The role of carbohydrate in the glycoenzyme invertase of Neurospora crassa. Data obtained concerning the carbohydrate moieties of the glycoenzyme invertase (EC 3.2.1.26, beta-D-fructofuranoside fructohydrolase) from Neurospora crassa were consistent with a linkage of some carbohydrate chains by O-glycosidic bonds to serine and threonine residues; the possibility of N-glycosylamine linkage of some of the carbohydrate to the amide group of asparagine is also indicated. The invertase was remarkably stable on storage at low temperatures. Oxidation of the carbohydrate residues in the enzyme by sodium periodate markedly affected the heat-stability of the enzyme. It is suggested that the carbohydrate moieties function as stabilizers of the tertiary structure of the glycoenzyme.


Effect of opiate receptor blockade on pain sensitivity in the rat. Blockade of opiate receptors by naloxone (2 mg/kg) was found to produce a significant increase in pain sensitivity as measured by the tail-flick test. This finding supports the view that endogenous opiate systems may play a role in the modulation of pain sensitivity. Naloxone, however, was found to have no effect on pain responsiveness as measured by tail-pinch. These findings, together with additional reports, suggest that endogenous opiate systems may exert differential actions on different sensory modalities.


Effect of lowered intracellular ATP and GTP concentrations on purine ribonucleotide synthesis and interconversion. The effects of lowered intracellular ATP and GTP concentrations on enzymes of purine ribonucleotide synthesis and intercoversion were studied using intact Ehrlich ascites tumor cells. The apparent rates of phosphoribosyl pyrophosphate synthetase (EC 2.7.6.1) and of inosinate dehydrogenase (EC 1.2.1.14) were increased in cells containing lowered purine nucleotide concentrations, but apparent activities of amidophosphoribosyltransferase (EC 2.4.2.14), the purine phosphoribosyltransferases, and other enzymes of purine ribonucleotide interconversion were not affected.


Uridine phosphorylase activity of isolated plasma membranes of rat liver. Plasma membranes were isolated from rat liver homogenates either by differential centrifugation or by fractionation in discontinuous sucrose density gradients. Both membrane preparations contained about 17% of the total uridine phosphorylase (EC 2.4.2.3) activity and 44% of the total 5'-nucleotidase (EC 3.1.3.5). The enrichment factor for uridine phosphorylase in the fractions prepared by differential centrifugation was about 2.8 and by the gradient method, as much as 11.0; the respective enrichment factors for 5'-nucleotidase were 1.8 and 9.5. Uridine phosphorylase activity of isolated plasma membrane fractions was stimulated 2.5-fold by 0.1% Triton X-100. Unlike the cytosol enzyme, uridine phosphorylase of plasma membranes showed little or no deoxyuridine-cleaving activity. Contamination of the membrane fractions by thymidine phosphorylase (EC 2.4.2.4) of the cytosol was negligible. The other subcellular organelles obtained by either procedure and characterized by marker enzyme activities were found not to contain significant uridine phosphorylase activity; the cytosol fractions contained just over 70% of the total uridine phosphorylase activity with an enrichment of only about 2.8-fold. The activity of the cytosol enzyme was not stimulated by Triton X-100.


Correlation between organophosphate poisoning, acetylcholinesterase inhibition, and increased cyclic GMP levels in malathion-treated insects. Organophosphate poisoning with malathion caused large increases (up to 125 and 440%, respectively) in the level of cyclic GMP in larvae of Mamestra configurata Wlk. and in the fly Sarcophaga bullata Parker. Cyclic AMP was little affected. The malathion-induced increase in cyclic GMP was time and dose dependent. Time-course studies with the head and thorax of S. bullata demonstrated that the increase in cyclic GMP level occurred precipitously after a lag period  of about 1 h, during which time the activity of acetylcholinesterase (EC 3.1.1.7) was progressively inhibited. The abrupt increase in cyclic GMP began when acetylcholinesterase activity had been inhibited to a sufficient extent to permit accumulation of acetylcholine. It is suggested that the accumulation of acetylcholine in the malathion-poisoned insects caused cyclic GMP levels to rise.  Cyclic GMP may have a role in cholinergic transmission in normally functioning insect neural tissue. Increased levels of cyclic GMP induced by organophosphate and organocholorine (Bodnaryk, R. P. (1976) Can. J. Biochem. 54, 957-962) insecticides appear to be a vital and previously unrecognized biochemical lesion in insects poisoned by these compounds.


The syntheses of alpha-D-glucopyranose-1-phosphates labelled with 18O in the phosphoryl or ester oxygen positions. Phosphoryl-oxygen-labelled alpha-D-glucopyranose-1-phosphate (Glc1P) has been prepared by the hydrolysis in 18O-enriched water of alpha-D-glucopyranose-1,2-cyclic phosphate catalyzed by extracts of Saccharomyces fragilis. Ester-oxygen-labelled alpha-D-galactopyranose-1-phosphate has been prepared by galactokinase-(EC 2.7.1.6)-catalyzed phosphorylation of D-[1-18O]-galactose by ATP, and it has been isomerized to ester-oxygen-labelled Glc1P using galactose-1-phosphate uridyltransferase (EC2.7.7.12) and UDPgalactose 4-epimerase (EC 5.1.3.2). These labelled glucose phosphates are for use in oxygen-exchange experiments.


Hypermodified alkali-stable dinucleotide sequences in each of the high-molecular-weight (26S and 18S) ribosomal RNA species of wheat. Two hypermodified, alkali-stable dinucleotide sequences, each containing a base modification in addition to sugar methylation, are known to be present in wheat embryo 26S + 18S rRNA (Gray, M.W. (1974) Biochemistry 13, 5453-5463). Quantitative analysis of unfractionated 26S + 18S rRNA had suggested that each of these sequences (Cm-psi p and psi m-Ap, where Cm=O2'-methylcytidine and psi m-O2'-methylpseudouridine) was present in either the 18S or the 26S rRNA species, but not the both, at a frequency of not more than once per chain. In the study reported here, the individual 32P-labeled 18S and 26S rRNA species were isolated from viable wheat embryos germinated in the presence of [32P]orthophosphate. From analyses of phosphodiesterase and alkaline hydrolysates of the separated [32P]RNAs, we conclude that psi m-Ap is confined to wheat cytosol 18S rRNA, whereas Cm-psi p is localized in wheat cytosol 26S rRNA. The presence of psi m in the 18S rRNA of wheat stands in contrast with the situation in animal cells, where this hypermodified nucleoside is located in the 28S rRNA (Khan, M.S.N. & Maden, B.E.H. (1976) J. Mol. Biol. 101, 235-254).


Primary structure and morphine-like activity of human beta-endorphin. The complete amino acid sequence of human beta-endorphin was obtained by automatic sequencing of a sulfonyl isothiocyanate derivative of this peptide, in combination with peptide mapping of a tryptic digest of the native molecule. It was found to be identical with the carboxy-terminal portion 61-91 of human beta-lipotropin (beta-LPH). The morphine-like activity of beta-endorphin is comparable both in the mouse vas deferens bioassay and in the opiate receptor binding assay. However, beta-LPH is not active up to concentrations of 10(-6) M.


Effects of antipsychotic drugs on action potential production in skeletal muscle. II. Haloperidol: nonspecific and opiate drug receptor mediated effects. The effects of haloperidol, an antipsychotic butyrophenone, on excitability and action potential production in frog's sartorius muscle fibers were studied. This drug produced a local-anestheticlike effect which developed slowly over 1 to 5 h with lower concentrations (2.7 to 5.3 X 10(-6 M) but was completely reversed by exposing the muscles to a drug-free solution. In studies with intracellular microelectrodes, evidence was obtained showing that haloperidol decreased excitability and depressed action potential production by inhibiting the specific increase in sodium conductance (gNa) which normally follows an adequate stimulus. Evidence also was obtained showing an inhibition of the secondary increase in potassium conductance (gK). Haloperidol is structurally related to meperidine and it was found that the inhibition of gNa produced by haloperidol is partially antagonized by low concentrations of naloxone (2.8 X 10(-8) and 2.8 X 10(-7) M); as was previously shown for meperidine. Thus haloperidol, like meperidine, suppresses action potential production by two mechanisms of action: one, a nonspecific local-anaestheticlike effect; and the other, a specific inhibition of gNa mediated by means of an opiate drug receptor associated with the muscle fiber membrane. Naloxone did not antagonize the effects of chlorpromazine on gNa.


Sequential changes in hepatic polyamine, deoxyribonucleic acid, and cyclic adenosine 3',5'-monophosphate metabolism after subacute exposure to cadmium in rats. Heavy metal treatment (2 X 1 mg/kg per day) for 3, 5, and 7 days resulted in progressive augmentation in the incorporation of [14C]thymidine into hepatic DNA. In contrast with the observed enhancement in DNA synthesis, cadmium exposure tended to produce a decrease in the activity of hepatic ornithine decarboxylase (EC 4.1.1.17) at 1, 3, or 5 days with the lowest (34% of control values) enzymic activity seen after 7 days. A similar reduction in the activity of S-adenosylmethionine decarboxylase (EC 4.1.1.50) was observed in livers of rats treated with cadmium for 1-7 days. Subacute exposure to cadmium significantly lowered the hepatic levels of spermidine and spermine whereas the endogenous concentrated of putrescine remained unaltered. In addition to the observed effects on the biosynthesis of polyamines and DNA, heavy metal treatment produced stimulation of the hepatic adenylate cyclase (EC 4.6.1.1)--cyclic AMP system. Significant increases in the activity of hepatic adenylate cyclase and endogenous cyclic AMP levels were detected as early as 1 day and the observed alterations persisted during the entire 1-week period of cadmium exposure. The depression in polyamine formation was accompanied by enhanced DNA biosynthesis as well as stimulation in the adenylate cyclase-cyclic AMP system of rat liver.


Action of a beta-bungarotoxin on autonomic ganglia and adrenergic neurotransmission. A beta-bungarotoxin was isolated from the venom of Bungarus multicinctus by column chromatography on Sephadex G-50 and SP-Sephadex. The toxin produced presynaptic effects on neuromuscular transmission with characteristics similar to those described by others. In a sympathetic ganglion, the toxin increased spontaneous acetylcholine (ACh) release and decreased ACh release evoked by preganglionic nerve stimulation. The toxin did not block the response of isolated ileum to cholinergic nerve stimulation, did not block the release of noradrenaline from the adrenergic nerve terminals of a nictitating membrane preparation, and did not alter the responses of smooth and cardiac muscle preparations to noradrenaline. It is suggested that the specificity of beta-bungarotoxin for certain nerve terminals is related either to selective binding of the toxin or to the selective presence of a necessary substrate for its action. An attempt to show selective binding of 125I-toxin to cholinergic nerve terminals in skeletal muscle was not successful.


