Morphological changes of pituitary gonadotrophs and thyrotrophs following treatment with LH-RH or TRH in vitro.

To investigate the relationship between LH-RH and TRH and the formation of castration cells and thyroidectomy cells, pituitary glands of 14-day old female rats were cultured with LH-RH or TRH for 10 days. Observed in these glands were external and internal zones, the former containing active, healthy appearing cells, and the latter consisting of many degenerative and necrotic cells. Gonadotrophs and thyrotrophs were readily demonstrated in the external zone of the organs by immunocytochemistry. Hypertrophic gonadotrophs (castration cells) and thyrotrophs (thyroidectomy cells) were observed only in the external zone by electron microscopy. Neither the typical signet-ring gonadotrophs nor intracisternal granules of thyroidectomy cells were seen in the present study. However, the hypertrophic gonadotrophs or thyrotrophs were similar to castration cells or thyroidectomy cells observed in the pituitary gland following castration or thyroidectomy. The results indicate that LH-RH and TRH directly stimulate the secretion of hormones and alter the morphological features of pituitary target cells.


Stability of the "two active X" phenotype in triploid somatic cells.

We examined triploid cells of XXY karyotype heterozygous for glucose 6 phosphate dehydrogenase (G6PD) electrophoretic variants with regard to the stability of their X chromosome phenotype. Clonal populations of cells derived from these human fibroblasts maintained a precise 1:2:1 ratio of A:heteropolymer:B isozymes throughout their life span, indicating stability of the two active X chromosomes in these cells. To determine the influence of the autosomal complement on X chromosome expression, we attempted to perturb the relationship. Fusion of these triploid cells with human diploid fibroblasts carrying a novel G6PD variant (B') resulted in heterokaryons exprssing a novel heteropolymer, presumably indicating that all three parental X chromosomes were active. However, no derepression of the inactive X chromosome was observed. Analysis of interspecific hybrids derived from triploid cells and mouse fibroblasts confirmed that activity of parental X chromosomes is maintained. Some human mouse hybrid clones, however, expressed only a single human G6PD isozyme, probably attributable to segregation of the pertinent X chromosome, but elimination of a relevant autosome cannot be excluded. The triploid cells transformed by SV40 showed alterations in LDH pattern and an approximately 10-20% decrease in chromosome number, but maintained the precise G6PD phenotype of the untransformed cell. These studies provide evidence for the stability of the X chromosome phenotype in triploid cells.


Immunoreactivity for beta-endorphin in LH-RH neurons of the fetal human hypothalamus.

A peptide immunochemically related to beta-endorphin was detected in some LH-RH neurons of the fetal human hypothalamus by comparison of adjacent sections stained for beta-endorphin and for LH-RH. In the same section, by successive staining and after antibody elution, both peptides were again revealed in the same neuron. The significance of the presence of the beta-endorphin-like material in LH-RH neurons is discussed.


Regulation of the microtubule steady state in vitro by ATP.

ATP increases microtubule steady state assembly and disassembly rates in vitro in a concentration-dependent manner. Bovine brain microtubules, composed of 75% tubulin and 25% high molecular weight microtubule-associated proteins (MAPs), were purified by three cycles of assembly and disassembly in the absence of ATP. When assembled to steady state, these microtubules add dimers at one end and lose them at the other in a unidirectional assembly-disassembly process. In the presence of 1.0 mM ATP the unidirectional flow of tubulin from one end of the microtubules to the other increases as much as 20 fold, as revealed by loss of 3H-GTP from uniformly labeled microtubules under GTP chase conditions and by the rate of disassembly following addition of 50 microM podophyllotoxin. UTP, CTP and 5' adenylylimidodiphosphate (AMP-PNP) cannot substitute for ATP in producing this effect. Furthermore, the increase in steady state flow rate persists afer ATP is removed. Thus microtubules assembled in ATP and centrifuged through sucrose cushions to separate them from nucleotides continue to exhibit increased rates in the next assembly cycle in the absence of ATP. It is possible that an ATP-dependent microtubule protein kinase is responsible for the observed increase in tubulin flow rate. A kinase activity associated with brain MAPs has been reported to be cAMP-dependent (Sloboda et al., 1975). We have found an adenylate cyclase activity associated with these microtubules. Whether the adenylate cyclase is a contaminant or due to a specific microtubules-associated protein, and whether its activity is functionally linked to the increased rate of assembly and disassembly in the presence of ATP, remain to be determined.


Two-dimensional gel analysis of cyclic AMP effects in cultured S49 mouse lymphoma cells: protein modifications, inductions and repressions.

In this study, we used two-dimensional gel electrophoresis to analyze the responses of cultured S49 mouse lymphoma cells to incubation with analogs or inducers of cyclic AMP (cAMP). Putative phosphorylations were detected by charge alterations in proteins labeled with 35S--methionine and, in some cases, confirmed by labeling with 32P--phosphate. We assessed the relative stabilities of proteins affected by cAMP, the periods of susceptibility of proteins to cAMP-dependent modification and any cAMP-mediated changes in protein synthesis or stability. Five proteins (of about 650 resolved) behave as expected for "orthodox" substrates of a cAMP-activated protein kinase: both newly synthesized and prelabeled forms of these proteins are subject to modification; this modification involves an acidic charge shift of about one unit; and cAMP-mediated conversion of these proteins to their modified forms is virtually complete. The acidic forms of at least three of these proteins also exhibit cAMP-mediated increases in 32P--phosphate incorporation. Each protein comprised less than approximately 0.005% of cellular protein. Under basal conditions they appear to be phosphorylated to an extent about 20--30% of that found in fully stimulated cells. Nine proteins show cAMP-dependent changes in rates of synthesis with six inductions and three repressions. Most of these changes are of a magnitude of about 3 to 5 fold, and reach their maximal extents after about 4--5 hr of exposure to dibutyryl cAMP. In addition to the phosphorylations, inductions and repressions mentioned above, approximately 12 other reproducible cAMP-dependent changes in protein patterns are observed. Mutant cell lines deficient in catalytic activity of cAMP-dependent protein kinase show none of the changes in protein pattern attributable to cAMP.


Plus and minus strand leader RNAs in negative strand virus-infected cells.

Sendai virus and VSV minus strand genome RNAs, labeled specifically at their 3' ends with RNA ligase, were used as probes to detect leader RNA--that is, short transcripts (approximately 50 nucleotides) complementary to the exact 3' end of the minus strand genome. These probes have allowed the detection of plus strand leader RNAs in both Sendai virus and VSV-infected cells as well as in the virion transcriptase reactions. The use of a similar probe, prepared from the self-complementary ends of DI genome RNA and containing the 3' end of the plus strand antigenome RNA, has allowed the detection of a minus strand leader RNA of identical size in VSV-infected cells. Since the presence of DI genomes could not be detected by analytical sucrose gradient centrifugation in these VSV-infected cells, this minus strand leader RNA is apparently synthesized on the template formed by the exact 3' end of the antigenome RNA.


Characterization of a new isolate of poliovirus defective interfering particles.

An independent isolate of poliovirus defective interfering particles has been analyzed. These particles, designated DI(A), are apparently analogous to the DI particles described by Baltimore and co-investigators. Electron microscopic heteroduplex analysis reveals that the DI(A) isolate is a mixture of deletion mutants which changes with passage level. The DI(A) population consists of at least five distinct deletion mutants, including one double deletion. Electron microscopic mapping of the deleted regions indicates that most, if not all, of the viral capsid region can be deleted. Despite this heterogeneity, the mutant genomes are quite similar in physical size. We propose a model which suggests that the observed properties of poliovirus DI genomes reflect selective pressures extant during the amplification of the mutant genome. According to this model, only those deleted genomes which retain a minimal size and the capacity to synthesize a functional viral polymerase will replicate successfully in a mixed infection. Furthermore, this model proposes a mechanism for the enrichment of poliovirus DI genomes and an explanation for the low level of complementation observed in mixed infections of picornaviruses.


Differences between the endogenous and exogenous DNA sequences of Rous-associated virus-O.

DNA sequences related to the endogenous retrovirus of chickens, Rous-associated virus-O (RAV-O), have been examined using site-specific DNA endonuclease analysis of cellular DNA derived from line 15 and line 100 chickens. Individual embryos from both inbred lines were used as a source of embryonic fibroblasts from which cellular DNA was isolated. Analysis of DNA containing either endogenous RAV-O sequences alone or both endogenous and exogenous RAV-O sequences produced identical patterns of RAV-O-specific DNA fragments after digestion with the endonucleases Eco RI, Hind III, BgI II, Bam HI or Xho I. Similar analysis with endonucleases Hinc II or Hha I, however, produced several RAV-O-specific DNA fragments which were derived from cellular DNA containing both endogenous and exogenous RAV-O sequences but not from cellular DNA containing only endogenous sequences. Although some differences exist between the DNA fragments specific for the endogenous viral sequences of line 15 and line 100 cellular DNA, the DNA fragments specific for the exogenous viral sequences were identical between the two inbred lines. Cleavage of an unintegrated linear RAV-O DNA molecule with Hinc II or Hha I produced DNA fragments identical to those specific for the exogenously acquired RAV-O provirus. This suggests that these characteristic fragments contain no cellular DNA. The potential DNA junction fragments containing both viral and cellular DNA, identified after analysis of DNA that contains both endogenous and exogenous viral sequences, were identical to those observed after analysis of DNA containing only endogenous viral sequences. These results support the following conclusions. First, exogenous proviral sequences are integrated into chicken cell DNA following an interaction between viral and cellular DNA that is specific with respect to the virus and nonspecific with respect to the cell. Second, both the free linear RAV-O DNA intermediate and the newly integrated exogenous provirus contain specific endonuclease sites that are not found in endogenous RAV-O DNA sequences. These results suggest that the formation of the exogenous DNA provirus involves specific alteration of the endogenous viral DNA sequences before reinsertion of the sequences as the exogenous RAV-O DNA provirus. It is possible that newly integrated exogenous RAV-O sequences are characterized by specific differences in the pattern of base methylation and a limited sequence arrangement.


Stimulation of collagenase production by collagen in mammalian cell cultures.

In this study, we investigated the possible regulatory role of collagen in collagenase production by cultured human skin fibroblasts and human and rabbit synovial cells. Addition of types I, II or III collagen in solution to the culture media markedly stimulated trypsin-activable collagenase activity in these cultures. In the human cell cultures the stimulatory effect of collagen was further enhanced by a soluble factor isolated from human monocyte culture media (Dayer, Russell and Krane, 1977). Both native and denatured forms of collagen stimulated enzyme production; their relative efficacy varied among the different types. The native form of both types I and II collagen showed a greater effect on collagenase production than the corresponding denatured form, whereas with type III collagen the denatured form was more effective.


