Incidence and outcome of posttraumatic respiratory failure. From 1972 through 1975, a total of 6,196 patients were admitted to San Francisco General Hospital Trauma Service, of whom 908 required admission to an intensive care unit and 390 required mechanical ventilation. Duration of mechanical ventilation was one week or less for 76%, two weeks or less for 90%, and four weeks or less for 97%. Survival in patients ventilated seven days or less was 64%, in patients ventilated 8 to 14 days it was 55%, and in patients ventilated 15 to 30 days it was 55%. The mortality for ventilated patients aged 20 to 49 years was 23% while for ventilated patients older than age 60 it was 53%. The average age of ventilated patients was 43. Use of controlled-pressure soft-cuff endotracheal tubes has eliminated tracheal-esophageal fistula and tracheal stenosis as causes of morbidity and mortality. Appropriate ventilator alarms have minimized fatalities due to mechanical equipment failure. Complications related to positive pressure ventilation, such as pneumothorax and subcutaneous emphysema, still occur in 12% to 18% of patients. In our patients who survived the acute respiratory distress syndrome, recovery of lung function was universal, and permanent disability was less than 1%.


Effects of interferon preparations on rabbit corneal xenograft. High doses of rabbit interferon (2 X 10(6) and 2 X 10(5) units/ml, a drop of about 0.1 ml administered topically into the eye twice daily) suppressed the rejection of rabbit corneal xenograft, whereas a low dose (2 X 10(3) units/ml) enhanced it, However, neither heat-inactivated rabbit interferon nor mouse L cell interferon showed any effects on rabbit corneal grafts.


Supercoiled DNA of nuclear polyhedrosis virus of Galleria mellonella L. Supercoiled infectious DNA was isolated from nuclear polyhedrosis virus infecting great wax moth Galleria mellonella L.) Covalently closed DNA molecules constitute approximately 10--30 per cent in our preparations. These molecules dissappear during storage. Electron micrographs of supercoiled and open circular molecules are presented. Length of the open rings is about 50--52 micron. Infectivity of different DNA forms is discussed.


Timolol. A beta-adrenergic blocking agent for the treatment of glaucoma. Timolol maleate, a beta-adrenergic blocking agent, was studied in 30 patients with glaucoma. Significant intraocular pressure lowering was achieved with 0.5% and 1.5% solution of the drug. At seven hours following treatment, the IOP was lowered 50% from the pretreatment pressure with both strengths of timolol. There were no subjective or objective ocular or systemic side effects detected. Visual acuity and pupil response were unchanged from pretreatment levels throughout the study. beta-Adrenergic blocking agents and specifically timolol may be an important breakthrough for the medical management of glaucoma.


Timolol, dose response and duration of action. Timolol maleate, a beta-adrenergic blocking agent, was used in a study of 20 adult patients with chronic open-angle glaucoma. This single-dose study demonstrated a dose response for this drug using concentrations of 0.1%, 0.25%, 0.5%, and 1.0%. Timolol, 0.5%, appears to give the maximal ocular hypotensive effect; however, each concentration of the drug produced an ocular hypotensive effect for at least 24 hours. No local or systemic side effects were discovered throughout the study. Timolol may be an effective, innocuous, once-a-day, topical agent for the treatment of glaucoma.


Penetrating keratoplasty in herpes simplex keratitis. Recurrence in grafts. A review of 231 penetrating keratoplasties performed for corneal disease due to herpes simplex keratitis shows that clear grafts may be obtained in 75% and satisfactory visual results in about 70% during a long-term follow-up. Over a three-year follow-up period, recurrence of the herpetic disease in the graft was observed in 12%. Over a longer period of follow-up, up to 15 years, the rate of recurrence increased to 47%. About half of the corneas with recurrence achieved clear grafts with treatment. Recurrence did not appear to be less frequent in those cases which circumscribed scars as compared with those in which there was diffuse scarring. The state of activity of the disease preoperatively could not be demonstrated to influence greatly the results of the keratoplasty.