Biochemical reactions involved in pancreatic enzyme secretion. 4. Effects of cytochalasin B on functions of the exocrine pancreas. These experiments were performed to evaluate the effects of cytochalasin B on pancreatic enzyme secretion and thus perhaps establish a role for microfilaments in the exocytosis process. The alkaloid at a concentration of 1 microgram/ml (2micron) inhibits amylase secretion induced by urecholine or cholecystokinin-pancreozymin (CCK-PZ) but does not modify that induced by dibutyryl cyclic AMP. The inhibitory effect of the drug is reversible after a 30-min washing out period. It does not affect O2 consumption, basal calcium efflux, or efflux caused by CCK-PZ. Amino acid accumulation in the tissue and their incorporation into proteins are not modified. It is suggested that cytochalasin B inhibits pancreatic enzyme secretion, probably through an effect on the microfilament system.


Angiotensinase activity and angiotensin receptor binding in guinea pig aorta. The angiotensinase (EC 3.4.99.3) activity of the subcellular fractions of guinea pig aorta has been studied in relation to their [14C]angiotensin binding capacity. The enzyme activity occurs in the following decreasing order: supernatant greater than plasma membrane fraction greater than 105 000 X g pellet greater than mitochondrial fraction. The specific binding of [14C]angiotensin to these fractions follows the same pattern. Pretreatment of the subcellular fractions at 47 degrees C for 20 min was performed in an attempt to differentiate binding of angiotensin to the pharmacological receptor from binding to the destroying enzymes. This procedure decreased the angiotensinase activity in the plasma membrane fraction only whereas the specific binding of [14C]angiotensin to this fraction was not significantly decreased, suggesting that the plasma membrane angiotensinase is a thermolabile enzyme.


Urinary cyclic AMP excretion during volume expansion in the rat. We have studied the effect of volume expansion with 0.9% saline solution (10% body weight in 1 h) on urinary cyclic AMP excretion in rats. In nine normal rats urinary cyclic AMP excretion (picomoles/minute) was 113.9+/-9.9 during the control period, rose significantly to 171+/-20 during the first 10 min of expansion, came back to control levels for the next 20 min, and finally decreased significantly to 48.8+/-13.2 at the end of the expansion period. In nine parathyroidectomized rats control cyclic AMP excretion was 78.0+/-4.5, did not rise during expansion, and decreased significantly (to 33.7+/-4.1) at the end of the expansion period. The transient increase in cyclic AMP excretion observed in normal rats could be due to parathyroid hormone release. Our studies do not allow us to explain the subsequent decrease noted in both groups.


Dopamine beta-hydroxylase and cytochrome oxidase activities in brown adipose tissue of newborn rats following sympathectomy with 6-hydroxydopamine. Dopamine-beta-hydroxylase (DBH) (EC 1.14.17.1) activity is present in the interscapular brown adipose tissue (IBAT) as early as 2 days of age in the white rat. The specific and the total activities of this enzyme, as well as those of cytochrome oxidase (COX) (EC 1.9.3.1) in IBAT increase up to at least 20 days of age. Daily administration of 6-hydroxydopamine (6-OHDA) between the second and the twelfth day after birth does not significantly alter IBAT weight gain relative to untreated controls, but the increase in protein content with age is reduced to about half the normal value at the end of the treatment. The treatment with 6-OHDA also results in a drastic lowering of DBH specific and total activities, and a much smaller rate of increase of COX specific and total activities with age in IBAT compared with controls. These results provide additional evidence for a previously proposed role of sympathetic nervous system activity in the development of the thermogenic potential of IBAT in the newborn rat.


ICMSF methods studies. VIII. Comparative study for the enumeration of Clostridium perfringens in foods. Four methods were compared in an international comparative study for the enumeration of Clostridium perfringens: the SFP (Shahidi-Ferguson perfringens) agar (A), TSC (tryptose-sulfite-cycloserin) agar (B), SC (sulfite-cycloserine) agar (C), and neomycin blood agar (D) methods. The confirmed C. perfringens counts were slightly lower for D than for A-C. The percentages of presumptive colonies confirmed as C. perfringens were essentially the same in each method. The relative numbers of nonspecific colonies were the lowest in C, followed by B, D, and A. The methods were also compared for simplicity and for aspects associated with the recognition and selection of presumptive colonies.


Nitrous oxide as end product of denitrification by strains of fluorescent pseudomonads. Growing cultures of several strains of Pseudomonas fluorescens and Pseudomonas chlororaphis produced N2O as the only detectable gaseous product of denitrification, and other strains produced N2 as the gaseous end product of denitrification. All of the nitrogen in NO3- or NO2- added to cell suspensions of the N2O-producing strains P. fluorescens PJ 185 and P. chlororaphis B-560 was recovered as N2O. All of the nitrogen in NO3- or NO2- added to cell suspensions of the N2-producing strain P. fluorescens PJ70 was converted to N2. Cell extracts of P. fluorescens PJ 70, PJ 185, and P. chlororaphis B-560 exhibited NO3- reductase activity when sodium succinate was the electron donor. Reduced nicotinamide adenine dinucleotide and flavine adenine dinucleotide were required to demonstrate NO2- reductase activity in cell extracts.


Enterotoxin formation by Clostridium perfringens type A studied by the use of fluorescent antibody. Fluorescein isothiocyanate-conjugated antibody to purified enterotoxin of Clostridium perfringens was used to study the intracellular formation of enterotoxin by this organism. Enterotoxin was detected at 4 h of growth at the end of the cell containing forespore. With the development of the spore, enterotoxin accumulation continued and involved the entire length of the cell until its lysis with the release of enterotoxin and mature spore. The spores did not contain demonstrable enterotoxin. Only a certain number of the sporulated cells of the enterotoxigenic strains studied produced this toxin. The amount of enterotoxin produced varied with sporulation percentage, and between strains and individual cells.


Malignant parotid tumors: natural history and treatment. Sixty-seven cases of histologically proven malignant parotid tumors are presented. Forty percent of the tumors were differentiated adenocarcinoma, while malignant mixed tumors (or pleomorphic adenocarcinomas) formed 18%. The remaining types of malignant salivary tumors are also represented in the series. The natural history and spread of these tumors are studied in detail, with lymph-node metastasis occurring in 25% of the cases and distant metastasis in 20%. Malignant tumors involve largely the retromandibular portion of the parotid gland in over 40% of the cases and the preauricular portion in about one-fifth of the cases. Both regions are affected in a further fifth of the cases. A combination of surgery and radiotherapy was the method employed for curative therapy, with radiotherapy alone reserved mainly for palliation. While the overall five-year survival was 42%, late recurrences constitute a serious problem with eventual demise of about half of these patients. Various factors affecting the prognosis, including histological type of tumor, method of treatment and response to radiotherapy are discussed.


Radiation-induced cranial nerve palsy.

Radiation-induced cranial nerve palsy. Twenty-five patients with 35 cranial nerve palsies were seen at the Fondation Curie during follow-up after radical radiotherapy for head and neck tumors. The twelfth nerve was involved in 19 cases, the tenth in nine, and the eleventh in five; the fifth and second nerves were involved once each and in the same patient. The twelfth nerve was involved alone in 16 patients and the tenth nerve alone in three, with multiple nerves involved in the remaining six patients. The palsy was noted from 12 to 145 months after diagnosis of the tumor. The latency period could be correlated with dose so that the least square fit equation representing NSD vs delay in NSD = 2598 - Delay (in months) X 4.6, with a correlation coefficient of -0.58. The distinction between tumor recurrence and radiation-induced nerve palsy is critical. It can often be inferred from the latency period but must be confirmed by observation over a period of time.


Lipid cell tumor of the ovary: an ultrastructural study. An ovarian lipid cell tumor without Reinke's crystalloids in a woman with secondary amenorrhea, minimal hirsutism, and elevated 17-ketosteroid excretion was studied by light and electron microscopy. Tumor cells were found in small clumps or scattered singly within a collagenous matrix. The cytoplasm of the tumor cells contained abundant smooth endoplasmic reticula, numerous mitochondria with tubular cristae, lipid droplets, lysosomal dense bodies, and concentric membranous whorls, characteristic of steroidogenic cells. In addition, "peripheral canalicular systems" were found at the outer margins of the nests of the tumor cells. These "peripheral canalicular systems" were bordered by the cell membranes and the surrounding collagenous stroma into which microvilli projected. Since the intercellular canalicular system present between the tumor cells was continuous with the "peripheral canalicular system," both systems probably have a common function related to steroid metabolism. The intercellular and "peripheral" canalicular systems and cytoplasmic microfilaments found in this tumor suggest that this ovarian lipid cell tumor was derived from the ovarian stroma.


Malignant giant cell tumor of soft parts. An ultrastructural study of four cases. Four cases of malignant giant cell tumor of soft parts (MGCT) were studied ultrastructurally. Most of the cells in three cases were identified as undifferentiated mesenchymal cells and mononuclear monohistiocytic cells. The osteoclastlike cells seemed to arise from fusion of monohistiocytic cells. Immature fibroblastic cells, occasionally containing filaments with densities, were also present. They were scarce in three cases and predominant in the fourth. It was concluded that MGCT is a mesenchymal sarcoma with a bimorphic--monohistiocytic and fibroblastic--differentiation. MGCT is therefore considered a special variant of malignant fibrous histiocytoma.


Malignant fibrous histiocytoma. An electron microscopic study. Sixteen cases of malignant fibrous histiocytoma are presented. Electron microscopy of 15 cases demonstrated fibroblast-like and mononuclear and multinucleated histiocyte-like cells. A small capillary was at the center of all storiform areas examined. Ultrastructural examination can be diagnostically useful within the context of a narrow differential diagnosis by conventional microscopy and the ability, by electron microscopy, to eliminate other mesenchymal cell types. In 13 cases, follow-up information was available from 18 months to 9 years following histological diagnosis. Five patients are alive and 8 patients have died, including two non-tumor related deaths. In 3 cases follow-up was less than 4 months. The biologic behavior of the tumor in this series was generally not related to histopathological parameters. The issue of histogenesis is largely unresolvable. Ultrastructural studies of various types of fibrous histiocytomas, suggesting cells of origin other than histiocytes, give credence to the concept that the histiocyte may represent a morphologic state of a given mesenchymal cell rather than a particular cell type.


A new histochemical approach to olfactory esthesioneuroma. A nasal tumor of neural crest origin. A close collaboration between E.N.T. surgeons and pathologists permitted us to obtain 6 cases of olfactory esthesioneuroma in which fresh tumor tissue was available. Histochemical and biochemical studies were made in comparison with neuroblastoma. Different reactions and assays performed revealed a similarity with sympathetic tumors. Moreover, the presence of catecholamines in esthesioneuroma provides a great help in diagnostic as does electron microscope examination.