Pleiotropic mutations within two yeast mitochondrial cytochrome genes block mRNA processing.

The mRNAs from two yeast mitochondrial genes cob-box (cytochrome b) and oxi-3 (cytochrome oxidase 40,000 dalton subunit) are processed from large (7-10 kb) precursors. Certain mutations in each gene block the maturation of the RNAs from both genes at a variety of specific steps. The pleiotropic cytochrome b mutants seem to lack a functional trans-acting RNA required for the processing of both messengers. In contrast, the oxi-3 mutants may act by producing an activity that inhibits specific steps.


Splicing and the formation of stable RNA.

To determine whether RNA splicing plays an obligatory role in gene expression, we have constructed a series of SV40-transducing viruses carrying various combinations of splice junctions derived from the viral genome and a mouse globin gene. All of the viruses that retain at least one functional splice junction, derived from either the viral or the mouse genome, encode stable hybrid RNAs. In contrast, a virus from which all the splice junctions have been removed fails to produce any detectable stable RNA. These results suggest that splicing is a prerequisite for stable RNA formation.


An activity phosphorylating tyrosine in polyoma T antigen immunoprecipitates.

Polyoma T antigen immunoprecipitates contain a protein kinase-like activity which preferentially phosphorylates material of 50-60,000 daltons molecular weight. Phosphorylation is not diminished in extracts of polyoma tsA mutant-infected cells shifted to the nonpermissive temperature late in infection, conditions which inactivate the large T antigen. Phosphorylation is reduced or absent in cells infected with polyoma host range nontransforming (hr-t) mutants, which have defective small and medium T antigens. The major acceptor of phosphate is not the heavy chain of immunoglobulin, but appears to be the polyoma medium T antigen. The large T antigen is also phosphorylated, but usually to a lower specific activity. In terms of acid and alkali sensitivity and electrophoretic and chromatographic mobility in one and two dimensions, the phosphorylated residue behaves identically to phosphotyrosine and differently than phosphorylated serine, threonine, lysine and histidine.


Phosphorylation of polyoma T antigens.

The T antigens of polyoma virus have been examined for phosphorylation in vivo and associated protein kinase activities in vitro. The 100K "large" T antigen is the major phosphoprotein among the T antigen species in vivo as determined by labeling virus-infected cells with 32P-orthophosphate. Hr-t mutants show normal phosphorylation of their 100K T antigens. The wild-type 56K plasma membrane-associated "middle" T antigen is also phosphorylated in the cell, but to a lesser extent than the 100K; this low level phosphorylation is also observed in the presumably altered 56K protein induced by hr-t mutant NG59 and in the 50K truncated "middle" T of hr-t mutant SD15. Addition of dibutyryl cyclic AMP to the medium does not affect labeling of either large or middle T antigens in wild-type- or mutant-infected cells. Thus no differences are observed in T antigen phosphorylation in vivo between wild-type virus and hr-t mutants. Hr-t mutants are defective in a protein kinase activity assayed in vitro by adding gamma-32P-ATP to T antigen immunoprecipitates. In the case of wild-type virus, the 56K protein is the major phosphate acceptor in the in vitro kinase reaction, with a somewhat lower level of phosphorylation observed in the 100K band. Hr-t mutants NG59 and SD15 show no labeling of the altered 56K or 50K, respectively, but do show detectable levels of 32P in the 100K bands. A wild-type virus carrying a small deletion affecting the 100K and 56k bands shows a normal level of kinase activity associated with the truncated T antigens. Ts-a mutants appear to be normal with respect to the middle T antigen-associated kinase. Photoaffinity labeling of infected cell extracts with 8-azido cyclic AMP shows that the two major classes of regulatory subunits of cyclic AMP-dependent protein kinases are present in the immunoprecipitates. Phosphorylation of histone H1 occurs when this substrate is added to immunoprecipitates of either mock-infected or virus-infected cells, again demonstrating the presence of cellular kinases. Further experiments will be required to determine whether the middle T antigen of polyoma virus is itself a protein kinase or simply a substrate for one or more cellular kinases.


Structure of the Abelson murine leukemia virus genome.

Virions produced from cells transformed by A-MuLV contain a 30S, 5.6 kb RNA that can be translated in a cell-free system to form the characteristic A-MuLV protein. This RNA was mapped by heteroduplex methods using DNA probes from M-MuLV, the presumed parent of A-MuLV. The overall organization of the RNA was determined by using full-length M-MuLV reverse transcribed DNA and visualizing the heteroduplexes in the electron microscope. This showed that A-MuLV and M-MuLV have homologous sequences at both ends of their RNAs but that the central portion of the A-MuLV genome is not homologous to sequences in M-MuLV RNA. A precise measure of the lengths of the shared regions was obtained by using S1 nuclease to digest hybrids between 32P-labeled M-MuLV DNA and A-MuLV RNA; the resulting fragments were analyzed for their length by electrophoresis. The regions of homology were shown to be 1320 nucleotides long at the 5' end and 730 nucleotides long at the 3' end. Thus approximately 6200 nucleotides of the approximately 8300 in M-MuLV RNA were deleted when the A-MuLV genome was formed, but an insert of 3600 nucleotides, presumably derived from the normal murine genome, was inserted in place of the deleted region.


The genome of human papovavirus BKV.

The complete DNA sequence of human papovavirus BKV(Dun), consisting of 5153 nucleotide pairs, is presented. We describe the segments of the genome which correspond to the replication origin, the tandem repeated sequences, the 5' and 3' ends of the mRNAs, the splice sites, the early and late viral proteins and the putative viral polypeptides. These BKV DNA sequences are compared with analogous regions in the SV40 and Py virus genomes in an attempt to localize viral functions for lytic growth and transformation.


Changes in cyclic AMP binding and protein kinase activities during growth and differentiation of Blastocladiella emersonii.

Multiple protein kinases in the water mould Blastocladiella emersonii are described. A cyclic AMP-independent protein kinase which prefentially phosphorylates casein remains unchanged during vegetative growth of the cells and in the two phases of differentiation: germination and sporulation. In contrast, cyclic AMP-dependent protein kinase activity and cyclic AMP binding components are induced during the sporulation.


An example of how patient selection can influence survival in glioblastoma multiforme.

Drawing on experience in evaluating postsurgical treatment of glioblastoma multiforme, it is concluded that results of clinical trials programs can be greatly affected by the status of patients who are selected for study. Investigations report results only on patients considered "evaluable," a portion of the total which may be less than 20%. Improvement in patient survival should not be conceived to be due to aggressive treatment unless patient selection has been accounted for.


Intrapulmonary metastases from bronchogenic carcinoma and their radiographic detection.

Autopsies on 102 patients dying with bronchogenic carcinoma over an 11-year period revealed that 37.3% had pulmonary metastases. Review of the pathological evidence in questionable cases supported the validity of the diagnosis. This rather high figure is paralleled by the findings of others quoted in the literature. Dissemination in the lungs appears to take place early with respect to the time of diagnosis. Seventeen of 49 patients (34.7%) who died within 1 month of tissue diagnosis already had additional pulmonary foci. It was possible to identify intrapulmonary spread radiographically in over half of the cases despite the prevalence of concomitant pulmonary disease. Hence, x-ray examination is a useful means of establishing the existence of such metastases in pursuing clinical trials.


Adjuvant systemic therapy of lung cancer.

Most patients with lung cancer subjected to surgical resection are likely to have residual tumor burdens which lead to clinical relapse and death. Unfortunately, none of the systemic therapies for squamous cell, large cell and adenocarcinoma of the lung have demonstrated curative potential either in the advanced disease or in the surgical adjuvant setting. Interest in clinical trials of adjuvant therapy in lung cancer have been rekindled by three factors: 1) reports indicating the value of immunotherapy, 2) preliminary encouraging experience with new chemotherapy programs, and 3) methodologies including stage- and cell type-specific clinical trials leading to better interpretation of results. These concepts have stimulated new treatment protocol studies within the NCI-sponsored Lung Cancer Study Group, and clinical cooperative groups.


Adriamycin and cis-dichlorodiammineplatinum in nonresectable and metastatic carcinoma of the lung.

The combination of adriamycin and cis-dichlorodiammineplatinum (DDP) was evaluated in 20 patients with nonresectable and metastatic carcinoma of the lung. Both drugs were administered at a dosage of 60 mg/m2 intravenously every 3 weeks. The overall objective response rate was 40% (eight of 20 patients) with two complete responses (CR) and six partial responses (PR). Four of eight patients with small cell carcinoma responded (1 CR and 3 PR) and four of 12 patients with nonsmall cell carcinoma responded (1 CR and 3 PR). The median duration of response was greater than 36 weeks. The median survival of responders was 54 weeks. Nausea and vomiting were major side effects but rarely lasted longer than 2 days. Renal and bone marrow toxicities were generally minimal and controlled by dosage reduction. However, there was one death secondary to severe myelosuppression.


Controlled study with BCNU vs. CCNU as adjuvant chemotherapy following surgery plus radiotherapy for glioblastoma multiforme.

From September, 1972 to December, 1976, 102 consecutive patients operated on for glioblastoma multiforme were randomized, after total or subtotal tumor resection, to receive irradiation alone, irradiation plus BCNU or irradiation plus CCNU. BCNU and CCNU adjuvant chemotherapy was repeated every 6--8 weeks as long as the patients remained in complete remission. Patients were comparable for median age, type of surgery, and histological grade III and IV. Radiotherapy was administered at the tumor dose of about 5000 rads in all three groups. The percent of optimal dose administered was 96% for BCNU and 93% for CCNU. In the group treated with radiotherapy alone (32 cases) the median survival was 10.5 months, while in the groups treated with BCNU (34 cases) and CCNU (36 cases) the median survival was 12 and 16 months, respectively. Both relapse-free (P = 0.05) and total survival (P = 0.03) were significantly improved only in patients who were treated with radiotherapy plus CCNU compared to patients receiving radiotherapy alone after surgery. Present results show that in resectable glioblastoma multiforme, a slightly improved survival rate can be achieved by the prolonged use of adjuvant CCNU following maximal surgical resection and radiotherapy. The cure rate was not improved.


Streptozotocin: an inactive agent in small cell carcinoma.

Fifteen patients with advanced small cell carcinoma of the lung were treated with streptozotocin. No objective reponses were seen. A review of the literature reveals no objective response in a total of 56 patients, indicating that this agent is inactive as a single agent in small cell carcinoma.


Results of surgical therapy for lung carcinoma.

