Proteins induced in BHK cells by infection with foot-and-mouth desease virus. Although the infection of BHK cells with foot-and-mouth disease virus did not cause a marked inhibition of cellular protein synthesis, the proteins synthesized gradually changed from host-specific to virus-specific. The synthesis of at least thirteen virus-induced proteins was demonstrated by polyacrylamide electrophoretic analysis of the infected cells. Only a small proportion of the virus-induced proteins was incorporated into the mature virus particles. The addition of amino acid analogues caused an accumulation of the larger mol. wt. proteins, suggesting that in infected cells, some cleavages occurred, giving rise to the smaller mol. wt. proteins. The operation of the precursor-cleavage mechanism was more clearly shown in pulse-chase experiments, in which it was found that the proteins were degraded from larger to smaller mol. wt.


Determination of the molecular weight of bovine enterovirus RNA by nuclease digestion. The mol. wt. of a [32P]-labelled bovine enterovirus RNA has been determined by digesting with pancreatic RNase and separating the resulting oligonucleotides using a two-stage fractionation method on DEAE-Sephadex in 7 M-urea at pH 7-6 and pH 3-0. We have estimated the number of nucleotides as 8612 plus or minus 55. This corresponds to a mol. wt. of 2-93 plus or minus 0-02 X 10-6 which is in aggrement with estimates obtained by sedimentation and gel electrophoresis techniques.


The pathogenesis of pseudorabies in mice: virus replication at the inoculation site and axonal uptake. Three-week-old mice were inoculated in the right ear pinna with pseuforabies virus. Ears were surgically removed at various times after inoculation and changes from the normal pathogenesis were observed. Virus replication in the ear tissue and cervical dorsal root ganglia was also monitored. Followed inoculation with a small dose of virus, local multiplication of the virus was necessary before the infection spread to the nerves. With larger infecting doses there was probably direct uptake of virus from the inoculum into the nerve endings. After these larger doses virus was first detected in the dorsal root ganglia 17 h agter infection, suggesting a retrograde axonal flow rate of at lease 1-7 mm/h.


Virus DNA synthesizing ability of T antigen-forming defective SV40 produced by successive undiluted passages. SV40 stocks prepared by successive undiluted passages in African green monkey kidney (gmk) cells contain particles which produce T antigen but not capsid antigen (T partcles). The DNA of the T particles was shown to replicate during abortive infection of GMK cells, as indicated by immunofluorescence assay for antigen forming capacity of virus DNA. It was also shown that the synthesis of t particle DNA was induced in heterokaryons formed by fusion of 3T3 cells transformed by T particles with susceptible GMK cells.


Unidirectional replication of a minority of polyoma virus and SV40 DNAs. Poyoma DNA replication is initiated predominatly at a site which is 29% from the EcoRI cleavage site. Molecules replicating from this site, after digestion with EcoRI appear as linear structures with a double stranded loop centered at the origin of replication. These forms constitute 90% of all replicating intermediates. Approx. 10% of the replicating intermediates of polyoma and SVP40 DNAs occur as Y-forms after treatment with EcoRI. These structures have probably resulted from unidirectional replication initiated at an additional origin of DNA replication which is located near the EcoRI cleavage site on the genomes of these viruses.


Isolation and characterization of 'dense particles' from poliovirus-infected HeLa cells. Dense poliovirus particles (buoyant density 1.44 g/ml in CsCl) isolated from infected HeLa cells contain the normal four structural polypeptides VP1 to VP4, and 35S poliovirus RNA. In addition, small amounts of VPo and single-stranded RNA sedimenting slower than the poliovirus genome are present. Dense particles have a low specific infectivity, are neutralized by type-specific poliovirus antisera, and are detected during growth as early as normal virus but disappear when virus production stops.  They appear to represent a different, more open, conformation of the normal virus capsid.


Distinct molecular species of human interferons: requirements for stabilzation and reactivation of human leukocyte and fibroblast interferons. Human fibroblast interferon preparations were completely stabilized to 100 degrees C by sodium dodecyl sulphate (SDS) in the presence of mercaptoethanol, but only a minor fraction of their activities were stabilized by SDS without mercaptoethanol. On the contarary, human leukocyte interferon preparations were completely stabilized to 100 degrees C by SDS in the absence of mercaptoethanol, but only a minor fraction of their activities were stabilized by SDS in the presence of mercaptoethanol. Furthermore, human fibroblast interferon preparations whose activities had been destroyed by boiling at 100 degrees C were completely reactivated by SDS under reducing conditions, but only a minor part of their activities were restored by SDS in the absence of reduction. On the contrary, human leukocyte interferon preparations whose activities had been destroyed by boiling at 100 degrees C were completely reactivated by SDS in the absence of reduction, but only a minor part of their activities were restored by SDS under reducing conditions. These data suggest that there are distinct molecular species of human interferons.


Electroconvulsive shock and postsynaptic catecholamine effects: increased psychomotor stimulant action of apomorphine and clonidine in reserpine pretreated mice by repeated ECS. Male mice were administered electric convulsive shocks (ECS) once daily for one (ECS X I), three (ECS X III) or seven days (ECS X VII). One (day 1), three (day 3) or six days (day 6) later they received reserpine 10 mg/kg, followed 2 hours later by clonidine, 1.5 mg/kg, and/or apomorphine, 1.5 mg/kg. Control animals received the same drug treatments but no ECS. The motor activity was recorded for 1 hour, starting immediately after the last drug injection. The behavioral depression induced by reserpine was equally pronounced in ECS-pretreated animals as in the control animals. The psychomotor stimulant effects of clonidine and/or apomorphine, given after reserpine was, however, enhanced by pretreatment with ECS X VII, DAY 1. Pretreatment wit- ECS X VII increased the motor activity also in animals given reserpine, apomorphine and clonidine at day 3 or day 6. ECS X III was also effective in this respect at day 1, whereas ECS X I, day 1, was ineffective. Mice given ECS X VII but no drug treatment showed at day 1 an increased motor activity during the initial 10 min and a decreased activity during the last 10 of the 1 hour recording period. At day 3 and day 6 after ECS X VII the motor activity was increased in comparison to that in untreated control animals during the whole 1 hour period. Animals given repeated ECS furthermore showed increased irritability and reduced body weight. The results indicate that repeated ECS increase the sensitivity of postsynaptic catecholamine receptors in the brain or alter neuronal structures which are connected to these receptors.


Congenital granular-cell myoblastoma.

Congenital granular-cell myoblastoma. The clinical and pathologic features of congenital granular-cell myoblastoma in five infant girls are reported. One lesion, treated expectantly, progressively decreased in size and after 3 yr and 9 mo could not be detected, while two lesions which were imcompletely excised did not recur. It is suggested that congenital granular-cell myoblastoma is caused by an intrauterine stimulus, and that this stimulus may possible be production of estrogen by the fetus. Congential granular-cell myoblastoma should be treated expectantly or by limited excision, and has an excellent prognosis.


Papillary lymphoid hyperplasia of the terminal ileum: an unusual cause of intussusception and gastrointestinal bleeding in childhood. Papillary lymphoid hyperplasia of the terminal ileum is a benign condition associated with abdominal pain, intussusception, and gastrointestinal hemorrhage. It appears to represent a distinct clinicopathologic entity, separate from the usual idiopathic intussusception of infancy and childhood. The lesions are reasonably well circumscribed, localized in the submucosa of the terminal ileum, and composed of lymphoid tissue with prominent germinal follicles. Management by ileocolectomy resulted in complete cure with no postoperative complications in our six cases. However, many authors recommend conservative therapy. The cause is not known but there may be some relationship between these cases and intestinal adenovirus infection.


Dopaminergic mechanisms of opiate actions in brain. We have studied the interactions between morphine and a dopamine-blocking agent (haloperidol) and a dopamine precursor, L-3,4-dihydroxyphenylalanine (L-dopa). We found that haloperidol potentiated morphine-induced analgesia and enhanced morphine tolerance. Morphine-tolerant mice exhibited enhanced sensitivity to the locomotor excitatory actions of L-dopa. We propose that morphine exerts some of its central nervous actions by first interfering with dopamine-mediated synaptic transmission and then initiating compensatory changes that superficially resemble denervation supersensitivity. These compensatory changes may underlie the excitatory actions of morphine.


Experimental tests of hypotheses about neurochemical mechanisms underlying behavioral tolerance to the anticholinesterase, diisopropyl fluorophosphate. Neurochemical and psychopharmacological studies of rats were designed to examine four hypotheses which have been proposed to account for the development of behavioral tolerance to the anticholinesterase, diisopropyl fluorophosphate (DFP). The fact that the activity of the enzymes, adenosine triphosphatase, alkaline phosphatase and cytochrome oxidase, did not change concomitantly with behavioral measures during chronic treatment with DFP suggests that nonspecific metabolic changes are unlikely mechanisms of behavioral tolerance. Similarly, a lack of change in choline acetylase activity coupled with constantly high acetylcholine levels (140%) and low cholinesterase activity (28.5%) tends to eliminate end-product inhibition of acetylcholine synthesis as a primary mechanism of tolerance to DFP. Alpha-Methyl-p-tyrosine in doses to 150 mg/kg affected the behaviors of control and DFP-treated rats to a comparable degree, offering no support for the hypothesis that a redundant adrenergic system may replace the cholinergic system during the development of tolerance to DFP. In contrast to these various negative findings, pilocarpine was less effective in suppressing the responding of rats tolerant to DFP than that of control subjects. This confirms other evidence indicating that a decreased sensitivity of cholinergic (muscarinic) receptors is one mechanism underlying the development of tolerance to DFP.


Mechanism of blockade of neuromuscular transmission by pentobarbital. The mechanism of block of neuromuscular transmission by pentobarbital has been studied in the frog sciatic nerve-sartorius muscle preparation by means of intracellular microelectrode and voltage clamp techniques. The resting membrane potential was decreased by pentobarbital only to a small extent (less than 15 mV) in both end-plate and non-end-plate regions. Both sodium and potassium components of end-plate current underwent drastic changes after application of pentobarbital. The peak amplitude was decreased with an apparent dissociation constant of 0.9 mM for both currents. The maximum rate of rise of end-plate current was reduced, with apparent dissociation constants of 0.9 and 1.2 mM for sodium and potassium currents, respectively. The times for sodium and potassium end-plate current to reach their peaks were shortened only to a negligible extent. The falling phase of end-plate current was greatly accelerated, sodium current being affected more than potassium current. The transient end-plate depolarization induced by iontophoretic application of acetylcholine was suppressed more effectively than end-plate potential by application of pentobarbital. The falling phase of the former was also shortened. The quantal content of end-plate potential tended to increase at 0.5 mM, but underwent no appreciable change at 1.0 and 1.4 mM. Pentobarbital has a dual action on both quantal content and end-plate membrane depending on the concentration, and the block of neuromuscular transmission is due primarily to a suppression of the end-plate sensitivity to acetylcholine. The differential effect of pentobarbital on sodium and potassium components of end-plate current is compatible with the notion that these two ionic conductances are separate entities.


