[General adaptation syndrome (Selye) in cattle. 6. Influence of stress conditions on antibody levels after active and passive immunization as well as on the topographic distribution of various groups of pathogens in the gastrointestinal canal]. The action of experimental and natural stressors on the humoral immune response of calf was studied. Particular attention was given to the H-antigen of S.dublin and equine erythrocytes,the degradation rate of passively acquired humoral antibody, as well as the quantity and topographic distribution of certain groups of germs in the gastrointestinal tract. The following results were obtained:1. Antibody formation was impededby repeated or lasting stressor effect (ACTH injections). 2. The immunological reactions of the calves involved to antigen injection immediately after transport into the rearing unit were stronger than those to antigen application three days from transfer. Immunisation of animals transferred to rearing or fattening units, therefore, should be applied immediately after arrival in the new accomodation, but no interval of three or four days should be allowed. 3. Antibody formation was no longer impaired in calves immunised two weeks from transfer, as compared to those immunised immediately after arrival in the rearing unit. This seemed to suggest that by that time adaptation of the animals to their new environment had been almost complete. 4. Lasting stress (slow drip infusion of ACTH, cortisol, colibacteria or coli-endotoxin) led to no detectable by paper electrophoresis. 5. Calves that had been given three weekly dosesof 1 IU ACTH per kilogram of live weight through four weeks,did not differ,withthe authors'method,from the controls regarding the decomposition rate of passively acquired humoral antibody. 6. ACTH slow drip infusions of calves over several days caused higher concentrations of colibacteria throughout the intestinal tract, including those proximal sections of the small intestine in which little or no colibacteria should occur under physiological conditions in calves of that age.


Hyporesponsivity of chronic schizophrenic patients to dextroamphetamine. Among the evidence supporting the dopamine hypothesis of schizophrenia is the finding that both amphetamine and methylphenidate hydrochloride, potent releasers of dopamine, can cause exacerbation of symptoms in the acute schizophrenic patient. The present report describes three experiments in which the effects of amphetamine in chronic schizophrenic patients were studied. In one of the experiments, orally administered, daily doses of 20 mg of dextroamphetamine sulfate given at 8 PM had little or no effect on the sleep duration of the subjects. In the other two experiments, doses up to 40 mg given orally also had little or no effect on the performance of the subjects on a variety of behavioral tests. There was no evidence of an exacerbation of the disease process in any of the subjects. The most consistent amphetamine effect was a dose-related increase in blood pressure. These results indicate that the chronic schizophrenic patient may be hyporesponsive to amphetamine and suggest that if the dopamine hypothesis is correct, then it must be modified to take into account these findings in the chronic patient.


Diagnosis and treatment of minimal brain dysfunction in adults: a preliminary report. Minimal brain dysfunction (MBD) has long been considered a disorder limited to childhood. A number of longitudinal and adoption studies suggest that MBD may persist into adult life where its existence is concealed by the application of a variety of diagnostic labels. In order to test the hypothesis that MBD does persist into adulthood, 15 putative MBD adults were identified on the basis of current MBD-like complaints, self-description of MBD characteristics in childhood, and a parental rating (on a standardized form) of "hyperactivity" in childhood. Eleven of the 15 subjects were given a double-blind trial of methylphenidate hydrochloride, and all 15 were given an open trial of pemoline, imipramine hydrochloride, or amitryptiline hydrochloride. Eight of the 11 showed a significant response to the double-blind trial of methylphenidate. Of the 15, eight showed a good response to stimulants or tricyclic antidepressants, two showed a moderately favorable response, and five were unresponsive to drug therapy.


Prediction of tricyclic antidepressant response: a critical review. This article reviews all the prospective, double-blind controlled studies that have evaluated the prediction of response to imipramine hydrochloride and amitriptyline hydrochloride in depressed patients. Despite widely divergent methodologies, an attempt is made to extract clinically useful conclusions from these data. Critiques of each study and the criteria used in their evaluation are presented, with suggestions for future research included. The predictors of positive response to imipramine and amitriptyline are as follows: upper socioeconomic class, insidious onset, anorexia, weight loss, middle and late insomnia, and psychomotor disturbance. The predictors of poor response are the following: neurotic, hypochondriacal, and hysterical traits, multiple prior episodes, and delusions. Pretreatment urinary 3-methoxy-4-hydroxyphenylglycol (MHPG) levels may some day be useful in predicting to which of these two tricyclic antidepressants a patient will respond.


The antitussive and expectorant drug Duopect evaluated by the preferential test. The effect of the preparation Duopect (narcotine hydrochloride + glyceryl guiacolate) on intensity and frequency of cough and expectoration was evaluated by the preferential test in 62 patients with chronic bronchitis. The preparation was compared to narcotine and to a placebo in a double-blind study. Duopect was found to be an effective antitussive and expectorant medicine with only minimal and transient side effects.


Botulinum antitoxins and antibacterial IgM and IgG antibodies in sera of persons immunized with botulinum polytoxoid combined with cholera vaccine. I. Response to botulinum toxoid. Two groups of subjects were immunized with combined vaccine containing aluminum hydroxide-adsorbed botulinum toxoids, types A, B, and E, and 4 mld of formaldehyde-inactivated V. cholerae Inaba and Ogawa organisms. The first group included laboratory workers who were previously immunized against cholera and had professional contact with botulinum toxins and viable V. cholerae organisms. The second group included young men who were never vaccinated against botulism or cholera. The three-dose immunization schedule with combined vaccine resulted in clear-cut antitoxin response; after the third dose, the A, B, and E antitoxin level ranged from 0-2 to 10 IU/ml. Immunity against botulinum toxins lasted at least one year. Distribution of the antitoxins among IgM and IgG globulin classes resembled that in the case of response to other toxoids; 21 days after the third immunization antitoxin activity was found in IgG globulins.


Use of antilymphocyte globulin for leukemia induction in mice. Biological activity of rabbit antimouse lymphocyte sera (ALS) and their IgG fractions was studied. Several ALS and ALG pools were cytotoxic in vitro for normal and leukemic mouse lymphocytes, causing temporary depletion of circulating leukocytes, prolongation of skin allografts, and progressive growth of L-1210/V leukemia transplanted across an H-2 barrier. As a rule, splenomegaly and a significant increase in the number of spleen cells were observed after i.p. injection of ALG. In experiments on the influence of prolonged treatment with ALG on tumor incidence in mice, one ALG pool appeared to be highly and repeatedly leukemogenic. The possible mechanism of leukemia induction by ALG was discussed.


Infections complicating orthotopic liver transplantation: a study emphasizing graft-related septicemia. In 93 recipients of 102 orthotopic liver homografts, the incidence of bacteremia or fungemia exceeded 70%. The graft itself was usually an entry site for systemic infection after both immunologic and nonimmunologic parenchymal injury, especially if there was defective biliary drainage. The role of the homograft itself as the special infectious risk factor has prompted increased use of defunctionalized jejunal Roux limbs to reduce graft contamination. It has also stimulated very aggressive postoperative diagnostic efforts to rule out remedial mechanical complications of the transplant.


Altered reticuloendothelial function in long-term renal allograft recipients. Using the clearance of microaggregated iodinated human serum albumin reticuloendothialial system (RES) phagocytic function was tested in 48 long-term renal allograft recipients and was found to be defective at the time of testing in 70%. Depression of RES phagocytosis could be related to total steroid dosage in the previous year and to the patients liability to bacterial infections. Evidence from this test does not suggest an immunosuppressive effect of cytomegalovirus. However, three patients are discussed who have developed chronic active hepatitis that is not due to type B virus.


Determination of mumps and influenza antibodies by haemolysis-in-gel. A stabilized modifications of the single radial haemolysis-in-gel (HIG) technique was developed. Crude mumps or influenza virus preparations were coupled to erythrocytes with CrCl3 and mounted in agarose gel containing diluted guinea pig serum. Serial serum samples taken from 204 conscripts before and after vaccination with a killed mumps vaccine were analysed with the HIG technique. The results showed that the sensitivity of the HIG test in detecting seroconversion was higher than that of the conventional haemagglutination inhibition test (HI). The correlation between the diametre of the rings of haemolysis and antibody titres measured by the standard complement fixation (CF) technique was fairly good. 62 pairs of sera from patients with suspected influenza were tested by both the HIG-method and the conrentional CF and Hi techniques. The HIG test appeared to be as sensitive as the common methods in detecting increases in antibody levels. The HIG  test was found to be insensitive to the nonspecific inhibitors of haemagglutination and aggregated IgG while it was slightly affected by the rheumatoid factor. These features together with the technical ease of performing the test, make the HIG test superior to the conventional HI and CF techniques in mumps and influenza diagnostics.


Alterations in corneal morphology following thermokeratoplasty. Fifteen corneal buttons were removed at keratoplasty from patients with keratoconus who had previously undergone thermokeratoplasty (TKP). The buttons were studied with light microscopy, in an attempt to analyze the type of changes produced by TKP. The principal histologic findings in patients with retarded epithelial healing (8 of 15) included epithelial thinning, bullous keratopathy, thickening of the epithelial basement membrane, and frequent destruction of Bowman's membrane. More severely affected patients showed asceptic stromal necrosis and fibrinous iritis with hypopyon. A poor visual result in patients with normal epithelial healing (7 of 15) was caused by superficial stromal scarring or persistent inflammatory infiltrate. Vascularization was seen in only one patient.


The relief of anxiety through relaxing meditation. Our sensory input derives from sources in the environment, in our body and in the mind itself. When the sensory input reaches a critical level it is incompletely integrated, and anxiety results. A logical understanding of the cause of anxiety has no therapeutic effect. But the mind itself has the ability to reduce anxiety if suitable circumstances are provided. This can be quite easily achieved in the stillness of mind induced in a simple meditative experience known as Mental Ataraxis. The patient is first shown complete physical relaxation in global fashion. He is then brought to experience the relaxation as part of his whole being so that his mind fully participates in the process. He practises this, starting in a position of slight discomfort which eases as the meditative experience develops. The approach does not involve the patient in doing less work. The lessening of anxiety reduces nervous tension, psychosomatic disorders and defensive distortions of the personality.


The construction and analysis of sucrose gradients for use with zonal rotors. The rate of sedimentation of a particle in a sucrose solution depends on the viscosity and density of the medium. These two variables are related to the sucrose concentration and the temperature of the medium by new simple equations. These equations were used in a rapid iterative procedure that relates the distance moved by a zone in a continuous sucrose gradient to its sedimentation coefficient. It is shown by comparison with experiment that this iterative method allows the distance moved by a zone to be calculated rapidly. The method may therefore be used to optimize the separation of particles in a sucrose-gradient-centrifugation experiment. The method also allows the unknown sedimentation coefficients of several zones to be measured from a single sucrose-gradient-centrifugation experiment.