Serum glutamic oxalacetic transaminase/glutamic pyruvic transaminase ratios in hepatocellular carcinoma. Serum enzyme activities were studied in 131 cases of hepatocellular carcinoma (HCC), 76 cases of metastatic liver carcinomas (MLC) and 234 cases of hepatic cirrhosis. SGOT was elevated above SGPT in most of the time in these patients, SGOT/SGPT was greater in HCC compared with other groups, and that this ratio increased during the preterminal period more markedly in patients with HCC because of the significant increase of SGOT in the face of relatively stable SGPT. Preterminal rises of alkaline phosphatase and LDH activities were more pronounced in MLC. Leucine aminopeptidase activity exhibited no characteristic feature of diagnostic value. Of the five enzymes, SGOT changes were more closely correlated with the growth of HCC; SGPT reflected more of the liver parenchymal damage while SGOT was probably accounted for in part by tumor-derived GOT. Other clinical and pathological implications are discussed.


Cellular immunity to herpes simplex virus in man. VII. K-cell activity to HSV1 infected target cells in disease. Herpes simplex virus-infected target cells, sensitised with antiviral antibody can be lysed by non-immune human lymphocytes (K-cells). This antibody dependent cell mediated immunity (ADCC) is a very efficient technique for destroying foreign target cells and may play a role in maintaining the localization of recurrent herpetic infections. We have used 51Cr labelled herpes virus infected target cells to examine the K-cell activity of peripheral blood lymphocytes in patients who have either malignant reticuloendothelial diseases or who are receiving cytotoxic and steroid therapy for other reasons. K-cell activity was generally within the normal range in such patients and was surprisingly stable despite the occasional use of relatively large doses of cytotoxic agents. It is unlikely, therefore, that a defect in ADCC is responsible for the dissemination of herpes virus infections that may be seen in association with the above drugs and diseases.


Clinical report of the treatment of locally advanced lung cancer. This paper discusses the results of the treatment of 345 patients entered in the Veterans Administration Lung Group Protocol 13L. The study was activated March 1972, and closed for the patient accesion March 1975. All patients had a histological diagnosis of primary lung cancer considered clinically non-resectable or inoperable. Patients were equally randomized into two groups, radiotherapy alone or radiotherapy with chemotherapy. The analysis of the data included: treatment regimen, radiation dose, initial performance status, performance status change, cell type, duration of survival, quality of survival and age. The strongest influence on median survival was the level of radiation dose. The small cell carcinoma patients treated with radiotherapy and chemotherapy showed significant improvement in the median survival (38.2 weeks) over the patients treated with radiotherapy alone (20.6 weeks). The patients treated with radiotherapy and chemotherapy also showed improvement in performance status more frequently than the patients treated with radiotherapy alone. Other parameters of the analysis will be presented.


The viral etiology of cancer: a realistic approach. The etiology of cancer resembles that of many other diseases in that multiple factors may be required. Because of this, the role or viruses in the etiology of human cancers is especially difficult to assess. When animal tumor systems were used as models, the roles of various predisposing characteristics in virus oncogenesis were elucidated. Extrapolation of these findings to the human diseases suggests the importance of genetics, age, hormones, immune competence, and stress in determining susceptibility to tumor development in individuals infected with an oncogenic virus. The importance of cofactors in induction of those human tumors most strongly associated with virus infection, including Burkitt's lymphoma, nasopharyngeal carcinoma, cerviccal carcinoma, acute myelogenous leukemia, and breast cancer, is reviewed. Understanding of the role of these cofactors in virus carcinogenesis may lead to disease prevention through elimination of one or more of the cofactors.


Diet and cancer of endocrine target organs. The role of diet in the etiology of cancer of the breast and endometrium is considered. Geographic variation in rates, effects of migration, consequences of exogenous hormone use, and other epidemiologic factors are reviewed. Cancer of the endometrium is related to diet probably through simple caloric excess. The excess is reflected in obesity and the consequent overproduction of estrone, a natural but carcinogenic human estrogen. Diet may also influence breast cancer risk somewhat through the same mechanism. But this mechanism probably would not explain a large proportion of the disease. It is possible that, in addition, some specific dietary factor, for example, saturated fat, is causally related to breast cancer. Such a relationship would probably be mediated through an endocrine mechanism.


The application in radiation therapy of substances which modify cellular radiation response. Compounds to be considered as potential adjuvants in radiotherapy should indicate a strong mechanistic rationale for a differential response between tumor and normal tissues. Drugs which selectively radiosensitize hypoxic cells might be exploited in radiotherapy to increase the sterilization of resistant hypoxic areas of solid tumors which have outgrown their vascular supply. Several compounds have been shown to selectively radiosensitize hypoxic mammalian cells in vitro but their application in clinical studies in presently limited by their toxicity at the high drug dosages required to effect the sensitization. Phase I and Phase II clinical studies have now been completed with the sensitizer, metronidazole. This drug was tolerated by patients at dosages of 6 g/m2 three times a week for 3 weeks with a minimum of side effects, and survival of patients with glioblastoma multiforme treated with fractionated radiotherapy was significantly increased when the sensitizer was used in combination with the radiation. Drugs which selectively radioprotect oxygenated cells might also be exploited in therapy be permitting the use of higher radiation doses and thereby incurring more damage to the resistant hypoxic cells in tumours. Again, drug toxicity appears to be a limiting factor with this approach. If toxicities are not additive, combinations of radiosensitizers and radioprotectors might prove more effective than either individual approach.


Altered cobalamin distribution in rat hepatomas and in the livers of rats treated with diethylnitrosamine. The distribution of cobalamin cofactors was investigated in the livers and tumors of rats bearing transplanted Morris 7777 or 7800 hepatomas, in the livers of rats treated with the hepatocarcinogen diethylnitrosamine, and in normal rats. There was a significant increase in the proportion of methylcobalamin both in livers and tumors from rat bearing the hepatomas 7777 and 7800 compared to the proportion of methylcobalamin in the livers of normal rats. The total cobalamin content of the hepatomas was significantly lower than that of host or control livers. Similarly, the total cobalamin content of the livers from the tumor-bearing rats was less than that in control animals. The administration to rats of an acute dose of diethylnitrosamine led to an 84% increase in the hepatic concentration of methylcobalamin. Chronic administration of diethylnitrosamine slightly increased the hepatic methylcobalamin concentration, but this was not statistically significant. Liver weight was reduced, and the hepatic content of total cobalamin fell to 55% of that in control animals.


Changes in phosphatidylinositol metabolism correlated to growth state of normal and Rous sarcoma virus-transformed Japanese quail cells. Second-passage Japanese quail embryo cell cultures, normal or quantitatively transformed by Rous sarcoma virus, were investigated for phospholipid composition and metabolism. Cells cultivated at low and high population density as well as in the presence or absence of serum, have been compared by chemical analysis and in pulse-chase experiments. No differences in the lipid compositions between the normal and the tumor cells or between cells under different culture conditions were detected. In no case was the metabolism of phosphatidylserine or sphingomyelin affected by culture conditions. The metabolism of the choline and ethanolamine glycerophospholipids, however, differed according to culture conditions, whether cells were normal or transformed. Significantly, in normal cells, the breakdown of [32P]phosphate-labeled phosphatidylinositol was slowed when cell growth was restricted, i.e., at high population density or in medium without serum. This effect was not observed in tumor cells under such culture conditions, and cells were not growth inhibited. Hence, release of [32P]phosphate from phosphatidylinositol is the only parameter in the metabolism of phospholipids observed to correlate with growth.


A putative role for nicotinamide adenine dinucleotide-promoted nuclear protein modification in the antitumor activity of N-methyl-N-nitrosourea. Incubation of HeLa cells with the anticancer agent N-methyl-N-nitrosourea (MNU) results in: (a) depression of intracellular nicotinamide adenine dinucleotide levels; (b) stimulation of the chromatin-associated, chromosomal protein-modifying enzyme polyadenosine diphosphoribose [poly(ADP-ribose)] polymerase, which uses nicotinamide adenine dinucleotide as substrate; and (c) some fragmentation of cellular DNA. DNase treatment of HeLa nuclei in vitro also stimulates poly(ADP-ribose) polymerase activity, but not in nuclei derived from MNU-treated cells unless they have been subsequently incubated to allow for recovery from MNU damage. DNA polymerase activity is stimulated in vitro by poly(ADP) ribosylation of nuclear proteins. By using intact nuclei derived from MNU-treated HeLa cells, the repair via elongation of single-strand DNA breaks is demonstrated in vitro. This repair is dependent on DNA polymerase activity and is enhanced by adenosine diphosphate ribosylation of histones. Inhibition of poly(ADP-ribose) polymerase with nicotinamide results in extensive degradation of MNU-damaged DNA. Taken as a whole, these results suggest that poly(ADP-ribose) polymerase may play a role in the repair of alkylation damage to cellular DNA and that the inhibition of this enzyme in vivo might be exploited to potentiate the antitumor and carcinogenic activities of MNU.


A new in vitro cell line established from human oat cell carcinoma of the lung. A new tissue culture line, OAT-1975, has been established from explant cultures of human oat cell carcinoma of the lung, characterized and maintained for more than 1 year. The cells grew in a monolayered sheet with 24 hr of population-doubling time and are capable of clonal growth with about 60% plating efficiency. The cells still retain unique morphological and structural characteristics of oat cell carcinoma and showed "tumor takes" when transplanted into nude mice and conditioned young Syrian Golden hamsters. The chromosome constitution of the cells is hypertriploid with a modal peak at 74. The permanent growth integrity of the present cell line made it possible to utilize it for cell biological studies. Cell inactivation experiments indicated that the cells are sensitive to X-rays and Mitomycin C when compared to the responses of HeLa S3 cells.


N,N-Dimethylformamide-induced morphological differentiation and reduction of tumorigenicity in cultured mouse rhabdomyosarcoma cells. N,N-Dimethylformamide treatment of cell cultures established from a transplantable murine rhabdomyosarcoma-induced morphological differentiation and a marked reduction in the tumorigenicity of the sarcoma cells. Fourteen of 17 CE/J mice receiving injections of inducer-treated cells did not develop tumors after 6 months, whereas all 21 mice receiving inocula of untreated sarcoma cells died of disease between 11 and 31 days. The drug-treated cells did not grow in soft agar; untreated tumor cells grew in the semisolid medium. The untreated tumor cells showed a reduced serum requirement and had a higher saturation density compared to drug-treated cells. Thus the reduction in tumorigenicity of N,N-dimethylformamide-treated cells correlated with certain in vitro growth properties that are more characteristic of normal, mesenchymally derived cells than of sarcoma cells.