Hormonal influences in the development of the hypermetabolic state of the liver produced by chronic administration of ethanol. Chronic administration of ethanol to rats, either in liquid diets as the only source of food or by gastric intubation while the animals are fed ad libitum, leads to the development of a hypermetabolic state of the liver. This hypermetabolic condition of the liver can be observed independently of the feeding state of the animals. The calorigenic effects produced by ethanol in the liver, as measured in liver slices, could be reproduced by a single large dose of epinephrine. Oxygen consumption by liver slices of animals given a 2-mg/kg dose of epinephrine bitartrate increased by 40 to 50 percent. In these livers all the extra oxygen consumption, but not the basal respiration, could be abolished by ouabain, an inhibitor of the sodium pump. Dinitrophenol did not affect the respiratory rate in the liver of epinephrine-treated animals while markedly increasing that in controls. In the liver of treated animals, the activatory effect of dinitrophenol could be recovered in the presence of ouabain. The calorigenic effect of epinephrine in the liver was found to be completely abolished by phentolamine (alpha adrenergic blocker) but was not modified by DL-propranolol (beta adrenergic blocker). Also, the calorigenic effects produced by epinephrine could not be seen in thyroidectomized animals or by incubating the liver slices in a calcium-free medium. Thyroidectomy and administration of phentolamine markedly reduced and adrenalectomy completely abolished the hypermetabolic state produced in the liver of rats by chronic administration of ethanol.


Potentiation of carbon tetrachloride-induced hepatotoxicity in alloxan- or strepto- zotocin-diabetic rats. Studies were performed to examine the effects of alloxan- or streptozotocin-induced diabetes on carbon tetrachloride (CCl4) liver injury. Male rats were pretreated with single i.v. injections of alloxan monohydrate (40 or 80 mg/kg) or streptozotocin (65 mg/kg). A challenging dose of CCl4 (0.1 ml/kg i.p.) was given to rats 4 days after alloxan pretreatment or 5 days after streptozotocin pretreatment, and the animals were sacrificed 24 hours later. Biochemical and morphologic evidence was obtained to show that pretreatment with the diabetogenic agents markedly enhanced CCl4-induced hepatotoxity. The challenging dose of CCl4 had no effect on the serum glutamic pyruvic transaminase (SGPT) activity in control rats. However, the administration of this dose of CCl4 to rats pretreated with 40 and 80 mg/kg of alloxan as well as to rats pretreated with streptozotocin resulted in 11-, 68-, and 32-fold increases, respectively, in SGPT activity. Hepatic triglyceride concentrations in the diabetic rats were also markedly elevated above control values after CCl4 challenge. Alloxan- or streptozotocin-pretreatment alone did not enhance these biochemical parameters of liver injury. Hepatic glucose-6-phosphatase activity, which increased in the rats given a diabetogenic agent, was lowered as a result of CCl4 injection. Insulin treatment of rats given alloxan (80 mg/kg) markedly protected against CCl4-induced hepatotoxicity. The severity of the morphologic changes in diabetic rats given CCl4 correlated with the biochemical findings.


Influence of prostaglandins E1 and F2alpha on pulmonary vascular resistance, isolated lobar vessels and cyclic nucleotide levels. The effects of prostaglandins E1 (PGE1) and F2alpha) on the pulmonary vascular bed were studied in the intact dog under conditions of controlled pulmonary blood flow. PGF2alpha increased lobar arterial and venous pressure when injected or infused into the lobar, artery. The pressor response was dose-related and doses as low as 0.03 and 0.1 mug, which established concentrations of 0.1 to 0.3 ng/ml in lobar arterial blood, increased pulmonary vascular resistance. PGF2alpha also increased airway resistance in the left lower lobe. However, the effects of this substance on the vascular bed were not related to its effects on bronchomotor tone since similar pressor responses were observed in normal and nonrespiring lobes, PGE1 decreased pressure in the lobar artery and vein when infused into the lobar artery and the effects of PGE1 and PGF2alpha on the pulmonary vascular bed were similar when the lung was perfused with dextran or with blood. PGF2alpha increased isometric tension in isolated helical segments of lobar vein 3 to 5 mm in diameter but was without effect on arterial segments of the same diameter. The increase in isometric tension in the venous segments with PGF2alpha was associated with a significant increase in intracellular levels of guanosine 3',5'-monophosphate (cGMP) but no change in adenosine 3',5'-monophosphate (cAMP) levels. PGE1 decreased isometric tension in both arterial and venous segments and the decrease in tension was accompanied by a significant elevation in smooth muscle cAMP levels and a small but significant reduction in vein cGMP. Results of the present study indicate that PGF2alpha increases pulmonary resistance by constricting lobar veins and to a lesser extent vessels upstream in the precapillary bed whereas PGE1 dilates lobar veins and upstream vessels. These results suggest that PGE1-induced vasodilation may be mediated by an increase in cAMP levels while PGF2alpha-induced venoconstriction may be related to increased smooth muscle levels of cGMP.


Two descending nerve pathways activated by distension of guinea-pig small intestine. 1. Intracellular recordings were made from myenteric neurones of the guinea-pig small intestine in preparations which had a synaptic input from an orally situated segment of intestine. 2. Excitatory synaptic potentials could be evoked in most neurones by distension of the attached intestinal segment. 3. It was possible to distinguish two distinct firing patterns of synaptic potentials in response to distension. A transient short latency discharge was recorded from some neurones. From the others, a persistent synaptic discharge was recorded only after a long latency (2-11 sec). 4. Distension of intestinal segments evoked short latency transient inhibitory junction potentials in the circular muscle layer followed by excitatory junction potentials in both the circular and longitudinal muscle layers. 5. It is suggested that distension may cause both descending inhibition and, after a delay, descending excitation of the guinea-pig small intestine.


Estimates of statistical release parameters from crayfish and frog neuromuscular junctions. 1. Transmitter release at crayfish and frog neuromuscular junctions was studied by recording synaptic potentials with extracellular or intracellular glass micro-electrodes. 2. The binomial release parameters n and p were calculated using the experimental observations for the mean number of quanta (m) released in a series of trials, the variance (var) of the quantum content distribution and the number of transmission failures (n-o). 3. In one series of experiments on frog neuromuscular junction, action potentials were blocked by tetrodotoxin. Transmitter release was limited to a circumscribed part of the nerve terminal by focal stimulation with a glass micro-electrode. Values for m were between 0-6 and 5-7 and calculated values for n ranged from 3 to 23. p was between 0-05 and 0-48. In a second series of experiments on frog neuromuscular junction transmitter release was evoked by conventional stimulation of the nerve trunk in Mg-Ringer. m was mostly between 20 and 60 and in the same experiments n was calculated to be between 70 and 125. Values for p ranged from 0-25 to 0-48. 4. It can be concluded from the present results that transmitter release at frog neuromuscular junction is described by binomial statistics as is the case for neuromuscular junction of crayfish. 5. In comparing the values for n for the two series of experiments on frog neuromuscular junction it appears that n is dependent on the length of the activated synaptic contact. The numbers for n calculated from frog neuromuscular junction are of the same order of magnitude as the number of 'active zones' seen in the corresponding length of the synaptic nerve terminal.


Nursing staff training for an effective geropsychiatric environment. In describing an early training program of the Gerontological Manpower Project, we have attempted to demonstrate that short-term, on-site training with a frim theoretical and empirical base, can be an effective vehicle for translating research into application. In this case, research and theory based on the importance of the environment as therapeutic, and directed to maximizing geriatric patient independence was incorporated into a geriatric training programmthis was delivered to personnel working with the elderly in two institutional settings, differing in geriatric patient population and service orientation. To improve the effectiveness of future training programs, extensive evaluative research was included in the training processes. Although results were predictably tentative, they did yield much useful information both for subsequent program design as well as reinforcing a need for this process itself. Finally, a very satisfying and beneficial result of this project, as specified by the research staff and all participants, is that effective application of research results via appropriate training mechanisms are easily within reach of the nurse-practioner, and the oft-desired bridge between academia and the service delivery system is truly a collaborative reality.


The dysfunction of DSM-II (Diagnostic and Statistical Manual of Mental Disorders). The task of seeking alternatives to a classification system such as DSM-II that will maximize its benefits and minimize its limitation is a complex and arduous one. There are no clear-cut answers. Certain issues had been elaborated on here. The task at hand is one that must be approached on a multiprofessional basis seeking to integrate a variety of theoretical bases into a functional conceptual framework that will accomplish the purpose of a classification scheme. There is also need to conduct systematic data collection in an effort to determine operational definitions related to the phenomenon that is being studied. There is a movement by some professionals to have more emphasis placed on the social functioning level of the individual and his significant othersmif a concerted effort is not begun to look at this problem it will remain unchanged for the most part or other interested professionals will have changed it with little impact from nursing.


Stimulation of protein synthesis in vivo in immature mouse testis by FSH. Human pituitary FSH was found to increase the incorporation of tritiated lysine into testicular protein in prepubertal mice in vivo. Radioactivity was measured in washed trichloracetic acid precipitates prepared from crude testicular homogenates. The time of maximum response was 8 to 16 hr after subcutaneous injection of the hormone. This was considerably later than the maximum response in vitro reported by other workers. Neither HCG nor dibutyryl cyclic 3',5'-adenosine monophosphate had a significant effect on the incorporation of lysine.


Ultrastructural relationships between decidua, trophoblast and lymphocytes at the beginning of human pregnancy. Between the 20th and 40th day of pregnancy, human decidual cells in the endometrial connective tissue come into direct contact with the trophoblastic cells eroding the stromal tissue. Fibrillar deposits of maternal origin were randomly distributed between the maternal and fetal sites and did not form a continuous barrier. Structural features were prominent in the maternal and embryonic cells. Large decidual cells were intermingled with trophoblastic cells, and there was little intercellular material. Small developing decidual cells arose around the maternal precapillary arterioles and capillaries and were closely accompanied by lymphocytes. Fully developed decidual cells exhibited a fine structure characteristic of a state of active secretion, and trophoblastic cells gave structural indications of producing protein. The aggregates of decidual and trophoblastic cells in the human endometrium during early pregnancy ('deciduotrophoblastic complex') may function as a local humoral regulator releasing chemical factors and partaking in the establishment of immunological privilege at implantation.


