokinetic adaptive reactions of the adrenal cortex. Rather the subcapsular blastema represents a reserve area which after the destruction of the endocrine parenchyma through specific pathogens the organism enabled to the regeneration of the adrenal cortex.


[Efficacy of angiography for the diagnosis of trophoblastic tumors (author's transl)]. Angiography has proved valuable as a method for visualizing the position and extent of trophoblastic tumors. Angiography together with the determination of gonadotropin makes a more exact diagnosis possible and, therefore, an individual therapy. The treatment must be intentionally carried out's the age of the patient plays an important role here. For many cases today, purely conservative therapy with cytostatic drugs is possible. This method was used in 11 female patients with suspected or histologically established trophoblastic tumors. In 4 women, the tumor was suspected a because of the reincrease in HCG excretion following a hydatidiform mole. A metastasizing mole was established histologically in 3 patients and a choriocarcinoma, in 4 patients. A good correspondence was found between angiographic and macroscopic findings in 6 women who underwent surgery. In evaluating control angiograms following chemotherapy, it should be noted that tumors do not recede completely in every case.


Hormone independent in vitro erythroid colony formation by mouse bone marrow cells. B.M. cells of RLV-infected BALB/c mice can proliferate in methylcellulose in the absence of E.P., while normal B.M. cells cannot (12). Not only the more primitive BFU-E shows hormone-independency (18). This phenomenon is in favour of the view that the Rauscher virus induced erythroblastosis is a true neoplasia although transplantation experiments failed so far. The experiments in which transformation in vitro of B.M. cells by RLV is established (19) show that the CFU-E can serve as a target for the virus. Treatment of normal mice with CFA leads to a rapid increase in CFU-E in the bone marrow (18). Splenomegaly of RLV-infected mice is enhanced by CFA-treatment probably due to an increase in targets. Transfection with proviral DNA also can transform the CFU-E of BALB-c mice. This approach allows in vitro studies on the resistence of mouse strains to RLV in vitro. The studies are of interest for the human disease in two aspects. In vitro transformation assays are needed to study the oncogenic potential of putative human leukemia viruses. Furthermore the studies have yielded some new insight in the pathogenesis of virally induced erythroblastosis. This might serve as a model for e.g. acute myeloid leukemia in man.


Restricted addition of proviral DNA in target tissues of chickens infected with avian myeloblastosis virus. Proviral DNA is synthesized within an hour after infection of chicken cells with an avian oncornavirus and is integrated into nuclear cellular DNA within a short time. The viral DNA appears to be synthesized as double-stranded molecules of approximately 6 X 10(6) daltons some of which are converted into supercoiled cricles perhaps as a requisite for integration. The endogenous v-DNA in normal chicken cells and both the endogenous and amv v-DNA in leukemic chicken myeloblasts are covalently linked with chromosomal DNA. There is no detectable free DNA either circular or linear present in leukemic cells several weeks after infection. The endogenous v-DNA which is transmitted vertically from parents to offspring is uniformly and stably distributed in all chicken organs. There are about 1-2 copies of endogenous provirus per haploid genome of all normal cells. This DNA is very closely related to RAV-O RNA. After infection with AMV it seems that target cells such as leukemic myeloblasts, RBC and nephroblasts acquire complete copies of AMV DNA. Interestingly, only these target cells can be converted to neoplastic cells in the chicken as well as in vitro. The target cells acquire 1-2 copies of AMV specific DNA per haploid genome in addition to the endogenous v-DNA. All the available evidence shows that leukemic and kidney tumor cells have acquired AMV v-DNA. It remains to be elucidated whether the newly added viral DNA is alone responsible for neoplastic changes or does so in conjunction with endogenous viral information.