The accessibility of protein-bound dinitrophenyl groups to univalent fragments of anti-dinitrophenyl antibody. A series of N-(N-dinitrophenylaminoalkyl)maleimides were sythesized with alkyl-chain lengths of two, four and six carbon atoms. When these compounds reacted with the thiol group of mercaptalbumin, the tryptophan fluorescence of the protein was quenched. This change in fluorescence was used to determine the rate of reaction of the Dnp (dinitrophenyl)-maleimides with mercaptalbumin. The second-order rate constants were similar to those observed in reactions between low-molecular-weight thiol compounds and maleimides. When N-(N-Dnp-aminoalkyl)succinimidomercaptalbumins were added to univalent fragments of anti-Dnp antibody the antibody fluorescence was quenched. Florescence-quenching titrations showed that the protein-bound Dnp groups were fully available to the antibody even when the alkyl chain was short. The apparent dissociation constants were significantly greater than that of the interaction between anti-Dnp antibody and the free hapten, 6-(N-Dnp)-aminohexanoate. The antibody fluorescence was quenched efficienty by [dnp-Lys41]ribonuclease A, also with an increased dissociation constant. It could be concluded from the increase in dissociation constant that the Dnp group spent no more than 0.1% of its time in the dissociated state, available to antibody. The second-order rate constants for the association between the Dnp-mercaptablumins and the antibody were determined and were similar in magnitude to those observed in other interactions between protein and anti-protein antibody.


Solubilization and other studies on adenylate cyclase of baker's yeast. 1. Adenylate cyclase of Saccharomyces cerevisiae was sedimented from mechanically disintegrated preparations of yeast over an unusually wide range of centrifugal forces. 2. The enzyme was readily solubilized by Ficoll and by Lubrol PX. Lubrol caused a 2-fold activation. 3. Both particle-bound and Lubrol-solubilized enzyme had an apparent Km for ATP of 1.6 mM in the presence of 0.4 mM-cyclic AMP and 5 mM-MnCl2 at pH 6.2 and 30 degrees C. 4. The Lubrol-solubilized enzyme behaved on gel filtration as a monodisperse protein with an apparent mol.wt. of about 450000.


[The matched-pairs differences-W-test for evaluating clinical treatment effects in matched-pair samples (author's transl)]. Starting from the U-test (see Buck 1975), for 2 independent samples of differences (e.g. pre-post differences), a corresponding test for 2 dependent samples of differences of pre- and post-measurements of 2 treatments (e.g. verum vs. placebo) is described (matched-pairs differences-W-test). Clinically relevant applications (verum vs. placebo) of the matched-pairs differences-W-test are mentioned and discussed as to their restrictions.


Clinical significance of immune deposits in the skin in SLE. The lupus band test (LBT) was evaluated in 53 SLE patients. There was no significant correlation between the presence of a positive LBT and active clinical nephritis, renal histology, hypocomplementemia, or elevated anti-nDNA antibodies. In 14 patients the classes of immunoglobulins in the skin and the glomeruli were directly compared and differences were present in 11 patients. Repeat LBTs frequently became positive in patients after successful suppression of disease activity.


Simpson as a teacher--a students notebook. This paper gives an account of a book of lecture notes taken by a medical student who attended Simpson's course during 1848-9. The notes include an early account of the use of chloroform in midwifery, and describe the obstetric practice of the period as taught by Simpson.


Leukaemogenic action of phorbol in intact and thymectomized mice of different strains. Phorbol, the unesterified parent alcohol of the skin promoter TPA, was administered i.p., twice weekly, throughout the lifetime of mice of 7 inbred strains: males and females of AKR/J, C3Heb and BALB/c, and females of SJL/J, DBA/2, SWR and C57BL. A striking difference in strain response was observed, with a pronounced leukaemogenic effect in SWR, a signficiant shortening of the latent period for spontaneous reticulum cell sarcomas (RCNB) in SJL/J, and no demonstrable effect in the other strains. When mice of 3 of the above-mentioned strains (SWR, SJL/J and AKR/J) were thymectomized prior to the beginning of phorbol treatment, different patterns of response were again observed. Thymectomy did not influence the leukaemia incidence in SWR mice, slightly inhibited RCNB development in SJL/J mice and converted phorbol into a leukaemogenic agent for AKR/J mice.


Smooth-muscle-associated contractile protein in renal mesenchymal tumour cells and in transformed cells from DMN-injected rats. Cryostat sections and established in vitro cultures of dimethylnitrosamine(DMN)-induced renal mesenchymal tumours and monolayer cultures of transformed kidney cells derived from rats treated with a carcinogenic dose of DMN were examined by indirect immunofluorescence with human serum containing smooth muscle antibody. Eight mesenchymal tumours examined showed filamentous cytoplasmic staining of spindle cells infiltrating between renal tubules, whilst in normal kidneys interstitial cells were only weakly positive. In established in vitro cultures from 6 mesenchymal tumours, different patterns of staining were observed in morphologically different cell forms, ranging from fine filamentous staining in giant cells to diffuse cytoplasmic fluorescence in small bipolar cells, and cell outline staining in polygonal cells. In addition filamentous staining of microvillous projections and nucleolar staining were observed in some tumour cells. Monolayer cultures of transformed kidney cells showed strong staining of coarse, randomly-orientated cytoplasmic filaments, whilst fibroblasts cultured from normal rat kidney demonstrated an ordered array of fine, parallel filaments. Specificity of the immunofluorescent staining reaction was established by failure to obtain staining with normal serum, with smooth muscle antibody serum neutralized by homogenates of smooth muscle or extracts containing actin derived from smooth muscle. These results indicate that there is an apparent increase of actin-like contractile microfilaments in transformed cells and in renal mesenchymal tumours. The cytoplasmic contracile microfilaments in these cells may play a role in tumour cell mobility and invasion.


Membrane-associated phosphatidylglycerophosphate synthetase from Escherichia coli: purification by substrate affinity chromatography on cytidine 5'-diphospho-1,2-diacyl-sn-glycerol sepharose. The membrane-associated cytidine 5'-diphospho-1,2-diacyl-sn-glycerol (CDP-diglyceride):sn-glycerol-3-phosphate phosphatidyltransferase (EC 2.7.8.5) from Escherichia coli has been solubilized wiTriton X-100 and purified 6000-fold to 85% of homogeneity. The major purification was attained using several modifications of the the CDP-diglyceride Sepharose affinity chromatography system described by Larson et al. (Larson, T.J., Hirabayashi, T., and Dowhan, W. (1976), Biochemistry 15, 974). The native enzyme in Triton X-100 had an apparent molecular weight of over 200 000, as judged by Sepharose 6B gel filtration. The apparent size of the native enzyme appeared to be due to its association with Triton X-100, as judged by sucrose gradient centrifugation, polyacrylamide gel electrophoresis, and the lack of affinity for ion-exchange resins. The minimum subunit molecular weight of the enzyme, determined by sodium dodecyl sulfate polyacrylamide gel electrophoresis, was 24 000. This low molecular weight is consistent with the stability of enzyme to heat, urea, or sodium dodecyl sulfate denaturation. The purified enzyme had an absolute requirement for magnesium ion (KM = 50 mM) and Triton X-100 (0.5-6%) for activity when either CDP-diglyceride or dCDP-diglyceride was used as substrate. Kinetic analysis of the enzymatic reaction indicated an ordered sequential Bi-Bi reaction with the liponucleotide forming a dead-end complex at high concentration, which inhibited both the forward and reverse reactions. The enzyme would not hydrolyze the pyrophosphate bond of its lipid substrate or the phosphate esters of its lipid product but would catalyze a cytidine 5'-monophosphate dependent exchange reaction between glycero-3-phosphate and phosphatidylglycerophosphate.


The biology of talc.

The biology of talc. Data are presented on the effects on health of talc dusts from exposure in industry and use of talc-containing health products. The mineralogy of talc and the composition of cosmetic and industrial grade talc dusts are described. Studies in animals are reviewed, and epidemiological data are considered in relation to exposures that occur during industrial and consumer uses of talc dusts. Hamsters exposed to 8 mg/m3 of respirable cosmetic grade talc dust for up to 150 minutes a day for 300 consecutive days showed no difference in incidence or nature of pathological lesions from those observed in a group of untreated animals. A retrospective study of the causes of death of 227 talc mine millers exposed to cosmetic grade talc at the threshold limit value for talc (20 million parts per cubic foot) for an average of 15-8 years showed that the causes of death were no different from those in a control cohort not exposed to talc dust. The available data indicate that talc dust exposure in the modern mining of cosmetic grade talc does not appear to be injurious to health. The significantly lower dust exposure in the normal use of cosmetic grade talc dusts in talc-containing health and cosmetic products confirms that their use is not a hazard to health.


Double-stranded DNA in methanol-ethanol-buffer solvent system. DNA in a solvent system consisting of roughly equal volumes of methanol and ethanol and 5% buffer has a conservative circular dichroism (CD) spectrum of very low intensity above 220 nm and an increase of epsilon258 comparable to that of denatured DNA (about 40%). A direct comparison of this spectrum with the CD of single-stranded DNA reveals many differences, indicating DNA in this solvent system has a conformation different from that of denatured DNA. When the alcohols are removed, the B form conformation and normal epsilon258 are restored in native DNA, while single-stranded DNA remains denatured. A double-stranded structure of DNA in the methanol-ethanol-buffer solvent system is confirmed by the neutral cesium chloride density gradient centrifugation of DNA in which one chain is labeled with [14C]thymidine and the other [3H]5-bromodeoxyuridine. The doubly labeled DNA exposed to the alcohol solvent system has a centrifugal pattern identical with that of control DNA; the two radioactivities cosediment and form a superimposing band, distinctly different from that of single-stranded DNA; 3H-labeled (thymidine) chains sediment further than 14C-labeled chains (5-bromo-deoxyuridine). Denatured DNA exhibits varying CD spectra depending on solvents. It is suggested that single-stranded DNA in different solvent systems assumes different modes of base stacking.