The coagulation of insoluble and basic proteins from rat siminal vesicle secretion with vesiculase: influence of collagenase-like peptidase from rat testis. Proteins from the secretion of rat seminal vesicles were fractionated to yield (i) insoluble protein, (ii) basic proteins (soluble in the presence of urea) and (iii) neutral plus acidic proteins. The first two are necessary for the formation in vitro of a coagulum which is very similar to the copulatory plug. Collagenase-like peptidase from rat testis degrades both protein fractions and prevents coagulum formation.


Implantation in ovariectomized mice treated with dibutyryl adenosine 3',5'-monophosphate (dibutyryl cyclic AMP). Experiments are described that demonstrate that uterine intraluminal injection of a 1-25 mM-solution of dibutyryl cyclic AMP (dcAMP) in phosphate buffered saline (PBS) induced implantation in ovariectomized pregnant mice. Pretreatment with progesterone was essential for this effect. When PBS was injected alone, it did not induce implantation in mice treated with progesterone. Bilateral adrenalectomy had no effect on the ability of dcAMP to substitute for oestradiol, showing that the effect was not due to dcAMP-induced oestrogen synthesis in the adrenal cortex. It is suggested that the dcAMP may act at the level of the uterus, the embryo, or both.


Potential inhibitors of L-asparagine biosynthesis. 2. Chemistry and biological activity of beta-hydroxyaspartic acid and its derivatives. Several derivatives of erythro-beta-hydroxy-DL-aspartic acid (1) were prepared as a potential inhibitors of L-asparagine synthetase (ASase) from rat Novikoff hepatoma. Benzylation of 1 gave the dibenzyl ester 2 which upon coupling with carbobenzoxyglycine afforded the blocked dipeptide 3. Deblocking of 3 gave glycl-erythro-beta-hydroxyl-DL-aspartic acid (4) which could not be diazotized. The dimethyl ester of 1 was coupled with carbobenzoxyglycine to give the blocked dipeptide 7a which was deblocked to give dimethyl glycel-erythro-beta-hydroxy-DL-aspartate hydrochloride (8). Diazotization of 8 gave impure diazo compound 9 which on reaction with HCl gave the chloro compound 10. The methods of isolation, assay, and inhibition of ASase are discribed. At 10 mM concentrations 10, 1, and its D and L enantiomers inhibit ASase by 45, 47, 36 and 66 percent, respectively.


Adrenocorticotropin. 47. Synthesis and biological activity of adrenocorticotropic peptides modified at the tryptophan position. Three biologically active peptides, [9-beta-(1-naphthyl)alanine]-ACTH-(1-19), [9-Ni-formyltryptophanl-ACTH-1(1-19), and (8-lysine,9-phenylalaninel-ACTH-1(1-9), have been synthesized by the solid-phase method. All of the synthetic peptides showed diminished biological activity compared to ACTH-(1-19). It was also shown that the steroidogenic and lipolytic activities of ACTH-(1-19) were not inhibited by [8-lysine,.-phenylalanine]-ACTH-(1-19).


Distinction between Duchenne and other muscular dystrophies by ribosomal protein synthesis. Ribosome concentration, ribosome distribution on sucrose density gradients, and in-vitro ribosomal amino-acid incorporation (noncollagen and collagen synthesis) were studied in muscle biopsy samples obtained from 30 patients with Duchenne muscular dystrophy, seven patients with Becker muscular dystrophy, and 10 with facioscapulohumeral muscular dystrophy. Ribosome concentration was normal in Duchenne and facioscapulohumeral and decreased in Becker muscular dystrophy. Distribution of ribosomes in sucrose density gradients showed abnormalities (sharp monosomal peak and fewer polyribosomes) only in Duchenne muscular dystrophy and was normal in the other two types. In-vitro amino-acid incorporation of ribosomes in Duchenne muscular dystrophy revealed high collagen and low noncollagen synthesis of the heavy polyribosomes. This abnormality is controlled by an undetermined enzymatic factor belonging to the soluble enzyme fraction. Supplementation of the dystrophic heavy polyribosomes with normal soluble enzymes restored the synthesis of collagen to that of the controls. Heavy polyribosomes extracted from normals or from carriers produce proportionately more collagen in the presence of soluble enzyme fraction from Duchenne muscular dystrophy than in the presence of their homologous enzymes. In Becker muscular dystrophy, both noncollagen and collagen synthesis of the heavy polyribosomes were increased, under the influence of ribosomal factors. The different protein synthesis in Duchenne and Becker muscular dystrophies suggests that these conditions are non-allelic. In facioscapulohumeral muscular dystrophy the changes in protein synthesis occurred only in the early stage of the disease and consisted of increased noncollagen synthesis of the light polyribosomes, while the heavy polyribosomes had normal activity including collagen synthesis. This reaction was controlled by ribosomal factors.


Bilayer structure in phospholipid-cytochrome c model membranes. Lipid protein interactions in biological membranes differ markedly depending on whether the protein is intrinsic or extrinsic. These interactions are studied using lipid spin labels diffused into model systems consisting of phospholipid bilayers and a specific protein. Recently, an intrinsic protein complex, cytochrome oxidase, was examined and the data suggest there is a boundary layer of immobilized lipid between the hydrophobic protein surfaces and adjacent fluid bilayer regions. In the present study, a typical extrinsic protein, cytochrome c, was complexed with a cardiolipin/lecithin (1:4 by weight) mixture. The phospholipids in the presence and absence of cytochrome c exhibit typical bilayer behavior as jedged by four spin-labeling criteria: fluidity gradient, spectral anisotropy of oriented bilayers, response to hydration and the polarity profile. Any effects of cytochrome c on the ESR spectra of lipid spin labels are small, in contrast to the effects of intrinsic proteins. These data are consistent with electrostatic binding of cytochrome c to the charged groups of the phospholipids, and indicate that the presence of extrinsic proteins will not interfere with measurements of boundary lipid in intact biological membranes.


Membrane proteins and urea and acetamide transport in the human erythrocyte. Previous studies have shown that urea and acetamide traverse the erythrocyte membrane by way of facilitated diffusion. The nature of this selective pathway is unknown. The present studies investigate the effects of proteolytic enzymes and crosslinking agents on amide transport. Cleavage of the erythrocyte membrane surface by pronase or trypsin had no effect on urea and acetamide permeability or inhibition by phloretin. These findings suggest that the sialoglycopeptide segment of the sialoglycoproteins is not critical to urea and acetamide transport. In addition, extensive crosslinking of membrane proteins with glutaraldehyde had no effect on amide transport in the absence or presence of phloretin.


Isolation and characterization of the plasma membrane from Yoshida hepatoma cells. Plasma membranes isolated from Yoshida ascites hepatoma AH-130 by a modification of the method of T.K. Ray (Biochim. Biophys. Acta 196:1, 1970), were subfractionated into three fractions having densities (d) 1.12, 1.14 and 1.16 by discontinuous sucrose density-gradient. Membrane subfractions were characterized by electron-microscopy, by assay of marker enzymes and by lipid composition. All subfractions appeared to be essentially free from whole mitochondria, lysosomes and nuclei. Subfraction d 1.16 had the highest 5'-nucleotidase, Mg++-ATPase and (Na+ +K+)-ATPase activities; cytochrome c oxidase was undetectable in any fraction and glucose-6-phosphatase was measurable only in fraction d 1.14 and 1.16. Cyclic AMP phosphodiesterase was nearly equally distributed in the fractions. Adenylate cyclase, 5'-nucleotidase and Mg++-ATPase activities of tumor membrane were lower with respect to liver plasma membrane, while cyclic AMP phosphodiesterase and (Na" +K+)-ATPase were found to have similar activities in the two membrane preparations. With respect to liver membrane, hepatoma membrane contained a higher amount of glycolipids and a higher amount of phospholipids accounted for mainly by sphingomyelin, phosphatidylserine and phosphatidic acid. The possible significance of the decrease of adenylate activity in the hepatoma membrane is briefly discussed.


Primary tissue culture of human wart-derived epidermal cells (keratinocytes). The formation of human skin warts may depend on transformation by human papilloma virus or result from a productive virus infection. To explore this problem, methods were developed for the primary tissue culture of epidermal cells (keratinocytes) derived from wart tissues and normal skin. Wart-derived keratinocytes continued to proliferate for periods exceeding 3 months, but the patterns of cell growth were identical to normal keratinocytes. Electron microscopic and immunohistologic examination of wart-derived cultures did not evidence virion assembly.


Differences between soluble complement-fixing antigens derived from P3HR-1, RAJI, and Hk-Ly-28 cell lines. Soluble complement-fixing (CF) antigens of the virus-producing and virus-nonproducing P3HR-1 and RAJI Burkitt lymphoma cell lines and Hk-Ly-28 nasopharyngeal cancer cell line, identified by use of viral capsid antigen-positive human sera, could readily be distinguished by differences in their stability to chemical and physical conditions. The P3HR-1 soluble CF antigen lost titer when heated or exposed to acid perchlorate under conditions in which RAJI and Hk-Ly-28 soluble antigens were stable. Differences in solubility were also found. These results contribute not only to the interpretation of the relationship of Epstein-Barr virus (EBV)-associated components of RAJI to P3HR-1 and Hk-Ly-28 cell lysates but also to the selection of conditions for the development of identification tests and purification procedures for CF antigens and antibodies of Burkitt's lymphoma, nasopharyngeal cancer, and other EBV-associated tumors.


Pathogenic and serologic studies of Japanese quail infected with JM strain of Marek's disease herpesvirus. Japanese quail were inoculated with Marek's disease herpesvirus at 1 day of age, and a second group of the same age was infected by contact with inoculated quail. A third group raised in our laboratory as a source of hatching eggs was accidentally infected with Marek's disease (MD). All quail were observed and examined for periods up to 360 days; the usual clinical signs of MD were not observed. However, the birds were susceptible to MD infection because of the evidence that: 1) The MD-specific agar gel-preceipitation (AGP) antigen was found in the feather tips or feather follicular epithelium of some of them; 2) the MD-specific AGP antibody was found in 6 of 57 exposed to MD; and 3) the virus was recovered from 4 MD-exposed birds by subpassage of their kidney cultures into quail embryo fibroblasts. The specificity of the AGP antigen or antibody in MD-infected quail was examined by the line of identity and cross-absorption tests. The antigen specificity was tested against similar to 700 sera from chickens and quail. Although most of these sera reacted with quail-positive antigen (QPA) prepared from MD-infected quail, some sera from infected or uninfected normal chickens reacted nonspecifically with QPA and with an antigen prepared from normal quail. By gel filtration on Sephadex G-150, the specific antigenic component could be separated from the nonspecific component(s) in QPA.