Sequences and functions of Rous sarcoma virus RNA. A procedure has been developed to map the genetic elements of avian tumor virus RNA, which has a molecular weight of about 3 X 10(6) daltons and a poly(A) sequence at the 3' end. For this purpose, about 30 RNase T1-resistant oligonucleotides were ordered relative to the 3'-poly(A) terminus of the RNA, to construct an oligonucleotide map of viral RNAs. A cluster of seven envelope gene (env)-specific oligonucleotides, identified by their absence from the otherwise very similar oligonucleotide map of an envelope-defective deletion mutant (which lacks the major viral glycoprotein), mapped at a distance of 0.9 to 1.6 X 10(6) daltons from the poly(A) end of sarcoma virus RNA. A cluster of three sarcoma gene (src)-specific oligonucleotides, identified by their absence from the otherwise nearly identical oligonucleotide map of a transformation-defective deletion mutant mapped at a distance of 0.2 to 0.6 X 10(6) daltons from the poly(A) end of sarcoma virus RNA. The oligonucleotide maps of sarcoma viruses and of related deletion mutants were the same from the poly(A) end up to 0.2 X 10(6) daltons and included one terminal oligonucleotide, termed C, which is found in all avian tumor viruses tested so far. Preliminary mapping experiments ordering the src-specific and env-specific oligonucleotides of recombinants, selected for sarcoma and envelope genes of different parents, agree with those obtained by comparing maps of wild type viruses and deletion mutants. A partial genetic map consistent with these results suggests that the src gene maps between the env gene and the 3'-poly(A) end of viral RNA. This map reads: poly(A)-src-env-(pol, gag).


Control of peptide chain initiation in uninfected and virus infected cells by membrane mediated events. Initiation of protein synthesis in tissue culture cells is rapidly inhibited or blocked by addition of either DMSO, ethanol, TPCK, cytochalasin B, or sucrose to the growth medium. In contrast, these agents do not interfere with the initiation of protein synthesis in cell-free extracts to a comparable extent. These results support the hypothesis that protein synthesis in tissue culture cells can be influenced by membrane mediated events. Translation of viral mRNA in RNA virus infected cells is resistant to a number of these inhibitors of peptide chain initiation and proceeds under conditions where translation of host mRNA is almost completely suppressed. It appears that viral mRNA possesses a greater ability than host mRNA to form mRNA-ribosome initiation complexes when the overall rate of peptide chain initiation is reduced. This observation has led to a number of predictions concerning the strategy of virus directed suppression of host mRNA translation. Under optimal growth conditions protein synthesis appears to be regulated mainly, but not exclusively, by the amount of the mRNA available for translation. However, when cellular growth and/or the overall rate of peptide chain initiation is restricted, control of protein synthesis at the translational level becomes decisive with the translation of each mRNA species proceeding with its own characteristic efficiency most probably as a result of inherent differential affinities of individual mRNA species for ribosomes.


Proliferation kinetics and cyclic AMP in human leukaemic cells: their relevance to therapy. In the cells of acute leukemia the duration of the cell cycle and its phases was determined in vitro by autoradiographic techniques. Tests were run simultaneously for bone marrow and peripheral leukaemic cells and the intracellular levels of cAMP were measured. The results suggest a correlation to exist between cAMP concentration in leukaemic cells and the growth rate of the cell population. Hence, cAMP assays appear to be suited for monitoring cellular proliferation kinetics and could provide a useful parameter for planning cell-phase-specific therapy.