Blepharoptosis correction with the sutureless Fasanella-Servat operation. The Fasanella-Servat operation for correction of blepharoptosis in cases where the function of the levator muscle is good can be performed effectively without the use of sutures, thus eliminating the possible complication of corneal abrasion in the postoperative period. The author has performed 12 sutureless operations with satisfactory correction in 11 cases.


Rhomboid flap.

Rhomboid flap. The rhomboid flap is a practical and efficient method of repairing small- to moderate-size surface defects of the head and neck. This flap has the advantage of combining the lateral shift and rotational methods of transfer that aid in the closure of the donor site as the flap is transposed. A description of this flap, the principle of its design, its clinical application, and potential pitfalls are presented.


Pyrimidine biosynthesis in Serratia marcescens: polypeptide interactions of three nonsequential enzymes. Orotidine-5'-monophosphate pyrophosphorylase (OMPppase, E.C. 2.4.2.10) and orotidylate decarboxylase (OMPdecase, E.C. 4.1.1.23) were purified from Serratia marcescens HY. These enzymes required physical association for maximal catalytic activities and formed a fragile complex with dihydroorotase (DHOase, E.C. 3.5.2.3). OMPppase reversibly lost 50% of its activity upon separation from DHOase. The kinetic characteristics of OMPppase were modified by this separation. In the presence of DHOase, the Kms for PRPP and orotate were stoichiometric: 2.3 X 10(-6) M and 2.6 X 10(-6) M, respectively. Following separation, the Kms were significantly different: 1.3 X 10(-6) M for PRPP and 4.1 X 10(-6) M for orotate. OMPppase and OMPdecase could be reversibly separated by acrylamide gel electrophoresis, but the separation was accompanied by a loss of catalytic efficiency for both enzymes. DHOase readily associated into multiple molecular forms and could not be purified. The DHOase-OMPppase-OMPdecase interactions demonstrate that a weakly aggregated, multifunctional enzyme complex participates in the biosynthesis of pyrimidine nucleotides in S. marcescens. This unique association of non-sequential biosynthetic enzymes may represent a larger complex which provides a channeling or regulatory unit.


The catalytic activity of monomeric yeast hexokinase A. It has been suggested [Williams, D.C. & Jones, J.G. (1976) Biochem. J. 155, 661-667] that monomeric hexokinase isoenzyme A is not catalytically active. We here present data from reacting-enzyme sedimentation, dissociation experiments and from previous studies which are consistent with the monomeric form possessing catalytic activity.


Inhibition of the reconstitution of the haemolytic activity of the first component of human complement by a pepsin-derived fragment of subcomponent C1q. 1. A fragment of subcomponent C1q, which contained all the collagen-like features present in the intact molecule, was isolated by pepsin digestion as described by Reid [Biochem. J. (1976) 155, 5-17]. 2. The pepsin-derived fragment of subcomponent C1q did not bind to antibody-coated erythrocytes under conditions where complete binding of sub-component C1q took place. 3. The peptic fragment blocked the reconstitution of C1 haemolytic activity by competing with intact subcomponent C1q in the utilization of a mixture of the other two subcomponents, C1r and C1s. 4. Reduction and alkylation of the interchain disulphide bonds in the pepsin fragment did not markedly affect its inhibitory effect, whereas heating at 56 degrees C for 30min completely abolished the effect. 5. Lathyritic rat skin collagen and CNBr-derived peptides of pig type II collagen showed no ability to mimic the inhibitory effect of the pepsin fragment when tested over the same concentration range as used for the peptic fragment. 6. The peptic fragment was unable to block efficiently the reconstitution of C1 haemolytic activity unless it was added to the mixture of subcomponents C1r and C1s before the attempt to reconstitute C1 haemolytic activity, in solution, or on the surface of antibody-coated erythrocytes. 7. Evidence was obtained that suggested that subcomponent C1q bound the subcomponent C1r-C1s complex more efficiently when the subcomponent C1q was bound to antibody than when it was free in solution.