Tumor-associated antigen in bovine and ovine lymphosarcoma. Specific tumor-associated antigens were found on the membrane and in the cytoplasm of lymph node cells and peripheral blood lymphocytes (PBL) from cattle and sheep with lymphosarcoma by immunofluorescence tests. Materials from 15 cattle with the adult form of lymphosarcoma were examined. Cytoplasmic antigen was detected in fixed tumor cells from all 15 cases and in PBL from 9 cases tested. Membrane antigen was detected in living cells from 10 of the cases tested. In 3 calf-type cases, cytoplasmic antigen was found in a few (1 to 3%) of the tumor cells, while 1% of the cells from 2 thymic cases had cytoplasmic tumor antigen. In 15 cattle infected with bovine leukemia virus (BLV) but with no evidence of tumor, PBL from 3 cattle had the tumor-associated antigen in the cytoplasm. Negative results were obtained with similar tests done with 9 normal cattle that had no detectable BLV or BLV antibody. Cells from tumors induced with BLV in 5 sheep also had cytoplasmic antigen and membrane tumor-associated antigen. Tumor-associated antigen was found in PBL from 1 or 7 BLV-infected sheep with no clinical evidence of tumor. Similar tests were negative on 4 normal sheep.


Nuclear protein phosphokinases in normal and neoplastic tissues. Protein phosphokinases were isolated from the nuclei of normal and fetal liver and neoplastic tissues. Chromatography on phosphocellulose columns resolved the normal and fetal liver kinases into five reproducible fractions. Each of the fractions differed in optimal divalent cation and substrate requirements. Hepatic proliferation was accompanied by quantitative changes in the kinase activity profiles (with endogenous phosphoprotein as natural substrate). An additional phosphoprotein kinase activity stimulated by Mn2+ was found in the nuclei of malignant cells. This tumor-specific kinase could not be detected either in tumor cytoplasm or in fetal or regenerating liver nuclei. Mn2+-dependent phosphoprotein kinase from Novikoff hepatoma phosphorylated only one major protein band detectable by polyacrylamide gel electrophoresis. This substrate could not be detected in chromatin of normal tissues.


Partial purification and characterization of the defective cyclic adenosine 3':5'-monophosphate binding protein kinase from adrenocortical carcinoma. Previous studies with isolated adrenocortical carcinoma 494 cells from this laboratory have indicated that the lack of cyclic adenosine 3':5'-monophosphate (cyclic AMP) control in steroidogenesis in the tumor may be due to the defective cyclic AMP-dependent protein kinase enzyme system. This paper describes the partial purification of such an enzyme. Purification was achieved by precipitation of the tumor homogenate with 30 and 45% ammonium sulfate, adsorption on 3% calcium phosphate gel, and chromatography on DEAE-cellulose. Four major protein peaks were isolated. Peak 2 showed cyclic AMP-binding activity and was investigated further for its kinetic properties. In contrast to the cyclic AMP-dependent protein kinase enzyme found in the normal adrenal gland, the enzyme specifically bound cyclic AMP but failed to phosphorylate exogenous histone. It is postulated that lack of the cyclic nucleotide-dependent kinase activity of the protein kinase enzyme may be responsible for the loss of cyclic AMP-regulated corticosterone synthesis in adrenocortical carcinoma cell. It is further shown that the tumor cyclic AMP-binding enzyme undergoes endogenous phosphorylation, which indicates that it has kinase activity but it is independent of cyclic AMP.


Biological effects of inhibition of guanine nucleotide synthesis by mycophenolic acid in cultured neuroblastoma cells. Mycophenolic acid, an inhibitor of inosinate dehydrogenase, had cytostatic and cytotoxic effects on cultured neuroblastoma cells. Proliferation was inhibited by 50% when cells were incubated with 0.07 micrometerM mycophenolic acid, and cell viability was reduced by 83% when cells were treated with 10 micrometerM mycophenolic acid for 24 hr. Treatment of monolayer cultures with mycophenolic acid reduced intracellular concentrations of guanosine triphosphate by 70% within 3 hr, whereas cytidine triphosphate and uridine triphosphate concentrations were significantly elevated, and adenosine triphosphate concentrations were increased only slightly. Reduction of cellular guanine nucleotides had differential effects on rates of macromolecular synthesis: incorporation of radioactive thymidine into acid-insoluble material was inhibited by mycophenolic acid to a much greater extent than was that of adenosine and leucine. Although proliferation of neuroblastoma cells was inhibited, differentiation, as judged by formation of neuronlike processes in serum-free medium, was unaffected by decreased intracellular concentrations of guanosine triphosphate.


Extraction and analytic procedures for cytosine arabinoside and 1-beta-D-arabinofuranosyluracil and their 5'-mono-, di-, and tri-phosphates. A qualitative and quantitative comparison of perchloric acid (PCA) and trichloroacetic acid (TCA) extraction procedures of biologic material containing cytosine arabinoside (Ara-C) and 1-beta-D-arabinofuranosyluracil (Ara-U) and their biologic 5'-phosphate derivatives revealed definite superiority of the PCA procedure over the TCA procedure. High-pressure liquid chromatography provides rapid and accurate quantitative and qualitative separation of both Ara-C and Ara-U and their 5'-mono-, di-, and tri-phosphates (mu Bondapak NH2 column), or their 5'-mono-, di-, and tri-phosphates without nucleosides (ABX Permaphase columns).


Physiologic disposition of cytosine arabinoside and its derivatives in man. Enzymatic and clinical pharmacologic studies indicate that 2,2'-anhydro-1-beta-D-arabinofuranosyl-5-fluorocytosine (AAFC) is relatively resistant to enzymatic deamination and is not phosphorylated but slowly releases 1-beta-D-arabinofuranosyl-5-fluorocytosine. The latter is deaminated rapidly to 1-beta-D-arabinofuranosyl-5-fluorouracil. After iv injection of labeled cytosine arabinoside (Ara-C), 2,2'-anhydro-1-beta-D-arabinofuranosylcytosine, and AAFC into patients, radioactivity appears in substantial amounts and persists for a prolonged time in the saliva. AAFC slowly penetrates into the cerebrospinal fluid, reaches significant levels, and is retained there for a long time. Ara-C, due to rapid deamination in plasma, does not provide sustained levels of Ara-C in the cerebrospinal fluid. It is likely that this difference between AAFC and Ara-C is due to reduced polarity of the former compound.


Isolation of fat cell precursors from adult rat adipose tissue. The possible existence of adipocyte precursors in adult rat adipose tissue was investigated. Cells were isolated from the stromal fraction of adipose tissue and were grown in culture. Skin fibroblasts were used as controls. The stromal fraction cells were initially fusiform and proliferated; in culture, they accumulated lipid inclusions, became rounder and acquired an eccentric nucleus. In contrast, the skin fibroblasts from the same rat grown under identical culture conditions, did not exhibit any appreciable lipid accumulation. The doubling time for both the stromal fraction cells and skin fibroblasts was 40-60 h. At confluency, the stromal fraction cells contained 5-7 times more glyceride-glycerol than skin fibroblasts. Thus, adipose tissue of adult rats contains cells with the potential to proliferate and acquire morphological characteristics similar to those of adipocytes.


Comparative ultrastructural investigations of the uterine epithelium in the viviparous Salamandra atra Laur. and the ovoviviparous Salamandra salamandra (l.) (amphibia, urodela). The uterine epithelium of the viviparous Salamandra atra and the ovoviviparous Salamandra salamandra was studied in non pregnant and ovulating females and in females during different stages of pregnancy. The epithelium of both species is organized in a monolayer. The epithelial cells are characterized by a moderate secretory activity, a variable amount of apical granules which include PAS-positive material and by some apical and basal exo- or endocytotic vesicles. Adjacent cells are joined by junctional complexes. The lateral surfaces form a tortuous boundary with adjoining cells which suggest that the epithelium is involved in transport. Sporadic light cells possess highly variable cytoplasmic inclusions and are not joined with neighbouring cells. Possibly they represent migratory cells. The entire epithelium, except for a small cranial portion of the uterus in S. atra, undergoes no remarkable morphological changes during the different physiological stages examined except that flattened cells seem to be more numerous in pregnant females. The results are discussed with regard to the possible supply of the developing young by the mother.


Ultrastructure of the respiratory epithelium in the lungs of the tortoise, Testudo graeca. The respiratory epithelium in the lungs of the tortoise (Testudo graeca) has been studied by electron microscopy. The epithelium consists of a mosaic of two different cell types (here called "pneumonocytes"). Type I pneumonocytes are roughly squamous and possess attenuated flanges of cytoplasm which extend over the septal capillaries. Localized cytoplasmic expansions are often present near the periphery of these flanges. Most of the organelles are concentrated in the perinuclear region; the most prominent of these are the mitochondria and osmiophilic inclusions. In contrast, type II pneumonocytes are cuboidal and are richly endowed with organelles including large Golgi complexes, extensive endoplasmic reticulum and numerous inclusion bodies. The morphological evidence suggests that type I pneumonocytes are involved in the secretion of osmiophilic material (presumed to be pulmonary surfactant) and in maintaining the integrity of the air-blood barrier. Type II pneumonocytes appear to be concerned solely with the production of surfactant.


Investigations on the turnover of adrenocortical mitochondria. VIII. A stereological study of the effect of chronic treatment with ACTH and dexamethasone on the morphology of mitochondria in zona glomerulosa cells of the rat. The effects of chronic administration of ACTH and dexamethasone on the morphology of mitochondria in zona glomerulosa cells of the rat adrenal cortex were investigated by stereological techniques. It was found that the volume of the mitochondrial compartment as well as the surface of the outer and inner mitochondrial membranes were significantly increased or decreased in relation to the number of days of ACTH- or dexamethasone-treatment. In ACTH-administered rats, the average volume of individual mitochondria decreased significantly up to the 6th day of treatment and then showed a conspicuous increase from the 6th to the 15th day, whereas in dexamethasone administered animals this parameter, after a small increase during the first 6 days of treatment, displayed a significant decrease. The number of mitochondria per cell showed a dramatic increase during the first 6 days of treatment with ACTH and continued to increase, but only slightly, with the subsequent treatment. In contrast, this parameter showed a parabolic decrease as a function of the duration of treatment in animals receiving dexamethasone. In the light of evidence showing that dexamethasone blocks ACTH-release, these findings are discussed and interpreted to indicate that ACTH is involved in the maintenance and stimulation of the growth and proliferative activity of mitochondria in rat adrenal zona glomerulosa.


Phosphomonoesterases in growth cartilages of the rat. Epiphyseal plate cartilage, epiphyseal cartilage, synchondroseal cartilage and mandibular condylar cartilage were studied morphologically and histochemically in 14 days old rats. Ordinary decalcified paraffin sections were stained with hematoxylin & eosin, van Giesons connective tissue stain, or toluidine blue, and used for morphological studies of the different cartilaginous structures. Undecalcified cryostat sections were used for demonstration of acid and alkaline phosphatase. The enzyme activity was tested for at regular intervals during incubation from 15 sec to 120 min. The morphologic study revealed that a marked similarity of construction exists between epiphyseal plate cartilage and synchrondroseal cartilage. The construction of epiphyseal and condylar cartilage differ from that of the other two structures and also differ mutually. With small variations the reaction for both alkaline and acid phosphatase was found to be identical in the zones of erosion, hypertrophy and maturation of the four structures. Intercellularly, acid phosphatase is present in all zones in the synchondroseal and the epiphyseal plate cartilage, while in the epiphyseal and condylar cartilages it is only present in the zones of erosion, hypertrophy and maturation. The identical reaction for acid phosphatase in the epiphyseal and the condylar cartilage is thought, in all likelihood, to be accidental. When kinetic conditions are taken into account, epiphyseal cartilage seems to react like epiphyseal plate and synchondroseal cartilage, while the condylar cartilage takes up an exceptional position among growth cartilages.