[Mechanism of anti-inflammatory action of 2-(2-fluoro-4-biphenylyl) propionic acid (flurbiprofen)]. Anti-inflammatory mechanism of 2-(2-fluoro-4-biphenylyl) propionic acid (Flurbiprofen, FP-70) was studied by various analysis in comparison with other drugs. It was found in the test of rat edema induced by various phlogists that carrageenin and yeast-induced edemas were markedly inhibited by FP-70, whereas dextran, formalin, serotonin and bradykinin-induced edemas were scarcely inhibited by FP-70. The action of FP-70 was similar to that of soy bean trypsin inhibitor. However, FP-70 showed no effects on kinin synthetase and kininase. FP-70 showed a marked inhibition on prostaglandin synthesis. The inhibitory effect of FP-70 was 10.1, 96.5 and 2280.6 times as large as indomethacin, ibuprofen and acetylsalicylic acid, respectively. FP-70 did not inhibit the permeability of dye induced by prostaglandin E2 in the rat skin. FP-70 inhibited the acid phosphatase and beta-glucuronidase activities of isolated lysosome of rat liver and also suppressed the release of acid phosphatase from the lysosome. These effects were similar to those of indomethacin. On the other hand, FP-70 suppressed markedly the heat-induced hemolysis of dog erythrocytes. The effect was similar to that of indomethacin and was 10 times stronger than those of ibuprofen, ibufenac and phenylbutazone. Activation of rat liver mitochondrial ATPase by FP-70 at a concentration of 10 muM was 74.7%, while indomethacin showed 37.8% activation at the same concentration. FP-70 as well as ibuprofen and phenylbutazone uncoupled the oxidative phosphorylation in rat liver mitochondria. From the above and previously reported results, it is suggested that the potent anti-inflammatory action of FP-70 is the result of the following effects; inhibition on the protein and leucocyte migration, inhibition on the prostaglandin synthesis, stabilization of the cell membrane and activation of ATPase.


Angiographic findings in some rare pancreatic tumors. Angiographic findings in one giant cell carcinoma, one cystadenocarcinoma, one poorly vascularized mucinous cystadenocarcinoma, as well as in two avascular (gastrin- and glucagon-producing) islet-cell tumors of the pancreas are described. Two hypervascularized islet-cell tumors are presented for comparison and a case of tumorous chronic pancreatitis in a child is reported because ot its rarity. The aggressiveness of the giant cell carcinoma of the pancreas was demonstrated by its expansive growth. In the case of cystadenocarcinoma angiography revealed the tumor with hepatic metastases not diagnosed at explorative laparotomy. The relative hypovascularity in the case of mucinous cystadenocarcinoma was unusual. Both avascular islet-cell tumors simulated a pancreatic pseudocyst and the final diagnosis was made only by immunoassay. Chronic pancreatitis in a child presented with marked hypervascularization.


Who gets aftercare? A study of patients discharged from state hospitals in Kentucky. A cohort of 579 patients discharged from state mental hospitals in Kentucky during a one-year period was studied to determine the amount and nature of after-care services provided to them. A total of 319 received some form of attercare from community mental health centers during the two to three years after discharge. The majority received at least one service that could be classified as individual therapy or chemotherapy, about one-third received evaluation or rehabilitation services, and only a few had family interviews or group therapy. The majority of those receiving services had contact with psychiatrists, nurses, or other mental health professionals; less than half had contact with social workers or psychologists. In general the aftercare services tended to be medically oriented, brief, and limited in number. They were provided to a majority of the patients within their first month in the community.


Community aftercare of patients discharged from Utah state hospital: a follow-up study. To find out whether patients discharged from mental hospitals to the community are following recommended courses of aftercare treatment, the authors, with the assistance of five medical students, studied a sample of patients discharged from Utah State Hospital during 1973 and 1974. The discharge plans of 143 patients admitted from Salt Lake County indicated that 125 of them should have received aftercare services from a community mental health center. A review of CMHC records and interviews with the patients indicated that only 100 actually had contact with a center. Only 43 of the patients were still receiving aftercare services in the summer of 1975.


Aftercare of psychiatric patients and its relation to rehospitalization. In their study of the effects of posthospital treatment on psychiatric patients the authors found that those who entered aftercare were rehospitalized less often than those who did not. They found the rehospitalization rate was higher for schizophrenic patients than for patients in other diagnostic groups, and that schizophrenic patients who did not enter aftercare were rehospitalized at a higher rate than those who did. They found no difference between rehospitalization rates of patients who continued with their hospital therapist after discharge and those patients who worked with a new therapist.