A series of 300 pulmonary resections in patients with lung carcinoma is presented. Total survival rate of the series since the operation, including surgical mortality, was 33% at 3 years and 24% at 5 years. The survival rate and surgical criteria were correlated, having better results when standard surgery was performed. The authors emphasize that the surgical figures of the series are of great value as the surgical indications were large and nonselective, with 85% of resectability in the thoracotomies.


[Transfer of genetic information from T to B human lymphocytes during an immune response to herpes simplex virus].

Human blood lymphocytes carrying different allotypes were divided into B and T subpopulations and cultured in presence or in absence of ultraviolet inactivated Herpes Simplex Virus. Isolated B or T cells did not produce any antiherpetic activity. The B lymphocytes cultured in presence of 1 or 50% of the supernatant collected from virus exposed T cells synthesized on antiherpetic antibody with some allotypic markers of the T cell donor.


[Importance of protein-protein interactions for the structural integrity of membrane framents from Torpedo marmorata electric organ].

Electron spin resonance and electron microscopy were used to show that alkaline extraction of a 43,000 dalton protein from torpedo membrane induces structural destabilization of the membrane and rotational diffusion of the cholinergic receptor protein.


[Diptera of Calliphora genus, potential carriers and spreaders of swine vesicular disease virus].

Swine Vesicular Disease virus ingested by the adult fly Calliphora persists several days in the digestive tract of the insect and is eliminated in feces. The virus ingested by the insect at larval stage has been recovered from the digestive tract and feces of adult flies. Thus, the dissemination of the virus, even in a limited fashion, seems to be possible and attention is chiefly centered on the second process.


[New data on the intervention of cyclic monophosphate in the cardiac automatogenous process].

We present here some new holistic titrations related to the demonstration of the intervention of threshold endogenous values of cAMP as a trigger biochemical factor of heart automatogenic function.


[Visualization of opiate receptors in the locus coeruleus of rats: high resolution radioautography after administration of a tritiated met-enkephalin analog].

Opiate receptor sites were visualized by high resolution radioautography in the locus coeruleus of the Rat following intraventricular injection of the met-enkephalin analogue FK 33-824 tritiated with a high specific activity (3H-FK). After administration of a tracer dose of 3H-FK, more than 75% of the radioactivity detected in the locus coeruleus is specifically bound to opiate receptor sites. These are distributed both between and inside neuronal perikarya. After administration of 3H-FK at high concentration, electron microscope radioautography shows the presence of specific opiate binding sites at the level of axo-somatic and axo-dendritic synaptic junctions. These synaptic binding sites could correspond to receptor sites for endogenous enkephalins.


In vitro differences between the lymphocytes of normal subjects and atopics.

Recently evidence has accumulated that atopic disease is associated with a deficiency of thymus-derived (T) cells. This deficiency appears to be primary, rather than secondary to treatment or manifestations of the disease. Results of in vitro studies indicate that the deficiency is most pronounced in certain subpopulations of T cells, and therefore a disturbance of the balance between subsets of T cells, notably suppressor and helper T cells, may develop in vivo in atopics. Some results suggest that there is indeed a relative deficiency of suppressor T cells in atopic diseases, and thus the hyperproduction of IgE which is associated with these diseases may be explained. The cause of the T cell deficiency in atopy may be a basal cellular abnormality, manifested e.g. as increased sensitivity to inactivation by physiological substances, notably to agents which increase intracellular levels of cyclic AMP. It is equally possible that the T cell deficiency is a direct consequence of subnormal production of certain thymic hormones.


Determination of cadmium in whole blood and urine by Zeeman atomic absorption spectroscopy.

Direct determination of cadmium in whole blood and urine can be achieved using Zeeman effect flameless atomic absorption spectroscopy. Within-run precision studies for whole blood cadmium determinations gave relative standard deviations of 11.3% and 6.3% for 0.53 micrograms/l and 3.16 micrograms/l, respectively. Within-run precision studies for the urine analyses yielded relative standard deviations of 11.3% and 5.2% for 0.62 micrograms/l and 2.48 micrograms/l, respectively. The detection limit is 0.12 micrograms/l in the diluted specimens. Thus, this methodology may be used to quantitate normal and toxic cadmium levels in whole blood and urine.


Experience with human growth hormone in Great Britain: the report of the MRC Working Party.

The Working Party on human growth hormone (hGH) has during the past decade developed a system for the evaluation and treatment of patients suffering from hGH lack. Today there are nineteen measurement centres in the United Kingdom at which patients are assessed and where the effects of therapy are monitored. The current supply of hGH, which is prepared from pituitary glands collected by pathologists in the National Health Service, is just enough to meet demand, but research conducted on behalf of the Working Party suggests that hGH deficiency is more common than has been thought and that the prevalence may be as high as one in 10 000. If, as is hoped, patients are diagnosed younger and more patients with partial deficiency are recognized, demand may soon outstrip supply. Work is in progress to define better methods of hGH production and optimal dose regimens, both of which will help to minimize the problem of supply and demand. A few children have anti-hGH antibodies, which block growth as a result of treatment. Improved hGH production techniques may result in a less antigenic product and the resolution of this problem. Many of the Working Party's activities began as research and have evolved into service. Because of this shift in emphasis, and although much research is still to be done, responsibility for provision of treatment with hGH transferred from the Medical Research Council to the Department of Health and Social Security in July 1977.


Induction of puberty: long-term treatment with high-dose LHRH.

Gonadotropin releasing hormone 500 mcg was administered twice daily to four patients (two male, two female) with hypogonadotrophic hypogonadism due to LHRH deficiency, for a minimum of 1 year. Despite initially encouraging biochemical responses and physical changes in all four patients, gonadotrophin responsiveness waned, sex steroid levels were not maintained and pubertal development has not occurred. This diminishing responsiveness appears to be a severe limitation on the use of LHRH as a therapeutic agent.


Radioimmunoassay of plasma 18-hydroxy-11-deoxycorticosterone and its response to ACTH.

A radioimmunoassay for the measurement of 18-OH-DOC in plasma was developed using an antiserum raised against the gamma-lactone derivative. The steroids with the greatest degree of cross reaction were 18-OH-corticosterone-gamma-lactone and aldosterone-gamma-lactone which showed cross-reactivities of 1.96% and 0.47% respectively. These and other interfering steroids were eliminated by chromatography of the extracts on columns of Sephadex LH-20. The lowest limit of detection of 18-OH-DOC in 1 ml of plasma corresponded to 33 pmol-1. The intra-assay precision was 9.7, 4.8 and 2.6% at 102.0, 316.1 and 1144.0 pmol 1(-1) respectively and the interassay precision was 15.3 and 5.4% at 71.3 and 404.7 pmol 1(-1) respectively. The amount of 18-OH-DOC measured (y) which showed a high degree of correlation (r = 0.999) with the amount added (x) to plasma in the range 240--1920 pmol 1(-1) could be predicted from the linear least squares equation y = 1.006x + 31.3. The concentration of 18-OH-DOC in ten normal subjects was 172.1 +/- 39.1 pmol 1(-1) at 09.00 h, 100.9 +/- 16.9 pmol 1(-1) at 12.00 h and 95.8 +/- 33.3 pmol 1(-1) at 16.30 h. Plasma 18-OH-DOC and cortisol levels were measured after various intravenous doses of ACTH in three patient with esential hypertension. Lower doses of ACTH caused similar percentage increases in both hormones but higher doses caused considerably greater increases in 18-OH-DOC. These results confirm the ACTH dependancy of 18-OH-DOC secretion.


The circadian variation of immunoreactive lipotrophin and its relationship to ACTH and growth hormone in man.

There is a clear circadian rhythm of plasma immunoreactive LPH in man with the trough occurring between 22.00 h and 03.00 h and the peak between 07.00 h and 08.00 h, immediately after waking. At all times circulating LPH and ACTH levels follow each other closely. However, no correlation was observed between LPH levels and either plasma GH or the stage of sleep.


Intermittent Cushing's disease with spontaneous remission.

A patient with well documented Cushing's syndrome, associated with excessive secretion of ACTH, was studied over a 20 month period. During this time there were three episodes of spontaneous biochemical remission and except for a 7 day period of exacerbation the patient has had normal cortisol production for the last 15 months. During this recent prolonged remission period, repeat pituitary function testing demonstrated persistent abnormalities of the hypothalamic pituitary axis, in that ACTH failed to respond to a hypoglycaemic stimulus. However in all other respects adrenal function was normal with preservation of diurnal rhythmicity of cortisol and ACTH, and intact dexamethasone suppressability of cortisol. The additional unusual features of normal GH and TSH responses to stimulation, during either remission or exacerbation, suggest a subtle and localized hypothalamic defect in this case.


Lp(a) phenotypes, other lipoprotein parameters, and a family history of coronary heart disease in middle-aged males.

In a study of 95 presumably healthy, 40-42-year old males from Northen Sweden, the Lp(a) phenotype distribution differed between those who had, and those who did not have one or more close relatives (parent or sib) with coronary heart disease. In the former group, 60% of the males were Lp(a+), as opposed to 28% in the latter group. Thus, in the homogeneous population sample studied, analysis of the normal inherited Lp(a) variation permitted the identification of distinct subpopulations, with respect to familial occurrence of coronary heart disease. None of a series of other parameters distinguished such sub-populations. The results reported are in agreement with our previous finding of a close association between phenotype Lp(a+) and risk of contracting coronary heart disease.


Preliminary experience with parenteral mecillinam.

Case reports are reviewed of 26 patients, mainly with severe Gram-negative infections, treatment with parenteral mecillinam. Twenty-three patients received other antibiotics in addition to mecillinam. In 19 patients, potentially synergistic antimicrobial therapy was given. Overall, a beneficial clinical effect was recorded in 17 (68%) of the 25 patients assessed. A better response was observed in those patients who had not received other antibiotics before treatment with mecillinam was instituted. It is concluded that parenteral mecillinam may have an important role in the treatment of severe Gram-negative infections.


Exogenous DNA transcription in cells with their native DNA inhibited. 1. DNA incorporation into homologous and heterologous cells.

The incorporation of mouse S-EAC DNA into homologous normal cells (mouse embryo secondary cultures), and into heterologous cancer cells (TC-SV40 line), with both systems having their native DNA blocked by BrUdR incorporation, was studied. 3H-TdR-DNA was inoculated with DEAE-D to protect it and to potentiate its incorporation, the process being autoradiograohically controlled. The amount of incorporated DNA was radioisotopically determined, and the incorporation process was studied by analysing the fractions obtained after density gradient centrifugation separation of the inoculated cells DNA. Receptivity was greater in those cells inoculated with DEAE-D-protected DNA. The incorporation was slightly greater for cells whose DNA had been blocked by BrUdR incorporation, and for homologous with respect to heterologous cells. In those cells inoculated while the DNA blockade was incomplete, part of the inoculated DNA became incorporated into the cell genome (L-H chains). However, in the completely blocked cells it could not be determined if the incorporation occurred in a lysogenic-like or in an episomic-like form.