Conformational changes of 30S ribosomes measured by intrinsic and extrinsic fluorescence. The intrinsic tryptophan fluorescence and the fluorescence of N-(3-pyrene)maleimide, a covalently bound sulfhydryl-specific extrinsic probe, have been used to study the conformation of the 30S ribosomal subunit of Escherichia coli. (a) The tryptophan fluorescenct spectrum of the free ribosomal proteins is shifted to shorter wavelengths than that of free tryptophan. When the proteins are incorporated into the organized structure of the ribosome, there is a small additional blue shift and the emission band becomes narrower. In 6 M urea, the spectrum of the proteins, whether free or in the ribosome, becomes identical with that of the amino acid, reflecting exposure of previously shielded tryptophan residues. (b) When magnesium-depleted ribosomes are unfolded at low ionic strength, the tryptophan fluorescence spectrum changes, although circular dichroism shows no change in alpha-helix content of the proteins. (c) Intrinsic and extrinsic fluorescence were both found to be sensitive to a limited and fully reversible transition that takes place when ribosomes are incubated under conditions that increase their activity in vitro. This suggests that both probes may be of use in monitoring conformational changes that occur under conditions consistent with activity. The kinetics of the concurrent changes in extrinsic fluorescence and aminoacyl-tRNA binding activity were compared. (d) Conditions are described for labeling ribosomes with N-(3-pyrene)maleimide without impairing their activity.


Demonstration of ribosome-dependent photoinduced chain breakage of the 16S ribosomal ribonucleic acid component of the Escherichia coli 30S ribosomal subunit. The effects of 253.7-nm radiation on the structural integrities of free and ribosome-bound 16S ribosomal ribonucleic acid (rRNA) have been elucidated. Exposure of aqueous solutions of Escherichia coli 30S ribosomal subunits to 253.7-nm radiation results in RNA-chain scission and the formation of single-stranded breaks in double-stranded regions of the ribosome-bound 16S rRNA. The minimum doses of  incident 253.7-nm radiation required for the first detection of the two types of RNA chain breaks are 2 X 10(20) quanta for single-strand breaks in double-stranded regions of the ribosome-bound 16S rRNA,and at least 5 X 10(20) quanta for RNA-chain scission. In contrast to the photosensitivity of ribosome-bound 16S rRNA toward chain breakage, free 16S rRNA seems to be resistant toward photoinduced chain breakage at doses of incident 253.7-nm radiation up to at least 10(21) quanta.


Mutations affecting the reduced nicotinamide adenine dinucleotide dehydrogenase complex of Escherichia coli. A strain carrying a point mutation affecting the NADH dehydrogenase complex of Escherichia coli has been isolated and its properties examined. The gene carrying the mutation (designated ndh) was located on the E. coli chromosome at about minute 23 and was shown to be cotransducible with the pyrC gene. Strain carrying the ndh- allele were found to be unable to grow on mannitol and to grow very poorly on glucose unless the medium was supplemented with succinate, acetate or casamino acids. The following properties of strains carrying the ndh- allele were established which suggest that the mutation affects the NADH dehydrogenase complex but apparently not the primary dehydrogenase. Membrane preparations possess normal to elevated levels of D-lactate oxidase and succinate oxidase activities but NADH oxidase is absent. NADH is unable to reduce ubiquinone in the aerobic steady state and reduces cytochrome b very slowly when the membranes become anaerobic. NADH dehydrogenase, measured as NADH-dichlorophenolindophenol reductase is reduced but not absent. NADH oxidase is stimulated by menadione although not by Q-3 or MK-1 and in the presence of menadione, cytochrome b is reduced normally by NADH. Further mutants affected in NADH oxidase were isolated using a screening procedure based on the growth characteristics of the original ndh- strain. The mutantions carried by these strains were all cotransducible with the pyrC gene and the biochemical properties of the additional mutants were similar to those of the original mutant. The properties of the group of ndh- mutants established so far suggest that they are affected in the transfer of reducing equivalents from the NADH dehydrogenase complex to ubiquinone.


An evaluation of N-phenyl-1-naphthylamine as a probe of membrane energy state in Escherichia coli. Colicin El and the uncoupler of oxidative phosphorylation, trifluoromethoxy-carbonylcyanidephenylhydrazone (FCCP), cause an increase in the fluorescence intensity of N-phenyl-1-naphthylamine bound to whole cells of Escherichia coli. It has been shown elsewhere that this fluorescence increase correlates well with de-energization. Addition of glucose causes a large cyanide-sensitive decrease of intensity, tentatively associated with energization, with the emission spectrum almost returning to the original trace with a peak at 417 nm. These data suggest that there may be a measurable competition between de-energization and energization of the cell membrane, and that the probe fluorescence intensity may be a general indicator of membrane energy level. The conclusions reached about cellular energy level from measurements of the probe fluorescence intensity correlate partly (a, b below, not c) with the energy level assayed physiologically through rates of active transport; (a) FCCP is found to be a poor inhibitor of proline transport if cells are first incubated with glucose, showing eutger cinpetition between the processes of energization and de-energization or an increase in the envelope permeability barrier to FCCP caused by glucose addition. (b) Cyanide blocks the fluorescence decrease caused by glucose and inhibits proline and serine transport, consistent with the decrease in probe fluorescence intensity indicating an increase in membrane energization. However, (c) it appears that the amplitude of the fluorescence intensity decrease caused by glucose addition in the presence of FCCP and colicin E1 greatly exaggerates the extent of real membrane energization. Glucose added after uncoupler can cause only a small increase, and after colicin, a negligible increase in the proline transport rate, indicating that the magnitude of the fluorescence intensity decrease after glucose addition is not a true measure of membrane energization, but rather seems to amplify this energization greatly. Glucose addition does not cause a decrease in fluorescence intensity in cells treated with EDTA to remove lipopolysaccharide and an apparent barrier to the probe. The rotational relaxation time of the probe in intact cells appears to correlate somewhat better with the cellular energy level than does intensity.


Changes in membrane proteins of Escherichia coli K12 mediated by bacteriophage IKe-specific plasmids. At least 3 minor proteins have been detected by sodium dodecyl sulfate polyacrylamide gel electrophoresis in the unseparated (31 000 daltons) and inner membranes (53 000 and 86 000 daltons) of a non-piliated strain of E. coli K12 bearing the phage IKe-specific resistance plasmid RM98 (Tra+); these proteins are lacking in its Tra- derivative and in the plasmidless host. In contrast, in another K12 host carrying each of different IKe-specific colicinogenic factors, an extra 30 000 dalton protein is observed in the outer membranes; a number of minor proteins seen in the inner membranes appear to be different from those observed in the RM98+ strain. The evidence suggests that the 30 000-31 000 dalton protein is transfer function specific.


Physicochemical studies on the creatine kinase M-line protein and its interaction with myosin and myosin fragments. The M-line protein which is identical to the muscle form of creatine kinase was purified from rabbit skeletal muscle using ion exchange chromatography. Gel electrophoresis in the presence and absence of sodium dodecyl sulfate revealed the protein to be homogeneous. Sodium dodecyl sulfate gel electrophoresis gave 44 000 +/- 2000 as the minimum molecular weight while low speed sedimentation equilibrium experiments yielded a molecular weight of 84 000 +/- 4000, suggesting that the parent molecule is a dimer. Circular dichroism spectra revealed the presence of two negative dichroic bands located at 218 and 208 nm suggesting the presence of some beta-structure. Ellipticity values at these two wavelengths were -8000 +/- 400 and -9000 +/- 400 deg-cm2-dmol-1. Circular dichroism measurements indicated the protein to interact with myosin, heavy meromyosin and heavy meromyosin subfragment 1 (S1). The Ca2+-activated ATPase activities of myosin, heavy meromyosin and subfragment 1 were inhibited by the addition of M-line protein. When the protein was mixed with subfragment 1 in a 1:1 mole ratio in 0.15 M KC1, 50 mM Tris pH 8, low speed sedimentation equilibrium studies gave a molecular weight of 205 000 +/- 10 000 for the complex, indicative of an interaction of the two components. Both circular dichroism and sedimentation equilibrium studies indicated no interaction of M-line protein with light meromyosin.


UDP-glucose dehydrogenase from Escherichia coli. Purification and subunit structure. UDPglucose dehydrogenase from Escherichia coli has been purified 330-fold with an overall yield of 27%. A single homogeneous subunit was demonstrated by ultracentrifugation in 6 M guanidium chloride and by dodecyl sulfate-polyacrylamide gel electrophoresis. Since the molecular weight of the intact dehydrogenase is in the order of 86 000 and the subunit weight determined by the dodecyl sulfate-polyacrylamide gel electrophoresis is 47 000, the enzyme consists of two polypeptide chains. The sole amino terminal acid shown by the dansylation technique was arginine. Forty-four tryptic peptides were obtained by peptide mapping, in agreement with the number of arginine and lysine residues/mole protein [43] determined by amino acid analysis. The data are consistent with the presence of two identical or very similar polypeptide chains in E. coli UDPglucose dehydrogenase.


End group analysis of yeast fatty acid synthetase. The purified yeast fatty acid synthetase complex has been subjected to amino and carboxyl terminal amino acid end group analysis. Amino end groups were studied by Edman degradation and by dansylation of the sodium dodecyl sulfate- or urea-denatured complex. No N-terminal amino acid could be identified by either method. C-terminal amino acids were investigated by tritium labeling and by digestion of the complex with carboxypeptidases A and B. By both methods, the two amino acids valine and lysine were consistently identified as the C-termini of two different polypeptide chains. After separation of the fatty acid synthetase subunits A and B by sodium dodecyl sulfate polyacrylamide gel electrophoresis lysine was identified as the C-terminus of subunit A and valine as that of subunit B. The results are interpreted as evidence that the yeast fatty acid synthetase complex is basically composed of two nonidentical and multifunctional polypeptide chains.


Cell wall synthesis regulation in Saccharomyces cerevisiae. Effect of RNA and protein inhibition. In this investigation the regulation of wall formation in Saccharomyces cerevisiae ts-136 (Hutchison, H.T., Hartwell, L.H. and McLaughlin, C.S. (1969) J. Bacteriol. 99, 807-814) was analyzed by following the inhibition of RNA and protein synthesis. Lomofungin, thiolutin and 8-hydroxyquinoline at the concentrations needed to inhibit RNA synthesis also produced inhibition of glucan and mannan synthetases. The synthesis of RNA was also blocked in S. cerevisiae ts-136 by incubation at the non-permissive temperature (37 degrees C). Mannan formation decreased steadily but glucan synthesis remained after 4 to 5 h. After a few minutes of blocking protein synthesis with cycloheximide mannan synthesis was also blocked whereas glucan formation was unaffected by the presence of the drug. These results suggest a high degree of stability for glucan synthetases. S. cerevisiae ts-136 after 2 h of incubation at the non-permissive temperature (37 degrees C) showed a preferential formation of wall materials (mannan and glucan) indicating that the RNA messengers  which codify wall mannan peptides have a slower decay rate than those of the cytoplasmic proteins. The data presented indicate that the existence of stable glucan synthetases and RNA messengers of the wall mannan peptides of slow decay rate results in the continuous synthesis of glucans and mannoproteins of the yeast wall throughout the cell cycle.