Transitional care: a new approach to aftercare. The transitional-care program incorporates the principles of behavior modification and group and family therapy in order to help recently discharged psychiatric patients maintain the positive changes that occurred in the hospital. Patients meet for approximately three hours a week, for up to 12 weeks, with nursing staff who worked with them while they were hospitalized. Each patient has an individual treatment program and goals, and during meetings with his team he discusses his progress and problems in meeting those goals. Staff members encourage patients to accept responsibility for each other and not to become too dependent on them. The program has been successful in preventing rehospitalization.


Pharmacological aftercare for homogeneous groups of patients. In a study to evaluate the effectiveness of specific pharmacological treatment groups, former VA inpatients who have similar diagnoses and respond satisfatorily to drug treatment join a Prolixin group, a lithium group, or an affective disorder group. In monthly meetings patients learn about their illness, the importance of the taking medication as prescribed, and possible side-effects; they also receive psychological support. The 85 patients who joined the groups during the first 12 months attended 87.3 per cent of the sessions. Twenty per cent of them were rehospitalized, about half of them because they did not take medication as prescribed. The authors believe the groups are an economical and effective way to provide aftercare.


Spontaneous contractions of dispersed vascular muscle in cell culture. Dispersed vascular muscle cells from chick omphalomesenteric vessels maintained in primary cell culture contracted spontaneously. Six methods which produced contracting isolated muscle cells are described and compared. The combination of dispersion method and culture conditions to produce contracting muscle cells was more critical for vascular than for heart muscle. These findings of continuing pacemaker function demonstrate that functional integrity of isolated vascular muscle cells is possible to maintain. Further indication of the full functional state of the isolated vascular muscle cells was demonstrated by the sensitivity to norepinephrine at a physiological concentration (0.1 muM). Spontaneous contraction frequencies were similar to the range found in situ, and spontanious or norepinephrine-induced contractions had time courses corresponding to intact vessel contractions. This is the first report that isolated vascular muscle cells in primary cell culture retain functional characteristics found in situ and are suitable for pharmacological characterization of individual muscle cells.


Banded karyotypes of H-4-IIE-C3 rat hepatoma cells grown in vitro. Mitotic cells from the H-4-IIE-C3 rat hepatoma tissue culture line showed a range of 45 to 53 chromosomes per cell with 75% of the cells displaying a chromosome mumber between 49 and 52. Analysis of Wright's-Giemsa banded karyotypes of 22 cells revealed considerable cell to cell variation. Twenty-one structurally abnormal chromosomes were identified in these cells; the origin of nine of the 21 chromosomes were identified in these cells; the origin of nine of the 21 chromosomes could be determined. Of the structurally abnormal chromosomes detected, only one (M-1) occurred with a sufficiently high frequency to be of general use as a marker for these cells. This marker appears to be a Robertsonian translocation involving chromosome number 2 and chromosome number 10.


Mechanism of action of Pseudomonas aeruginosa exotoxin Aiadenosine diphosphate-ribosylation of mammalian elongation factor 2 in vitro and in vivo. Previous studies showed that Pseudomonas aeruginosa exotoxin A (PA toxin) catalyzes nicotinamide adenine dinucleotide (NAD)-dependent inhibition of protein synthesis in a rabbit reticulocyte lysate and transfer of radioactivity from [14C]adenine-labeled NAD to a protein having the same molecular weight as elongation factor 2 (EF-2) (B.H. Iglewski and D. Kabat, 1975). Such an inhibited protein-synthesizing lysate was restored to activity by addition of a protein from normal mouse liver which co-purifies with EF-2. In addition, EF-2 activity was almost totally absent in livers of mice which had been injected 24 h earlier with PA toxin. On the contrary, EF-2 concentrations were only partially reduced in other organs and were normal in brains of intoxicated mice. Studies using NAD labeled in various positions show that PA toxin, like fragment A of diphtheria toxin, catalyzes transfer of the adenosine 5'-diphosphate-ribosyl moiety of NAD. Furthermore, reversal occurred when the modified protein was incubated with excess concentrations of PA toxin and nicotinamide, and NAD was identified as a product of the reverse reaction. The protein modification catalyzed either by PA toxin or by fragment A of diphtheria toxin could be reversed by incubation with other toxin. These results support the proposal that these two toxins adenosine 5'-diphosphate-ribosylate and same amino acid of EF-2 in a stereochemically identical fashion. Furthermore, PA toxin inactivates EF-2 in intoxicated mice to an extent which would ultimately result in death.