On the pathogenicity of Entamoeba histolytica. Part I: The role of bacterial associate on the modification of virulence of E. histolytica.

Of the 3 strains of Escherichia coli used, only Milner A strain was found capable of modifying the virulence of Entamoeba histolytica. None out of twenty-four hamsters inoculated with either 5 X 10(5) of axenically-cultured E. histolytica of NIH: 200 strain, or 1 X 10(7) of Esch. coli (A, B or C strains), was found to have amebic liver abscess. Whereas one out of six hamsters inoculated with the same number of amebae preincubated for 12 hrs with Esch. coli of Milner A strain was found to have abscess. The role of bacterial associate seems to be nothing but provides a more suitable environment for amebae, thus enable them to survive longer and endow them more time to adapt themselves to the given new environment. From liver abscess E. histolytica was recovered and successfully reaxenized. These amebae were capable of producing liver abscess, therefore the virulence seemed to be inheritable.


Adrenergic receptor function is different in male and female patients with essential hypertension.

As plasma norepinephrine (NE) levels may be similar in hypertensive and normotensive subjects, the sensitivity of adrenergic receptors was investigated in patients with essential hypertension and normotensive subjects of similar age and sex. Alpha-adrenergic receptor sensitivity was measured in platelets by the specific binding of [3H]dihydroergocryptine and the NE inhibition of prostaglandin E1 (PGE1)-stimulated cyclic AMP (cAMP) production. The number of alpha-adrenergic receptors in platelets from hypertensive women was 1.5 times that in the platelets from normotensive ones, with no differences between hypertensive and normotensive women or between men and women in the affinity of the alpha-adrenergic receptor for [3H]dihydroergocryptine. PGE1-stimulated cAMP production was half as great in hypertensive as in normotensive men, while NE inhibition of PGE1-stimulated cAMP production was similar in hypertensive and normotensive men and women. [3H]Dihydroergoeryptine binding in female hypertensives, and PGE1-stimulated cAMP in male hypertensives did not differ from that in sex-matched controls. The sensitivity of the beta-adrenergic receptor, measured by [3H]dihydroalprenolol binding and cAMP production was similar in hypertensive and normotensive subjects.


[Interactions of low density lipoproteins with glycosaminoglycans and foam cells within the atherosclerotic aorta (author's transl)].

Very low and low density lipoproteins (VLDL and LDL) isolated from aorta's intima in salt solution were predominantly, associated in complex with glycosaminoglycans (GAG): hyaluronic acid, chondroitinsulfate A(C), heparin. Heparin was detected mainly in complex, isolated from fibrosis plaque. Studying of interaction between LDL and foam cells showed that cells in situ as far as under their incubation in medium with llow LDL concentration primarily accumulated lipid of lipoprotein particles. Incubation foam cells under increased concentration of lipoprotein in medium led to greater entry of radioactivity into the cells. However, there was less tendency for primarily lipid accumulation. Increase of LDL concentration correlated with larger cell's CO2 production. GAG prevented this effect of LDL. Besides LDL uptake was decreased in this condition too. It is possible that GAG effect lipoprotein properties and in such a way the character of their interaction with cells.


[Population studies of adenylate kinase (AK) in Bulgaria].

The author carried out studies on 600 Bulgarians from the south Bulgaria (the town of Plovdiv) and 232 gypsies. There was gene frequency of AK1=0.9633 and AK2=0.0367 in the Bulgariana and respectively AK1=0.9784 and AK2=0.0216 in the gypsies as the differences between both populations were insignificant. All data, published up to now, were summarised connected with the examination of the Bulgarians for AK (1278) and the estimated mean values for gene frequencies were: AK1=0.9648 and =K2=0.0352). Gene frequencies of the Bulgarians, compared with other populations, revealed that they were in the ranges of the mean European values for gene frequencies and differed substantially from those of the nergos and some Asian nations (Japanese, Chinese). The theoretical probability for excluding fatherhood by means of the system of Ak for the Bulgarian was 3.28%.


[Effect of biological agents on glucose-14C utilization by erythrocytes varying in age].

The authors examined young, adult and old erythrocytes from rabbits, aged 15,35 days and 3--4 years. They measured the amount of glucose 14C transfere and metabolized by the erythrocytes during 2-hour incubation at 37 degrees C in a suitable medium. A series of erythrocytes from the three age groups were examined without treatment with chemical agents. Other series of erythrocytes were treated 1 degree minutes before incubation and during its performance with various biological active substances insulin, ACTH, adrenaline, STH, 3AMP, ClCl2. There were differences due to the age in the nontreated erythrocytes. Erythrocytes increased their permeability for grucose and thus its amount was raised in the cells after treatment with insulin. ACTH inhibited this process. Adrenaline, STH, 3AMP and CaCl increased the amount of glucose in the young erythrocytes, obtained from the young animals. The data showed differences in the transfer of glucose 14C and its metabolism in accordance with the chemical and biological properties of the biological agents as well as in accordance with the age of erythrocytes and the animals.


[Cesium].

Cesium is an alkaline metal close to Rubidium, which is studied in psychopharmacology in the prospect of a possible antidepressive effect. It has stimulating properties of the motor activity on the animal, antagonizes some sedative substances but does not modify either self-stimulation behaviour or induced agressivity. Given in large doses, Cesium presents hypertensive effects bound with a stimulation of the adrenal Epinephrine secretion. As for the synapses Cesium increases the neuro-transmitter release. However the Norepinephrine turn-over is not modified in the central nervous system while that of Serotonin is increased as well as the cerebral concentrations of Tryptophan, Serotonin and 5.HIAA. These very fragmentary data about the toxico-pharmacological properties of Cesium, as well as the very prolonged half-life of this ion (from 50 to 100 days), explain that contrary to Rubidium it has not still been used in man.


Effects of (Bu2)-cAMP and clomiphene on serum concentrations of several proteins and hormones in gonadectomized monkeys.

Sixteen mumol kg-1 of dibutyryl cyclic adenosine 3',5'-monophosphate [(Bu)2cAMP] given intramuscularly to gonadectomized green monkeys (Cercopithecus aetheops sabeus) for two-week periods, produced a significant elevation in the serum concentrations of thyroxine-binding globulin (TBG), fibrinogen, and haptoglobin. No changes were noted in cortisol-binding golbulin (CBG), cortisol, thyroxine, transferrin, total proteins, and albumin. Clomiphene citrate (12.5 mg orally) given concomitantly with (Bu)2cAMP blocked partially the (Bu)2cAMP-induced fibrinogen increment, but not the change in TBG concentration.


Preventive effect of diethyldithiocarbamate on experimental diabetes produced by dithisone and 8-oxychinoline.

Diethyldithiocarbamate sodium (DDC)--a compound forming complexes with metal ions-did not change the structure of endocrine pancreas and did not affect the glycemia after the repeated administration of a dose of 250 mg kg-1 while a single dose of 500 or 1000 mg kg-1 increased the glycemia for 2 to 4 h in most of rabbits tested. DDC injected in similar doses prevented a diabetogenic action of dithisone and of a number of 8-oxichinoline derivatives. This effect appeared very rapidly and persisted for several hours being dependent on a dose of DDC and on the interval between its administration and the injection of diabetogenic compounds. It was concluded that DDC forms stable complexes with zinc ions in pancreatic beta-cells which appear to be resistant towards the action of diabetogenic compounds directed selectively to the blocking of zinc ions too and thus to the development of experimental diabetes.


Study of peptides inhibiting the angiotensin-converting enzyme of human seminal plasma.

Several peptides were investigated for their inhibitory capacity against dipeptidyl carboxypeptidase (angiotensin-converting enzyme) from human seminal fluid. The strongest inhibitor was the nonapeptide SQ 20881. A marked inhibition was also shown by the compounds Phe-Ala-Pro and Boc-Phe-Ala-Pro, which behaved as competitive inhibitors. Among the peptides related to angiotensin, angiotensin III was the strongest inhibitor, followed by angiotensin II and the C-terminal hexapeptide of angiotensin II. The results indicate the dipeptidyl carboxypeptidase of human semen is very similar to pulmonary dipeptidyl carboxypeptidase in its susceptibility to peptide inhibitors. In view of these and other previously reported similarities, it is possible that both enzymes are identical.


Chemical and enzymatic characterization of the collagenase from the insect Hypoderma lineatum.

The collagenase from the larvae Hypoderma lineatum, with a molecular weight of 24 000 and isoelectric point of 4.1, was obtained in homogeneous form by ion-exchange chromatography. It is stoichiometrically inhibited by diisopropylfluorophosphate. On the other hand it is unaffected by ethylenediaminetetraacetate, p-chloromercuribenzoate, dithiothreitol, N-tosyllysine chloromethyl ketone, N-tosylphenylalanine chloromethyl ketone and ovomucoid trypsin inhibitor. The enzyme which degrades native collagen in its helical parts, has a specific activity on thermally reconstituted collagen fibrils of 150 micrograms collagen degraded x min-1 x (mg enzyme)-1 at 37 degrees C. It hydrolyses casein but has no esterolytic activity characteristic of trypsin, chymotrypsin nor elastase. It has no action on the synthetic peptide 4-phenylazobenzyloxycarbonyl-L-prolyl-L-leucyl-L-glycyl-L-prolyl-D-arginine. The amino acid composition of Hypoderma collagenase indicates a distinct similarity with the serine proteinases of the trypsin family and with another athropode serine collagenase, that of the fiddler crab Uca pugilator. This suggests that eucaryotic collagenases with digestive rather than morphogenic function represent a new category of members of the trypsin family.


Microbial production of methyl ketones. Purification and properties of a secondary alcohol dehydrogenase from yeast.

Cell-free extracts derived from yeasts Candida utilis ATCC 26387, Hansenula polymorpha ATCC 26012, Pichia sp. NRRL-Y-11328 Torulopsis sp. strain A1 and Kloeckera sp. strain A2 catalyzed an NAD+-dependent oxidation of secondary alcohols (2-propanol, 2-butanol, 2-pentanol, 2-hexanol) to the corresponding methyl ketones (acetone, 2-butanone, 2-pentanone, 2-hexanone). We have purified a NAD+-specific secondary alcohol dehydrogenase from methanol-grown yeast, Pichia sp. The purified enzyme is homogenous as judged by polyacrylamide gel electrophoresis. The purified enzyme catalyzed the oxidation of secondary alcohols to the corresponding methyl ketones in the presence of NAD+ as an electron acceptor. Primary alcohols were not oxidized by the purified enzyme. The optimum pH for oxidation of secondary alcohols by the purified enzyme is 8.0. The molecular weight of the purified enzyme as determined by gel filtration is 98 000 and subunit size as determined by sodium dodecyl sulfate gel electrophoresis is 48 000. The activity of the purified secondary alcohol dehydrogenase was inhibited by sulfhydryl inhibitors and metal-binding agents.