In vitro and in vivo studies on cytodifferentiation of pituitary clonal cells derived from the epithelium of Rathke's pouch. A clonal strain of anterior pituitary cells was derived from Rathke's pouch of the rat. These cells were shown to secrete ACTH, growth hormone and prolactin but no glycoprotein hormones, when grown in vitro. Cells from the 2A8 clone were implanted for one month under kidney capsules or into hypothalami of hypophysectomized female rats. Under the kidney capsule, prominent prolactin cells and poorly developed cells of other types were differentiated as seen in usual pituitary grafts. In hypophysiotrophic areas of the hypothalamus, the grafts were cytodifferentiated into various types of anterior pituitary cells with rich vascularization. These cells had the ultrastructural features indicative of hormone secretion. Increases in body and ovarian weights reflected the secretion of somatotrophic and gonadotrophic hormones. The results obtained indicate that implants of 2A8 clonal cells may differentiate into all types of anterior pituitary cells under the influence of hypothalamic hormones or perhaps some unknown factors present in the general systemic circulation of the rat.


Mechanism of formation of pseudotypes between vesicular stomatitis virus and murine leukemia virus. Pseudotypes of vesicular stomatitis virus (VSV) and Moloney murine leukemia virus (MuLV), defined by their resistance to neutralization by anti-VSV antiserum, are released preferentially at early times after infection of MuLV-producing cells with VSV. At later times, after synthesis of MuLV proteins has been inhibited by the VSV infection, neither MuLV virions nor the VSV (MuLV) pseudotypes are made. Infection of MuLV-producing cells with mutants of VSV having temperature-sensitive lesions in either G or M protein does not generate pseudotypes at nonpermissive temperature, indicating that both proteins are needed for pseudotypes to form. Although the pseudotypes resist neutralization by anti-VSV serum, they are inactivated by anti-VSV serum plus complement, and they can be precipitated by rabbit anti-VSV serum plus goat anti-rabbit IgG. These results, coupled with experiments using a temperature-sensitive mutant of VSV G protein grown at partly restrictive temperature, suggest that small numbers of VSV G protein are obligately incorporated into VSV(MuLV) pseudotypes. There appears to be a stringent requirement for recognition of the viral core by homologous envelope components as the nucleating step in the budding process. Only after such a nucleation can the envelope components of the second virus substitute into the membrane of the budding particle.


Properties of mammalian cells transformed by temperature-sensitive mutants of avian sarcoma virus. Fibroblasts from European field vole (Microtus agrestis) and from normal rat kidney (NRK) have been infected by avian sarcoma virus mutants which are temperature-sensitive for the maintenance of transformation. These cells are transformed at 33 degrees C, but show normal cell characteristics in morphology, colony formation in agar, saturation density, sugar uptake and membrane proteins at 39 degrees C and 40 degrees C, the nonpermissive temperatures. Ts mutant virus was rescued from most of the ts transformed cell lines. NRK cells infected by avian sarcoma virus ts mutants and kept at the nonpermissive temperature can be transformed by wild-type avian sarcoma virus. The susceptibility of the temperature-sensitive NRK lines to this transformation is higher than the susceptibility of uninfected NRK at either permissive or nonpermissive temperature.


Specific folding and contraction of DNA by histones H3 and H4. We demonstrate that the arginine-rich histones H3 and H4 can introduce torsional constraints on closed circular DNA with a concomitant compaction of the nucleic acid. SV40 DNA I complexed with H3 and H4 appears relaxed in electron micrographs and contains particles of 75 +/- 10 A in diameter along the DNA. SV40 DNA I is contracted 2.75 +/- 0.25 fold by all the four smaller histones and 2.6 +/- 0.4 fold by H3 and H4 alone. The arginine-rich histones can cause the topological equivalent of unwinding the DNA close to one Watson-Crick turn per particle formed. Spherical nucleoprotein complexes morphologically similar to isolated nu bodies or nucleosomes are obtained by association of H3 and H4 with 140 base pair length DNA isolated from chromatin core particles. These reconstituted particles sediment at 9.8S, as compared to 10.8S for native core particles, and contain a tetramer of the arginine-rich histones. None of these specific alterations in DNA structure is seen om complexing the slightly lysine rich-histones H2A and H2B to DNA. Our data provide further evidence indicating that the arginine-rich histones are the major determinants of the architecture of DNA within the chromatin core particle.


Distribution of concanavalin A in fibroblasts: direct endocytosis versus surface capping. Indirect immunofluorescence of intact or acetone-extracted cells has allowed us to distinguish concanavalin A (Con A) which is associated with the plasma membrane of CHO cells from Con A which has been interiorized. We find that Con A is directly endocytized by these cells with no intervening stage of plasma membrane aggregation. The lectin accumulations observed by direct fluorescence are actually cytoplasmic collections of pinosomes which contain Con A. Only in a small fraction of CHO cells are true plasma membrane aggregates, or caps, found. This predominance of direct pinocytic interiorization over capping was not affected by dibutyryl cAMP or by treatments which can disrupt microtubules, including cold shock or exposure of the cells to anti-mitotic agents. Cytochalasin B, however, inhibited the uptake of Con A and at the same time promoted the formation of large surface aggregates of the lectin, or minicaps. Capping may reflect a competition between aggregation in the plane of the membrane and direct interiorization of bound lectin. Surface cap formation may be a characteristic process of cells with very low endocytic rates, such as lymphocytes.


[Intestinal vasoactive peptide receptors in enterocytes : specific binding and stimulation of cyclic AMP]. Receptors for the vasoactive intestinal peptide (VIP) were characterized on enterocytes isolated from Rat small intestine. Native VIP inhibited competitively the binding of 125I-VIP to enterocytes and strongly stimulated cyclic AMP production; both these effects were observed for concentrations of VIP as low as 0.5-1.0 ng/ml (0.15-0.30 nM) which are compatible with the VIP concentration in the gut.


[Biological and biochemical characterization of a new isolate of baboon Papio papio endogenous type C virus]. A new strain of Baboon Papio papio endogeneous type C virus was isolated from the testis of an animal of this species. The strain was characterized biochemically and biologically. It presents a host-range different from that of the prototype strain of the same virus.


A simple and sensitive method for estimating the concentration and synthesis of 5-phosphoribosyl 1-pyrophosphate in red blood cells. A method is presented for the determination of 5-phosphoribosyl 1-pyrophosphate (PRPP), which is based on the release of 14CO2 from [carboxyl-14C]-orotic acid by the consecutive action of orotate phosphoribosyltransferase and orotidine-5'-monophosphate decarboxylase. The assay is simpler and less time-consuming than most methods currently employed and is equally sensitive. The method proved to be suitable for measuring low concentrations of PRPP such as found in human erythrocytes and fibroblasts. An increased PRPP concentration was observed in erythrocytes from patients with partial or complete deficiency of hypoxanthine-guanine phospho-ribosyltransferase. frp, sp,e (but not all) gouty patients and from a patient with deficiency of purine nucleoside phosphorylase. PRPP synthetase activity was measured with a method similar to the assay for PRPP. In erythrocytes with an increased PRPP concentration, PRPP synthetase activity was found to be normal at both optimal and suboptimal substrate concentrations.


Studies on a beta-migrating high density lipoprotein. An unusual serum lipoprotein (Lp) profile was detected in a Japanese family. A double beta-Lp was observed when serum was subjected to polyacrylamide gel electrophoresis. The slower migrating beta-Lp was identified as a subfraction of high density lipoprotein (HDL). It was present in the d 1.063--1.21 fraction, migrated to the position designated as the midband L.1 (sinking pre-beta-lipoprotein) by Mead, M.G. and Dangerfield, W.G. (1974) (Clin. Chim. Acta 51, 173--182) [1], and reacted against human anti-beta-Lp antiserum. This lipoprotein contained greater amounts of triglyceride than the usual beta-lipoprotein and could not be clearly detected by paper electrophoresis. Individuals exhibiting this high density midband lipoprotein appeared to be heterozygous for an autosomal dominant gene. Although other reports have indicated the possibility of a positive association between the occurrence of serum lipoproteins with unusual eletrophoretic mobility and premature ischemic heart disease, no such correlation was demonstrable in these subjects.


Lipoprotein concentrations in untreated adult onset diabetes mellitus and the relationship of the fasting plasma triglyceride concentration to insulin secretion. Plasma lipoprotein concentrations in 64 untreated adult onset diabetics were compared to the lipoprotein levels found in non diabetics. No significant difference was found. The relationship of the fasting plasma triglyceride concentration to the plasma immunoreaction insulin response to 50 g glucose orally was investigated. Using correlation analysis a significant and insulin response in the non diabetics but not in the diabetics.


'Big ACTH' and calcitonin in an ectopic hormone secreting tumour of the liver. A young man presented with rapidly developing Cushing's syndrome which was due to the ectopic secretion of ACTH and beta-MSH-like material from hepatic tumour deposits, possibly originating from biliary radicals. This association of the ectopic ACTH syndrome has not previously been described. During the 22 month course of the illness the plasma immunoreactive ACTH and 'beta-MSH' concentrations rose by logarithmic progression. The plasma calcitonin concentration was also raised but did not change during the last 12 months. At any stage of the illness the plasma concentration of the ecotopically produced hormones was stable except that after hydrocortisone there were inconstant variations. During the course of the illness the ectopic ACTH became biologically less potent. This ineffectural ACTH was present in the plasma, in the tumour, and in the medium in which the tumour was cultured, in a large molecular weight form. This 'big ACTH' differed from the normal ACTH found in the patient's pituitary and from authentic ACTH in its immunochemical character: the C-terminal antigenic determinant (33-39 region of ACTH) was masked in the large molecular weight form but was uncovered after extraction in neutral buffer and this 'big ACTH' was more readily extracted from the tumour at pH 7.0. The tumour tissue also contained immunoreactive beta-MSH-like material and immunoreactive calcitonin which resembled calcitonin M chromatographically.


Intractable diarrhoea of infancy.