Frentizole, a novel immunosuppressive, and azathioprine: their comparative effects on host resistance to Pseudomonas aeruginosa, Candida albicans, herpes simplex virus, and influenza (Ann Arbor) virus. Frentizole, 1-(6-methoxy-2-benzothiazolyl)-3-phenyl urea, a new immunosuppressive agent, and azathioprine were administered subcutaneously at predetermined immunosuppressive dose levels of azathioprine and up to 50 times an immunosuppressive dose level of Frentizole. After 10 days of treatment at these dose levels, the experimental groups were inoculated intraperitoneally with Pseudomonas aeruginosa or herpes simplex virus, inoculated intraveneously with Candida albicans, or infected by aerosol with Ann Arbor influenza virus. The results of these series of experiments indicate that Frentizole, even at super immunosuppressive doses, does not predispose the hose (mice) to Pseudomonas aeruginosa, Candida albicans, herpes simplex virus, or Ann Arbor influenza virus.


Herpes simplex virus infections in guinea pigs deficient in the fourth component of complement. Separate groups of normal and C4-deficient guinea pigs were inoculated with herpes simplex virus by intradermal (i.d.) and intraperitoneal (i.p.) routes. Virus infection, confirmed by clinical, virological, and serological criteria, did not last longer and was not more severe in C4-deficient guinea pigs than in normal guinea pigs. Serum C component levels were measured before, during, and after herpes simplex virus infection. In normal gruinea pigs there was no evidence for C4 utilization after either i.d. or i.p. inoculation. In both normal and C4-deficient guinea pigs, C1 and C3-9 levels remained unchanged in spite of i.d. or i.p. infection. These data suggested that C4 and the classical C pathway were not important for virus clearance.


Stable L-forms of Clostridium perfringens: growth, toxin production, and pathogenicity. Growth and toxin production of stable L-forms of Clostridium perfringens grown in a mini-fermentor were monitored. A gradual but steady increment in viable count occurred over a 7-h period, followed by death. The peak of viability preceded the optical density peak by 3 h. Theta, alpha, kappa, and lambda toxins were measured, with theta toxin appearing first in the culture supernate. Growth of the parent bacillus form of C. perfringens was compared under similar conditions. Toxin levels achieved by the bacillus culture exceeded those of the L-form culture four- to eightfold; however, based upon viable count, the L-form organism produced 8 to 16 times as much toxin as did the bacillus. The amounts of extracellular toxin produced by both forms were similar when related to cell protein rather than cell number. Guinea pig inoculation showed that the L-form of C. perfringens did not produce gas gangrene, although it was not entirely without effect. Both guinea pig and human sera were inhibitory to these L-forms, a fact attributable to a heat-liable component in the sera, most likely complement.


Interferon induction by radioprotective mercaptoalkylamines and derived thiophosphates. Radioprotective thiols (five mercaptoalkylamines and four derived thiophosphates) induced interferon and resistance to virus infection. Interferon production occurred in human and mouse nonlymphoid cell cultures. One of the thiols, S,2-aminoethylisothiourea, given intraperitoneally, protected mice against two unrelated viruses--Semliki forest virus and Herpesvirus hominis type 1. Two structurally different radioprotective thiols--the disulfide cystamine and L-cysteine--were unable to induce the firus resistance state or interferon.