Biochemical and physiochemical characterization of pepsin-solubilized type-II collagen from bovine articular cartilage. Solubilization of collagen from bovine articular with pepsin requires the preliminary extraction of proteoglycans from the ground substance. Biochemical and physiochemical properties of this pepsin-solubilized collagen are independent of the pretreatment (extraction with 1.5M-CaCl2, 5M-guanidinium chloride or 0.2M-NaOH) and of the age range (2-4-year-old and 2-month-old animals). Characterization of the de-natured components, of the CNBr peptides and of the amino acid and cross-link composition shows that the collagen of the hyaline cartilage is all type II. Electrical birefringence measurements showed the presence of tropocollagen molecules (length 280nm) and molecules whose length is slightly less than twice that of the tropocollagen molecules. This latter molecule may be a dimer composed of two monomers linked by intermolecular head-to-tail bonds and whose theoretical length (530nm), according to the quarter-stagger theory, is in good agreement with our measured values (510-530nm). We have verified that the beta-components of this collagen are formed of two alpha-chains linked by the stable intermolecular bond, dehydrodihydroxylysinonorleucine. These dimeric molecules are absent from solutions of skin collagen whose beta-components possess only aldol-type intramolecular cross-links. Although reconstituted fibres from solutions of skin and cartilage collagen are similar, the segment-long spacing crystallites formed with pepsin-solubilized cartilage collagen present a symmetrical and dimeric form corresponding to the lateral aggregation of two monomers with an overlap (90nm) of the C-terminal ends.


L-serine dehydratase from Arthrobacter globiformis. 1. L-Serine dehydratase (EC 4.2.1.13) was purified 970-fold from glycine-grown Arthrobacter globiformis to a final specific activity of 660micronmol of pyruvate formed/min per mg of protein. 2. The enzyme is specific for L-serine; D-serine, L-threonine and L-cysteine are not attacked. 3. The time-course of pyruvate formation by the purified enzyme, in common with enzyme in crude extracts and throughout the purification, is non-linear. The reaction rate increases progressively for several minutes before becoming constant. The enzyme is activated by preincubation with L-serine and a linear time-course is then obtained. 4. The substrate-saturation curve for L-serine is sigmoid. The value of [S]0.5 varies with protein concentration, from 6.5mM at 23microng/ml to 20mM at 0.23microng/ml. The Hill coefficient remains constant at 2.9.5 The enzyme shows a non-specific requirement for a univalent or bivalent cation. Half-maximal activity is produced by 1.0mM-MgCl2 or by 22.5mM-KCl. 6. L-Cysteine and D-serine act as competitive inhibitors of L-serine dehydratase, with Ki values of 1.2 and 4.9mM respectively. L-Cysteine, at higher concentrations, also causes a slowly developing irreversible inhibition of the enzyme. 7. Inhibition by HgCl2 (5micronM)can be partially reversed in its initial phase by 1mM-L-cysteine, but after 10 min it becomes irreversible. 8. In contrast with the situation in all cell-free preparations, toluene-treated cells of A. globiformis form pyruvate from L-serine at a constant rate from the initiation of the reaction, show a hyperbolic substrate-saturation curve with an apparent Km of 7mM and do not require a cation for activity.


Alloxan cytotoxicity in vitro. Microscope photometric analyses of Trypan Blue uptake by pancreatic islet cells in suspension. Suspensions of islet cells were prepared by shaking pancreatic islets from non-inbred ob/ob mice in a Ca2+-free buffer. The cells were incubated with or without 20 mM-alloxan, and subsequently with Trypan Blue. The uptake of Trypan Blue by cell nuclei was analysed by microscope photometry and by counting the frequency of cells appearing stained on visual inspection. Cells classified as stained or unstained by inspection showed no overlap in nuclear absorbance. Suspensions not exposed to alloxan contained 70-80% of unstained cells. Alloxan markedly decreased the frequency of unstained cells, an effect counteracted by 5 or 20 mM-D-glucose. The spectrum of Trypan Blue in islet-cell nuclei was red-shifted by about 20 nm. A similar red-shift was observed on adding the dye to solutions of albumin or histones, but not on mixing the dye with DNA. Binding to basic proteins may explain the concentrative uptake of Trypan Blue in dead cells and contribute to the oncogenic transformation of phagocytotically active cells. Beta-Cells in vitro are killed by alloxan and hence represent a valid model for studying the diabetogenic action of the drug.