The absence of DNA photoreactivation enzyme in yeast mitochondria. Mitochondria isolated from haploid yeast cells by spheroplast lysis were purified by flotation on renografin gradients. Electron micrographs and respiratory control ratios revealed that the purified mitochondria were still intact and functional. Assays for photoreactivation enzyme using as substrate [3H]-thymine-labeled Escherichia coli DNA were performed on crude and purified mitochondrial preparations. While the crude preparation contained high amounts of photoreactivation enzyme, it appeared to be associated with contaminating nuclei. The purified mitochondria lacked any photoreactivation enzyme activity. We suggest that yeast mitochondria do not normally contain photoreactivation enzyme.


Rifampicin and chloramphenicol effects on DNA replication in ultraviolet-damaged Escherichia coli B/r WP2 thy trp. The antibiotic rifampicin, which blocks specifically RNA synthesis, limited DNA replication in Escherichia coli strain B/r WP2 thy trp after an increase of about 50%, when added to the incubation medium at the time of replication initiation after ultraviolet fluences of 20 J/m2...


Interaction of microbial DNA with cultured mammalian cells. Binding of the donor DNA to the cell surface. Cultured fibroblasts grown in monolayer were incubated for a short time with radioactively labeled microbial DNA and diethylaminoethyl-dextran (DEAE-dextran), poly-L-lysine, or calcium phosphate, agents previously demonstrated by others to markedly enhance transfection. Immediately after such treatment of 1 by 10(-6)-1.5 by 10(6) cells with DNA, approx. 0.05-0.15 mug of donor DNA representing 10-30% of the input DNA became cell associated. In contrast, when the cells are similarly treated with only a DNA solution approx. 0.5-5% of the donor DNA was retained by the cells. More than 95% of the cell-associated donor DNA was shown to be bound to the surface of cells treated with polycation. It was also shown that in the absence of polycation treatment, most of the cell-associated donor DNA was bound to the cell surface.


Ultraviolet-induced inhibition of ribosomal RNA synthesis in yeast strains differing in radiation sensitivities. The ultraviolet-induced inhibition of rRNA synthesis has been measured during the first hour after irradiation for stationary yeast cells differing in radiation sensitivity. rRNA was isolated and separated on an agarose-polyacrylamide gel. The wild type and a mutant which is possibly defective in recombinational repair show a sigmoidal inhibition curve, an excision-deficient mutant shows an exponential one. From these curves it is deduced that a pyrimidine dimer acts as a transcription terminating lesion as was shown for bacteria. During the first hour after irradiation the excision repair system decreases the number of transcription terminating lesions by 22% in the wild type and 25% in the mutant defective in recombinational repair. An approximation of the repair efficiency gives a value of 7500-10 000 transcription terminating lesions per cell being removed during the first hour after irradiation by excision. Ultraviolet-induced lesions of this kind can partially be removed by photoreactivation. The inhibition coefficients are the same for 26 S and 18 S rRNA in stationary cells, whereas exponentially growing cells show different inhibition coefficients for 26 S and 18 S rRNA leading to the suggestion that the processing of the ribosomal precursor RNA is different in stationary and exponentially growing cells.


Characterization of the RNA transcribed in vitro from native mammalian DNA by Escherichia coli RNA polymerase. High-molecular-weight native mouse DNA was transcribed with Escherichia coli RNA polymerase under low salt conditions, and the nature of the DNA sequences transcribed determined by molecular hybridization. The results indicated that E. coli RNA polymerase does not transcribe the sequences in native mouse DNA randomly under these conditions. First, hybridization with a large excess of mouse DNA showed that no more than 5% of the RNA synthesized had been transcribed from repeated sequences in the DNA. Second, hybridization with tracer amounts of labelled non-repeated mouse DNA indicated that the bulk of the RNA had been transcribed from less than 1% of the non-repeated sequences and only about 10% had been transcribed from a further 25% of these sequences; the remaining non-repeated sequences in the DNA, amounting to 50% of the genome, were not represented in the RNA synthesized in vitro to any detectable extent. Third, the proportion (40%) of complementary DNA transcribed from mouse-liver nuclear polyadenylated RNA which hybridized with the RNA synthesized in vitro was significantly greater than would have been expected if transcription had been random. The data have also been interpreted as indicating the presence of two types of initiation site for E. coli RNA polymerase in the non-repeated sequences in mouse DNA. The frequencies of their occurrence have been calculated to be one per 150 000 base-pairs and one per 500 base-pairs, respectively.


Effect of stringent and relaxed control on transcription of the tryptophan operon from the ptrp promoter and the PL promoter in trp phage. Tryptophan (trp) mRNA synthesis from the authentic trp promoter (Ptrp) is apparently arrested upon translation blockage, while trp mRNA synthesis as a result of read-through from the Pl promoter of the N gene in trp phage is not so affected. When translation is blocked at a nonpermissive temperature in the temperature-sensitive mutants of Escherichia coli rel carrying altered ribosomal elongation factors G (strain CP78G) and Ts (strain HAK88), CP78G and HAK88 show relaxed and stringent phenotypes respectively in control of RNA synthesis. Under conditions causing translation blockage in both mutants, Pl-promoted synthesis of trp mRNA is not depressed while Ptrp-promoted synthesis of trp mRNA is blocked. The insensitivity of Pl-promoted transcription of the translocated trp operon to stringent control is also confirmed by using a strain 10b6s rel carrying a temperature-sensitive valyl-tRNA synthetase. In contrast to the above observation, transcription of the trp operon from either the Pl and Ptrp promoters in a 10b6r rel infected with trp is inhibited greatly at the nonpermissive temperature. This effect occurs even if the level of trp transcription observed at 30 degrees is already low, thus suggesting that translational machinery is intrinsically abnormal in the rel strain.


The sucrose gradient and native DNA S20,W, an examination of measurement problems. Sedimentation coefficients of T7, T2H AND T4 DNA were determined with isokinetic sucrose gradients in both 0.1 M and 1 M NaCl. The s values were completely equivalent to those measured by analytical ultracentrifuge and no reduction of s20,w was observed due to the presence of sucrose (anomalous sedimentation). s20,w values are calculated on the basis of both partial specific volume (v) and apparent specific volume (0). Using the latter value s20,w molecular weight relations are derived for 0.1 M and 1 M NaCl solvents. The glucosylation of T2H and T4 DNA appears to influence s20,w in a manner disproportionate to the molecular weight added by glucose.


Freeze-fracture ultrastructural alterations induced by filipin, pimaricin, nystatin and amphotericin B in the plasmia membranes of Epidermophyton, Saccharomyces and red complex-induced membrane lesions. The effects of chemically different polyenes on fungal membranes (Epidermaphyton floccosum, a human pathogenic fungus, and Saccharomyces cerevisiae) and human red blood cell membranes were studied by freeze-fracture electron microscopy in order to elucidate the interaction of these antibiotics with ergosterol. Each type of neutral, small amphoteric and large amphoteric polyenes produces a distinct morphoneutral, small amphoteric and large amphoteric polyenes produces a distinct morphological effect on the fungal membranes: (1) Pit formation type. Filipin, a neutral polyene, produces 250-300 A diameter "pits" or "invagination" both in ergosterol-containing fungal plasma membranes and cholesterol-containing red blood cell ghost membranes. (2) Network particle aggregation type. The small amphoteric polyene, pimaricin, produces a network of membrane particle aggregation which encloses 1000 A diameter particle-free areas in fungal membranes. These areas are slightly elevated toward the outside of the cell. (3) Random particle aggregation type. The large amphoteric polyenes, amphotericin B and nystatin, cause a random segregation of the fungal plasma membrane and the red blood cell ghost membranes into particle-free and aggregated areas. It is concluded that these morphological differences are due to different mechanisms of polyene-sterol interactions in which the different size of the mocrolide ring in the antibiotic structure may be involved. Since all of these antibiotics, except filipin, cause no alterations on whole red blood cells detectable by negative staining and freeze-fracture electron microscopy, it is possible that they have a higher affinity to ergosterol than cholesterol in membranes.


Interaction between tetanolysin and Mycoplasma cell membrane. 1. A partially purified tetanolysin preparation lysed the sterol-requiring Mycoplasma capricolum cells but had no effect on M. capricolum cells adapted to grow with no or very little cholesterol. The sterol-non-requiring Acholeplasma laidlawii cells grown either in a cholesterol-rich or a cholesterol-poor medium were unaffected by the tetanolysin preparation. 2. The lysis of M. capricolum cells by the tetanolysin preparation was temperature dependent, inhibited by cholesterol, sublytic concentrations of lucensomycin, and Mg2+. The sensitivity to lysis was greatly affected by the age of the culture, being highest in cells from the early logarithmic phase of growth and declining sharply thereafter. 3. Isolated M. capricolum membranes were capable of binding large amounts of the tetanolysin activity (up to 30 hemolytic units per mug membrane protein), 20 times as much as membranes of the adapted strain. The binding of tetanolysin activity to membranes was almost the same at 4,22, or 37 degrees C, and was very little affected by the age of the culture. The binding capacity of the membranes was not affected by the removal of 60-70% of membrane proteins by pronase digestion but markedly decreased with the removal of membrane lipids. 4. Of the five polypeptide bands detected in electrophorograms of the partially purified tetanolysin preparation, two bands (mol. wt. 44 000 and 42 000) were found to bind to the cholesterol-containing mycoplasma membrane preparation. EPR spectrometry revealed that the freedom of motion of fatty acid spin labels in the tetanolysin-treated membranes was markedly higher than that in untreated membranes. 5. The concept that tetanolysin interacts specifically with membrane cholesterol resulting in the shielding of cholesterol from its interaction with membrane phospholipids is discussed.


Release of outer membrane fragments from normally growing Escherichia coli. A complex containing lipopolysaccharides, phospholipids and proteine separated from the medium by gelfiltration on Sephadex G-200 or by centrifugation. Electron microscopy revealed that this material is released as vesicles and membrane fragements. To determine the origin of these fragments, they were compared to outer and cytoplasmic membranes with respect to keto-deoxyoctulosonic acid, phospholipid, and protein content, phospholipid composition, fatty acid composition, protein distribution on sodium dodecyl sulfate-polyacrylamide gels, buoyant density, and content of several membrane marker enzymes. The results of this comparison indicate that the membrane fragments found in the culture supernatant of normally growing Escherichia coli consist of practically unmodified outer membrane. Possible mechanisms as to the cause of the release of outer membrane fragments, and its relationship to cell-division, are discussed.