Studies of FBJ osteosarcoma virus in tissue culture. II. Autoinhibition of focus formation. A high-titered non-focus-forming virus, FBJ-MuLV (murine leukemia virus), present in FBJ tumor preparations, inhibited significantly the expression and production of FBJ-MuSV (murine sarcoma virus) in tissue culture. This "autoinhibition" was comparable to that observed when a 3- to 4-log excess of infectious MuLV was added to standard MuSV. The degree of inhibition was influenced by the tropism of the MuLV (or the ease of spread and propagation of MuLV in certain cells), multiplicity of infection by MuLV, amount of excess MuLV, and ability of the MuSV-transformed cells to replicate independently. The FBJ MuLV-MuSV complex may be a model system for the detection of sarcoma viruses in spontaneous tumors in various animals where inhibition by excess nontransforming virus could be an important biologic phenomenon.


Feline oncornavirus-associated cell membrane antigen. II. Antibody titers in healthy cats from household and laboratory colony environments. Antibody titers to the feline oncornavirusassociated cell membrane antigen (FOCMA) were determined for 447 healthy cats from laboratory colony and household environments. Only 2.7 percent of 221 cats from colony environments were antibody positive as compared to 50.4 percent of 256 cats from household environments. Incidence of FOCMA antibody and geometric mean antibody titer for pet cats from New York City representing single cat apartment habitats were substantially lower than values for unscreened cats from the Boston, Glasgow, and Detroit suburban environments. Geometric mean antibody titer for young adults in the Boston population was significantly higher than titers for kittens or aged cats. This may be due to greater mobility resulting in increased exposure to other cats. In contrast to the high frequency of FOCMA antibody positivity in pet-cat populations, less than 2 percent of the same groups were positive for virus group-specific antigen in peripheral blood leukocytes and platelets. This was interpreted as an indication that many more cats became infected with feline leukemia virus under natural conditions that the number developing persistent virus infection and/or clinical leukemia.


Feline oncornavirus-associated cell membrane antigen. III. Antibody titers in cats from leukemia cluster households. Serum samples from 182 healthy cats residing in environments known to have a natural exposure to feline leukemia virus (FeLV) were examined for the presence of antibody to the feline oncornavirus-associated cell membrane antigen. The number included 138 cats from leukemia-"cluster" households. Such cats previously showed greatly increased frequencies of FeLV infection, as determined by the virus group-specific antigen (gasa) in peripheral blood leukocytes and platelets. The geometric mean antibody titer for all 182 cats with known exposure to FeLV was 4.69, more than four times higher than the mean titer for healthy pet cats from the same geographic areas but with no known FeLV exposure. About 92 percent of the cats in exposure environments were positive for FeLV gsa or feline oncornavirus-associated cell membrane antigen (FOCMA) antibody, and gsa-positive cats had lower FOCMA antibody geometric mean titers than gsa-negative cats. In the exposure environments, no differences were seen for cats of different sexes, but a higher geometric mean antibody titer was observed for cats 5 years and over when compared to younger groups. These results prove that previous reports of increased incidences of leukemia in cluster households were not due to chance alone, but rather to increased infection rates of cats in the FeLV environment. Also, they suggest that horizontal transmission of FeLV is highly efficient under crowded conditions, and that most cats naturally exposed to virus elicit an active immune response.


Infection of capuchin monkeys (Cebus albifrons) with Herpesvirus saimiri. Herpesvirus saimiri (HVS) induced persistent, clinically inapparent infections of long-term duration in capuchin monkeys (Cebus albifrons). The infections were characterized by development of antibody to HVS-associated antigens and recovery of low levels of virus-genome-carrying lymphocytes in the peripheral blood. Peripheral lymphocyte counts remained in low-normal to normal ranges and no physical signs of lymphoma were evident. Prednisolone treatment caused immunosuppression in one monkey; this was accompanied by a progressive loss of humoral antibody to HVS-associated antigens, but neoplastic disease did not develop.


Depression in lymphocyte response to general mitogens by owl monkeys infected with Herpesvirus saimiri. The effect of Herpesvirus saimiri (HVS) infection on the response of owl monkey lymphocytes to general mitogens was examined during the development of neoplastic disease. The reactivity of the lymphocytes was then correlated with the clinical condition of the infected monkeys and the content of virus rescued from the peripheral blood lymphocytes (PBL). Eight monkeys developed lymphoma which, in six monkeys, was accompanied by lymphocytic leukemia. All animals that died of HVS-induced neoplasia consistently showed a lack of mitogenic response to phytohemagglutinin and concanavalin A. The response to pokeweed mitogen, while always reduced, was generally less markedly affected than the response to the other two mitogens. Lymphocytes from five of the leukemic animals demonstrated an elevated level of spontaneous DNA synthesis in culture late in the disease. This increased spontaneous DNA synthesis tended to correlate with the rescue of HVS from the PBL as demonstrated by the infective center assay. Although mitogenic hyporesponsiveness corresponded with HVS rescue from PBL in six of nine monkeys, the impairment of normal lymphocyte responsiveness sometimes preceded virus recovery.


Effect of bacillus Calmette-Guérin immunization in marmosets infected experimentally with Herpesvirus saimiri. Eight white-lipped marmosets immunized with BCG and 3 sham-immunized marmosets were studied after inoculation with Herpesvirus saimiri (HVS). BCG immunization had no significant influence on the incidence of infection by HVS, incidence of fatal malignant lymphoma, time of leukemia onset, development or titer of HVS antibodies, or average survival time. One BCG-immunized, HVS-infected marmoset failed to develop malignant lymphoma, whereas the remaining 10 HVS-infected marmosets died of malignant lymphoma. Prolonged survival occurred also in 1 marmoset immunized with BCG 100 days after HVS inoculation. The development and disappearance of lymphocyte reactivity to tuberculin were followed in 4 BCG-immunized marmosets.


Virus expression and immunoprophylaxis of a murine lymphoma. CBA mice tested with a rapidly lethal transplanted lymphoma could survive challenge when pretreated with allogeneic lymphoma cells or other material expressing murine leukemia virus (MuLV). Protection resulted when recipients were given injections of AKR and C3H transplanted lymphomas or selected normal AKR and SJL tissues, filtrates of which give positive assays for MuLV by in vitro and/or in vivo tests. There was no protection when recipients were given injections of C3H lymphomas or C3H normal tissues that failed to have positive assays for virus.


Antitumor activity of bacterial infection. II. effect of Listeria monocytogenes on growth of a guinea pig hepatoma. Growth of a guinea pig hepatoma was suppressed when tumor cells were mixed with viable Listeria monocytogenes (LM) before intradermal (id) injection into syngeneic recipients. Heat-killed LM were less effective than viable organisms in suppressing tumor growth. A vaccine containing oil droplets and LM cell walls lacked antitumor activity. Intratumor injection of viable LM on the 7th day after id injection of tumor cells prolonged survival of guinea pigs that did not succumb to LM infection. After intratumor injection of 0.6 times 10-8-1.0 times 10-8 LM, 5 of 22 guinea pigs died from acute infection (23 percent). In the 17 survivors, 3 tumors regressed completely (18 percent). Animals surviving injections of LM and tumor cells were immune to a second challenge with tumor cells. Immunization ofguinea pigs with an intravenous injection of LM decreased the mortality from intratumor injection of LM, but the intratumor injection of LM failed to cure a significant fraction of LM-immune animals bearing 7-day hepatoma transplants. BCG was more effective than LM in producing tumor regression. Synergism between LM and BCG was not observed, and simultaneous intratumor injection of BCG and LM was no more effective than intratumor injection of BCG alone in the treatment of 12-day tumor transplants.


Aberrant viruses in cells infected with murine sarcoma virus-feline leukemia virus. This study describes an unusual type of virus seen when a feline leukemia virus (FeLV) pseudotype of murine sarcoma virus (MuSV) obtained by cocentrifugation procedures infected feline embryo cells (FEF) and two Crandell cat cell lines (CrFK1, CrFK2). When all three cell cultures were infected with MuSV-FeLV, only FEF and CrFK2 were transformed and only these showed normal and aberrant virus. The CrFK1 infected with MuSV-FeLV did not transform but did replicate normal type-C virus with a 50-A intermediate coat. The virus replicated in the two transformed lines showed three particles; a normal particle with a 50-A intermediate coat, a normal particle with a 100-A intermediate coat, and an aberrant particle with a 100-A intermediate coat.


Moloney sarcoma virus-induced tumors in athymic (nude) mice: growth pattern and antibody responses. Tumor growth and antibody production were evaluated in nude (nu/nu) mice and their heterozygous normal (+/nu) littermates after inoculation of Moloney sarcoma virus (MSV). Sarcoma-bearing nude mice developed progressively growing tumors, whereas 53 percent of their normal counterparts showed tumor regressions. By indirect membrane fluorescence, significant amounts of IgG antibody to MSV could be detected in thymus-bearing but not in nude mice.


Intercurrent cytomegalovirus colitis in a patient with ulcerative colitis. Acute intercurrent CMV colitis developed in a patient with UC who was receiving prednisone.  CMV infection was suggested by light and electron microscopic study of a rectal biopsy taken during the acute episode and was confirmed by serology done nine months later.  The microscopic studies of plastic-embedded tissues demonstrated that infected cells were concentrated in a subendothelial location and were presumably macrophages.  Epithelial and endothelial cells were not involved.  Steroid therapy and the inflammation and repair process (granulartion tissue) of active UC may have predisposed the present patient to CMV colitis.  CMV infection has been reported to be more common in patients with UC than in the general population.  Detection of CMV colitis in patients with UC could be of special importance since alteration of immunosuppressive therapy may be indicated.


Renal action of cholera toxin: II. Effects on adenylate cyclase-cyclic AMP system. The effects of cholera toxin (CT) on the adenylate cyclase-adenosine 3',5'-cyclic monophosphate (cAMP) system(s) in renal cortex were examined using the isolated renal cortical tubules of rat. Unlike parathyroid hormone, catecholamines or prostaglandins, CT had no immediate effects on cAMP production by the tubules or on adenylate cyclase activity. However, after 30 min of incubation at 37 C, cAMP production by the tubules started to rise and reached a plateau between 60 and 90 min. This rise in cAMP production was not abolished by protein synthesis inhibitors (actinomycin D and cycloheximide) nor by the inhibitors of prostaglandin synthesis (acetyl-salicylate and indomethacin). Repeated washings of the tubules exposed to the toxin for five minutes at 0 or 37 C did not abolish the effect of CT to stimulate cAMP production. Assays of adenylate cyclase activity using homogenates prepared from isolated tubules which were incubated for 60 min with CT revealed an increase in the basal adenylate cyclase activity without any change in NaF-sensitive enzyme activity. It is concluded that CT binds to renal tubule cells rapidly, possibly through energy-independent process. CT stimulates adenylate cyclase activity and increases cAMP production by the renal tubule cells after a latent period of 30 min. The stimulatory effects of CT are not due to new protein synthesis or prostaglandin formation.