Pathways of complement activation in chronic discoid lupus: serologic and immunofluorescence studies. To assess the relative roles of the classic and alternative pathways of complement activation in chronic discoid lupus erythematosus (CDLE), serum levels of properdin, C3, and C4, and deposition of these proteins in the dermal-epidermal junction (DEJ) of 20 CDLE patients were compared to the findings in patients with clinically active and inactive SLE. Properdin was demonstrated in the DEJ of 10 of 14 (71%) histologically typical skin lesions from patients with CDLE, usually in association with deposits of immunoglobulin, C3, and C4. Properdin levels in CDLE patients were significantly increased (137 +/- 34%) (P less than 0.05) when compared to normal controls (101 +/- 18%) or to patients with clinically active SLE (89 +/- 32%). C3, C4, DNA-binding, and antinuclear antibody tests in CDLE were indistinguishable from those in normals, but significantly different from patients with active SLE (P less than 0.05). The complement profiles of patients with clinically inactive SLE resembled those of CDLE patients more closely than those of active SLE patients.


Skin basement membrane immunofluorescence in rheumatoid arthritis: lack of diagnostic correlation. Thirty-nine patients with rheumatoid arthritis were studied for the presence of skin basement membrane immunofluorescence. Punch biopsies from normal sun-exposed skin of the forearm were negative for basement membrane immunofluorescence in all cases, except one which was read as questionable. No correlation with serum antinuclear antibody or lupus erythematosus cells was observed. Skin immunofluorescence studies are helpful in differential diagnosis when patients with a clinical picture of rheumatoid arthritis present with serum antinuclear antibodies and lupus erythematosus cells. Positive basement membrane immunofluorescence is strong evidence of systemic lupus erythematosus.


Antiglobulins in juvenile rheumatoid arthritis. With initial hopes of establishing a diagnostic test for juvenile rheumatoid arthritis (JRA), five types of immunoadsorbents were used to search for occult antiglobulins in sera from children. The immunoadsorbents included glutaraldehyde-aggregated globulin, heat- and glutaraldehyde-aggregated globulin, Affi-Gel-bound globulin, Sepharose-bound globulin, and bromozcetyl cellulose-bound globulin. The glutaraldehyde-aggregated immunoadsorbents retained nonspecific protein. The Affi-Gel, Sepharose, and bromoacetyl cellulose absorbents were effective and specific, but the Affi-Gel and bromoacetyl cellulose were impractical for routine use. Separose-bound globulin was selected for use in a survey of 52 patients with JRA and 8 normal children. Materials resembling occult antiglobulins, measured as acid elutable protein, were found in normal children as well as in children with JRA. Only a small proportion of children with JRA had statistically elevated levels of such occult anti-globulin consisted of IgM, IgG, and IgA, and had latex fixation activity in 2 of 5 tested sera from normal children and in 27 of 52 sera from children with JRA. Thus the presence of antiglobulins per se is not diagnostically helpful in distinguishing children with JRA.


Single-blind evaluation of hypnotic activity of chlordesmethyldiazepam in No-placebo-reactor medical patients. 69 patients hospitalized for chronic respiratory diseases and complaining of moderate to severe insomnia were treated with placebo for 2 days, 36 subjects (52.17%) were discarded for their placebo responsiveness. To the remaining patients who suffered from severe insomnia, 7-chloro-5-(2-chlorophenyl)-1,3-dihydro-2H-1,4-benzodiazepin-2-one (chlordesmethyldiazepam, 2 mg orally) was administered for 7 consecutive evenings. The onset, duration, quality of sleep and the state on awakening were scored on the basis of patient subjective measurements. High scores of the four considered parameters were obtained during chlordesmethyldiazepam administration. Three treated patients complained of diurnal drowsiness. Results are discussed with regard to methodological problems in the evaluation of hypnotics.