[Effect of polyene antibiotics on protein synthesis by free and membrane-bound ribosomes of Candida albicans]. Polyene antibiotics (levorin, nistatin, amphotericin B) inhibit protein synthesis at concentrations decreasing 14C-amino acid incorporation into Candida albicans protoplasts by 30--60%, the depression of membrane permeability beginning earlier than protein synthesis inhibition. Fractionation of protoplast lysates revealed that protein synthesis by free ribosomes was inhibited by antibiotics stronger than in case of membrane-bound ribosomes. It is supposed that different response of two ribosome classes for polyenes-induced damages is due to different sensitivity of free and membrane-bound ribosomes to the decrease of intracellular K+ concentration.


[Effect of streptomycin on RNA synthesis in E. coli cells]. Under streptomicin administration (10-200 mug/ml), initiation of translation is blocked at any steps before the formation of tightly bound subunits. Incomplete supression of protein synthesis with streptomycin stimulate rRNA synthesis in rel+ CP78 E. coli cells. Thus, partial uncoupling of translocation and transcription promotes compensatory synthesis of rRNA synthesis directed by rel-gene, but on addition to pre-starved cells, antibiotic can not stimulate rRNA formation. It is shown, that in contrast with many other ribosomal antibiotics streptomycin do not penetrate into the cell during aminoacid starvation.


[Equine pepsins].

[Equine pepsins]. 6 forms of pepsin are found in horse gastric juice. Amino acid sequence is determined of N-terminal (most variable) part of polypeptide chain of main pepsin components. Equine pepsines, which have pI 2.1 and 2.3, are found to have identical amino acid sequence at least for 31 amino acid residues. The same sequence is observed in the component with pI 2.6 for 10 first residues. The sequence of equine pepsin with pI 3.2 has 3 substitutions for 33 amino acids, when compared with pepsines having pI 2.1 and 2.3. The forms of equine pepsin studied are more similar than the other isoenzyme pair, human pepsin and gastricsin.


How chemicals may induce cancer.

How chemicals may induce cancer. Recent studies which lead to a better understanding of chemical carcinogenesis are reviwed. Almost all carcinogens require an activation by metabolic processing in order to operate "in vivo" or "in vitro". Some chemicals and drugs induce the enzymes which metabolize carcinogens. This induction seems to be of fundamental importance to the development of human cancers resulting from environmental pollution. Nearly all the lesions in cellular DNA caused by chemical carcinogens can be repaired by a DNA repair system. Failure to repair some lesions or the action of "error prone" repair can lead to mutations. The mutagenic effect of some chemicals provides a rapid screening test for the detection of their carcinogenic activity. Such tests are described and analyzed in detail. Finally, a general and overall scheme for possible mechanisms of action of chemical carcinogens is proposed.


Immunofluorescence of synovial membrane multifactorial analysis of the results. Synovial membrane taken by needle biopsy from the knee joint of 61 patients with various rheumatic diseases were studied using immunofluorescence methods. Staining techniques and their controls were detailed. Classical statistical tests and principal components multifactorial analysis of the data emphasized some differences between the pathological groups. Connective tissue diseases seemed to be characterized by plasma cells fluorescence and mixed immunoglobulins and complement deposits. These were mostly localized to extracellular spaces in sero-positive rheumatoid arthritis and to blood vessels in sero-negative rheumatoid arthritis and systemic lupus erythematosus. On the contrary, isolated immunoglobulins without complement were mostly found in the other inflammatory arthritis, while negative results were obtained in non inflammatory arthropathy. Immunoglobulin classes did not seem to have any diagnostic value. On the contrary, rheumatoid factor was specific for rheumatoid arthritis, whatever the serological pattern was, and it was particularly frequent in patients suffering from rheumatoid arthritis associated with a Sj5AOGREN SYNDROME. A strict relationship between classical histological findings and immunofluorescence results was not always found; so, immunological methods can be aquivocal.


Distribution of erythrocyte receptors at the surface membrane of human T-lymphocytes. During incubation at 4 degrees C, the binding sites for neuraminidase-treated human erythrocytes (nHRBC) and sheep red blood cells (SRBC) were diffusely distributed on the surface membrane of most rosette-forming normal human blood lymphocytes. Increasing the temperature to 37 degrees C resulted in a rapid rosette dissociation which was accompanied by a relative increase in cap formation and a corresponding decrease of ring-like rosettes. The conversion into caps was inhibited by reincubation of the cells at 4 degrees C. The results indicate a movement of the nHRBC and SRBC receptors in the plane of the human T lymphocyte plasma membrane.


Prognostic value of mouse red cell rosette formation in chronic lymphocytic leukaemia. Mouse red cell (MRC) rosette formation was studied in 20 malignant haemopathies. Only cells from chronic lymphocytic leukaemia (CLL) and hairy cell leukaemia (HCL) formed MRC rosettes with a high frequency. However, cells from 2 cases of CLL could not be demonstrated to form MRC rosettes, although they were undoubtedly found to be of B origin. These 2 cases presented as typical CLL, but they were observed to transform into a sarcomatous type after 5-6 months of evolution, suggesting that the study of MRC rosette formation might be of prognostic value in CLL.


Rosette forming T-cells in human lymphomas: relationship with age and clinical parameters. Peripheral blood spontaneous rosette forming T cells (RFC) were studied in 40 Hodgkin's Disease (HD) and 20 lymphosarcoma (LS) patients, classified according to the histological type and spread of disease and to their age, treatment, clinical condition and evolution. LS patients had near normal RFC values; in HD, the lowest values of RFC corresponded to the most severe histological picture. In all patients, differences could be seen according to the spread of the disease and the clinical condition, but none in relation with treatment nor with the clinical evolution in a one year period after the test. While in controls and LS patients, RFC were higher in younger than older subjects, in HD, values were higher after 50 years of age. The usefulness of RFC test in the evaluation of lymphoma patients is discussed.


Leucocyte migration fibrinolysis technique (LMFT): description of a method. Human leucocytes can migrate in a gel medium consisting of fibrin, 10% horse serum and tissue culture medium 199. The cells migrate within the fibrin gel mass. The 24 h. areas of migration depend upon the volume and concentration of the cell suspension applied on the fibrin gel. The amount of cells per mm2 migration culture area is less than half the amount used in the leucocyte migration agarose test. The results are reproducible, the standard variation of the migration areas is below 10%. The method is potentially useful for several purposes such as for measuring fibrinolytic and migratory activity of granulocytes, monocytes and lymphocytes and modification of these functions by lymphokines.


A review of the satatistical considerations involved in the treatment of isotope dilution calibration data. The use of linear regression analysis for the reduction of isotope dilution data is reviewed. The calculation of linear regression statistics is based upon four assumptions: zero variance in the independent variable, equal variance for all values of the dependent variable, linearity and continuity. Unfortunately, isotope dilution data often violate one or more of these assumptions, which results in the calculation of an inaccurate calibration line. The inaccuracies can be avoided through careful inspection of the data, including analyses of variance and linearity. Large differences in the variances of the dependent variable require the use of a weighted linear regression. Nonlinearity necessitates either discarding data in the nonlinear portion of the calibration or the calculation and use of atom % excess and dilution instead of the sample isotope ratios.


Fetal erythropoiesis following bone marrow transplantation. "Fetal" erythrocytes are present in older children and certain adults with hematologic disorders. To determine if regenerating bone marrow produces such cells, we examined the blood of seven allogeneic bone marrow transplant recipients. Six patients were engrafted with donor cells, while on e patients recovered autologous bone marrow after rejection of a marrow transplant. All seven patients had fetal hemoglobin levels of up to 10% by 100 days after transplant. In three patients, the Ggamma to Agamma ratio in the fetal hemoglobin was "newborn", while in one it was "adult". Gamma chain synthesis in blood and bone marrow never exceeded 20% of total non-alpha globin synthesis. The fetal hemoglobin was heterogeneously distributed in the cells. High titer i antigen also appeared. All fetal characteristics declined by 200 days. Erythropoiesis during bone marrow recovery appears to be associated with an accelerated, albeit partial, recapitulation of ontogeny.


[Trial prevention of mammary epithelioma metastases by a progestagen-estrogen combination]. The authors studied the effects of a combined oestrogenprogestogen treatment on the natural history of mammary tumours during a maximum period of seven and a half years. The group consisted of 135 patients divided into two on an age basis, sixty-six women still menstruating, and sixty-nine menopausal. A statistical study has shown that there is no significant difference between the treated and non-treated groups in the two groups of women studied. One can therefore deduce that the treatment has no effect on the evolution of mammary tumours whatever the age of the patient.


A postauricular flap to reconstruct facial defects. The postauricular skin may be transferred as a pedicled flap by making use of reverse flow in the superficial temporal artery when suitable delayed. The range of the flap is shown in figure 14.


The effect oa a new anti-inflammatory drug, flurbiprofen, on the respiratory, haemodynamic and metabolic responses to E. coli endotoxin shock in the cat. 1 The intravenous administration of E. coli endotoxin (2.0 mg/kg) in cats anaesthetized with sodium pentobarbitone resulted in immediate pulmonary hypertension and reductions in lung compliance and systemic arterial PO2. These effects were abolished, or greatly reduced, by the prior intravenous administration of flurbiprofen in doses (100 and 250 mug/kg and 1.0 mg/kg) which were devoid of cardiovascular or metabolic effects. Flurbiprofen is thus the most active antipyretic-analgesic drug so far examined in this experiment model. 2 Production of lactate, characteristic of the severe, secondary endotoxin shock phase, was delayed only by the highest dose of flubiprofen; hypotension, hypoglycaemia and the reduction in cardiac output which occurs during this phase, were unaffected. 3 These findings are discussed with reference to the treatment of the (shocked lung" syndrome of human septicaemia.


Centrally induced hypotension and bradycardia after administration of alpha-methylnoradrenaline into the area of the nucleus tractus solitarii of the rat. 1 In anaesthetized rats, bilateral injections of alpha-methylnoradrenaline, noradrenaline or adrenaline into the area of the nucleus tractus solitarii (NTS) of the brain stem caused dose-dependent decreases of systemic arterial blood pressure and heart rate. The effects of alpha-methylnoradrenaline were most pronounced and lasted longest. 2 The cardiovascular effects of alpha-methylnoradrenaline appeared to be restricted to the medio-caudal part of the NTS. 3 Prior administration of the alpha-adrenoceptor blocking agent, phentolamine, reversed the fall in blood pressure and heart rate induced by alpha-methylnoradrenaline into an increase. 4 Systemic administration of atropine combined with vagotomy potentiated the inhibitory effects of alpha-methylnoradrenaline on the cardiovascular system.


Pancreatic scanning.