Intractable diarrhoea of infancy. The intractable diarrhoea syndrome of infancy continues to be a major diagnostic and therapeutic challenge to the paediatrician and paediatric gastroenterologist. A carefully organized, staged approach to diagnosis will provide the best method of identifying those infants in whom a specific aetiology exists and for whom specific therapy is often available. Regardless of aetiology, however, the early use of appropriate nutritional support will not only reduce morbidity and mortality in these infants, but will prevent the development of many of the secondary consequences of malnutrition. The physician must compulsively pay attention to the details of daily management and provide an organized approach to diagnosis and treatment in order to improve the outcome of infants with intractable diarrhoea.


Interaction of isolated Lp(a) lipoprotein with calcium ions and glycosaminoglycans in vitro. The interaction between the lipoprotein carrying the Lp(a) antigen, i.e. the Lp(a) lipoprotein, and agarose gels substituted with glycosaminoglycans, as well as the precipitation of the Lp(a) lipoprotein by Ca++ were studied. Comparisons between Lp(a) lipoprotein and other serum lipoproteins were conducted. Very low density lipoprotein (VLDL) and low density lipoprotein (LDL) were bound to the tested glycosaminoglycans at low ionic strength of sodium chloride, but no binding was found with the Lp(a) lipoprotein. However, Ca++ as a divalent buffer cation gave a precipitating Ca++-Lp(a) lipoprotein complex even at a physiological Ca++ concentration. VLDL and LDL were not precipitated under these conditions. These findings may be of interest in relation to the previously reported higher frequency of the phenotype Lp(a+) in subjects with coronary heart disease.


Unchanged hormone sensitivity of rat fat cell adenylate cyclase in uremia. The sensitivity to hormones of the fat cell adenylate cyclase system was tested in uremic rats and in pair-fed control animals. Basal enzyme activities averaged 1.25 nmoles of cAMP formed per mg protein per 15 min in controls compared to 1.30 nmoles cAMP/mg protein/15 min in fat cell ghosts obtained from uremic rats. NaF caused an approximately 4-fold stimulation of enzyme activities in both systems. It was shown that parathyroid hormone should be included amongst the hormones which act as stimulators of the enzyme system. The responsiveness of the rat fat cell adenylate cyclase system towards saturating concentrations of ACTH, glucagon, epinephrine and parathyroid hormone was not altered in the presence of chronic renal failure.


The role of prostaglandins in experimental arthritis in the rat. In the experimental model of streptococcal CFE-induced arthritis, PGE and PGF concentrations rise significantly. This is due to an increase both in the substrate concentration and the enzyme activity for PG biosynthesis. Indomethacin therapy inhibited PG synthesis and diminished some of the gross, but not the histological, features of the inflammatory response. These results lend further support to the contention that the beneficial effect of indomethacin in arthritis is due to inhibition of PG biosynthesis.


The effect of apomorphine and clonidine on locomotor activity in mice after long term treatment with haloperidol. 1. Mice were given haloperidol (approximately 3 mg.kg-1 day-1) or vehicle for 21 days and then withdrawn from the drug. All tests were performed 4 days after withdrawal. 2. Haloperidol treated mice (premedicated with reserpine plus alpha-methyl-p-tyrosine) displayed an increased locomotor response to apomorphine and to apomorphine plus clonidine, but neither haloperidol- or vehicle-treated animals revealed any stimulant response to clonidine. 3. In mice which had not been pretreated with reserpine plus alpha-methyl-p-tyrosine, clonidine produced a significant stimulation of locomotor activity in animals withdrawn from haloperidol but not in those withdrawn from the vehicle. Phenoxybenzamine blocked the locomotor stimulat difference between these two groups, but did not completely antagonized the stimulant effect of clonidine in mice withdrawn from haloperidol. Pimozide was largely effective in blocking the clonidine-induced stimulation. Co-administration of phenoxybenzamine and pimozide was completely effective in blocking the stimulant effect of clonidine in mice withdrawn from haloperidol. 4. The evidence for a change in catecholamine receptor sensitivity was supported compared to the vehicle-treated animals. 5. The data suggest that there is a change in the functional responsiveness of both adrenergic and dopaminergic receptors after withdrawal from long term haloperidol treatment.


Synthesis of estradiol-17 beta by Sertoli cells in culture: stimulation by FSH and dibutyryl cyclic AMP. Sertoli cells isolated from the testes of 18 to 20-day old rats synthesize estradiol-17 beta when grown in primary culture in the presence of testosterone (5 X 10(-7) M). After an initial lag phase of about 2 hours, follicle-stimulating hormone (FSH) stimulates synthesis of estradiol up to 50-fold, synthesis being approximately linear for 24 hours when medium is changed every 2-6 hours. Luteinizing hormone (LH) causes only a marginal stimulation, at concentrations consistent with contamination with FSH. Dibutyryl cyclic 3',5'-adenosine monophosphate (dbcAMP) (10(-4)M) mimicks the effect of FSH. No significant synthesis occurs in the absence of steroid substrate or in the presence of pregnenolone (5 X 10(-7) M). When cells are pre-incubated for 24 hours with FSH in the absence of testosterone, addition of testosterone results in an immediated increase in estradiol synthesis without a lag phase, suggesting that FSH or dbcAMP are capable of inducing the conditions necessary for stimulation in the absence of testosterone. These observations provide the first direct evidence of estradiol-17 beta synthesis by Sertoli cells from normal animals, and offer evidence that synthesis of this steroid is regulated at the level of the aromatizing enzyme system by FSH and cyclic AMP.


Angiofibromata mimicking viral warts.

Angiofibromata mimicking viral warts. A case is presented of an otherwise healthy woman with a relatively normal childhood and adult life and no evidence of any congenital anomaly, who at the age of twenty-seven, suddenly, almost explosively, had development of verruca-like lesions of the face. It is well to remember that the clinically obvious is not always the obvious, things are not always what they seem, and verrucae are not always viral warts.


[Hypertrophic obstructive cardiomyopathy and lentiginosis (author's transl)]. Progressive cardiomyopathic lentiginosis was first described in 1972 by Polani and Moynahan. It is characterised by pigmentation of the skin (multiple symmetrical lentigines), hypertrophic obstructive cardiomyopathy and retarded growth. There may also be mental retardation. All characteristics of the syndrome were present in a 46-year-old woman. In addition there were also other features: lentigo developed at only 35 years of age, there was hypertelorism, thoracic kyphosis, and intermittent severe depression. Cyclic adenosine monophosphate in plasma was raised to 90.5 nmol/1.


[Histological results of lumps excised from the breast after aspiration (author's transl)]. In 614 breasts tumours assumed to be harmless cysts on palpation were aspirated. In 430 cases aspiration was the definitive treatment. In the other 184 cases excision was necessary in order to obtain histology. 11 cysts were radiologically suspect after air filling. In one case excision demonstrated an undifferentiated milk duct carcinoma. 12 cysts had radiologically suspicious areas outside their margins. In 4 cases lobular carcinoma in situ was present. 161 excisions were necessary because no fluid was aspirated at puncture of the tumour. In 10 cases histology showed malignancy: half of these were metastases from a known primary tumour and half were primary breast carcinomas. Out of 17 precancerous states neoplasia could be demonstrated in two cases in addition to papillary and proliferative duct changes. We have classified this neoplasia as lobular carcinoma in situ.


[Oestrogen treatment of girls with increased growth (author's transl)]. Forty-one pubescent girls of constitutionally tall stature were given conjugated oestrogens for a continuous period averaging twenty-two months. Oral progestin was also given, daily for five days during the second half of each menstrual cycle. Average predicted height of the girls was 186.5 cm, average final height was only 179.2 cm. In half of the girls in whom treatment had been started before menarche the height reduction was 2 cm greater than in those started after menarche. There were no severe side effects. Extreme care should be taken to limit strictly the indications for this treatment. Only girls with a growth prognosis above 183 cm should be considered. Therapeutic overstimulation of the endometrium must be avoided. Earlier views that the cessation of growth under oestrogen is due to early epiphyseal closure have been shown to be wrong. Growth frequently ceases long before epiphyseal closure. Reduction of somatomedin production was found to be the cause. The average serum somatomedin level fell to 56.9% of basal concentration after six months' treatment. At the same time, the circadian profile of growth hormone indicated a threefold increase above basal level. This phenomenon probably reflects a negative feedback mechanism between growth hormone and somatomedin.


Hexoprenaline: a review of its pharmacological properties and therapeutic efficacy with particular reference to asthma. Hexoprenaline1, N,N-[2-(3,4-dihydroxyphenyl)-2-hydroxyethyl] hexamethyl-enediamine, sulphate is a selective beta2-adrenoreceptor agonist which is active in man as a bronchodilator by the oral or intravenous routes and by inhalation. It is indicated for use in the treatment of bronchospasm associated with obstructive airways diseases, including asthma, bronchitis and emphysema. Clinical experience and double-blind studies have established that hexoprenaline is an effective bronchodilator. It major advantage over many other many other brochodilators of equal efficacy is its generally low production of side-effects, particularly tremor, palptitations, and tachycardia. In comparative trials, it has generally been rated as superior to orciprenaline or trimetoquinol, but comparisons with salbutamol have provided equivocal results. Oral hexoprenaline was superior to fenoterol as long-term maintenance therapy is asthma, principally because its somewhat lesser bronchodilatory effects were more than compensated for by a lesser incidence of side-effects.


[Aspects of bonding and fusing of materials meltable by lasers (caries prophylaxis and therapy)]. Experiments are described which aim to combine a protective cover with the enamel. Borium proved to be a good absorbtive mediator and phosphate glass was well suited as ground substance.


Stability of nuclear triiodothyronine binding sites. Rat liver nuclei have recently been shown to possess limited, high affinity binding sites for triiodothyronine, only a fraction of which are believed to be filled in vivo. We have found that preincubation of nuclei in the absence of added triiodothyronine leads to a rapid decrease in binding capacity. This loss of binding sites is considerably faster at 37 C than at 4 C. On the other hand, once binding equilibrium is reached between nuclear bound and free triiodothyronine the level of binding does not change during the time in which a considerable loss of binding had occurred when nuclei were preincubated in the absence of hormone. Also the rate of dissociation of triiodothyronine from nuclei is slower than the rate of loss of binding sites during preincubation in the absence of added hormone. Thus, it appears as if unfilled sites are being lost during the preincubation period. Administration of cycloheximide or actinomycin D, inhibitors of protein and RNA synthesis, respectively, to rats caused a decrease in the amount of triiodothyronine which could be bound in the in vitro binding assay to liver nuclei.