Effect of hypotonicity on cyclic adenosine monophosphate formation and action in vasopressin target cells. We have shown previously that overhydration of toads renders their urinary bladders less responsive to the antidiuretic action of vasopressin (AVP). The present study investigates the relationship between osmotic swelling of vasopressin target cells and their sensitivity to AVP and dibutyryl cyclic adenosine monophosphate (db-cAMP). Conditions which engender osmotic swelling of toad bladder epithelial cells, such as immersing bladders on both surfaces in hypotonic Ringer's fluid or subjecting them to a net mucosal-to-serosal volume flux, markedly inhibited the effectiveness of db-cAMP in raising bladder permeability to water. This inhibitory phenomenon was seen both with serosal and mucosal applications of the nucleotide. Examination of isolated epithelial cells by phase contrast microscopy showed them to behave as osmometers, doubling their volume as the effective osmolality of the incubation medium was halved. AVP was found to increase the total content of cAMP about 3.5-fold both in the swollen and the normal cells, so that the actual concentration of cAMP may have diminished as the cell volume increased. Consistent with this suggestion was the observation that increasing exogenous db-cAMP abolished, in part, the inhibitory effects of hypotonicity. These observations indicate that homeostasis of body fluids in the toad depends in part upon the osmotic regulation of anti-diuretic homone action, and that intracellular cAMP may participate in coupling changes in cell volume to the altered state of responsiveness of the vasopressin target cell.


[Age-dependent biochemical studies on the extrarenal effect of furosemid (author's transl)]. The present study investigated the activities of lysosomal enzymes of the liver after administration of Furosemid. 10 weeks and 1-year old male albino rats were treated with 40 mg Furosemid for 4 subsequent days. According to the method devised by de Duve a sediment rich in lysosomes was produced by fractionated centrifugation and subsequently the enzyme activity of beta-glucuronidase, beta-acetylglucosaminidase, cathepsin D and a collagenolytic enzyme was measured in the sediment as well as in the corresponding lysosomal supernatant. The protein content served as a reference for the enzyme activities. In addition, we investigated the activities of cytoplasmic enzymes such as GOT, GPT, gamma-GT and the alkaline phosphatase. The enzyme activity changes were age-dependent. With Furosemid treatment the activities of beta-glucuronidase and cathepsin D increased in the lysosomal supernatant and the lysosomal sediment of the 1-year old rats, whereas the activities of the collagenolytic enzyme increased in the lysosomal sediment of the same group. In the lysosomal sediment of the 10-weeks old rats a decrease of beta-glucuronidase, beta-acetylglucosaminidase and cathepsin D was observed. These results are discussed in the light of reports from the literature.


[Carcinofetal antigens. I. alpha-fetoprotein (author's transl)]. Alpha-fetoprotein (AFP) is an alpha1-glycoprotein (M.W. about 65000) appearing in the fetal serum of most mammals including man during the early stages of pregnancy; 4 weeks after birth it disappears altogether or exists at very low concentrations as in the normal adult. AFP is formed in the yolk sac, the fetal liver and the gastro-intestinal tract. One of its physiological functions in fetal life is supposed to be the protection of the fetus from maternal oestrogens (oestrophilic property). The clinical significance of AFP is based on the regular and increasing production in primary liver cell carcinoma, less frequently in teratogenetic tumors where it serves as a control of therapy and course of the disease. Less frequent, minor and temporary increases in the AFP serum level occur in several primary tumors with secondary liver involvement, and in inflammatory gastro-intestinal diseases, e.g. of the liver (hepatitis, cirrhosis). AFP has an increasing importance in gynecology (gestational age, fetal distress syndrom, malformations, hydatidiform mole/chorion carcinoma). The physico-chemical properties of AFP are widely known. Both fetal and tumor AFP appear to be immunologically and biochemically identical, as are that of tissue and biological fluids. The differences observed (variants, microheterogeneity) depend mainly on the different content of sialic acid. An antigenetic relationship exists, between the AFP of most species. The immunodiffusion (Ouchterlony) is the most frequently used but relatively insensitive test (1-5 mug/ml) in finding AFP, whereas the radioimmunoassay is the most sensitive one (up to 0,25 ng/ml) and permits the determination of normal serum levels in adults (below 20 ng/ml). The serum concentration in healthy pregnant women lies up to 500 ng/ml, in patients with hepatitis, liver cirrhosis and other liver diseases mostly under 3 mug/ml, whereas in those with primary liver cell carcinoma levels up to and above 600 mg-percent have been found.


[Gas exchange and blood indices in thyroid gland dysfunction]. An exposure of intact rats to a hyperoxic atmosphere of 2 atm for one hour a day during 20 days induced no changes in the gas exchange or hemoglobin and red blood cells content. The concentration of free radicals tended to decline. Thyroidectomized rats kept in a normoxic atmosphere showed anemia and gas exchange decrease. The concentration of free radicals in the tissues declined slightly. These parameters returned to the norm after the hyperoxybaric exposure. During a normoxic exposure thyroxine injections accelerated gas exchange, producing no significant changes in the red blood composition. The free-radical activity tended to increase. After a hyperoxybaric exposure similar changes took place.


Short term culture of dissociated rat submandibular gland cells. Submandibular glands of 4-week-old rats were dissociated by a procedure involving digestions with collagenase and hyaluronidase, chelation with ethylenediaminetetraacetic acid, and mechanical force. The isolated cells were purified by centrifugation in a Ficoli solution and were maintained in culture for 36 hours. On the basis of trypan blue exclusions, about 70 per cent of the  dissociated cells were viable. Electron microscopic observations indicated that the isolated acinar cells and intercalated and striated duct cells retained their essential in situ ultrastructural characteristics. During a 36-hour culture period the number of viable cells declined to about 40 per cent, and the various cell types formed mixed aggregates. The ultrastructural features of the intercalated and duct cells changed relatively little, but the acinar cells revealed several structural alterations. These included a decrease in the number of the secretory granules, fusions of the secretory granules, and an increase in the rough surfaced endoplasmic reticulum. In general, the polarity of acinar cells became less distinct. The endogenous peroxidase activity in the acinar cells gradually diminished during the culture. Isoproterenol when added to the cultured cells failed to stimulate the incorporation of radioactive thymidine or the discharge of the secretory material from the acinar cells.


Dissociation of rat parotid gland.

Dissociation of rat parotid gland. Rat parotid gland was dissociated by sequential collagenase and hyaluronidase digestions, chelation with ethylenediaminetetraacetic acid, and mild shearing force to yield predominantly single cells. The isolated acinar cells retained their morphologic characteristics and their amylase activity. The functional integrity of the isolated cells was assessed by measuring their secretory response to isoproterenol, epinephrine, and carbamylcholine and by their ability to incorporate radioactively labeled leucine and thymidine. The discharge of amylase from the dissociated cells was not effected by isoproterenol or norepinephrine and the response to carbamylcholine was minimal. The data indicate a destruction or perturbation of hormone receptors during the dissociation procedure. The maintenance of the cells in culture for up to 18 hours failed to restore the responsiveness of the isolated parotid gland acinar cells to isoproterenol. The isolated cells incorporated 14C-leucine into proteins at a linear rate between 30 and 180 minutes. Chromatographic and electrophoretic profiles of newly synthesized proteins indicated that all major proteins synthesized in vivo were also synthesized by the isolated cells. The isolated cells incorporated tritiated thymidine into DNA. Furthermore, stimulation of DNA synthesis by isoproterenol in vivo was reflected by a higher rate of thymidine incorporation by the isolated cells as compared with controls. The dissociated parotid gland cells offer a convenient system for studying various cellular processes, particularly the synthesis of macromolecules with high specific activity. However, some functions, notably the response to beta- adrenergic agonists, are lost during the dissociation procedure.


Viruses: causative agents of cancer.

Viruses: causative agents of cancer. Evidence accumulated over a period of 60 years has clearly established that a number of different viruses cause neoplasia (of a broad spectrum) amongst a variety of animals. Although it is known that viruses do produce self-limiting proliferative human diseases, complete verification of a human viral carcinogen remains to be provided. This paper presents an overview of the conceptual and practical tools for the detection of tumor viruses, which have derived from experimental model systems. It indicates how these are being applied to studies of human malignant disease, with special reference to assessing a possible viral etiology for laryngeal cancer.


Development of hyperbetalipoproteinemia in pigs fed atherogenic diet. Hormel miniature pigs were studied over a period of 24 weeks to observe the changes in serum lipoprotein pattern, cholesteryl ester, free cholesterol, and triglyceride in the atherogenic-fed pigs. These pigs were compared to age-related control animals in our breeding herd. Pigs fed the atherogenic diet (20% tallow, 3% cholesterol, and a bile supplement) exhibited a heterogeneous response but showed mean increases in cholesteryl ester (571 mg/dl) and free cholesterol (226 mg/dl), a slight increase in triglyceride (58 mg/dl), and a severe hyperbetalipoproteinemia. Three animals with the highest cholesteryl ester (all above 600 mg/dl) had resolvable beta components in their 1.006 g/ml very low density lipoprotein fraction (type III), as well as huge increases in the Sf 12-20 low density lipoprotein fraction. The other four animals had substantial increases in Sf 0-20, and the three highest had much of their low density lipoprotein in the Sf 12-20, or "remnant" fraction. The test pigs all showed gross lesions in the aorta with an increase in cholesteryl ester and free cholesterol in the tissue as compared with control animals.


[The situation of suicide prevention (author's transl)]. Suicide research has always been especially concerned with the social connections. Its results however have not been sufficiently used for the benefit of suicide prevention. Suicide prevention is linked to a somatically oriented medicine, which is not able to provide with an after care program. To show the present status of practical suicide prevention, an investigation was carried out in Hanover, the result of which postulates for an intesification of social services and psychiatric consulting services; it also demands a support of counseling services by telephone, special information about the possibilities of suicide prevention, and the initiation of suicide prevention centers in big towns, that work with an inter-disciplinary staff.


[Synthetic depot-ACTH, plasmacortisol levels, and asthmatic syndrome (author's transl)]. Plasmacortisol levels were measured in 33 patients with asthmatic syndrome before and after they received treatment with 6 mg of depot-ACTH (Tetracosactid) over seven days. 75 percent of the patients showed improved VC and FEF after therapy. This effect was not in any way correlated to the basal level of cortisol, nor to the increment during treatment.