Inactivation of gluconeogenic enzymes in glycolytic mutants of Saccharomyces cerevisiae.

Yeast mutants blocked at different steps of the glycolytic pathways have been used to study the inactivation of several gluconeogenic enzymes upon addition of sugars. While phosphorylation of the sugars appears a requisite for the inactivation of fructose 1,6-bisphosphatase and phosphoenol-pyruvate carboxykinase, malate dehydrogenase is inactivated by fructose in mutants lacking hexokinase. The normal inactivation elicited by glucose in a mutant lacking phosphofructokinase indicates that the process does not require metabolism of the sugar beyond hexose monophosphates. A possible role for ATP in the inactivation process is suggested.


Studies on a mutant form of Escherichia coli citrate synthase desensitised to allosteric effectors.

Naturally occurring citrate synthases fall into distinct molecular and catalytic types. Gram-negative bacteria produce a 'large' enzyme, allosterically inhibited by NADH and, in the facultative anaerobes such as Escherichia coli, also by 2-oxoglutarate. On the other hand, Gram-positive bacteria and all eukaryotes produce a 'small' citrate synthase which is insensitive to these metabolites. As a complement to structure-function studies we have explored the possibility of genetically altering one type of citrate synthase to the other. By mutagenesis and suitable selection we have succeeded in isolating a mutant of E. coli whose citrate synthase is both 'small' and insensitive to NADH and 2-oxoglutarate. Some characteristics of the enzyme are described. Such mutant enzymes offer a novel approach to the study of citrate synthase, its regulation and its natural diversity.


A model for the chemical interactions of adenosine 3':5'-monophosphate with the R subunit of protein kinase type I. Refinement of the cyclic phosphate binding moiety of protein kinase type I.

The cAMP receptor site in the regulatory subunit of adenosine 3':5'-monophosphate (cAMP)-dependent protein kinase type I was mapped using analogues of cAMP in which the ribose phosphate moiety was systematically modified. Electronical alteration of the cyclophosphate ring at the 3' and 5' positions by sulfur and nitrogen decreased the affinity of these analogues towards the kinase. Substituents at these positions are not tolerated. Testing the separated diastereomers of derivatives in which one of the exocyclic oxygens at the phosphorus has been substituted by sulfur, it was found that one diastereoisomer is preferentially recognized. Based on these results it is proposed that the hydrophylic cyclic phosphate-ribose moiety of cAMP is bound to the kinase via its 3' and 5'-oxygens, the 2'-hydroxy group and the negative charge in a fixed position. Based on our and other published results it is further proposed, that the adenine moiety is bound in a hydrophobic cleft without any hydrogen bond interactions. The chemical interactions between cAMP and the R subunit of protein kinase type I differ from those found for the binding of cAMP to the chemoreceptor of Dictyostelium discoideum [18].


Bile salt binding to serum components. Taurocholate incorporation into high-density lipoprotein revealed by photoaffinity labelling.

1. Photoaffinity labelling of human serum albumin with the sodium salts of (3 beta-azido-7 alpha,12 alpha-dihydroxy-5 beta-cholan-24-oyl)-2-amino[2(-3)H (N)]ethanesulfonic acid, (7,7-azo-3 alpha,12 alpha-dihydroxy-5 beta-cholan-24-oyl)-2-amino[2(-3)H (N)]ethanesulfonic acid and (11 zeta-azido-12-oxo-3 alpha,7 alpha-dihydroxy-5 beta-cholan-24-oyl)-2-amino[2(-3)H (N)]ethanesulfonic acid resulted, in each case, in a considerable covalent incorporation of radioactivity into the protein. 2. Photoaffinity labelling of whole serum, obtained from fasting test persons, revealed with all three photolabile derivatives of taurocholate at the physiological concentration of 2.1 microM the incorporation of radioactivity not only into albumin but also into high-density lipoprotein, as demonstrated by density gradient centrifugation and by immunological characterization. 3. The bulk of radioactivity incorporated into high-density lipoprotein by photoaffinity labelling of whole serum was found to have been associated with the lipids. Only 10-20% of the label was covalently bound to apolipoproteins, predominantly to the apolipoproteins A-I and A-II. 4. The interaction of taurocholate with high-density lipoprotein has been confirmed by density gradient centrifugation using 14C-labelled taurcholate. It is assumed that the interaction of taurocholate with high-density lipoprotein is physiologically of significance.


High number of high-affinity binding sites for (-)-[3H]dihydroalprenolol on isolated hamster brown-fat cells. A study of the beta-adrenergic receptors.

The beta-adrenergic receptors of hamster brown adipocytes have been characterised by binding of the radioactive ligand (-)-[3H]dihydroalprenolol, directly to isolated intact cells in suspension. The brown fat cell contains 57,000 specific and saturable binding sites which have a dissociation constant (Kd) for [3H]dihydroalprenolol of 1.4 nM as determined by Scatchard analysis. The kinetically derived Kd, determined from forward and reverse rate constants, is 5 nM. Both of these values are in agreement with the dissociation constant (Kd = 2.2 nM) for alprenolol, determined from competition studies with [3H]dihydroalprenolol in these cells. Beta-adrenergic agonists competed for the specific binding sites with a typical beta 1-adrenergic specificity. The order of potency of agonists agrees well with the ability of these agents to stimulate respiration in isolated brown adipocytes: 50% stimulation of respiration occurs with apparently less than 10% occupancy of binding sites. Both the high affinity and high number of specific binding sites of [3H]dihydroalprenolol in brown fat cells presumably reflect the generally accepted dominating role of catecholamines in the regulation of brown fat metabolism and non-shivering thermogenesis.


Characterization of the membrane-bound inorganic pyrophosphatase in Rhodospirillum rubrum.

The membrane-bound inorganic pyrophosphatase (EC 3.6.1.1) from Rhodospirillum rubrum has been investigated with the tools of enzyme kinetics, and with two amino acid reagents, N-ethyl-maleimide (MalNET) and 4-chloro-7-nitrobenzofurazan (Nbf-Cl). 1. The concentration of the true substrate, MgPPi, was varied with constant concentrations of free Mg2+ or PPi. It was observed that Mg2+ acted as an activator. 2. Heat inactivation of the enzyme at 62 degrees C was slowed down in the presence of Mg2+. 3. MalNET and Nbf-Cl bind to the enzyme, and inhibit its activity. The effect of both reagents is dependent on the temperature. 4. A model is proposed where the 1:1 complex of Mg2+:PPi acts as substrate and Mg2+ interacts directly with the enzyme as an activator. PPi can bind to the enzyme, but is not hydrolyzed in the uncomplexed form.


Glutathione reductase from human erythrocytes. Amino-acid sequence of a major fragment that links the FAD, NADP and interface domains.

A major CNBr fragment of glutathione reductase, peptide Q [Krohne-Ehrich, G., Schirmer, R.H. & Untucht-Grau, R. (1977) Eur. J. Biochem. 80, 65-71], was further fractionated by trypsin, chymotrypsin, thermolysin and clostripain digestion. The peptides were isolated and most of them were sequenced by solid-phase Edman degradation. The whole peptide Q was sequenced N-terminally up to position 51 by the same technique. A total sequence of 128 amino acids (28% of the whole protein) was obtained and could be localized in the electron density map [Schulz, G.E., Schirmer, R.H., Sachsenheimer, W. & Pai, E.F. (1978) Nature (Lond.) 273, 120-124] from position 259-387. This part of the polypeptide links and participates in all three domains of the flavoenzyme.


Nuclear mutants of Neurospora crassa temperature-sensitive for the synthesis of cytochrome aa3. Mitochondrial protein synthesis and analysis of the polypeptide composition of cytochrome c oxidase.

Three previously isolated mutants of Neurospora crassa, temperature-sensitive for the production of cytochrome aa3, have been further analyzed. These mutants have a slightly reduced capacity for mitochondrial protein synthesis when grown at 41 degrees C, although this relative deficiency appeared to be no greater than the deficiency in other cytochrome-aa3-deficient mutants. Thermolability studies revealed that the cytochrome c oxidase purified from each of the mutants grown at 23 degrees C is no more sensitive to heat inactivation than the enzyme isolated from wild-type cells. Sodium dodecylsulfate gel electrophoresis of immunoprecipitates obtained from the mitochondria of each of the mutants grown at 23 degrees C, using antiserum directed against holocytochrome c oxidase, indicated that all the subunits of cytochrome c oxidase were present in relative amounts similar to those found in mitochondria from wild-type cultures. However, when the mitochondria from mutant cultures grown at 41 degrees C were examined in the above fashion, only subunits 5 and 6 of the oxidase were detected. Nonetheless, the mitochondrially synthesized subunit 1, 2 and 3 polypeptides could be immunoprecipitated from mitochondria isolated from mutant cells grown at 41 degrees C and labelled with [3H]leucine in medium containing cycloheximide. Although subunits 4 and 7 could not be detected, because a suitable antibody was not available, the fact that five of the seven subunits were present, but not associated with each other, suggested that the genetic defects in these mutants may affect the process of cytochrome c oxidase assembly.


Nucleic acid binding and unfolding properties of ribosomal protein S1 and the derivatives S1-F1 and m1-S1.

The nucleic acid binding and unwinding properties of wild-type Escherichia coli ribosomal protein S1 have been compared to those of a mutant form and a large trypsin-resistant fragment, both reported recently [J. Mol. Biol. 127, 41-45 (1979) and J. Biol. Chem. 254, 4309-4312 (1979). The mutant (m1-S1) contains 77% and the fragment (S1-F1) 66% of the polypeptide chain length (approximately 600 amino acid residues) of protein S1. The mutant is active in protein synthesis in vitro; the fragment, although retaining one or more of the functional domains of S1, is inactive in protein synthesis. We find that m1-S1 is is almost as effective as S1 in binding to poly(rU), phage MS2 RNA and simian virus 40 (SV40) DNA, and in unfolding poly(rU) and the helical structures present in MS2 RNA and phi X174 viral DNA. S1-F1, however, binds to poly(rU) and denatured SV40 DNA, but not to MS2 RNA. It unfolds neither poly(rU), nor the residual secondary structure of MS2 RNA or phi X174 viral DNA. Thus, there appears to be a correlation between the loss in ability of S1 to unwind RNA and the loss in its ability to function in protein synthesis.