Pancreatic scanning. The 75Se-selenomethionine pancreas scan cannot differentiate between chronic pancreatitis and carcinoma of the pancreas and may--although chiefly in certain defined clinical conditions--give a false-positive result. It is also rather more time-consuming than other radioisotopic scanning procedures. Its value in clinical practice stems from the fact that all the other methods currently available for studying the pancreas have their own disadvantages. Because a normal scan is only rarely seen in patients with carcinoma of the pancreas, pancreatic scanning can be particularly useful in helping to decide which patients should, and which should not, be subjected to other procedures involving greater discomfort or risk.


Lumbar myelography with metrizamide-a new non-ionic contrast medium. In contradistinction to all currently available water-soluble contrast media, metrizamide (Amipaque) is not a salt, but a substituted amide and therefore does not dissociate in solution. This unique property results in solutions of high iodine content yet with low osmolality. Metrizamide probably has a lower neurotoxicity than any other known water-soluble contrast agent. A clinical trial of metrizamide lumbar myelography in 201 patients in three clinical centres represents the first clinical assessment of this new contrast medium in the United Kingdom. The technique of the radiological procedure and the adverse reactions encountered are presented and discussed. There were no serious adverse effects: no muscle spasm or epilepsy. Minor adverse reactions--headache, vomiting and nausea occur with approximately the same frequency as with meglumine iocarmate: 43 percent of patients complained of headache. In 118 patients, the lower dorsal subarachnoid space was also examined with no increase in toxic reactions. It is concluded that metrizamide is a safe contrast medium for lumbar and lower dorsal myelography. Water-soluble media will probably largely replace oil products for these investigations. Further clinical trials are being extended to include examination of the upper dorsal and cervical subarachnoid space.


Hypertension after renal transplantation. The incidence of hypertension (mean diastolic pressure above 90 mm Hg) was evaluated in 85 patients with renal transplants whose follow-up ranged from 3 to 84 months. Bilateral nephrectomy had been performed in 80 recipients. The proportion of hypertensive subjects rose during the first three months, subsequently stabilised around 50-60% for up to five years, and then decreased slightly during the next two years. Over the years hypertension fluctuated so that one-third of the initially hypertensive patients became normotensive, and over one-third of the initially normotensive patients became hypertensive. The main single aetiological factor was renal failure. A significant relation between steroid dosage and blood pressure was found in only a quarter of the hypertensive patients, and in another quarter no cause could be found.


New technique for investigating bacterial flora of female periurethral area. The bacterial flora of the female periurethral area was studied by impression cultures taken by polystyrene sponges, the results being recorded by photographic contact printing. With this technique it is possible to observe the preferential colonisation of different areas by different organisms and to detect persistent colonisation by pathogens. In assessing the possible aetiological importance of these results, and especially in comparing them with the findings of other workers, it is essential to use correct anatomical nomenclature.


Mild hypertension: a clinical trial conducted in hospital general practice. To compare findings in a hospital trial of hypotensive drugs with those in a general practice trial several patients with mild hypertension were studied at the same time in hospital and in general practice. They received bendrofluazide and potassium chloride or bendrofluazide, potassium chloride, and reserpine according to a double-blind crossover protocol, and blood biochemical values were studied over eight weeks and six months. When reserpine was withdrawn from nine women they followed a modified protocol comparing bendrofluazide and potassium chloride with potassium chloride alone. The blood pressure values measured by the general practitioners were similar to those measured in hospital. Both the diuretic alone and the diuretic with reserpine produced significant falls in blood pressures. Although plasma renin activity increased on diuretic treatment, continued treatment did not produce a further increase, and levels gradually declined towards normal.


Significance of variations in blood: breath partition coefficient of alcohol. A helium-neon laser was used to measure the alcohol content of breath from six volunteers at regular intervals over up to four hours. The corresponding blood values were calculated with a blood : breath partition coefficient of 2100. When these values were compared with those obtained by direct measurement it was obvious that substantial variations occurred from one person to another in the derived values and that even in the same person the use of the partition coefficient of 2100 led to significant differences between the direct and derived values for blood, and these differences changed with time. Thus the assertion that a constant partition coefficient of 2100 exists between alcohol in blood and that in breath is not supported by the evidence. Accordingly the use of such a partition coefficient to derive blood alcohol values for law enforcement is not justified.


Changes in alkaline phosphatase activity in periosteal cells in healing fractures. The quantitative changes in alkaline phosphatase activity in the periosteal cells close to the fracture in rat metatarsal bones has been measured during the first 5 days postfracture. This study has been made possible by two technological advances, firstly the development of cryostat microtomy for cutting unfixed, undemineralised bone, and secondly the use of scanning and integrating microdensitometry for quantifying the activity in each periosteal cell. The results showed a loss of alkaline phosphatase activity close to the fracture site, with activity rising to normal values 0.8-1.0 mm from the site. No alkaline phosphatase activity was found in the cells which proliferated from the periosteum. It is suggested that reduced glutathione could cause such inhibition.


In vitro accumulation of mineral components by invertebrate cartilage. In vitro mineralization of the gill cartilage of Limulus (horse-shoe crab) has been reported previously (Eilberg et al. 1975). The present study demonstrates that cranial cartilage of Loligo (squid) and odontophore cartilage of Busycon (marine snail) also mineralize in vitro in hydroxyapatite-metastable media, but not in carbonate-metastable media. In all three of these cartilages, light phase-contrast microscopy revealed that the mineral phase occured in the form of spherical or ovoid granules ranging between 2 and 9 mum in diameter. During mineralization, the granules appeared successively in the perichondrium, in the matrix, and finally, within chondrocytes. Mineralization occurred more rapidly in Eusycon odontophore cartilage which has a significantly higher content of phosphatidyl serine than in Limulus gill or Loligo head cartilages. In all tissues the mineralization process is related to temperature, taking place most actively at 37 degrees C and only weakly at 50 degrees C.


Synthesis of guanosine 5'-triphosphate,3'-diphosphate in a spo T strain of Escherichia coli. A spo T stringent strain of Escherichia coli rapidly accumulates guanosine 5'-triphosphate,3'-diphosphate (pppGpp) immediately after the onset of isoleucine starvation. Subsequently, its level rapidly falls, as guanosine 5'-diphosphate,3'-diphosphate (ppGpp) continues to rise to the maximum value, which is abnormally high compared with that in the spo T+ strain. The ppGpp level in the spo T strain never reaches a steady state as it does in the spo T+ strain. Immediately after starvation, pppGpp and ppGpp are labeled with [3H]guanosine at a similar differential rate in both the spo T and spo T+ strains, suggesting that the two strains synthesize these nucleotides by the same pathway. However, by 15 min after starvation, the synthesis of these nucleotides is nearly halted in the spo T strain, and is greatly reduced in the spo T+ strain. Since ppGpp is labeled with [3H]guanosine more slowly than pppGpp in the starved spo T+ strain, ppGpp cannot be a precursor of pppGpp. The kinetics of the GTP level during starvation suggests that GTP is a precursor of pppGpp. The observed differences between the spo T and spo T+ strains can be explained by postulating, firstly, that ppGpp negatively controls the conversion of GTP to pppGpp, which is subsequently converted to ppGpp; secondly, that a catabolite of ppGpp negatively controls the conversion of pppGpp to ppGpp; and thirdly, that the spo T mutation primarily reduces the rate of ppGpp catabolism.


The G and Q banding pattern of Ellobius lutescens. A unique case of sex determination in mammals. The karyotype of the microtine Ellobius lutescens Th., was analysed using the G and Q banding techniques. One of the chromsomes of the first pair has the same bands in both sexes while the other member of the pair is unique to each sex. We propose that the sex determining mechanism of Ellobius lutescens is located in these chromosomes. The genetic burden imposed on this species due to this constitution is discussed.


The significance of the serotype in the clinical and pathological features of naturally occurring porcine salmonellosis. Salmonellosis was the diagnosis in 63 of 327 consecutive porcine necropsy accessions during a one year period in Indiana, an incidence four to ten times that previously reported in other areas. A herd outbreak was usually either a septicemic disease characterized by sudden mortality. Little overlap between these two syndromes were seen. Animals dying with the septicemic form usually yielded Salmonella choleraesuis var. kunzendorf on culture. Salmonella typhimurium was the usual isolate from cases of enteric salmonellosis. Oral inoculation of SPF pigs with S. typhimurium resulted in a disease similar to naturally occurring enteric salmonellosis. The pathology of porcine salmonellosis due to S. typhimurium is described.


Escherichia coli strains isolated from diarrheic piglets in the Province of Quebec. During 1972 to 1974, 112 Escherichia coli strains isolated from diarrheic piglets were recieved from different parts of the Province of Quebec, Canada. Fifty-six strains elicited a positive gut loop response in three week old piglets and were then considered as Moon's class 1 enteropathogens, while four of the 56 remaining strains reacted only in ten day old piglets and were classified as class 2 enteropathogens. Forty-eight strains produced both a heat-labile and a heat-stable enterotoxin, while 12 isolates which included the four class 2 enteropathogens produced only a heat-stable enterotoxin. Fifty-one enterotoxigenic strains could be serogrouped using OK antisera against E. coli strains commonly associated with colibacillosis in piglets. The most common serogroups encountered were O157: K "V17"; 88a,c, O149:K91; 88a.c. O157:K"V1c, O149:K91; 88a.c, O157:K"V17"; 88 a,c or a,b and O45:K"E5"; 88a,c. No significant difference was observed in the fermentation patterns, antibiotic susceptibility, colicin production, production of a filterable hemolysin and transferable tetracycline resistance between the enterotoxigenic and the nonenterotoxigenic strains.


Pathological and microbiological observations made on spontaneous cases of acute neonatal calf diarrhea. The purpose of this report is to describe clinical signs, gross and microscopic lesions, bacteriological and immunofluorescence observations made on spontaneous cases of acute neonatal calf diarrhea (NCD) in dairy and beef herds. The following diagnostic tools were used: 1) direct smears of intestinal content, 2) Escherichia coli counts, 3) aerobic bacterial cultures of the small intestine and other organs (The O serogroup and the enterotoxigenicity of the E. coli isolated was determined), 4) detection of the two Nebraska NCD viruses (reo-like and corona-like) by the fluorescent antibody technique and 5) histological examination on different segments of the digestive tract. The following etiological diagnoses were suggested after post mortem examination of 55 cases of NDC (34 were submitted alive): reo-like virus only (1), reo-like virus + E. coli (4), reo-like virus + cryptosporidium (2), reo- + corona-like viruses (5), reo- + corona-like viruses + cryptosporidium (3), reo- + corona-like viruses + infectious bovine rhinotracheitis virus (1), coronavirus-like agent only (2), coronavirus-like agent + mycotic abomasitis (1), coronavirus-like agent + crytosporidium (1), E. coli only (6), cryptosporidium only (5), mycotic abomasitis (3), mycotic rumenitis + reticulitis (1) and undetermined (20). Most of the calves in the last group were submitted dead.