The mechanism of lipid-binding by plasma lipoproteins. In this article, we review the current state of knowledge about the structure of human plasma high density lipoproteins. Recent advances in this field, including amino acid sequencing and lipid-binding studies, suggest that the plasma lipoproteins have specific sites with special structural features that account for their ability to bind and transport lipid. A hypothesis which describes the structure of these lipid-binding sites and their interaction with lipid is presented.


[Results of a nine-year phenylketonuria (PKU) screening (author's transl)]. The percentage of children in Nordrhein-Westfalen who were tested for PKU free of charge and on a voluntary basis rose from 25.3% (1966) to 99.5% (1973). Among 1,431,999 newly born children, 262 children with suspected PKU were found, 225 of them diagnosed early and 37 late, 181 with "typical" and 81 with "atypical" PKU. Most of 102 children with late diagnosed PKU had an I.Q. of 80-50 or less, while in 95 children treated early, it was over 91 in 80% and below 90 in only 20%. There was a statistically highly significant difference in the I.Q. distribution between patients diagnosed early and those diagnosed late, while the mental development of phenylketonuria patients treated early was not different from that of a homogenous population group under the same conditions. After extending the test program in 1966, 1 maple syrup urine disease, 3 homocystinurias, 3 galactosemias and 1 galactokinase deficiency were found in addition to PKU.


Adaptive character of liver glucokinase. 1. Glucokinase is one of four glucose phosphorylating enzymes present in rat liver. Its distinctive features are a high K-m for glucose (high-K-m isozyme) and a rather narrow substrate specificity. In contrast, the other three enzymes, collectively called hexokinases or low-K-m isozymes, exhibit low K-m values for glucose and a wider substrate specificity. 2. Glucokinase is present in the liver os mammals (with some exceptions), amphibians and lower reptiles; It is absent from higher reptiles and birds. The presence or absence of glucokinase may represent an evolutionary adaptation to feeding habits and other physiological peculiarities. Differences in the immunological behavior and in the kinetic parameters of glucokinases from different taxa suggest the operation of divergent evolution. 3. The levels of glucokinase in rat liver depend strictly on the supply of carbohydrate in the diet. Glycogen phosphorylase and glycogen synthetase behave similarly, whereas other carbohydrate-metabolizing enzymes depend on the provision of either protein or protein plus carbohydrate. Glucokinase decays with a half-life of 33 hr when rats are starved or fed a carbohydrate-free diet, and is induced by the administration of glucose. The adaptive character is not exhibited by all mammals, indicating evolutionary discrimination within the same class and even within the same single order Rodentia. Enzyme adaptation in the liver may partially explain the condition known as 'hunger diabetes'. 4. The endocrine system plays a paramount role in glucokinase adaptation, since insulin is essential for glucose-dependent glucokinase induction and, on the other hand, glucagon, catecholamines and cyclic AMP prevent the induction. Glucocorticoids and some pituitary hormones modulate the rate of induction. The mechanisms underlying the hormonal regulation of glucokinase levels are not well known. 5. The variations in liver glucokinase correspond to changes in the amount of enzyme protein as assessed by immunochemical titration. This fact agrees with the effects of inhibitors of protein synthesis on glucokinase induction. 6. An antiserum against rat glucokinase reacts with the enzyme from mammals and turtles but not with the amphibian enzyme. It does not react with low-K-m hexokinases from different sources. 7. The saturation function for glucose is sigmoidal in mammalian and amphibian glucokinases but not in glucokinase from lower reptiles. The Hill's coefficient is very constant with values about 1.6. The K0.5 (concentration for half saturation) values in the different species studied vary between 1.5 and 8 mM. These kinetic parameters may be considered as another adaptive feature aimed to give maximal efficiency to the liver uptake of glucose at the changeable concentrations in the blood resulting from variations in the amount of dietary glucose.


[Diagnosis and surgical treatment of bronchial carcinoma - a report on 1000 cases (author's transl)]. One thousand cases of bronchial carcinoma, which had been treated in the Chirurgische Universitätsklinik Erlangen in 17 years, are reported. 63,2% were either primarily inoperable or found to be so after an exploratory laparotomy; Only 32,4% of all patients were admitted in the first 3 months, 17,6% after more than 12 months, Squamous epithelial carcinoma was the most frequent histological type (51,2%), then came small cell carcinoma (16,4%), dedifferentiated carcinoma (5,1%) and adenocarcinoma (4,1%). The best results of operative treatment were obtained with lobectomy in Stage I with a five year survival rate of 45%. In Stage II, pneumonectomy is the satisfactory operation (no five year cure with lobectomy, but 25,2% with pneumonectomy). Exploratory thoracotomy does not shorten the life expectancy and it should therefore be performed in doubtful cases.


[The importance of estrogen receptors in the pharmacotherapy of mammary carcinoma (author's transl)]. The physiological effect of estrogens on the estrogen-dependent target organs is mediated by estrogen receptors which are present in the cytoplasm of estrogen-sensitive cells. By determining the estrogen receptors in mammary carcinoma tissue, information may be obtained on the susceptibility of the tumor concerned to hormones. As a novel pharmacological principle, the anti-estrogens are able to blockade the hormonal effect of endogenous estrogens on the tissue of the mammary carcinoma. Possible relationships between prolactin and mammary carcinoma become apparent. From the pharmacologist's point of view, these questions are of particular importance in relation to the possibility of predisposition to mammary carcinoma through treatment with reserpine, which is at present under discussion.


Comparative analysis of deletion and base-change mutabilities of Escherichia coli B strains differing in DNA repair capacity (wild-type, uvrA-, polA-, recA-) by various mutagens. Dose-response curves were compared for deletions [ColBR (resistant to colicin B) mutations being more than 80% deletions] and base changes (reversion of argFam to prototrophy argplus) induced in the same set of E. coli strains (wild-type for DNA repair, uvrA-, polA- and recA-) by N-methyl-N'-nitro-N-nitrosoguanidine (NTG), ethyl methanesulfonate (EMS), hydroxylamine (HA), 4-nitroquinoline I-oxide (4NQO), mitomycin C (MTC, UV and X-rays. All these agents induced deletions as well as base changes in the wild-type strain. Thus chemical mutagenesis differed in E. coli and bacteriophages in vitro, for HA, NTG, EMS and perhaps UV produced only point mutations in phage Tr. The patterns of deletion and base-change mutability in E. coli were surprisingly similar. (I) The recombination less recA- strain was mutable by only three (NTG, EMS, HA) of the seven mutagens for either deletions or base changes. (2) The uvrA- strain, unable to excise pyrimidine dimers, was very highly mutable by 4NQO and UV but immutable by MTC for both deletions and base changes. (3) The polA- strain, defective in DNA polymerase I due to a non-suppressible mutation, was very highly mutable by HA and highly mutable by MTC and 4NQO for both deletions and base changes but was highly mutable only for deletions by UV and X-rays, remaining normally mutable by the other agents for both deletions and base changes despite its high sensitivity to their inactivating action. We conclude that errors in the recA-dependent repair of induced DNA damage (after 4NQO, MTC, UV and X-rays) or errors in replication enhanced by damage to the replication system or to the template strands (after NTG, EMS, and HA) give rise to deletions as well as to base changes. From a comparative analysis of 14 dose-response curves for deletions and base changes, we conclude that the order of mutagenic efficiency relative to killing is (EMS, NTG) greater than (UV, 4NQO) greater than HA greater than (X-rays, MTC), and that X-rays, 4NQO, HA and MTC induce more ColBR deletions than Argplus base changes, whereas UV and EMS induce ColBR deletions and Argplus base changes at nearly equal rates and the specificity of NTG is intermediate between these two types.


Viral hepatitis, type A. Identification by specific complement fixation and immune adherence tests. Hepatitis A antibody was detected by specific immune adherence and complement-fixation tests in a study involving 473 serum specimens from 20 patients who had viral hepatitis, Type A. In all 20 patients who had no detectable immune adherence antibody (less than 1:5) before onset of hepatitis high levels (greater than or equal to 1:1024) developed one to four weeks later, occasionally reaching peak levels exceeding greater than or equal to 1:81,920 several months thereafter. Five to 10 years later the immune adherence antibody levels ranged between 1:640 and 1:20,480. In general, the complement-fixation test was not as sensitive or as specific as the immune adherence test. These findings indicate that the immune adherence test should be a valuable tool for diagnosis, for epidemiologic surveys, for identification of susceptible and immune persons, for quantitative assays of gamma globulin and for identification of hepatitis A virus in attempts to propagate the virus in cell culture.


Plasma lipids and lipoproteins in vegetarians and controls. The possible effect of diet on plasma lipids was investigated in a group of vegetarians. Seventy-three men and 43 women who had adhered to a vegetarian diet for an average of three years were studied in the fasting state and compared with a randomly selected group of controls matched for age and sex. Mean cholesterol levels in milligrams per deciliter for vegetarians and controls, respectively, were 126 and 184; low-density lipoprotein, 73 and 118; very-low-density lipoprotein, 11.8 and 17.2; high-density lipoproteins, 42 and 49; and mean triglyceride levels 59 and 86. Mean weight and subscapular skinfold thicknesses were 58 kg and 6 mm for vegetarians and 73 kg and 17 mm for controls. Similar differences in lipid levels were found between subgroups of 42 vegetarians and controls with identical mean weights. Multiple regression analyses showed that consumption of dairy foods and eggs, but not body weight, was associated with the lipoprotein and cholesterol findings.


Reovirus antibody patterns in dogs: a trial for the application of principal component analysis to seroepidemiology. In 100 dogs in Morioka and its vicinity serologically surveyed for reovirus infection, there were significant correlations between hemagglutination-inhibiting and neutralizing antibody titers of the same virus types. In order to establish a proper index for the evaluation of infection, principal component analysis was applied to the analysis of data, including standard immune sera. Of nine samples of matrix examined, correlation matrix of 6 variables was suggested to afford the most proper result. Essential information from the original data was accounted for by factor loadings of the first 3 components. The 6 attributes of serum antibody were classified into three groups. In a scatter diagram serum samples were classified also into three major groups corresponding to the three reovirus types, and the serotypic pattern of infection was clearly visualized.