Properties of purified calf thymus poly(adenosine diphosphate ribose) polymerase. Comparison of the DNA-independent and the DNA-dependent enzyme.

The physicochemical properties of the purified calf thymus poly(ADP-ribose) polymerase were investigated. The enzyme purified to homogeneity was shown to contain about 10% DNA on a weight basis and its activity to be DNA independent. After removing this fragment of DNA, called the sDNA fraction, the enzyme becomes DNA dependent. The activity of this enzyme preparation was entirely dependent on, and completely restored by, added calf thymus DNA or sDNA. However, the calf thymus DNA concentration needed was a hundred times higher than that of sDNA. The properties of the two enzyme preparations, DNA independent and DNA dependent, were essentially the same. They both reacted against the specific antibody obtained with the DNA-independent poly(ADP-ribose) polymerase. The pH optimum was around 8; the activity was stimulated by Mg2+, Mn2+ and Ca2+, and inhibited by high ionic strength, p-chloromercuribenzoate, ADP-ribose, AMP and polylysine. Nicotinamide, thymidine and NADP were shown to be competitive inhibitors. The enzymatic activity was stimulated by histone H1 when the ratio of DNA to histone H1 was 2. Histones H2A, H2B, H3 and H4 had little effect on the DNA-independent enzyme activity, but were strongly inhibitory for the DNA-dependent enzyme. This inhibitory effect could be reversed by allowing the DNA-dependent enzyme to react with the sDNA fraction before adding histone subfractions. The apparent Km for NAD of the DNA-dependent poly(ADP-ribose) polymerase was shown to vary with the DNA concentration. It was minimum when the amount of sDNA was 10% of that of the enzyme. The ratio of the apparent Km for sDNA to the enzyme concentration was constant at any enzyme concentration. The minimum estimation of the number of base pairs of sDNA required for maximal activation of one enzyme molecule was 16. For calf thymus DNA, this estimation was of 640. These results suggest that the activation of the enzyme needs the formation of some complex between the protein and a specific part of the DNA. This complex was preserved in the DNA-independent enzyme preparation.


Comparative biochemical, histochemical and autoradiographic studies of Na+/k+-ATPase in the rectum of dragonfly larvae (Odonata, Aeshnidae).

Na+/K+-ATPase localization in the rectal wall of larval Aeshna cyanea (Insecta, Odonata) was studied with histochemical precipitation techniques and 3H-ouabain autoradiography in conjunction with biochemical measurements of enzyme activities and radiospectrometry of 3H-ouabain binding, respectively. The NPP-strontium and ATP-lead methods led to complete inhibition of Na+/K+-ATPase in this organ and hence to unreliable histochemical results. The 3H-ouabain binding technique revealed sodium pump sites at the basolateral plasma membranes of the absorptive rectal chloride epithelia.


The nuclear matrix of slowly and rapidly proliferating liver cells.

The nuclear matrix of slowly proliferating rat liver is compared with rapidly proliferating regenerating liver and Zajdela ascites hepatoma cells. While no differences are detected in overall ultrastructure, composition or polypeptide profiles of normal liver versus regenerating liver matrices, significant alterations are observed in the polypeptides of Zajdela hepatoma nuclear matrices.


Value of the brain angioscintigraphy in posterior view.

The results obtained with intravenous brain angioscintigraphy (IBAS) with 99 mTc-DTPA(Sn) are evaluated in 61 patients with clinical suspicion of cerebellar, cerebral or cranial lesions of posterior location or without focal signs or with a previous IBAS in anterior view being normal or of low significance. One hundred patients without demonstrable cerebral or cranial pathology were explored to define the normal characteristics and variation of the IBAS in posteior view. Of the 61 patients with pathology, 45 showed angioscintigraphic alterations. The following angioscintigraphic patterns were found: 1. Perfusion increased, 2. Perfusion diminished, 3. Flip-flop, 4. Mixed pattern, 5. Circulatory arrest, 6. Jugular venous reflux, 7. Normal IBAS associated with an abnormal morphologicall study, 8. Normal IBAS associated with a normal morphological study.


Two different primary tumours of the brain in a patient with breast cancer.

Some reports on multiple primary brain tumours have been published. When one or more tumours are found in brain scintigraphy they are often supposed to be metastases. Further investigations may thus be given up, especially if the patient has or has had a malignant tumour in some other part of the body. In this report a case is described where the patient began to have cerebral symptoms two years after she had been operated for breast cancer. In the scintigraphy a tumour was found in both brain hemispheres. The tumours were regarded as metastases. But when the patient died in a geriatric hospital it was recognized from the autopsy that one tumour was a meningioma and the other a glioblastoma multiforme.


Renal prostaglandins: relationship to the development of blood pressure and concentrating capacity in pre-term and full term healthy infants.

The relationships between urinary prostaglandins (PGs)E2 and F2 alpha and the postnatal development of blood pressure and renal concentrating capacity were investigated in 14 pre-term and 32 full term healthy infants. Mean PGE2 and PGF2 alpha excretion was 18.9 and 10.1 ng/h/1.73 m2, respectively, in pre-term infant. In full term infants mean urinary PGE2 was significantly lower (13.4 ng/h/1.73 m2) and PGF2 alpha significantly higher (22.2 ng/h/1.73 m2). The decrease of the PGE2/PGF2 alpha ratio (P less than 0.001) was accompanied by an increase in blood pressure. High PGE2 levels in pre-term infants were inversely correlated with urinary cAMP excretion. A decreasing PGE2/PGF2 alpha ratio in full term infants was associated with increasing urinary osmolality. After intranasal administration of antidiuretic hormone (DDAVP) in 8 full term infants the increase in urinary osmolality and cAMP excretion was accompanied by a drop in PGE2 excretion to less than half the basal values. These findings suggests that the postnatal changes in urinary PG excretion are associated with a concomittant increase in blood pressure and in the concentrating capacity of the neonatal kidney.


Hepatitis types A, B, and non A--non B in childhood.

199 children with acute hepatitis hospitalized between 1968 and 1978 were tested for serological markers of hepatitis A and B infection. In 24 out of 28 HBsAg-positive patients, hepatitis B infection was diagnosed because of the disappearance of the antigen during convalescence. The histories of the 171 HBsAg-negative children suggested acute hepatitis A infection in 69% of the patients. This diagnosis could be confirmed in 110 of the 116 tested cases (95%) by a more than fourfold increase in the anti-HAV titer or  by detection of anti-HAV of the IgM class. In the 55 HBsAg-negative patients without epidemiological clues as to the type of hepatitis, 40 children exhibited anti-HAV which could be related to acute A infection in 21 out of 22 tested cases. At least 11 patients had to be classified as having nonn A--non B infection. The results indicate that a combination of evaluation of the patient's history and selected serological tests will permit a fast preliminary diagnosis. This is important in the clinical management of patients and protection of contacts with immunoglobulin.


Development of children with early cytomegalovirus infection.

To find out whether cytomegalovirus (CMV) infection during the first months of life influences child development, developmental assessment at the age of 2 years was performed on 116 Finnish children chosen at random in a maternity hospital and followed from birth for the occurrence of CMV infection. Two of the children had congenital infection and 39 were infected after birth but before the age of 6 months. Another five children were infected between the ages of 6 and 12 months, and 70 remained non-infected. The developmental assessment, based principally on the Denver Development Screening Test (DDST), included 7 items for gross motor skills, 6 items for fine motor skills, and 4 items for language development. Significantly more perinatally infected children (7/39) were delayed in speech, compared with the non-infected children (3/70). Furthermore, some items assessing fine motor development were passed better by the non-infected than the perinatally infected children, but the difference in total score was not significant. The performance of the two children with congenital CMV infection did not differ from that of the non-infected children.


Pseudohypoparathyroidism type II and anticonvulsant rickets.

A patient treated with anticonvulsants showed hypocalcemia and hyperphosphatemia in association with increased serum parathyroid hormone, reduced serum 25-hydroxy D3, diminished response in phosphorus excretion to exogenous parathyroid hormone and normal response in cyclic AMP excretion. Oral administration of Vitamin D3 resulted in normalization of serum 25-hydroxy D3, calcium, and phosphorus. At this stage, phosphorus excretion after parathyroid hormone returned to normal. These findings suggest that the patient had pseudohypoparathyroidism type II with anti-convulsant medication as a complicating factor.


Neurophysiological comparison of dendritic cable properties in adolescent, middle-aged, and senescent rats.

Relative dendritic atrophy has been reported in hippocampal granule cells of senescent (25 month) as compared to young (3 month) rats. An electrophysiological study of synaptic response waveform was un;ertaken to determine whether these anatomical changes have physiological correlates of functional significance to the integrative capacity of the granule cells. The variable of interest was the rist time of the extracellularly recorded population synaptic response because this parameter (under appropriate conditions) has been shown to reflect relative electrotonic distance of active synapses from the spike generating zone located near the cell body. By selectively stimulating small subsets of perforant path fibers along its medio-lateral axis, synapses at increasing distances from the soma may be activated and the response waveform recorded. Systematic observations were made on both hemispheres of 3 adolescent (2 months), three middle-aged (12 months), and 3 senescent (28 months) rats. There were no significant differences in the mean, variance, or range of the frequency distributions of synaptic response rise times. However, the medians of the distributions revealed a small shift towards lower values with advanced age. Although the latter finding is consistent with the anatomical changes, the magnitude of the differences were so small as to make their functional significance dubious.


Meal induced increase in serum triglycerides has no effect on lymphocyte response to mitogens.

The effect of a standardized heavy meal on the lymphocyte transformations induced by phytohaemagglutinin (PHA), concanavalin A (Con A) and PPD tuberculin was studied. The meal significantly increased the serum triglycerides (P less than 0.01), while it had no effect on cholesterol or high density lipoprotein-cholesterol levels. The increase in serum triglycerides did not affect lymphocyte transformation induced by phytohaemagglutinin or concanavalin A in whole blood microcultures. A slight decrease was observed when lymphocytes were stimulated with one out of three concentrations of PPD tuberculin (P less than 0.05). However, there was no correlation with the increase of triglycerides and decrease in lymphocyte transformation. Our observation shows that physiological changes in serum triglycerides do not affect the capacity of lymphocytes to respond to mitogenic stimulation, and the whole blood micromethod for lymphocyte stimulation to screen the capacity of cell-mediated immunity does not depend on the meal schedule of the patients.


Lipid and carbohydrate metabolism in uraemia.