Modification of serum luteinizing hormone and prolactin concentrations by corticotropin and adrenalectomy in ovariectomized rats. The effects of ACTH and adrenalectomy (ADX) on serum concentrations of LH and prolactin were examined in ovariectomized (OVX) rats. In the first study ACTH (4 IU) was administered daily for one week. ACTH prevented an increase in serum LH in response to OVX and stimulated serum prolactin. ACTH also reduced serum LH in OVX+ADX rats and increased prolactin concentration. ADX was utilized in the second experiment to enhance secretion of endogenous ACTH. Two weeks after ADX serum prolactin was elevated above intact values. ADX also enhanced serum prolactin in OVX rats to the level found in intact animals. ADX markedly reduced the LH response to OVX but ADX alone had no effect on serum LH concentration. All of these findings indicate that ACTH acts directly to reduce serum LH concentration and to enhance serum prolactin. The influence of adrenal corticoids on serum concentrations of LH and prolactin as reported by other investigators appears to be mediated via their negative feedback effects on ACTH release. Whether ACTH has its action on adenohypophyseal cells or on hypothalamic sites has yet to be determined.


Regulation of gonadotropin receptors by luteinizing hormone in granulosa cells. In association with luteinization, LH induces a decrease in the content of receptors for FSH and LH and an increase in that for PRL. To elucidate if the mechanism by which LH regulates its own receptors involved occupancy of sites and/or loss of receptors the effects of a luteinizing dose of LH were examined in the preovulatory follicles of immature hypophysectomized rats primed with estradiol and FSH. The measurable LH receptor content declined by 82% 24 h after LH administration. Serum concentration of the hormone declined by 24 h to 1.4% of the concentration measured 2 h after LH administration. Administration of iodinated LH to demonstrate occupancy of sites in vivo, resulted in a decline in the amount of hormone bound in vivo, over a period of time. This decline in occupancy paralleled the decrease in the number of available sites as measured in vitro. Furthermore, a large dose of highly purified hFSH administered in lieu of LH induced luteinization and an associated loss of gonadotropin receptors. These results indicate that luteinizing doses of LH and FSH induce a loss in gonadotropin receptors by mechanisms other than occupancy.


Isolation of highly purified Leydig cells by density gradient centrifugation. Purification of specific endocrine cells from mixed populations after dispersion of target tissues is important for detailed analysis of mechanisms of hormone action. A simple method for rapid isolation of endocrine cells with retention of biological integrity, has been developed by centrifugation in density gradients formed with Metrizamide. By this procedure, highly purified Leydig cells retaining morphological and biochemical characteristics were obtained. Such preparations bound 20, 300+/-3, 100 molecules of hCG per cell with affinity of 1.1+/-0.25 X 10(10) M-1. During incubation with hCG, cyclic AMP and testosterone responses of purified Leydig cells were considerably increased, and hCG concentrations as low as 0.2 pM caused activation of cAMP-dependent protein kinase.


Identification of multiple protein kinases in normal human lymphocytes. The protein kinase activity of a 10,000 g supernatant of purified human lymphocytes can be resolved by DEAE-cellulose chromatography into six protein kinase fractions: three of them phosphorylate casein preferentially, and three histones. The same procedure with the corresponding nuclear fraction yields only two casein kinases. All these fractions, except one casein kinase of the cytosol, have been studied with respect to protein and nucleotide specificity, effect of salts and of cyclic nucleotides, sedimentation, etc. The results obtained indicate that the enzyme fractions of the cytosol have distinct characteristics, suggesting that they are different protein kinases, and that the nuclear kinases are similar to the two main casein kinases of the cytosol.


Characteristics of modulators and substrates binding to rat liver adenylate kinase. The binding of adenine nucleotides to liver adenylate kinase was dependent on Mg2+ ions. Citric acid enhanced the binding of all metal-chelated radioactive nucleotides and indicated two observable binding sites for Mg3H-ADP and Mg3-ATP and one-half binding site for Mg3H-AMP. Two binding sites of Mg3H-ADP and one binding site for Mg3H-ATP were also observed in the absence of citric acid. Stoichiometric binding of 14C-citric acid to liver adenylate kinase varied with additions of different nucleotides. AMP prevented whereas ADP and ATP enhanced the binding of 14C-citric acid.


New protein reagents. N-(4-Chloromercuriphenyl)-4-chloro-3,5-dinitrobenzamide and its use as a probe of the quaternary structure of yeast alcohol dehydrogenase. The new bifunctional reagent, N-(4-chloromercuriphenyl)-4-chloro-3,5-dinitrobenzamide (I) was used to investigate the quaternary structure of yeast alcohol dehydrogenase. The four essential - SH groups of the enzyme were substituted by the mercuriphenyl moiety of compound I in the course of the reaction of one mole of protein with four moles of the reagent (one molecule of compound I incorporated by yeast alcohol dehydrogenase monomer). In a second step only two of the four chlorodinitrophenyl fragments bound to the protein established intermonomeric cross-links with non-essential - NH2 groups. The resulting dimers could be re-dissociated with mercaptoethanol. This result suggests that the four protomers of the enzyme could be arranged as a dimer of dimers.


Proteolysis of L-(+)-lactate cytochrome c oxidoreductase (cytochrome b2) extracted from Saccharomyces cerevisiae and Hansenula anomala yeasts. The L-(+)-Lactate:cytochrome c oxidoreductase or cytochrome b2 from the yeasts Saccharomyces cerevisiae and Hansenula anomala were partially hydrolysed in various concentrations of trypsin. Conditions were found which allowed the isolation from the Hansenula enzyme of a 140 000 +/- 10 000-dalton flavoprotein. The prosthetic flavin groups were still reducible by substrate (spectroscopic evidence) but the flavoprotein was unable to form a complex with cytochrome c, the physiological acceptor in the enzymatic reaction. No such flavoprotein units could be found during proteolysis of the Saccharomyces enzyme. The heme prosthetic group of the Hansenula enzyme remained bound to a 15 500 +/- 1000-dalton protein unit which was larger than, but very similar to, the well known 'cytochrome b2 core' of the Saccharomyces enzyme. Moreover, the degradation of different enzyme samples by contaminated proteases allowed the isolation of a particular form of Hansenula enzyme: each tetramer had, on the mean, four bound flavins and only two heme groups. These molecules completely retained their ability to form a complex with cytochrome c.


Particle-bound enzymes in the bloodstream form of Trypanosoma brucei. We have screened the bloodstream form of Trypanosoma brucei for the presence of enzymes that could serve as markers for the microbodies and the highly repressed mitochondrion of this organism. None of seven known microbody enzymes were detected at all, but glycerol-3-phosphate oxidase, ATPase, isocitrate dehydrogenase, acid phosphatase and part of the hyperoxide dismutase and malate dehydrogenase activities were found to be particle-bound after fractionation of homogenates by differential centrifugation. Part of the ATPase activity was sensitive to oligomycin, an inhibitor of oxidative phosphorylation. This oligomycin-sensitive activity can serve as a specific marker for the mitochondria. More than 80% of the NAD+-linked glycerol-3-phosphate dehydrogenase in T. brucei was found to be particulate and latent. The enzyme could be activated by Triton X-100, by the combined action of sonication and salt, but not by salt alone, and partially by freezing and thawing. We conclude that the NAD+-linked glycerol-3-phosphate dehydrogenase is located inside an organelle.


A comparative analysis of the protein components of plasma membranes isolated from differentiated and undifferentiated mouse neuroblastoma cells in tissue culture. The plasma membranes of the cells of mouse neuroblastoma clone NB41A, were isolated without fixation by hardening procedures and were characterised by their enzyme activities and by their morphology in the light and electron microscopes. The membranes were prepared from two kinds of differentiated monolayer cultures; one in which the cells were induced to form long axon-like processes by the addition of N6,O2'-dibutyryladenosine 3':5'-monophosphate to the culture medium, and the other in which the cells were induced to form processes by the addition of 5-bromo-2'-deoxyuridine. The proteins from the solubilised membranes were compared with similar preparation from the membranes of undifferentiated cells, grown in suspension, by sodium dodecylsulphate polyacrylamide gel electrophoresis, using the incorporation of radioactive amino acids and L-fucose into proteins in the cultures to follow the differences in the patterns of polypeptide synthesis and glycosylation of the membrane proteins. The differentiated cells induced with either inducer show an increased incorporation of L-fucose into two distinct components with molecular weights of 60 000 and 70 000. The two types of induced cells differ from each other in that N6,O2'-dibutyryladenosine 3':5'-monophosphate stimulates both glycosylation and protein synthesis, with a relative increase in the low molecular weight proteins, but 5-bromo-2'-deoxyuridine stimulates mostly increased glycosylation of the membrane proteins.


On the interrelationship of prostaglandin endoperoxide G2 and cyclic nucleotides in platelet function. The prostaglandin endoperoxide G2 caused rapid aggregation and relase of ADP and [14C]serotonin in human platelets. Since the presence of the ADP phosphorylating system creatine phosphate/creatine phosphokinase markedly inhibited the aggregation caused by the endoperoxide, this effect seemed to be mediated mainly by ADP, which is instantaneously released by the endoperoxide. The prostaglandin G2 counteracted the increasing effect of prostaglandin E1 on the adenosine 3':5'-monophosphate (cAMP) levels in platelet-rich plasma. This effect of prostaglandin G2 was only observed when ADP was released by the endoperoxide. This finding indicates that the effect of prostaglandin G2 on the cAMP levels in platelet-rich plasma is principally mediated by ADP. The rapid release of ADP by prostaglandin G2 and the time courses for the effects of the endoperoxide and ADP on the level of cAMP give further evidence for this hypothesis. ADP also caused primary aggregation in the presence of indomethacin, and prostaglandin synthesis inhibitors did not influence the decreasing effect of ADP on the cAMP levels. N2,O2-Dibutyrylguanosine 3':5'-monophosphate did not influence the aggregation and release-reaction caused by ADP and no changes of the cGMP levels were observed after addition of prostaglandin G2.


Activity of protein kinase dependent on adenosine 3':5'-monophosphate and of its thermostable protein inhibitor in rat hepatoma (HTC) cells. Unlikely role in the permissive action of glucocorticoids. Normal expression of a variety of hormonal effects which depend on cyclic AMP (adenosine 3':5'-monophosphate) requires the presence of glucocorticoids. Our hypothesis was that glucocorticoids control directly or indirectly the activity of cyclic-AMP-dependent protein kinase. This has been investigated in cultured hepatoma (HTC) cells in which N6,O2'-dibutyryladenosine 3':5'-monophosphate increases the activity of tyrosine transaminase only after glucocorticoid treatment. In these cells, we have determined the concentration and half-life of protein kinase, the sensitivity of this enzyme in vitro to cyclic AMP and to its thermostable protein inhibitor, the state of dissociation of protein kinase holoenzyme in vivo and its sensitivity, in the intact cell, to dibutyryladenosine 3':5'-monophosphate and to the inhibitor diamide, and we have also determined the concentration of endogenous thermostable protein inhibitor of protein kinase. None of these parameters were influenced by glucocorticoids under conditions where these hormones stimulate the activity of tyrosine transaminase and restore sensitivity to dibutyryladenosine 3':5'-monophosphate. It is concluded that the permissive action of glucocorticoids probably results from a control of cyclic-AMP-dependent processes exerted at a level beyond the protein kinase system.