Expression of Aleutian mink disease antigen in cell culture. Infection of CRFK feline kidney cells with Aleutian disease vurus leads to production of virus-induced antigen(s) in the nucleus which could be demonstrated by the fluorescent-antibody technique. The number of fluorescent nuclei was lineraly dependent on the dilution of the inoculum, but rarely exceeded 20% of the cells. Aleutian disease nuclear antigen was only transiently detectable. The virus-induced antigen was detected after infection of cells of several divergent species; however, the CRFK line of feline kidney cells was the most susceptible. Inhibitor studies indicated that deoxyribonucleic acid synthesis, ribonucleic acid synthesis, and protein synthesis were required for viral antigen production. Cell growth was also a requirement for synthesis of viral antigen, An in situ radioimmune assay was used to measure binding of 125I-labeled mink anti-Aleutian disease virus to infected cells and competition with unlabeled sera. The system is suitable for quantitation of infectivity.


Cell-mediated immunity to Sendai virus infection in mice. The development of a cell-mediated immune response to Sendai virus infection in mice was examined by the use of a 51Cr release assay of cytotoxicity. A low level of "background cytotoxicity" to Sendai virus-infected L cells was found in the spleens of uninfected CBA mice. Spleen cells from Sendai-infected mice showed an elevated level of cytotoxicity against these target cells for a period of 5 weeks, commencing 4 days after infection of the mice. A more transient response was observed in the spleens of mice infected with a serologically distinct virus, the Kunz strain of influenza. This cross-reacting, cell-mediated immune response was intermediate between that observed in unsensitized and Sendai-sensitized spleen cells. The relevance of these cell-mediated immune responses to respiratory tract virus infections is discussed.


Increased histamine sensitivity in mice after administration of endotoxins. CFW mice given submicrogram doses of endotoxins intravenously became highly susceptible to the lethal effects of 0.5 mg of histamine given intraperitoneally 1 to 2 h later. The histamine-sensitizing effects of the endotoxins were transitory and disappeared within 6 to 8 h. L-Epinephrine administered intravenously immediately after histamine challenge protected mice from death, but aterenol and isoproterenol were ineffective. The histamine-sensitizing effect in endotoxins was precipitated by anti-endotoxin sera with a concomitant eightfold loss in activity. However, dissociation of the immune complex in 0.25 M acetic acid fully restored histamine-sensitizing activity. The transitory nature of the hypersensitivity produced by endotoxin and the high heat resistance of the active material prove that it is different from the histamine-sensitizing effects of pertussigen.


DNA binding and its relationship to carcinogenesis by different polycyclic hydrocarbons. Five different polycyclic hydrocarbons with different degrees of carcinogenicity in vivo were tested for their metabolism to water-soluble products and their binding to DNA, RNA, and protein in normal embryonic hamster and BHK cells. The compounds studied were 7, 12-dimethylbenz(a)anthracene, benzo(a)pyrene, 20-methyl-cholanthrene, dibenz(a,h)anthracene and dibenz(a,c)anthracene. All five compounds were metabolized to water-soluble produces in both types of cells and treatment of cells with aminophylline enhanced this metabolism. After and not before this enhancement of metabolism by aminophylline, there was a relationship between the degree of carcinogenicity and binding to DNA. There was no such relationship with binding to RNA or protein. The results, indicating a relationship between the degree of carcinogenicity and binding to DNA under appropriate conditions of metabolism, support the suggestion that DNA is the target for carcinogenesis by such carcinogens.