Lipid and carbohydrate metabolism variables were studied in twenty-eight patients with chronic renal failure (mean GFR 7.7 +/- 2.5 ml/min) and uraemic symptoms. 71% of the patients had hypertriglyceridaemia (greater than or equal to 2.2 mmol/l). Total serum cholesterol was normal while VLDL cholesterol was high and alpha-lipoprotein cholesterol was low. The fractional elimination rate of Intralipid was low and inversely correlated to serum triglyceride levels. Intravenous glucose tolerance was reduced with normal or slightly increased fasting blood glucose and insulin values before and during the test. Serum triglycerides were correlated to plasma insulin but not to residual renal function or serum urea levels. The cause of hypertriglyceridaemia and lowering of alpha-Lp cholesterol is not unequivocally clear. Present evidence indicate that retarded catabloism of triglyceride-rich lipoproteins is important but accentuated release of triglyceride-rich lipoproteins may have occurred in a number of cases. The commonly used treatment with beta-blocking agents for hypertension in chronic renal failure may accentuate certain of the metabolic responses in uraemia.


Glucose tolerance, plasma insulin and alpha-lipoproteins in young male myocardial infarction survivors compared with controls matched on serum cholesterol concentration.

In order to identify metabolic risk factors other than hypercholesterolaemia, all cases of acute myocardial infarction, diagnosed in males aged under 40 years, were studied over a period of 6 years in Göteborg, Sweden. Twenty out of twenty-four patients who were alive at the time of the study were compared with forty controls matched for serum cholesterol concentration. A previous report has noted lower apolipoprotein A (apoA) and a higher alpha-lipoprotein triglyceride concentration in these young myocardial infarction patients compared with the matched controls. Basal blood glucose, oral glucose tolerance and plasma insulin levels did not differ between the patients and controls, i.e. decreased glucose tolerance and elevated plasma insulin levels were not found to be additional risk factors for myocardial infarction in young males if serum cholesterol concentration was taken into consideration. This finding could be explained by the fact that patients and controls were all high cholesterol individuals and a difference between the patients and the general population is still possible. Low apoA was a risk factor independent of serum cholesterol as well as glucose intolerance and elevated plasma insulin levels.


Input-output properties of motor unit responses in muscles stretched by imposed displacements of the monkey wrist.

Reflex responses are "servo-like" where the output is graded with the input or are "triggered" where the output is independent of input once an input threshold is exceeded. Imposed displacements of monkey or human upper limb joints result in prolonged EMG output in the muscles stretched by the displacements. The longer-latency portions of the prolonged output have been variously reported to be servo-like or to be triggered in nature. In monkeys and humans, angular wrist displacements imposed by step loads result in three peaks (M1, M2 and M3) in the gross EMG recorded from the stretched muscles. Each gross EMG peak largely results from the firing of a separately-responding subpopulation of single motor units (SMUs). We studied the responses of SMUs to loads that were presented to the monkeys in a random order as to magnitude, duration and onset time. Average response histograms were constructed for the SMU responses for individual step load magnitudes. Averages were also constructed for the simultaneously-recorded gross EMG responses for each step load magnitude. The input parameters used were the initial velocity of displacement or the magnitude of step load, while the output was taken as the probability of firing/millisecond/presentation above baseline for the SMUs or the area under the response peaks above baseline for gross EMG. The results establish: 1) That it is not possible to unambiguously determine the input-output properties of the responses to imposed displacements utilizing the analysis of gress EMG activity due to the response characteristics of the various subpopulations of motor units contributing to the gross response. 2) That the SMU activity during all of the peak intervals is monotonically graded with increases in magnitude of the step load or the initial velocity of displacement. Hence, the long-latency portions of the EMG responses are servo-like in nature and are not preprogrammed or triggered responses. 3) That the gain (output/input) of the gross EMG responses almost entirely reflects the variation in the number of motoneurons recruited by changes in magnitude of the step loads rather than variation in the firing rates of motoneurons during the reflex responses.


Role of ACTH on the cytomorphology of testes of the immature rat.

The male gametogenic development of the immature rat shows inhibitory effect by the exogenous infusion of ACTH (0.25 IU and 0.50 IU; i.p.). Seminiferous tubular degeneration, Leydig cell atrophy and Sertoli cell regression are very conspicuous in the dosage of 0.50 IU ACTH (i.p.). Cytometrical and histological evidence confirms that ACTH could be inhibited by FSH during male gametogenic development which leads to cellular degeneration of testes in the immature rat.


Early changes of cyclic nucleotide levels in a mitogenic reaction in the rat mesentery.

By measuring simultaneously cAMP and cGMP we found a biphasic time course with regard to cGMP and the cGMP/cAMP ratio very early in a mitogenic reaction in vivo. This is a new finding.


Effects of dosage and cadmium pretreatment on the binding of cadmium in rat bile.

The effects of dosage and of cadmium pretreatment on the binding of cadmium in rat bile were studied. With increasing dose a higher cumulative biliary excretion of Cd was observed and a higher percentage of the Cd was excreted in a low-molecular-weight form. On the other hand, after cadmium pretreatment, a decrease in the cumulative biliary excretion of cadmium was observed but a greater percentage of that excreted into the bile was bound to high molecular weight compounds.


Sex-dependent action of prenatal fractionated X-irradiation in the mouse. Biochemical findings.

X-irradiation of pregnant NMRI-mice on gestational days 11-13 with 3 x 10.5 Gy increased postnatal mortality of the female offspring only. Weights, protein content and acetylcholinesterase, as well as Na,K-ATPase activities in the brains of all treated offspring, were changed. There were, however, no differences between females and males with respect to these parameters.


Changes in beta-2 adrenergic receptor sensitivity with maturation of erythroid progenitor cells.

Erythroid burst forming units (BFU-E) were much more sensitive to the beta-2 selective adrenergic drug, salbutamol, than erythroid colony forming units (CFU-E) in an in vitro study of erythroid progenitor cells.


Control of the hypothalamus-pituitary-adrenal axis in young and older rats injected with thyroxine.

Administration of T4 on alternate weeks for 30 weeks at a dosage which does not alter body weight depresses basal serum corticosterone levels in older rats (575 days), but not in young animals (260 days). Similar serum corticosterone response to HPA axis stimulation occurs regardless of age or T4 injection.


Fusion of L line daughter cells in the presence of Sendai virus.

Sendai virus induces the fusion of daughter cells in a population of murine fibroblasts (L cell line). Mitosis and the ensuing fusion of two daughter cells has been observed by means of time-lapse cinemicrography. The paper documents two types of cell fusion: a) two daughter cells become completely separated at the end of the mitosis, and fuse only after a certain period of time, or b) the daughter cells remain attached through a narrow bridge of cytoplasm which persists between the cells as a consequence of incomplete cytokinesis. Widening of the cytoplasmic bridge ultimately results in the fusion of cells. The interval between the termination of mitosis and the fusion ranged from 45 to 60 min. The biological significance of the resulting polyploid cells is discussed.


Effect of colchicine on PGE1 stimulation of cAMP1 formation in human lymphoblastoid and normal lymphocytes.

PGE1 increased cAMP level in human lymphoblastoid cells (RPMI 1788) after 5-60 min of incubation at 37 degrees C. A gradual decrease of cAMP concentration was found at the later time intervals. Colchicine significantly potentiated the stimulatory effect of PGE1, although it did not have any effect on cAMP level in control lymphoblastoid cells. The maximal effect of colchicine on PGE1 stimulation of cAMP formation was at the 0.1-1.0 microM level. Human lymphocytes also responded with increased cAMP formation to colchicine addition. In contrast, no stimulatory effect of colchicine was found in human granulocytes.


Guanyl nucleotide regulation of hormonally-responsive adenylyl cyclases.

A large number of hormones and neurotransmitters activate adenylyl cyclase [ATP, pyrophosphate lyase (cyclizing; EC 4.6.1.1.)] catalyzing the formation of cAMP and PPi from ATP in the presence of Mg2+. The cAMP formed is in turn responsible for eliciting the physiological responses of these hormones and neurotransmitters. In addition to hormones and neurotransmitters, fluoride ion, cholera toxin and guanyl nucleotides (GTP and GTP analogs such as GTP gamma S and GMP-P(NH)P) also stimulate adenylyl cyclase activity (Perkins, 1974; Birnbaumer, 1977; Gill, 1977). It has become evident that hormonally-responsive adenylyl cyclase is a multi-component system consisting of at least 3 physically distinct units. The first is the hormone receptor containing a specific site for a given hormone. The second is the catalytic moiety (C component) of adenylyl cyclase bearing the site responsible for catalysis of the cyclizing reaction. The third is the guanyl nucleotide regulatory subunit (G component) which binds guanyl nucleotide. Recently, a GTPase activity has been found to be associated with the G component of adenylyl cyclase (Cassel and Selinger, 1976; Cassel et al., 1977a, b; Lambert et al., 1979). In this review we will present information on the regulation of hormonally-responsive adenylyl cyclases. This is not intended to be a comprehensive review of the literature. Rather, it represents our views on the current status of the regulation of cAMP formation.


Regulatory functions of protein multisite phosphorylation.

In recent years it has become apparent that an increasing number of proteins can be phosphorylated at several different sites. In this article protein multisite phosphorylation is discussed with reference to the enzymes glycogen synthase, pyruvate dehydrogenase, and phosphorylase kinase. Each of these enzymes contains three or more different phosphorylation sites on one or more subunits. Activation and inactivation of the enzymes appear to correlate quite well with phosphorylation of a few key sites on the protein. The other phosphorylation sites may influence other kinetic properties of the enzymes or regulate the rates of dephosphorylation of the key sites by the appropriate phosphatase. Thus, multisite phosphorylation may represent an important mechanism for regulating several functions of complex proteins.


Central control of lactogenic receptors in liver membranes: effect of hypothalamic deafferentation.

The regulation by the central nervous system of a lactogenic receptor, present in the rat liver, was studied. The receptor, which is present in very low concentration or absent in the male rat, can be induced by anterior hypothalamic deafferentation, resulting in a typically female level of the receptor in lesioned male animals. Our observations indicate an important role of the brain in regulation of peripheral lactogenic receptors.


Preliminary characterization of in vitro synthesized hypothalamic precursor ACTH/beta-endorphin-like material.

We have previously shown that cultured bovine pituitary or hypothalamic cells incorporate 3H-labeled amino acids into high molecular weight glycoproteins containing the antigenic determinants of both corticotropin (ACTH) and beta-endorphin. We now report resolution of this 3H-labeled ACTH/beta-endorphin-like material into two predominant size classes upon SDS polyacrylamide-gel electrophoresis with apparent molecular weights (Mr) of 41 500 +/- 1600 and 36 000 +/- 1100. Isoelectric focusing revealed these components to be basic proteins with apparent isoelectric points greater than 8.5. Overnight trypsinization generated a 3H-labeled fragment comigrating with synthetic beta-lipotropin (61-69) [beta-endorphin (1-9)] upon paper electrophoresis which was immunoprecipitable with antibody directed against the corresponding synthetic fragment. Limited trypsinization of bovine immunoreactive ACTH/beta-endorphin extracted from freshly obtained bovine hypothalamic and anterior pituitary tissue, generated fragments which possessed ACTH bioreactivity. Both bovine pituitary and hypothalamic derived material are similar with respect to these foregoing physiochemical parameters and appear to be larger than the reported forms in mouse pituitary.