The prevalence of enterotoxigenic Escherichia coli in the feces of calves with diarrhea. A study was undertaken to determine the prevalence of enterotoxigenicity among Escherichia coli isolated from calves with diarrhea and from a control group of normal calves. The test organisms consisted of 200 E. coli recovered from scouring calves less than two weeks of age and 100 E. coli from normal calves. The enterotoxigenicity of the cultures was evaluated by three methods, namely, injection of ligated segments of piglet intestine, injection of ligated segments of calf intestine and oral inoculation of suckling mice. Live cutures of all the test organisms were used for the ligated intestine studies whereas sterile broth culture supernatants were used in the suckling mouse tests. Of the isolates from scouring calves, 36% were enterotoxigenic in the piglet intestine and 28% in the calf intestine. Amongst the isolates from normal calves, none was enterotoxigenic in the piglet intestine and one was enterotoxigenic in the calf test system. The ligated piglet intestine was considered unsuitable for determining the enterotoxigenicity of bovine E. coli, whereas the ligated calf intestine test was satisfactory and correlated completely with the suckling mouse test. The enterotoxigenic E. coli of bovine origin produced an enterotoxin that resembled the heat stable enterotoxin of typical porcine enteropathogenic E. coli.


Characterization of enterotoxigenic bovine Escherichia coli. Among 300 isolates of bovine Escherichia coli, 56 which had been found enterotoxigenic in calf gut loops were characterized on the basis of O and K antigens, colonial morphology and resistance to seven antimicrobial drugs. The 56 isolates enterotoxigenic in the calf were compared with the nonenterotoxigenic ones. Of the 56 enterotoxigenic E. coli the majority possessed the A type of K antigen and had OK groups, O9:K(PS274) or O101:K(RVC118). Fourteen of these isolates had the K99 antigen. None of 27 isolates found enterotoxigenic in the piglet but not in the calf possessed the K99 antigen or belonged to OK groups O9:K(PS274) or O101:K(RVC118). Comparison of the patterns of resistance to seven antimicrobial drugs showed that all enterotoxigenic and nonenterotoxigenic isolates were susceptible to nitrofurantoin and sulphachlorphyridiazine and that there was no significant difference in the patterns between the two groups. The majority of enterotoxigenic isolates were mucoid, whereas most of the nonenterotoxigenic isolates were nonmucoid.


Erythrocyte rosettes--a marker for bovine T cells. Many species of erythrocytes were investigated for their ability to form spontaneous rosette with bovine peripheral blood leukocytes and fetal thymocytes. Only sheep and chicken red blood cells gave rosettes. Using conditions shown optimum for the demonstration of human rosette forming cells, only low numbers of bovine rosettes were demonstrable. By changing culture conditions to include 100% fetal calf serum, neuraminidase treated erythrocytes and/or lymphocytes and optimizing the incubation times and temperature, up to 38% of peripheral blood leukocytes and 52% of thymocytes formed rosettes. A thymic origin of rosetting cells was ascribed to T cells for the following reasons: 1) thymocytes gave higher numbers than did peripheral blood leukocytes, 2) rosette forming cell numbers were increased in peripheral blood leukocyte subpopulations enriched in T cells by nylon column separation and 3) only very few rosette forming cells had surface immunoglobulin, a marker of B lymphocytes. The reasons why all T cells were not detected by the technique were discussed.


The mouse inoculation test in rabies diagnosis: early diagnosis in mice during the incubation period. Brain tissue from 64 rabies suspect specimens were inoculated intracerebrally into twenty 9-12 gm adult Swiss white mice. Two mice from each specimen were killed on specific days postinoculation and examined for the presence of rabies virus by the fluorescent antibody staining technique. In this way a positive diagnosis was made in the majority of cases between postinoculation days 4 and 12 when the incubation period of these same specimens ranged between eight and 20 days.


A direct fluorescent antibody test for large spirochetes in swine dysentery using hyperimmunized swine serum. A direct fluorescent antibody test was developed for the identification of large spirochetes which are considered to be the cause of swine dysentery. Sera from swine which had recovered from swine dysentery and had been hyperimmunized by the intravenous and intraperitoneal injection of filtered spirochetes were used for conjugation with fluorescein isothiocyanate. A bright greenish fluorescence of large spirochetes was observed with the conjugated serum from hyperimmunized pig No. 1 when diluted 1:8 and hyperimmunized pig No. 2 when diluted 1:2. Pig No. 1 had developed a serum titer of 1:64 using the indirect fluorescent antibody test for large spirochetes. The conjugated serum from the three swine which had recovered from swine dysentery fluoresced spirochetes only when undiluted. The conjugated serum from the two swine treated while having a hemorrhagic diarrhea did not fluoresce spirochetes. No immunofluorescence of Vibrio spp. was observed.


Immunofluorescence of spirochetes with serum from swine recovered from swine dysentery using an indirect fluorescent antibody test. Using an indirect fluorescent antibody test, immunofluorescence of large spirochetes was observed with serum from swine that had recovered from swine dysentery. The spirochetes were obtained from scrapings of the colonic mucosa on the first day of diarrhea which was the time when the spirochete population was observed to be the highest. Of 29 exposed nonmedicated swine which developed and recovered from a diarrhea characteristic of swine dysentery 27 had antispirochete serum titers which ranged from 1:2 to 1:16. None of the 50 nonexposes swine developed a titer. Of 19 swine with a serum titer and reexposed with infective swine dysentery inoculum, 18 did not develop a diarrhea and were presumed to be immune. Considering these findings it is possible that this test could be used to detect antispirochete antibody in unknown swine serum.


Nicolaus Friedreich and degenerative atrophy of the posterior columns of the spinal cord. A short outline is given of the pioneer efforts of Nicolaus Friedreich in the description of the spinocerebellar degeneration which now bears his name.


Antibody-coated bacteria in the urine of preschool and school-aged girls with asymptomatic bacteriuria. Urine samples from 3564 girls aged 2 to 13 years were screened for evidence of infection. Cultures were positive (bacteria count, more than 10(5)/ml) in 61 (1.7%) by the dipslide method and in 55 (1.5%) by standard culture techniques. In 13 (23.6%) of the 55, antibody-coated bacteria (ACB) were detected in the urine. The clinical, bacteriologic, radiologic and urinalysis findings in children with ACB were no different from those in children in whom the bacteria were not coated. Direct examination of uncentrifuged urine under high power revealed one or more bacteria per two high-power fields in 96% of infected urine samples and in only 7% of noninfected samples. Five or more leukocytes per high-power field in centrifuged urine were detected in 36.7% of infected urine samples but not in noninfected samples. The ACB test did not differentiate between asymptomatic bacteriuria with parenchymal scarring or vesicoureteral reflux or both and asymptomatic bacteriuria without these abnormalities.


[Localization of the site of urinary infection in children by an investigation of antibody-covered bacteria]. The demonstration by immuno-fluorescence of antibodies on the surface of urinary bacteria, a new method of determining the site of a urinary tract infection, was found to be as valuable in children as it is in adults. A clear correlation exists between a positive test result and renal parenchymal infection on one hand, and a negative result and lower urinary tract infection on the other. Moreover, immunoglobulins were still detectable in original positive urine samples that had been standing at 4degrees C for 7 weeks. The constant finding of IgA on bacteria suggests a particular synthesis for this class of immunoglobulin. A pathophysiologic role for complement would appear to be excluded by the facts that the serum concentrations of C3 were normal and that C3 was invariably absent from the bacterial surface.


Canadian clinical trial of antilymphocyte globulin in human cadaver renal transplantation. A multicentre, randomized clinical trial of antilymphocyte globulin (ALG) was conducted among patients who had undergone cadaver kidney transplantation; follow-up was continued for a minimum of 1 year. Of the 179 patients 92 were given conventional treatment only, while 87 were given in addition ALG (from a standardized, highly immunosuppressive, common pool of equine ALG), 20 mg/kg-d intravenously for 10 days after transplantation. The ALG-treated group had better accumulated graft survival, fewer nephrectomies, better graft function, less than half the number of acute rejection episodes and less prednisone use. There was a beneficial drug (ALG)-related effect in both the graft and the host during the first 3 months after transplantation.


Controlled trial of azathioprine in the nephrotic syndrome secondary to idiopathic membranous glomerulonephritis. In a controlled double-blind trial five patients with the nephrotic syndrome due to idiopathic membranous glomerulonephritis received azathioprine, 2.5 mg/kg-d, while four others received placebo. After 1 year of treatment there was no significant difference between the two groups with regard to the changes in leukocyte count, values for hemoglobin, serum creatinine, blood urea nitrogen or serum albumin, 24-hour excretion of protein in the urine, or creatinine clearance. In this study azathiprine appeared not to be useful in the treatment of idiopathic membranous glomerulonephritis.


Morphologic and immunologic studies of a human colon tumor cell line (SW-48). A permanent human tissue culture cell line (SW-48) has been established from an adenocarcinoma of the transverse colon. Cells in the center of early colonies were cuboidal and loosely bound; cells on the periphery of the same colonies were more columnar, the nuclei were displaced toward the basal region, and the free surface formed stunted microvilli. These columnar cells ofter aligned themselves to resemble normal absorptive tissue. Carcinoembryonic antigen (CEA) was identified by immunofluorescent microscopy on the surface membrane of the tumor cells. Significantly more CEA could be isolated from the culture medium than from the whole cells. On agar gel diffusion analysis with a monospecific anti-CEA serum (G.P. 32), SW-48 CEA and CEA preparations from solid tumors demonstrated complete identity.


Preoperative cell-mediated immune status of gastric cancer patients. The status of cell-mediated immunity was studied in 360 patients with gastric cancer before surgery. For the skin test, tuberculin and DNCB were employed. For the in vitro test, the blastogenesis of peripheral blood lymphocytes to phytohemagglutinin (PHA), the ratio of rosette-forming T cells to sheep blood red cells, and the macrophage migration inhibiotry factor against autochthonous tumor antigens were measured. As a result, it was found that in progressive gastric cancers the cell-mediated immunity decreased specifically or nonspecifically, especially the DNCB reaction; and the blastogenesis against PHA showed an inverse correlation to the advance of gastric cancer.