Further studies on ACTH secretion from pituitary grafts in the hypophysiotrophic area. Resting and stress-induced plasma corticosterone concentrations in adult male hypophysectomized rats bearing a pituitary homograft in the hypothalamus were determined fluorimetrically. Plasma corticosterone resting levels were maintained and a significant elevation of these levels in response to stress occurred only in those rats in which the graft was in the medial basal hypothalamus (hypophysiotrophic area, HTA) and the implant was connected with the median eminence. If these latter connections of the graft in the HTA did not exist, resting plasma corticosterone concentrations were reduced and the applied stress did not induce any elevation of the low pre-stress levels. On the basis of the present and previous findings, it is assumed that the site of CRF production is the median eminence region.


Adrenal cortical secretion in response to pilocarpine in dogs with hypothalamic lesions. Adrenal venous blood samples were collected from conscious and anesthetized dogs before and after an injection of pilocarpine and were analyzed for 17-hydroxycorticosteriods (17-OHCS). In conscious dogs the adrenal 17-OHCS secretion rate increased from a pre-injection secretion rate of 0.11 plus or minus 0.014 to 1.58 plus or minus 0.069 (mean plus or minus SE) mug/kg/min following an intravenous injection of pilocarpine (0.3 mg/kg); there was a return to the baseline level by 120 min. In anesthetized dogs a marked increase in adrenal 17-OHCS secretion was also observed, which was completely abolished by hypophysectomy A noticeable impairment of the adrenocortical response to pilocarpine was found to be produced by radio-frequency-current lesions of the anterior median eminence. A partial depression of the response was also observed in dogs with the anterior ventromedial hypothalamic lesions. Lesions destroying other hypothalamic areas were found to be without significant effect upon the adrenal cortical secretory response to pilocarpine.


Luteomas of pregnancy.

Luteomas of pregnancy. Since its original description in 1963, the pregnancy luteoma has been a controversial entity both clinically and pathologically. The most important questions which have been asked are: Is the luteoma a true neoplasm or does it represent simply an abnormal response to the hormones of pregnancy? What is the relation between the tumor and certain endocrinopathies that are associated with these lesions? This study of 20 cases collected from the files of the Emil Novak Ovarian Tumor Registry and the Pathology Section of the Baltimore City Hospitals directs its attention to these critical clinicopathologic features.


Prophylactic antibiotics in cesarean section. Using a random selection procedure, 100 patients were evaluated in order to determine the value of prophylactic antibiotics in cesarean section. All patients were evaluated for febrile morbidity, and specific infection sites were recorded. Significant differences were found between the treated and control groups and between cesarean sections performed prior to the onset of labor and after the onset of labor.


Virilization associated with bilateral luteomas of pregnancy. A case of maternal virilization associated with bilateral luteomas of pregnancy is described. Urinary 17-ketosteroids and plasma testosterone fell from markedly abnormal values to normal limits within 2 weeks of delivery. However, symptoms of virilization persisted in the postpartum state. A possible relation between polycystic ovarian syndrome in the nonpregnant state and luteoma of pregnancy is proposed.


A rapid solid-phase radioimmunoassay specific for human chorionic gonadotropin in gestational trophoblastic disease. A rapid solid-phase radioimmunoassay (RIA) specific for human chorionic gonadotropin (hCG) has been used for the measurement of serum hCG activity in patients with molar pregnancy and gestational trophoblastic disease (GTD). Serum hCG regression as determined by the specific RIA method after evacuation of uncomplicated molar pregnancy was noted to occur over a longer duration of time than previously reported from this Center using a nonspecific RIA system which measures human luteinizing hormone (hLH) and hCG simultaneously. Therapy for proliferative trophoblastic disease was withheld after evacuation of molar pregnancy while the serum hCG level regressed normally, but was instituted when the serum hCG level rose or plateaued for more than two consecutive weeks. Serum hCG levels in patients requiring chemotherapy for GTD were also more accurately monitored with the specific RIA method than with the nonspecific technic. Therapy was based solely on the hCG titer rather than the subsidence of toxicity, as has been our practice in the past. As a result, the duration of hospitalization, total dose of drug required for remission, and toxic side effects were substantially reduced without sacrificing the effectiveness of chemotherapy.


Thymidine kinase activity in cerebrospinal fluid of rabbits with Herpesvirus hominis encephalitis. The rabbit model of Herpesvirus hominis (HVH) encephalitis was utilized to determine whether thymidine (TdR) kinase (EC. 2.7.1.2.1), AN EARLY ENZYME IN THe HVH replicative cycle, is present in the cerebrospinal fluid (CSF) of rabbits with HVH encephalitis. The mean TdR kinase activity in CSF from infected rabbits was 2.30 plus or minus 1.69 pmol product/120 min/100 mul CSF, while the mean TdR kinase activity of uninfected CSF was 0.34 plus or minus 0.25. The enzyme appeared to be cell associated since the TdR kinase activity in the sedimented cell suspension was 5.08 plus or minus 2.93 (seven values) while the supernatant activity was 0.14 plus or minus 0.31 (seven values). To allow a comparison of TdR kinase from infected rabbit CSF with HVH and rabbit mononuclear cell TdR kinase, the mean ratio of TdR kinase activity at pH 6.0/pH 8.0, for infected rabbit CSF is intermediate between the ratio for HVH TdR kinase (P less than 0.001) and rabbit mononuclear cell TdR kinase (P less than 0.05).


Cyclic nucleotide metabolism coupled to cytodifferentiation of Blastocladiella emersonii. Cyclic GMP and cyclic AMP levels during growth and differentiation of B. emersonii were measured by use of chemical, biochemical, and immunologic assays. Of particular interest was the finding that net synthesis of cyclic GMP occurred during a single stage of its life cycle, sporulation, when intracellular levels increased 50- to 100-fold in a process requiring protein and RNA synthesis.


Mutant hamster cells exhibiting a pleiotropic effect on carbohydrate metabolism. Sixty-seven galactose negative (Gal minus) clones were established from survivors after treatment of Chinese hamster lung (V79) cells with 5-bromodeoxyuridine and black light. The mutational origin of these clonal isolates was inferred on the basis of the previously demonstrated mutagenic action of the combined treatment, persistence of the Gal minus phenotype in cell progeny in the absence of selection, conditional lethality of three isolates, interallelic complementation, and assignment of a homologous human gene to chromosome A2. In contrast to the parental cells, Gal minus mutants could not utilize exogenous galactose, mannose, fructose, galactose 1-phosphate, glucose 1-phosphate, or glucose 6-phosphate. Permeation of galactose into mutant cells appeared unimpaired. In intra- and interspecific cell hybrids, the mutation causing the Gal minus phenotype behaved like a recessive character. The pleiotropic nutritional response was due not to deficiency of any one specific enzyme in the Leloir pathway but to significant reduction in activities of phosphoglucomutase, NADP-dependent isocitrate dehydrogenase, and perhaps other enzymes. In temperature-sensitive Gal minus mutants grown at permissive temperature or in complementing intra- and interspecific cell hybrids, the activities of these enzymes were restored to normal levels, accompanied by a regained ability to use the particular hexoses or hexose monophosphates. We postulate that the change from Gal + to Gal minus phenotype in hamster cells could be due to mutations at a regulatory gene locus, or at a yet unknown locus with enzymic defect that causes secondary metabolic imbalances.


Demonstration that monocytes rather than lymphocytes are the insulin-binding cells in preparations of humah peripheral blood mononuclear leukocytes: implications for studies of insulin-resistant states in man. Insulin receptors have been demonstrated on mononuclear leukocytes prepared by centrifugation of buffy coats from normal blood donors on Ficoll-Hypaque gradients. The cell type that specifically binds insulin in this mixture of lymphocytes and monocytes has never been clearly identified, although it was assumed to be the lymphocyte since this cell constitutes about 80% of the population. In the present studies, insulin-binding assays were performed on the mononuclear leukocyte preparation before and after selective depletion or enrichment for monocytes using glass wool or Sephadex G-10 adherence columns. The amount of 125-I-labeled insulin specifically bound correlated significantly with the number of monocytes but not with the number of B or T lymphocytes. Approximately 90% of the specific insulin binding of this preparation could be accounted for by its content of monocytes. The amount of binding was unaffected by phagocytosis of latex particles or by metabolic inhibitors added to prevent endocytosis. Autoradiograms made on smears of whole peripheral blood and mononuclear leukocytes demonstrated that all of the cells that bound 125-I-labeled insulin were large mononulcear cells, 85-90% of which could be identified as monocytes by morphological criteria or by the functional criterion of latex particle ingestion. Since insulin receptor concentration may be altered in disease states in man, it is essential, when using this cell population for detecting such changes, to quantitate the number of monocytes in the preparation so that the insulin-binding data can be appropriately interpreted.


Colchicine affects kinetics of concanavalin a-mediated agglutination of hepatoma cells and plasma membranes from liver and hepatoma cells. Concanavalin A-mediated agglutination reactions of hepatoma cells [AH-130F(N)] and isolated liver cells, as well as of plasma membranes prepared from the liver and hepatoma, were investigated kinetically together with the effect of colchicine upon them. Concanavalin A-mediated agglutination of hepatoma cells at 25 degrees proceeded with biphasic kinetics (first and second stages of agglutination), while no appreciable agglutination of liver cells was observed in the presence of concanavalin A. The plasma membranes from the liver and hepatoma cells were similarly agglutinated with concanavalin A at 25 degrees but not at 0 degrees. The concanavalin A-mediated membrane agglutination proceeded with monophasic kinetics and was incomplete. Colchicine inhibited preferentially the second stage of concanavalin A-mediated agglutination of hepatoma cells, but it did not affect the concanavalin A-mediated agglutination of plasma membranes of both types of cells.


Regulation of development in Myxococcus xanthus: effect of 3':5'-cyclic AMP, ADP, and nutrition. An assay was developed to study the regulation of fruiting in Myxococcus xanthus. The nucleotides, adenosine 3':5'-cyclic monophosphate (cyclic AMP) and adenosine diphosphate (ADP), were found to greatly stimulate fruiting under the assay conditions. Very sharp concentration optima were observed. Even under conditions of starvation, these nucleotides greatly increased the number of aggregation sites. Nutrition was found to influence fruiting body morphology. The effect of amino acids on the nucleotide stimulation of fruiting was studied under our assay conditions. L-Methionine and L-isoleucine (1 mM) completely blocked either L-threonine or D,L-diaminopimelic acid synergistically enhanced the amount of fruiting in the presence of these nucleotides. The data presented suggest the existence of differentiation-related regulatory compounds in M. xanthus.