Immunization against poliomyelitis: risk/benefit/cost in a changing context.

Benefit/risk factors in immunization against poliomyelitis are examined from the viewpoint of the relative risk and efficacy of killed and live poliovirus vaccines in the currently changing contexts of poliomyelitis prevalence in developed and developing countries. Risk factors include virus of vaccine origin gaining access to the CNS, and failure of the vaccine to immunize. Data are presented to illustrate the degree to which the respective risks occur in developed and developing countries. The ultimate elimination of risk by eradication of wild and vaccine virus from the population is discussed.


Antibody to poliovirus in older children and young adults in Israel and the use of poliovaccines for the immunization of the seronegatives.

This report represents preliminary results of a more extensive study which is presently being carried out in Israel. The main aims of this study were twofold: (1) to carry out a survey of the state of immunity of children of various ages and of teenagers to the three poliovirus types and (2) to compare the antibody response of the seronegatives to the two kinds of presently available poliovaccines, the oral poliovaccine (O. P. V., Savin Vaccine) and the inactivated poliovaccine (I. P. V., Salk Vaccine).


Poliomyelitis epidemic in the Netherlands, 1978.

Some provisional data are given of the polio-epidemic in The Netherlands in 1978. Contrary to previous outbreaks, the present epidemic did not only affect areas with low vaccination acceptance rates, cases were also reported from many municipalities with acceptance rates of 90% and over. The epidemic caused by type 1 started mid-April 1978 in the centre of the country. Up to 1st November 110 cases were reported from a belt running from South-West (Zeeland) to North-East (Overijssel). Only one death occurred among the 80 paralyzed cases. The epidemic exclusively struck a population of some Protestant denominations which mostly reject vaccination on religious grounds. The majority of the cases belong to a population of about 66,000 individuals under the age of 27, who were not vaccinated on religious grounds. No case of polio, however, was found in about 384,000 individuals under the age of 27 who has not been vaccinated on other than religious grounds. No case of polio occurred in the vaccinated (inactivated poliovaccine) population. The exclusive spread of the epidemic in a poorly vaccinated but coherent population, whose members are thoroughy mixed in an extremely well vaccinated population (annual acceptance rate about 95%) is discussed.


Poliovirus antibodies in age groups: an assessment of obligatory vaccination in Belgium.

Since 1967, compulsory oral vaccination before 2 years of age has been nearly the only form of poliovaccination practiced in Belgium, and there have been almost no cases of polio. In recent sera from 2,225 Belgians, 2 year olds lacked demonstrable seroneutralizing antibody, in 31% for type 1, in 8% for type 2, and in 21% for type 3. The lack of antibodies grew as the children reached 11--12 years, at which age 65% lacked antibody for type 1, 15% for type 2, and 54% for type 3; only 19% had antibodies against all types. It is argued that this decline of antibodies is not due to insufficient vaccination in former years, but is a strong example of the waning of vaccine-acquired serum antibody. Antibody rates went up after age 12. The highest rates of types 1 and 3 antibodies were seen at age 20--59. There was some decrease of antibody after 60 years. Mean antibody titers paralleled positivity rates. Results of two laboratories studying different populations with different standard techniques were similar. The data support revaccination at school entrance.


Herpes simplex virus: benefit versus risk factors in immunization.

Vaccines developed against herpes simplex viruses (HSV) should be effective in two respects: (i) they should prevent primary infections and virus latency and (ii) if applied to individuals with recurrent herpetic lesions they should reduce the number of recurrences or at least mitigate the symptoms. The efficiency of vaccines which are presently available will be reviewed. Problems associated with the possible use of live attenuated or inactivated HSV vaccines will be discussed. Comparative aspects to existing vaccines against other herpes group viruses will be stressed.


Antibody and cell-mediated immunity to a DNA free herpes simplex subunit vaccine.

The immunogenicity of a DNA free herpes simplex subunit vaccine was evaluated in chimpanzees and rabbits. The results clearly demonstrate that 1 injection of 3 micrograms/kg elicited antibodies as well as cell-mediated immunity in all the animals studied. These antibodies persisted for at least 6 months. Furthermore the vaccine also protected 50% of the animals against an experimental infection and reduced the rate of latent infection in nervous sensory ganglia.


Guillain-Barré syndrome associated with Epstein-Barr virus in a cytomegalovirus-negative patient.

Epstein-Barr virus and cytomegalovirus have both been associated with Guillain-Barré syndrome after antibody investigations in several patients. In the teenage female patient in this present report, Guillain-Barré syndrome following infectious mononucleosis was associated with sero-conversion against Epstein-Barr virus but not cytomegalovirus. The findings are consistent with the hypothesis that Epstein-Barr virus might be an etiological agent for the Guillain-Barré syndrome and that infection with cytomegalovirus is not a requisite for the syndrome.


[Granulosa cell and thecacell tumors in 25 cases (author's transl)].

Granulosa cell tumors are derived from specific ovarion stroma and have the ability to produce hormones. In the 25 cases of granulosa cell tumors no concommitant endometrial carcinoma was found. Because of the varying degree of malignancy tumors which contain granulosa cells must be differentiated from pure theca cell tumors. Granulosa cell tumors are frequently malignant. Theca cell tumors are benign. The treatment of granulosa cell tumors is fundamentally the same as that of epithelial cancers of the ovary. The treatment of theca cell tumors is conservative.


[Hidradenoma of the vulva (author's transl)].

Five cases of hidradenoma of the vulva are reported. The differential diagnosis is discussed. Painstaking microscopic diagnosis is very important since the hidradenoma may be confused with an adenocarcinoma. Local excision of the small tumour is the only treatment necessary.


[Genetic determination of the response of the mouse hypothalamo-hypophyseo-adrenal system to cold and immobilization stress].

Peculiarities of the hypothalamo-pituitary-adrenal system (HPAS) reaction in mice of 8 inbred strains to a stress effect of the immoblization and of cold have been studied and a comparison of the effect with the reaction of the adrenal cortex to its direct stimulator--ACTH have been carried out. As a result of the study of the resting level of 11-hydroxycorticosteroids in peripheral plasma of inbred mice significant interstrain differences were found. A positive correlation between resting corticosteroids level and the reaction of HPAS to the stress in mouse strains under study was found, but none of them correlated with the reactivity of the adrenal cortex to ACTH.


Construction and characterization of a recombinant plasmid encoding the gene for the thymidine kinase of Herpes simplex type 1 virus.

We have constructed a hybrid plasmid by insertion of the thymidine kinase (TK) gene of Herpes simplex virus (HSV) type I at the BamHI site on Escherichia coli plasmid pBR322. The restriction endonuclease cleavage site map for the viral DNA fragment was determined for ten nucleases, and the insert in the recombinant plasmid has the same restriction nuclease digestion pattern as bona fide viral DNA. This result indicates that the plasmid contains an accurate copy of the viral DNA. The viral TK gene carried on the plasmid can be introduced into mammalian cells where it is expressed. This source of DNA with a selectable marker should be of considerable practical use in gene-transfer experiments in mammalian cells.


Hepatic prolyl hydroxylase and collagen synthesis in patients with alcoholic liver disease.

Hepatic prolyl hydroxylase activity and collagen synthesis were measured in patients with alcoholic liver disease to determine the feasibility of using the enzyme prolyl hydroxylase as a marker of hepatic fibrogenesis. Alcoholic patients with liver histopathology consistent with normal, steatosis, alcoholic hepatitis, early cirrhosis, or advanced cirrhosis were analysed for liver prolyl hydroxylase activity and in vitro collagen synthesis. Prolyl hydroxylase activity and the rate of in vitro collagen synthesis were correlated when these parameters were measured in samples of the same liver biopsy. Mean prolyl hydroxylase activity was significantly raised in all groups of alcoholic patients with alcoholic liver disease, except those with steatosis, when compared with alcoholic patients with normal morphology. Alcoholic patients with early cirrhosis had enzyme activity (mean +/- SE: 1.367 +/-0.162 mU/mg protein) significantly raised over all other groups. Mean enzyme activity was less raised (0.985 +/- 0.097 mU/mg protein) in patients with advanced cirrhosis. The percentage of collagen synthesis in patients with early or advanced cirrhosis was also raised compared with alcoholic patients with normal morphology. Prolyl hydroxylase activity and the rate of collagen synthesis are significantly correlated (r=0.62). These findings suggest that hepatic prolyl hydroxylase activity is a useful indicator of hepatic fibrogenesis and its measurement on available liver biopsy tissue should be a potent diagnostic tool reflecting active fibrogenesis and predicting progression of alcoholic liverdisease.


Vitamin D metabolism in acute and chronic cholestasis.

To study the effects of acute and chronic cholestasis on vitamin D metabolism we investigated six cases of acute extrahepatic obstructive jaundice and eight cases of primary biliary cirrhosis (PBC) (three supplemented with vitamin D). Plasma 25-hydroxyvitamin D (25OHD) was low in the patients with PBC unsupplemented with vitamin D but normal in obstructive jaundice. None of the patients with PBC showed radiological or histological evidence of osteomalacia. In PBC, dietary intake of vitamin D was low but response to ultra-violet irradiation of the skin was normal even in those with a considerably raised serum bilirubin. Patients with PBC or obstructive jaundice had low levels of 25 hydroxyvitamin D binding protein which correlated with the serum albumin. The half-life of intravenously injected (3)H vitamin D(3) ((3)HD(3)) and the subsequent production of (3)H 25OHD were normal in all the patients with obstructive jaundice and in most with PBC. The two patients with PBC who produced less (3)H 25OHD than expected were receiving vitamin D supplements. The urinary tritium ((3)H) excretion after the injection of (3)HD(3) correlated with the serum bilirubin. After the injection of (3)H 25OHD(3) the urinary excretion of (3)H was minimal and did not correlate with the serum bilirubin, suggesting that the radioactivity appearing in the urine after the (3)H vitamin D(3) injection was associated with vitamin D metabolites other than 25OHD. Factors contributing to the low plasma 25OHD in primary biliary cirrhosis may be a low dietary intake of vitamin D, inadequate exposure to ultra-violet light, and a tendency to urinary wastage of vitamin D metabolites.