[Studies on cardio-circulatory effects of carpipramine. Results of a double-blind study (author's transl)]. For eight weeks 30 psychotic patients with a healthy cardiovascular system received 3 X 50 mg 1-[3-(10,11-dihydro=5H-dibenz[b,f]-azepin-5-yl]-propyl]-4-piperidiono-piperdine-4-carboxamide dihydrochloride-monohydrate (carpipramine) per day with a placebo phase of three weeks either previous to the medication of succeeding it. In addition there was a group of similar patients without any medication. From this study it was concluded that carpipramide has no depressive effect on the cardiovascular system. The differences in blood pressure and heart rate after the change from medication to placebo indicate a certain vegetative lability in this phase. In the ECG, no irregularities of the atrioventricular and intraventricular conduction and its repolarisation in form of a lengthening were observed; only the WPW-syndrome noticed with one patient might be related to the medication since it occurred at the end of the three weeks during which carpipramine had been administered.


[Regulation of DNA replication (author's transl)]. Two aspects of the regulatory mechanisms of DNA replication are discussed: 1. When resting mammalian cells are stimulated to proliferate a genetic program is expressed which provides proteins and other replication factors; 2. the interaction of proteins with specific sections on the DNA leads to initiation; most probably, structural changes in chromatin are also involved. Once initiated, replication proceeds almost automatically through the elongation phase. Lymphocytes stimulated to proliferate by concanavalin A are used as an example to demonstrate the relative importance of the two aspects of DNA replication.


[IS-elements and transposons: new linkage of genes within bacteria (author's transl)]. IS-Elements are unique DNA sequences occurring in the DNA of E. coli and its chromosomes. They can be transposed within or between chromosomes in the same cell. Integration into a gene causes mutations and polar effects on adjacent genes. IS-Elements are integration sites for plasmids into chromosomes. IS-Elements also serve as parts of transposons (DNA structures capable to be transposed from one chromosome to another and carrying a gene for the resistance to an antibiotic or heavy metals).


[Genetic surgery: results of a new technique (author's transl)]. New techniques for cleaving DNA molecules at precisely defined points, and for successful DNA-transformation of E. coli have allowed genetic engineers to ligate DNA fragments of all possible origins and to convert them into replicating molecules sustained within bacterial cells. For this purpose, typically, segments of eukaryotic DNA are linked to bacterial "vectors", in particular plasmids carring their own replication machinery. This procedure allows to study the genetic organization of eukaryotic DNA and to detect whether or not the eukaryotic genes will function in a foreign environment. Work has just been initiated aiming at the complementary procedure, i.e., at introducing prokaryotic or foreign eukaryotic genes into eukaryotic cells which may carry hereditary defects.


[Gene expression in eucaryotic cells and its regulation (author's transl)]. Principles of 1. the organisation and compartmentalisation of the eukaryotic nuclear genome and 2. of the different processes involved in controlling its activity are outlined and the basic differences to prokaryotic cell systems are emphasized. The special composition and arrangement of the nuclear DNA of eukaryotes is demonstrated in terms of both redundancy classes (unique sequences, sequences of intermediate degrees of repetition, highly repetitive sequences, i.e., "simple sequences") and chromatin and chromosome organisation. The role of the nuclear envelope as an ubiquitous and characteristic structure involved in the compartmentalisation of the nuclear genome and its transcriptional machinery is illustrated. The diversity of the mechanisms of the controls of gene expression in the eukaryotic cells is discussed at different levels: a) chromatin elimination, b) polyploidisation, c) polytenisation, d) gene amplification, e) gene magnification, f) inactivation of genes by complexing with specific proteins and/or protamines, g) transcription, h) complex formation of transcriptional products with specific proteins, i) release of the transcriptional products from the template containing strands, j) processing of newly formed RNA's k) intranuclear degradation of newly formed RNA's, l) nucleocytoplasmic translocation, and m) various forms of RNA containing structures, including masked messenger RNA's and ribosome storages, in the cytoplasm. It is demonstrated that differences in transcriptional activity can be directly visualized and that a direct trnascriptional control of gene activity is indicated to exist at least in some specific cell systems.