Establishment in continuous culture of a new type of lymphocyte from a "Burkitt like" malignant lymphoma (line D.G.-75). The isolation and establishment in vitro of a hitherto undescribed type of lymphocyte designated D.G.-75 is reported. The original inoculum was derived from the pleural effusion of a child with a primary abdominal lymphoma, which clinically and histologically resembled Burkitt's lymphoma. In addition to the absence of the EBV genome and EBV receptors, this line possesses a number of other properties which distinguish it from previously described lymphoblastoid cell lines. It has different growth characteristics and morphology; does not form EAC or E rosettes (representative of B and T) cell surface markers, respectively); possesses IgM-kappa immunoglobulins on the cell surface (B lymphocyte), has an unusually high cap-forming ability and low agglutinability with fluorescent concanavalin A. One homologue of the No.14 chromosome pair possesses extra chromatin material as revealed on chromosome banding. This abnormal chromosome marker is similar to that described in biopsies and cultured tumor cells from patients with African Burkitt's lymphoma.


A case control study on immunity to two Epstein-Barr virus-associated antigens, and to herpes simplex virus and adenovirus in a population-based group of patients with Hodgkin's disease in Denmark, 1971-73. One hundred and eighty-five patients with untreated Hodgkin's disease (HD) comprising 86% of all new cases diagnosed in Denmark over a 2-year period, were individually matched with healthy controls of the same age, sex, and social class. In comparison to controls, HD patients showed significantly elevated mean antibody titres to Epstein-Barr viral capsid antierences in mean titres were found for adenovirus common antigen. Subdivision of the patients by age, sex, social class, HL-A antigens, stage of disease and histology did not alter this pattern, except that significant case-control differences in EB-VCA titres could be demonstrated only for the nodular sclerosis and lymphocyte predominance subgroups. After 1 year of treatment, a significant rise in EB-VCA mean titre had taken place. Splenectomy seemed to promote this titre elevation. Neither the initial titres to EB-VCA and EBV-EA nor changes in titres over time were related to prognosis. The results of our study, which are more representative of a population of HD patients and controls than previously reported studies, confirm that the reported relationship between EBV and HD exists, but that the elevated EBV titres are probably not of etiologic significance in most HD patients.


Detection of human C-type "helper" viruses in human leukemic bone marrow with murine sarcoma virus-transformed human and rat non-producer cells. Bone-marrow cells from two leukemic children were co-cultivated with the leukemic children A 7573. In early passages, C-type oncornaviruses were released as detected by extracellular reverse transcriptase assay. Co-cultivation of the infected canine cells with the non-producing cell lines R-970-5 (human) or K-NRK (rat) both transformed by Kirsten mouse sarcoma virus (MSV) yielded a new pseudotype of MSV that could transform rat embryo, rabbit SIRC and human kidney cells but not mouse embryo cells. The focur formation could be inhibited by an antiserum to the simian sarcoma virus but not by a serum directed against murine leukemia virus. A cell line derived from a focus of transformed cells became a highe virus is related to the simian sarcoma virus. It is concluded that the leukemic bone-marrow cells produce a C-type oncornavirus that can serve as a helper virus to the defective MSV.


Transformation of cultured rat adrenocortical cells by Kirsten murine sarcoma virus (Ki-MSV). Two-week-old primary cultures of normal adult rat adrenal cortex were exposed to Kirsten murine sarcoma virus (KiMSV). Within a week, the adrenal cells, which are normally fusiform and aligned in parallel, became pleomorphic and piled up extensively. Saturation density increased from 5-10 x 10(4) to 5-10 x 10(5) cells/cm2, population doubling time during exponential growth decreased from 36-40 to 16h, acid production increased and the growth rate became independent of a reduction in serum concentration from 10% to 1%. Inoculation of 2 x 10(6) of these transformed cells into immuno-depressed rats produced rapidly growing tumors within 1 week. Histologically, the tumors were pleomorphic carcinomas with areas ranging from anaplasia to near-normal, highly differentiated adrenocortical tissue. In addition to histologic evidence of differentiation, metabolic studies using 14C-prognenolone showed that the transformed cells were capable of 20alpha reduction and delta5,3beta dehydrogenation, both characteristic of normal steroid-secreting tissues. The transformed adrenocortical cells produced infectious C-type virus as indicated by electron microscopy, 3H-uridine incorporation, and focus formation in NRK (normal rat kidney) cultures. The neutralization pattern of this virus resembled that of authentic Ki-MSV. The transformation of adrenocortical cells by K-MSV demonstrates the capacity of this agent to induce carcinomas in differentiated cells after short-term culture, and widens the range of tissues known to be susceptible to K-MSV to include a secretory epithelium of mesodermal origin.