The major polypeptides of the murine-mammary-tumor virus isolated by plant-lectin affinity chromatography. Solubilized polypeptides of the murine mammary tumor virus (MuMT virus) were chromatographed on a column of immobilised concanavalin A. The unbound viral material was rechromatographed on phosphocellulose, resulting in the isolation of the major proteins with a molecular weight of 28000 (p28) and 12000 (p12) respectively. The adsorbed glycopolypeptides after elution with methyl alpha-D-mannopyranoside were subjected to gel filtration. The major glycoprotein with a molecular weight of 52000 (gp52) was obtained in an almost pure form. However, a considerable part of gp52 elutes together with a glycoprotein with a molecular weight of 36000 (gp36), suggesting that in addition to the free form of gp52 a complex exists of gp52 plus gp36.


Analysis of the relA gene product of Escherichia coli. The relA gene product, ATP: GTP 3'-pyrophosphotransferase (stringent factor) has been isolated in homogeneous form from an Escherichia coli strain polyploid for this gene at a yield of 1 mg/100 g cells and at a specific activity in a ribosome-activated assay at 37 degrees C of 120 mumol guanosine pentaphosphate formed min-1 mg protein-1. The specific activity in a methanol-activated assay at 25 degrees C was found to be 4 mumol guanosine pentaphosphate formed min-1 mg protein-1. These values are about 100 times higher than reported by others. Our further studies of this enzyme led to the following results. Antibodies raised against this enzyme inhibit the ribosome-activated synthesis of guanosine tetraphosphate and pentaphosphate but have no effect on the much slower synthesis, detected in the absence of ribosomes. The amount of stringent factor in the relA+ strain CP78 is estimated to about 1 copy per 200 ribosomes. The amount of antibody-binding material in CP79 (relA) is at least 5 times lower.


Ultrastructural features in a case of ureteric malakoplakia. A case of urinary malakoplakia involving both ureters and the bladder is presented. Ultrastructural data relating to the ureteric lesion are illustrated. Pale, dark and intermediate cells were involved in the pathological process. Features common and specific for these three types of cell are described. Dark cells are thought to be of a histiocytic nature, while the others are of either histiocyte or urothelial origin. Cytoplasmic inclusions were abundant in all three types in the form of single-or multicentred multilamellar bodies. It was not clear whether these were the product of secretion or phagocytosis. Michaelis-Gutmann bodies, on the other hand, were found to be due to the deposition of needle-like crystals inside pre-existing multicentred multilamellar bodies. Pathogens could not be demonstrated for certain. It is felt, therefore, that the aetiology of malakoplakia must be considered as debatable. Biochemical and immunological research directed to the detection of a possible enzyme deficiency is suggested, along with further ultrastructural investigation of the histogenesis of the pale and intermediate cells.


Post-nephrectomy changes in adrenal angiotensin II receptors in the rat; influence of exogenous angiotensin and a competitive inhibitor. 3H-angiotensin binding sites have been studied in a particulate fraction prepared from rat adrenal glands. This binding is rapid and reversible, of high affinity (KD29 degrees C = 3-5 X 10(-9) M) and with demonstrable specificity for the angiotensin II octapeptide. The number of binding sites varies with endogenous angiotensin levels: nephrectomy is followed by an increase in number of binding sites. This increase can be prevented by chronic angiotensin II administration and, to a lesser extent, by administration of Sar1,Ala8-angiotensin II, a competitive antagonist of the hormone. No variation in the equilibrium dissociation constant accompanied these changes in binding capacity. The post-nephrectomy increase in capacity is time-dependent, with a lag period of 24-40 h. The observed changes in receptor concentration do not appear explicable on the basis of receptor occupancy. Accordingly, angiotensin II receptors in the rat adrenal appear to be dependent on circulating angiotensin levels as previously reported for rat uterus.


The mechanism of action of two bradykinin-potentiating peptides on isolated smooth muscle. Bradykinin-induced contractions in the guinea-pig ileum were potentiated by the peptides A-VI-5 (Val-Glu-Ser-Ser-Lys) and BPP5a (Pyr-Lys-Trp-Ala-Pro), while the contractions induced by other agonists were not affected. Neither peptide added alone caused any response. Previous addition of the peptides shortened the latent period following the addition of bradykinin to a value corresponding to the contraction height with an equivalent dose of bradykinin added alone. Bradykinin in contact with a piece of ileum was inactivated at a relatively slow rate. This inactivation was not inhibited by either A-VI-5 or BPP5a in doses causing potentiation. Suppression of the cholinergic activity by cooling, atropine, morphine or tetrodotoxin did not influence the potentiating activity. Addition of the peptides at the moment a submaximal contraction due to bradykinin had been fully established, increased the contraction height within seconds. The two peptides caused a parallel shift to the left of the dose-effect curve of bradykinin, whereas the maximum bradykinin effect remained unchanged. It is concluded that sensitization of bradykinin receptors due to an increased affinity of the receptor for bradykinin is the hypothesis which best fits the experimental findings.


A study of binocular convergence in cat visual cortex neurons. The average latency of cortical neuronal responses to electrical optic nerve (ON) stimulation was 3.0 +/- 0.7 s.d. msec. No significant difference between latencies to ipsi- and contralateral ON stimulation was found. Binocularly excitable cells showed almost equal response latencies to stimulation of both nerves. The average latency of subcortically recorded geniculo-cortical fibers was 0.3 msec less, but showed the same variance as that of cortical cells, suggesting that in all cases direct monosynaptic excitation of cortical cells by fibers of either ocularity is possible. Classes of ocular dominance based on electrical stimulation were positively, but not 100% correlated with classes of ocular dominance to visual stimulation. An anatomical study revealed that in cat terminals of geniculo-cortical projection are segregated to a lesser degree into ocularity stripes than in monkey. Direct monosynaptic excitation of cells by fibers of either ocularity which was found physiologically would also on these grounds appear possible for all cells.


Maturation of evoked climbing fiber input to rat cerebellar purkinje cells (I.). An analysis of evoked responses of Purkinje cells in developing rat cerebellum to climbing fiber input was conducted to determine which identifying properties of this afferent system are established early in development and which specific features mature with age. Rat pups at various ages were anesthetized with 0.5% halothane and unit recordings made with glass micropipettes. By the third postnatal day, electrical stimulation of the sensorimotor cortex and limbs at low stimulation rates (less than 1/sec) could elicit distinct burst responses appearing at long latencies (180 msec), indicating that pathways of both ascending and descending climbing fiber systems are intact early in cerebellar cortical development. A distinctive feature maturing over the first 1-1.5 weeks was the characteristic of the all-or-none burst response since before about day 11 the interspike interval, amount of inactivation, and number of spikes in evoked burst responses all varied from stimulation to stimulation. Mean latencies decreased from 180 msec at dat 3 to 50 msec by day 10, but did not achieve the adult value of 20 msec until the fourth week. Typically, climbing fiber responses could only follow at stimulation rates of less than 0.2/sec at day 3, but by day 12 could follow up to 10/sec, which is the same as in the adult. The data indicate that the climbing fiber system establishes connectivity from diverse sources and exhibits identifying characteristics similar to the adult early in cerebellar development. Most aspects of the maturation of transmission can be explained if there were a decrease in the time scale of function of the synapses involved, mainly those in the inferior olive.


The buoyant density of synaptic plasma membranes from the cerebral cortex of neonatal rats. A fraction enriched in synaptic plasma membranes was prepared from neonatal (5-6 day old) rat cerebral cortex. The procedure was based on a method used to prepare synaptic plasma membranes from adult cerebral cortex. Critical steps were monitored by electron microscopy. Synaptic plasma membranes from neonatal cerebral cortex sedimented as a broad peak between 0.9 M and 1.1 M sucrose. In contrast, the majority of adult synaptic plasma membranes have been reported to sediment to 1.2 M sucorose. The activities of various enzyme markers were determined in subfractions of neonatal preparations in order to estimate contamination. The specific activities of these markers indicated substantial contamination of the neonatal synaptic plasma membrane fractions by microsomes and glia.


Evidence that catecholamines are not the afferent transmitter in the cochlea. Beta-receptor blocking agents (practolol, propranolol) and alpha-receptor blocking agents (phenoxybenzamine, phentolamine), when applied intracochlearly do not eliminate the afferetn discharges or compound action potential of the cochlear nerve. Under the assumption that the drugs used reach the synaptic site, it is therefore concluded that these drugs do not interfere with afferent synaptic transmission. Phentolamine,, however, has a toxic effect upon the cochlea independent from the synaptic process. Local application of catecholamines does not increase spontaneous activity of single fibres of the cochlear nerve. These findings show that catecholamines are not likely to be the afferent transmitter.


[Pharmacological analysis of monoaminergic effects of the amygdaloid complex on the food-procuring conditioned reaction of rats]. The produced by microinjections of dopamine, norepinephrine, serotonin and glutamic acid into the dorsomedial divisions of the tonsil on the food-procuring conditioned reaction of rats was studied. All of the studied substances were found to lengthen the latent time of the conditioned food reaction, the number of food-procuring movements and the scope of the reflex increasing under the effect of dopamine and serotonin, but diminishing following introduction of norepinephrine and glutamic acid. The dopamine effects materialize through the medium of the tonsil neuron receptors, which are sensitive of haloperidol or chloropromazine, and those serotnin--through D-serotonin-sensitive neuron system structures of the amygdaloid complex. The inhibitory effects of norepinephrine are not of an elective nature, for they correlate with the depression of the spontaneous motor activity of the rats. The modulating influence of glutamic acid on the food-procuring conditioned reflex reaction of the rats is not associated with the action on the dopamine, or serotonin-reactive neuron systems of the dorso-medial tonsil.


[Effect of ethyron on carbohydrate metabolism in the myocardium]. Tests staged on rats showed the hypertensive drug ethyron (ethylisothiuronium) to intensify the activity of the oxidative system and pentose cycle in the myocardium, to increase the pyruvate content, raise the redox potential of the lactate-pyruvate potential, to reduce the amount of lactate, heighten the activity of the lactate, malate dehydrogenases, glucose-6-phosphate and cytochrome-c-oxydase. In an isolated spontaneously contracting atrium ethyron abolished the suppressing effect of carbohydrate metabolism inhibitors (sodium fluoride and copper sulphate) on the intensity and frequency of contractions.


[Functional-morphological state of the digestive organs after administration of dianabol]. In tests set up on rats the effect of methandrostenolone on the external secretion of the liver, fermentative activity of the pancreatic tissue and histological and hisochemical findings subsequent to an investigation of internal organs were studied. Introduction of methandrostenolone led to changes in bilification and to a reduced concentration basic components of the bile, to a distinctly pronounced rise in the level of the trypsin inhibitor and to a depressed activity of trypsin in the pancreatic tissue. It also resulted in a change of histochemical and fermentative indices in the tissue of the liver, spleen and other organs.