Contribution of the peptide backbone to the action of oxytocin analogs. This investigation was undertaken to evaluate the functional contribution of the peptide backbone of oxytocin in its interaction with receptor. Corey-Pauling-Koltun models of (Gly-7) deaminooxytocin, deaminotocinamide, and their respective retro-D-analogs built in any specific conformation (e.g., the Walter-Urry model for oxytocin) have a quai-equivalent topochemical arrangement of amino-acid side chains; however, the CO and NH elements of the peptide backbone of the retro-D-analog are reversed. The retro-D-analogs of deaminotocinamide and (Gly-7) deaminooxytocin (prepared using D-alle for L-Ile) and their respective N-formyl derivatives were assayed for uterotonic activity relative to related L-peptides. All retro-D-analogs (tested at concentrations ranging from 10-10 to 10-5 M) were devoid of angonistic (or antagonistic) activity in the isolated rat uterus, except for the retro-D-(D-alle-3, Gly-7) deaminooxytocinamine, which retains a terminal NH-2 group on the tail; the latter is a partial agonist with very low affinity. The results obtained with retro-D-analogs indicate that one or more of the elements of the peptide backbone of the tocinamide ring are essential for "occupation" and "activation" of uterine receptors. Oxytocin action may be the resultant of multiple hydrogen-bonding interactions between CO, NH, NH-2, and OH groups of the hormone with complementary groups on receptor, made possible by appropriate hydrophobic bonding.


Distribution of membrane particles and gap junctions in normal and transformed 3T3 cells studied in situ, in suspension, and treated with concanavalin A. Freeze-fracture techniques were used to study the ultrastructural distribution of plasma membrane particles in cultures of normal Balb/c and Swiss 3T3 and simian virus 40- or murine sarcoma virus-transformed fibroblasts. No apparent differences were observed. Cultures fixed in situ show a seemingly random distribution of membrane particles both in normal or in transformed cells. Treatment of cell cultures in situ with concanavalin A does not result in an altered pattern of particle distribution. EDTA-induced suspension of normal or transformed cells does not result, per se, in modification of the type of membrane particle distribution seen in cells fixed in situ. However, upon further incubation, a proportion of normal or transformed cells in suspension show a varying degree of particle aggregation following a network pattern. Concanavalin A treatment of normal and transformed cells in suspension does not result in a specific change of the pattern of particle distribution. Because it has been established that treatment with concanavalin A of simian virus 40-transformed Balb/c 3T3 fibroblasts causes pronounced clustering of the concanavalin A receptors at the outer-surface, our results probably imply independence of membrane particles and concanavalin A receptors of these transformed cells.


Simian virus 40-related antigens in three human meningiomas with defined chromosome loss. Two out of seven meningiomas tested in early cell cultures by indirect immunofluorescence staining showed simian virus 40 (SV40)-related tumor (T) antigen. In one tumor 90% of the cells were positive. An additional SV40-related antigen (U) was found in 10% of cells of a third tumor. These findings indicate that the meningioma cells showing a positive reaction are transformed by a papova virus that has at least partly the same antigenic properties as SV40 virus. SV40-related viral capsid (V) antigen was absent in all the meningiomas tested. No virus infectious for African green monkey kidney (AGMK) cells could be isolated. The tumors positive for T and U antigens showed the chromosome aberration typical for human meningiomas, i.e., the loss of one chromosome, G-22. The T-antigen-positive tumors showed further hypodiploidization. Experiments to rescue virus from the T-antigen-positive tumors showed further hypodiploidization. Experiments to rescue virus from the T-antigen-positive meningioma cells were performed: fusion of cells pretreated with 8-azaguanine with cells premissive for SV40 led to a low percentage (0.01-0.05%) of V-antigen-positive nuclei in heterokaryon cultures. On the basis of these results, the possibility of a correlation between the meningioma, a relatively common intracranial tumor in man, and an SV40-related papova virus must be considered. It remains to be shown whether this virus is a causative agent for human meningiomas.


Translation of Rous sarcoma virus RNA in a cell-free system from ascites Krebs II cells. The template activities of the 60-70S RNA complex and of the 30-40S subunit RNA species of Rous sarcoma virus were tested in a cell-free protein-synthesizing system from mouse ascites Krebs II cells. Stimulation of protein synthesis over the endogenous background was about 2-fold with 30-40S viral RNA and about 1.3-fold with 60-70S viral RNA as template. Analysis by sodium dodecyl sulfate-gel electrophoresis showed that the predominant polypeptide synthesized in vitro in response to 30-40S RNA of Rous sarcoma virus had a molecular weight of 75,000-80,000. This polypeptide could be precipitated by antiserum against the group-specific antigens of the virus, although its molecular weight is higher than that of virion group-specific antigen proteins. Analysis of tryptic digests of the protein made in vitro indicates similarity to tryptic digests from authentic virion group-specific proteins. It is concluded that part of the RNA from Rous sarcoma virus is translated in vitro into a high-molecular-weight protein, perhaps a precursor of the virion group-specific proteins.


Viral-related information in oncornavirus-lik particles isolated from cultures of marrow cells from leukemic patients in relapse and remission. Characterization of ribonucleic acid content of particles released from cultures of marrow cells of leukemic patients indicates the presence of RNA molecules of size and base sequence characteristic of oncornarviruses. Seventeen marrow samples obtained from leukemic patients in relapse or in a chronic phase of the disease yielded particles containing high-molecular-weight RNA with a sedimentation velocity (about 70 S) similar to that obtained for murine oncornavirus RNA. Eight of nine marrow samples from non-leukemic patients did not yield detectable high-molecular weight RNA. Among patients in firm hematological remission, three of three samples from patients with acute lymphoblastic leukemia and three of nine samples from patients with acute myeloblastic leukemia were positive for high-molecular-weight RNA. The base sequence of the RNA in particles was characterized by synthesizing complementary (3-H)DNA in an endogenous reaction and hybridizing to excess RNA from known oncornaviruses. Hybridization of 40-60% of input complementary DNA to simian sarcoma virus RNA was detected. No monology was detected with an avian oncornavirus (Rous sarcoma virus) while an intermediate level of homology (10-30%) was detected in hybridization to murine sarcoma virus (Kirsten) and murine leukemia viruses (Rauscher, Moloney, and Gross).


Fatty acid chain flexibility in the membranes of normal and transformed fibroblasts. Fatty acid spin labels were used to measure the inherent flexibility of the lipid acyl chains in intact cell membranes of 3T3 mouse fibroblasts and several varieties of transformed 3T3 cells. No significant differences in inherent lipid flexibility were detected in the normal and transformed mouse fibroblasts. These results are compared with similar results obtained earlier for chick embryo fibroblasts. For the cells studied, a difference in the mobility of membrane glycoproteins of normal and transformed fibroblasts has either been demonstrated directly or inferred from differential agglutinability by other investigations. Therefore, a basis for correlating glycoprotein mobility and lipid motion in these cells was not found.


Blue dextran-sepharose: an affinity column for the dinucleotide fold in proteins. A procedure is described to utilize blue dextran-Sepharose as an affinity chromatographic column specific for the super-secondary structure called the dinucleotide fold, which forms the binding sites for substrates and effectors on a wide range of proteins. The procedure can be used to identify proteins, either purified or in crude cellular extracts, that possess the dinucleotide fold and to significantly improve the purification procedures for those proteins that possess the fold.


Initiation and maintenance of cell transformation by simian virus 40: a viral genetic property. The transforming ability in 10% serum medium of the temperature-sensitive mutants of simian virus 40 in the complementation group III (ts640 type mutants) was greatly reduced when the infected rat 3Y1 cells were incubated at the restrictive temperature of 40 degrees or incubated first at 40 degrees for 3 days and then shifted to the permissive temperature of 33 degrees. Transformation did occur efficiently after incubation at 33 degrees or after an initial incubation at 33 degrees for 5 days followed by a shift to 40 degrees. When growth properties of 3Y1 cells transformed at 33 degrees by the group III mutants were examined at 40 degrees, several aspects of the transformed state were rendered temperature-sensitive. These aspects were the ability of cells to grow in low serum (1.5%) medium and to make colonies, in 10% serum medium, on monolayers of untransformed 3Y1 cells and in soft agar. It is concluded that a simian virus 40 gene (cistron III) controls the initiation, as well as at least some aspects of the maintenance, of transformation and that the initiation reaction is a more heat-labile event than the maintenance reaction(s) under the experimental conditions.


Contact inhibition of phagocytosis in epithelial sheets: alterations of cell surface properties induced by cell-cell contacts. Contact inhibition of phagocytosis was found to be characteristic for epithelial sheets formed in cultures by several cell types: normal and transformed mouse kidney cells, and differentiated mouse hepatoma cells. In these sheets most central cells surrounded by other cells had very low phagocytic activity. In contrast, marginal cells having a free edge were able to perform an active phagocytosis. Contact inhibition of phagocytosis was absent in dense cultures of mouse embryo fibroblasts and in cultures of anaplastic mouse hepatoma 22a. The upper surface of epithelial sheets was nonadhesive for prelabeled epithelial cells and fibroblasts. In contrast, the upper surface of dense cultures of mouse fibroblasts was adhesive for these cells. These and other data strengthen the suggestion that contact inhibition of phagocytosis is a result of different adhesiveness of the upper cell surface and of the surfaces near the free edge. Agents inhibiting cell surface movements at the free edges of marginal epithelial cells (cytochalasin, azide, sorbitol, low temperature) prevented adhesion of particles to these edges. Possibly, the surface of actively moving cytoplasmic processes is the only cell part that has adhesive properties necessary for the formation of attachments with other cellular and noncellular surfaces. In epithelial sheets, in contrast to fibroblast cultures, Colcemid did not activate movements of immobile contacting cell edges. These results indicate that mechanisms of contact immobilization of cell surface may be different in epithelium and fibroblasts. Firm contacts formed between epithelial cells are sufficient for stable immobilization of the surface; additional stabilization of the surface by microtubules is not essential. Fibroblasts do not form firm contacts and the Colcemid-sensitive stabilization process is essential for maintenance of the immobile state of their surfaces. Differences in the stability of cell surface immobilization produced by cell-cell contacts may also explain different adhesiveness of the upper surfaces of dense fibroblastic and epithelial cultures.


Decrease in membrane-associated actin of fibroblasts after transformation by Rous sarcoma virus. The actin content of membranes prepared from cultured chick embryo fibroblasts has been measured on polyacrylamide gels. The actin was identified by tryptic peptide mapping. After transformation of the cells by Rous sarcoma virus, the amount of actin associated with the membranes is decreased by 30-50%. This result is not due to infection per se, since infection by a temperature-sensitive strain of the virus decreases membrane-associated actin only at the permissive temperature. A shift from the nonpermissive (41 degrees) to the permissive (36 degrees) temperature results in an increase in the percentage of total cellular protein synthesis devoted to actin production, so that the decrease in membrane-associated actin appears to be a selective displacement from the membrane rather than a general decrease in total cellular actin.