Horizontal transmission of feline leukemia virus under natural conditions in a feline leukemia cluster household. Ten post-weanling 4-month-old cats, designated "tracers", were placed in a feline leukemia cluster household to determine the efficiency of horizontal transmission of feline leukemia virus (FeLV). The tracer cats were confirmed as negative for prior exposure to FeLV. Following the placement in the leukemia cluster environment, the tracer cats were serologically monitored at intervals of 3-6 weeks for a total period of 1 year. The tests employed included the detection of FeLV using fixed-cell immunofluorescence and the detection and titration of antibody to : (1) the feline oncornavirus-associated cell membrane antigen (FOCMA), as detected by membrane immunofluorescence; (2) viable FeLV, using serum neutralization; (3) virion core protein p30, using radioimmunoprecipitation; and (4) virion glycoprotein gp70, using radioimmunoprecipitation. All of the tracers had evidence of horizontal infection by FeLV, by several criteria. Seven of the 10 had virus that could be isolated from plasma. All of these 7 developed a terminal illness within 18 months; 3 developed aplastic anemia, 3 infectious peritonitis, and 1 lymphoma. The remaining 3 were negative for FeLV by both virus isolation and fixed-cell immunofluorescence. These 3 did, however, develop high antibody titers by all four criteria and they remained healthy throughout the examination period. These results clearly indicate that unprotected pros-weanling cats brought into a leukemia exposure household environment have a high risk of becoming infected with FeLV. Furthermore, a large proportion of the cats are at risk for development of persistent viremia and FeLV-related diseases.


Treatment approaches to mania.

Treatment approaches to mania. Treatment of manic episodes is an urgency because of the lack of insight and excessive psychomotor activity. Over the past two decades various new modes of pharmacological treatment of this condition has been reported. Along with them, biochemical findings in manic patients have been exhaustively investigated. Etiologically based pharmacological treatment of manic episodes will help us therapeutically in improving the condition of the patient and scientifically, to confirm or explore biochemical basis of manic episodes. The new innovative approaches for the treatment of manic episodes need to be applied in treating therapy-resistant patients. Included in this paper are treatment approaches based on norepinephrine, dopamine, acetylcholine, serotonin, gamma-aminobutyric acid and permissive hypothesis, peripheral autonomic imbalance, endocrine abnormalities, electrolyte disturbances, paradoxical response, and cyclic AMP. In addition, use of antidepressants and pyrotherapy is described.


Prevention by medroxyprogesterone of perforation in the alkali-burned rabbit cornea: inhibition of collagenolytic activity. The incidence of perforation and deep ulceration of the alkali-burned rabbit cornea was substantially reduced by topical or parenteral administration of medroxyprogesterone. The mode of action of the hormone is probably at least in part related to its suppressive effects on production of tissue collagenase, as indicated by the considerable reduction in collagenolytic activity by living explants of the treated corneas.


The ocular anti-inflammatory action of imidazole. Imidazole administered intraperitoneally to albino rabbits at a dose of 250 mg. per kilogram inhibited the rise of aqueous humor protein concentration by approximately 50 per cent 30 minutes after paracentesis. Systemic imidazole administered daily to albino rabbits injected with intravitreal Shigella endotoxin decreased the conjunctival and iris hyperemia and reduced the anterior chamber cell and flare and the haziness of the optical media. Systemically administered imidazole had no effect on the aqueous humor concentrations of c-AMP or c-GMP in the rabbit. In vitro studies of rabbit ciliary body-iris phosphodiesterase activity indicated no effect of imidazole at a  concentration of 10(-3) molar.