[On the bronchodilatory effect of reproterol (author's transl)]. The results of an intra-individual double-blind study between placebo, orciprenaline and 7-(3-[2-(3,5-dihydroxyphenyl)-2-hydroxy-ethylamino]-propyl)-theophylline (reproterol, Bronchospasmin) were compared in 16 patients with chronic obstructive bronchitis by means of the findings of objective measurements as well as patients' comments. It was shown that the new monosubstance reproterol, a phenylethyl-aminoalkyl-xanthine, is a very potent bronchodilator whose effect persists for at least 6 h. This bronchlysis could be proven on the basis of total and specific airways resistance as having a significantly superior and longer-lasting effect when compared with placebo. Also, when compared with orciprenaline the total broncholytic effect (areas of Raw and SRaw for effect covering time) was significantly better. No essential cardiovascular or subjective side-effects were caused by any of the three preparations inhaled in from of an aerosol equipped with metering valve. Because of its assured broncholytic efficacy and good tolerance, as well as its different forms of application (ampoules, tablets, aerosol, inhalation-solution), reproterol is a recommendable alternative in recent beta2-adrenergic achievements.


Scanning electron microscopy: morphology of aortic endothelium following injury by endotoxin and during subsequent repair. A single injection of endotoxin P45 Poly Serratia marcescens was used to induce endothelial injury in rabbits. The aortic endothelium was examined by Scanning Electron Microscopy (SEM), at various times after administration of endotoxin, using the technique of silver staining and pressure fixation. Within one hour after injection, some endothelial cells were curled-up and spindle-shaped in appearance. Areas of aorta devoid of endothelial cover were occasionally observed and platelets were sometimes found adhering to these sites. Two and four weeks after initial injury no spindle-shaped cells were found. Instead, some endothelial cells were heavily stained with silver. Small denuded zones were still found and these were surrounded by brightly silver-stained cells. This study confirms that endotoxin rapidly causes endothelial injury and suggests that regenerating endothelial cells which were formed following injury are avidly stained by silver salts and appear as bright cells by SEM.


Postextrasystolic T wave changes and angiographic coronary disease. The significance of postextrasystolic T wave changes in beats following induced extrasystoles was assessed by angiography in 55 patients. These T wave changes were found in 81 per cent of coronary artery disease patients but also in 68 per cent of patients with normal coronary arteries (PNS). All patients with normal baseline electrocardiograms and normal coronary arteries showed postextrasystolic T wave changes. In electrocardiographic leads corresponding to the distribution of major coronary arteries, T wave changes occurred just as frequently when the artery was normal (54%) as when the artery was stenosed (55%). Left ventricular asynergy was not associated with an increased frequency of postextrasystolic T wave changes and in fact ejection fraction was greater end-diastolic pressure lower in patients with T wave changes. Thus, postextrasystolic T wave changes appear not to be useful in diagnosing or localising coronary artery disease.


Fibreoptic studies of the aortic valve in dogs. The study describes a method for visualisation of the aortic valve in the intact animal using a non-obstructive endoscopic system. Clearing of blood from the field of vision is achieved by the rapid injection of saline during a period of asystole.


Immunotherapy for acute myelogenous leukaemia: a controlled clinical study 2 1/2 years after entry of the last patient. One hundred and thirty-nine untreated patients with acute myelogenous leukaemia (AML) were admitted between August 1970 and December 1973 and allocated into two remission treatment regimens: one to receive chemotherapy alone and the other chemotherapy with immunotherapy. Of the patients who attained remission. 22 were in the chemotherapy group and in September 1975 2 remained alive, the median survival time being 270 days and after relapse 75 days. Twenty-eight patients received immunotherapy during remission, and 5 remained alive; the median survival time of the group being 510 days and after relapse 165 days. Ongoing acturial analysis precisely predicted early in the study the median survival of the two groups, but it took a 2-year follow-up after entry of the last patient before it became clear that there were very few long-term survivors. The increase in survival time produced by the immunotherapy is apparently made up of two components: prolongation of the first remission and length of survival after the first relapse. It must be notted that the chemotherapy for this study was devised 6 years ago and the results of the control arm (chemotherapy alone) may be poorer than those obtained in contemporary studies.