Systemic giant-cell histiocytosis: report of a case and a review of the adult form of Letterer-Siwe disease. The multiple classifications of histiocytic disorders, and the lack of specificity in their application, have degraded the utility of such diagnostic terms as "Letterer-Siwe Disease" and "Malignant Histiocytosis". A case is presented of a systemic histiocytic proliferation with erythrophagocytosis and giant-cell formation which does not fit into any presently accepted diagnostic niche. No similar case was found in the literature. Moreover, it appears that the entity "Adult form of Letterer-Siwe disease" has been uncritically accepted on the basis of a few poorly documented cases, and should be abandoned.


Acute renal insufficiency due to lymphomatous infiltration of the kidneys: report of six cases. Described are six patients with acute renal insufficiency due to histologically proven massive lymphomatous infiltration of the kidneys. All patients were admitted to a renal intensive care unit over a period of six years. Oliguria was the presenting symptom in two of the patients. This complication of lymphoma is suggested by enlargement of the kidneys and mild proteinuria in the absence of other causes of uremia and can be demonstrated by renal biopsy. Local radiation therapy performed in two patients produced improvement of renal function.


Evaluation of the mutagenicity of compounds of known carcinogenicity, belonging to the benz[a]anthracene, chrysene, and cyclopenta[a]phenanthrene series, using Ames's test. Fifty-four polycyclic compounds, 29 of the cyclopenta[a]phenanthrene series, 11 chrysenes, and 14 benz[a]anthracenes, have been tested for mutagenicity by Ames's method, using Salmonella typhimurium TA100. Without exception all 37 carcinogens and a known initiator were mutagens. Of the 16 noncarcinogens 7 were mutagenic, but none of these has yet been tested for initiating, as opposed to carcinogenic, activity. There appeared to be little quantitative correspondence between carcinogenic and mutagenic potency, however, and possible reasons for this are discussed. The aryl hydrocarbon hydroxylase inhibitor 7,8-benzoflavone strongly inhibited the mutagenicity of certain compounds when it was added to the incubations.


Detection of bacterial endotoxin in antitumor agents. The Limulus amebocyte lysate assay was used to detect the presence of bacterial endotoxin in a variety of chemotherapeutic agents currently in use. Results were quantitated using a standard Escherichia coli endotoxin curve. Relative concentrations of endotoxin ranging from 0.1 to 63 ng/ml were detected in individual lots of L-asparaginase, 5-azacytidine, bleomycin, DTIC, actinomycin D, adriamycin, and vinblastine diluted to normal concentrations for injection. When multiple lots of a given drug were tested, at least one lot produced a negative assay, indicating less than 0.1 ng/ml of endotoxin in 50% of these drugs. All lots of cytosine arabinoside, cyclophosphamide, daunorubicin, vincristine, and streptozotocin tested had less than 0.1 ng/ml of endotoxin. Inactivation kinetics of the Limulus-positive drug material in dilute alkali solutions were identical to those of E. coli endotoxin. The Limulus lysate test is a rapid and specific method for the detection of small amounts of bacterial endotoxin contaminating parenteral preparations of antitumor agents.


Calcium and ionophore A-23187 as initiators of DNA replication in the pluripotent haemopoietic stem cell. The proliferation rate of the haemopoietic stem cells of the mouse bone marrow can be stimulated by an increase in calcium influx. This key event may be brought about by raising the external calcium concentration or, at lower calcium levels, by use of A-23187 as calcium carrier, within definite dose ranges depending on the Mg2+:Ca2+ molar ratio in the incubation medium. The hypothesis is postulated of an accelerated influx of Ca+ as the possible common mediator of the effects of the various types of agents known to control the CFU cell cycle.


Light and electron microscopic study on the endocrine cells of the pancreas in a marine teleost, Fugu rubripes rubripes. The pancreatic endocrine tissue of Fugu rubripes rubripes consists of numerous round principal islets (Brockmann bodies) of various sizes scattered around the gall-bladder. The endocrine cells are divided into A-, B-, D-, and Ff-cells. Each cell type was identified by comparing thick and thin sections in both light and electron microscopy. Aldehyde-fuchsin positive B-cells contain numerous round secretory granules (average diameter 300 nm) each of which has a round compact core of moderate density; a narrow space exists between this core and the limiting membrane. Grimelius' silver positive A cells contain round secretory granules (average diameter 360 nm) with a hexagonal or tetragonal crystalline core (average diameter 170 nm) of high density; the silver grains perferentially appear in the space between the limiting membrane and the core. The crystalline core of each alpha-granule often contains an appendix like structure of variable shape. D cells blackened by the silver impregnation method of Hellman and Hellerström (1960) have round secretory granules (average diameter 320 nm) filled with a flocculent material of low density. The fourth cell type (Ff-cell) has a clear cytoplasm after differential staining for light microscopy. By electron microscopy, this cell has elongated fusiform secretory granules (520 nm average length x 230 nm average width) filled with numerous filaments arranged in parallel with the longitudinal axis. Figures suggesting granule formation in the sacs of the Golgi apparatus were obtained in all of islet cell types. Equivalents of emiocytotic release of secretory granules were encountered in the A and Ff cells.


The thyrotropic cells of the guinea pig pituitary. Electron microscopic study after characterization by immunocytochemical means. Three different immunocytoenzymatic techniques were used to identify and characterize the thyrotropic cells in the pituitary of normal guinea pigs at the ultrastructural level (superimposition technique, immunocytochemical technique using P.A.P. and indirect immunohistoenzymatic method before embedding). These cells are characterized  by a dark cytoplasm with granules ranging from 1500 to 2000 A in diameter. The appearance of these granules is very variable: some display a marked electron density and are homogeneous but some have a less marked electron density with a more electron dense peripherally situated region. The TSH molecules are essentially confined to the granules but when the immunocytochemical reactions are carried out before embedding, positive staining is also seen in the cytoplasm and the outer surface of most of the rough endoplasmic reticulum membranes. These results are discussed.


Ultrastructural localization of prolactin, growth hormone and luteinizing hormone by immunocytochemical techniques in the bovine pituitary. The technique of ultrastructural immunocytochemistry involving the unlabeled antibody and the soluble peroxidase-antiperoxidase complex was used to identify and describe the prolactin (P) cells, somatotropic (STH) cells and luteinizing hormone (LH) cells in the bovine anterior pituitary gland. This method was used to localize the three hormones at the electron microscopic level. Staining of varying intensity was found on the secretory granules and on the small granules and vesicles within the Golgi complex. No stain was found in nuclei, on mitochondria or in the endoplasmic reticulum.


Immunocytochemical study of the hypothalamo-neurohypophysial system. I. Localization of neurosecretory neurons containing neurophysin-I and neurophysin-II in the domestic pig. Antibodies raised against porcine neurophysin-I and porcine neurophysin-II using an injection regime in rabbits over a short time period, were used to localize neurophysin-I and neurophysin-II in hypothalamic neurosecretory elements of the domestic pig. In transverse section, neurophysin-II containing cells were more abundant in the dorsal medial region of the rostral supraoptic nucleus (SON) as compared with the distribution of neurophysin-I neurons. The main bulk of the cells of the SON were heavily stained for neurophysin-I with neurophysin-II containing cells positioned dorsal from the edge of the optic chiasma. Neurosecretory cells of the SON as seen in sagittal section also showed a differential staining for neurophysins-I and -II. Rostral regions of the pig paraventricular nucleus (PVN) contained magnocellular elements near the third ventricle which were stained predominantly for neurophysin-II. In regions corresponding to the caudal PVN there appeared two populations of neurosecretory neurons: (a) an area of cells adjacent to the third ventricle which contained neurophysin-II antigen and (b) a group of densely populated cells in the dorsal-lateral region which was stained for neurophysin-I. The results support the existence in the pig of at least two distinct populations of neurosecretory neurons corresponding to the neurophysin-I and neurophysin-II neurosecretory system.


Identification of gastrin-secreting cells and cholecystokinin-secreting cells in the gastrointestinal tract of the human fetus and adult man. Gastrin- and cholecystokinin (C.C.K.)-containing cells were detected by using anti-gastrin and anti-C.C.K. sera in the gastrointestinal tract of human fetuses and premature infants and in the stomach and duodenum of adult man obtained by biopsy from eight patients with normal gastro-duodenal endoscopy. The specificity of immunocytological reactions was ascertained by studying the inhibition of the reaction by gastrin, C.C.K., secretin, somatostatin, glucagon, insulin, serotonin, histamin, caerulein and octapeptide of C.C.K. In adult man, the gastrin cells are located only in the antrum and juxtapyloric region; C.C.K. was detected in the duodenum. In the human fetus, the first gastrin cells are seen in the antrum at 14 weeks of age and in the duodenum as early as 10 weeks; the C.C.K. cells are seen in the small intestine at 10 weeks of age.


Evaluation of a DNA polymerase-deficient mutant of E. coli for the rapid detection of carcinogens. Differential growth inhibition of two E. coli cultures was evaluated as a rapid screening technique for chemical carcinogens. Of the carcinogens tested, only "direct acting" carcinogens produced positive results. Furthermore, this test is not a quantitative assay in that neither was a dose--response relationship seen nor did potent carcinogens necessarily show a greater response than weaker carcinogens.


Radioimmunoassay and enzyme immunoassay compared for determination of digoxin. Patients' sera were analyzed for digoxin by using two different radioimmunoassays and an enzyme immunoassay. Quantitative results obtained by enzyme immunoassay (I) were compared to results obtained on aliquots of the same sample by the radioimmunoassays (II and III). The correlation coefficients were: I vs. II 0.90, n=108; I vs. III 0.94, n=102; and II vs. III 0.95, n=158. Day-to-day precision (10 days) on a low control (1.3 mug/liter) and a high control 3.0 mg/liter), expressed as coefficients of variation, were: I, 13% and 7.8%, II, 4.0% and 4.7%; and III, 8.9% and 4.2%. Ten digoxin-supplemented samples (0-8 mug/liter) were analyzed by the three methods. Correlation coefficients were: supplemented sample vs. I, O.99; supplemented sample vs. II, 0.97; supplemented sample vs. III, 0.98.


Polarisation fluoroimmunoassay of gentamicin. A polarisation fluoroimmunoassay has been developed for the routine determination of gentamicin levels in serum. The method employs fluorescein-labelled gentamicin which is easily, reproducibly and economically prepared and has excellent shelf-life. The assay is fast (minimum incubation time 2 min) and requires 1.25 mul of serum. There is no separation step prior to readout of results. Recovery experiments reveal no serum effects. Intra- and inter-assay coefficients of variation are of the order of 10% or less. Estimations of gentamicin levels in patient serum samples correlate closely with those obtained by bioassay (r = 0.93) and radioimmunoassy (r = 0.97).