Direct immunofluorescence in the diagnosis of scleroderma syndromes. Immunofluorescent study of the skin of nine patients with mesenchymal, inflammatory scleroderma (mixed connective tissue disease) revealed immunoglobulin and complement deposition at the basement membrane or within blood vessel walls. The skin specimens of ten patients with systemic scleroderma were negative for immunofluorescence. It is proposed that basement membrane or vascular (or both) immunofluorescence is an excellent means of identifying the infrequent patient who has scleroderma and myositis or lupus erythematosus in whom a corticosteroid response may occur.


An organ culture model for the study of pemphigus acantholysis. An in vitro model for the study of pemphigus acantholysis has been developed. The histological changes of pemphigus vulgaris were reproduced in vitro in organ culture by growing normal human skin in the presence of pemphigus vulgaris or pemphigus foliaceus sera. At 24 h a suprabasilar split was noted and at 72 h extensive suprabasilar acantholysis developed. Direct immunofluorescent tests demonstrated that pemphigus antibody became bound to the epidermal intercellular space antigen(s) during the first 6-12 h. As acantholysis increased the presence of tissue-fixed antibody decreased. The fixation of the pemphigus antibody to the skin prior to the development of acantholysis provides strong evidence for the pathogenetic role of this antibody in the production of acantholysis. The data suggest that complement is not required in this model for the production of the acantholytic changes of pemphigus since heating the serum for 30 min at 56 degrees C did not destroy the acantholytic activity and no complement (C3) could be detected by DIF of organ culture explants.


Measurement of dermal atrophy induced by topical steroids using a radiographic technique. A modified radiographic technique was used to assess dermal atrophy induced by topical steriods. The method proved reliable, and significant atrophy occurred with fluocinolone acetonide, clobetasone butyrate, hydrocortisone 17-butyrate and flurandrenolone, but not with hydrocortisone.


The immunotherapy of acute myelogenous leukaemia using intravenous BCG. In a 2-year period, 37 of 81 adults with acute myelogenous leukaemia achieved complete remission after repeated courses of Daunorubicin (DNR) and Cytosine Arabinoside (ARAC). They were randomized to maintenance treatment with monthly DNR/ARAC, or to identical chemotherapy plus intravenous BCG. Eighteen BCG treated patients had significantly longer survival times than 19 patients treated with chemotherapy only although no statistically significant difference can be seen in the remission duration of the two groups. Eleven patients in the BCG treated group who have relapsed, have received DNR/ARAC reinduction and five second and two third remissions have been obtained. Twelve control group patients have relapsed and 10 have received further reinduction treatment with DNR/ARAC but only one patient has entered a complete remission. Seven patients in the BCG treated group who survived for 75 weeks or more (76, 76, 96, 124, 125, 138 and 145 weeks) were either PPD positive before treatment or converted to PPD positivity after BCG treatment. Using a battery of skin tests it may be possible to define a good prognostic group of patients and design future treatment accordingly. The BCG group had a total of 198 intravenous treatments. All patients had pyrexia 6-12 h after injection lasting 12-72 h and occasionally headaches and muscle pains. Two patients had non-fatal anaphylactic reactions which did not recur when BCG was subsequently re-administered. Other complications of BCG therapy were not a problem and we have not needed to withdraw treatment for any patient.


Membrane changes in human platelets induced by lipopolysaccharide endotoxin. The unexplained occurrence of thrombocytopenia in cases of Gramnegative sepsis in man led us, in the light of animal experiments indicating the blood platelet as the target cell for endotoxin, to examine the effect of Salmonella enteritidis lipopolysaccharide B on human platelets. Human platelets were separated from a coat of plasma proteins by Sepharose 2B filtration or by a combined procedure of albumin gradient and Sepharose 2B filtration. The action of endotoxin on human platelets resulted in membrane changes manifested by dose-dependent release of [3H]serotonin and adenine nucleotides. Cytoplasmic marker, lactic dehydrogenase, and lysosomal marker, beta glucuronidase, were retained indicating that the release reaction was selective. Release of [3H]serotonin was specific for endotoxin since other particulates, zymosan and erythrocyte stroma, were without effect. Endotoxin, added to gel-filtered human platelets, induced a significant evolution of platelet factor 3 procoagulant activity. Preincubation of endotoxin with a membrane-rich homogenate of human platelets inhibited its labilizing effect on human platelets thus suggesting an interaction between endotoxin and the platelet membrane itself. Other plausible factors in this interaction [fibrinogen, adenine nucleotides, thrombin, sialic acid residues, and IgG] were eliminated on the basis of a series of control experiments. From the negligible effect of aspirin and indomethacin, we may infer that the interaction of endotoxin with platelets does not depend on the platelet prostaglandin synthesis pathway. The direct interaction of endotoxin with the human platelet membrane comprises a new mechanism which may help to clarify the pathogenesis of vascular and haemostatic disorders accompanying bloodstream infections due to Gram-negative bacteria.


A contribution from the field of psychodynamics to the controversy over reactive and endogenous depression. Comparisons of depressed with non-depressed subjects, and of endogenous depressives with reactive depressives, in respect of the quality and handling of aggression, suggest that endogenous depression is a distinct entity. Reactive depression is seen as essentially indistinguishable from other non-depressed psychiatric states.


Treatment of TRIC infection of the eye with rifampicin or chloramphenicol. An open trial was carried out on 63 patients in London to assess the efficacy of 1% rifampicin eye ointment in comparison with 1% chloramphenicol eye ointment in the treatment of sexually transmitted TRIC infection of the eye. Patients included were selected on the basis of positive cultures for Chlamydia trachomatis. Three weeks' treatment with rifampicin eye ointment used 3 times daily was not sufficient to cure the disease, but a 6 or 7 week course gave 90% clinical and microbiological cure rate. Treatment with chloramphenicol eye ointment 3 times daily for 4 to 6 weeks failed to cure the disease.


Immunopathology of the lens. II. Humoral and cellular immune responses to homologous lens antigens and their roles in ocular inflammation. It has been possible for the first time to demonstrate antibodies to homologous lens proteins in rabbits without the addition of adjuvant. By means of immunofluorescence and immunoperoxidase methods it has been possible not only to show for the first time that homologous lens antibodies cross-react with extraocular tissues but that the cross-reacting antigens are related to the cell mitochondria, microsomes, and the proteins associated with contractile organelles. The rabbits did not produce antibodies to gamma-crystallins even when the whole lens homogenate was injected with Freund's complete adjuvant. This suggests that gamma-crystallins are non-antigenic in homologous situations, and this may be related to both B- and T-cell tolerance. Alternatively, the failure of gamma-crystallins to induce antibody production may be due to intermolecular antigenic competition with other crystallins. The presence of mycobacteria in an adjuvant is essential for an antibody response to be detectable by agar diffusion techniques. The response to homologous lens antigens, both in magnitude as well as in duration, varied in different rabbits, which suggested to us that a central control mechanism involving the immune response (Ir) genes may plan an important role. Antibodies to homologous lens proteins as detected by immunofluorescence and immunoperoxidase methods were shown to be of the IgG class. This is the first time that the kinetics of the immune response to the same homologous lens antigen in saline with or without incomplete or complete adjuvant has been examined and their relative merits compared. Systemic homologous immunisation followed by discission of the lens led to a marked Arthus type reaction in and around the lens, but a typical granulomatous phakoallergic endophthalmitis was not produced. It seems likely that the rabbit is not suitable for the production of an experimental model of this condition.


Studies on the mechanism of Escherichia coli glucosamine-6-phosphate isomerase. Escherichia coli glucosamine-6-phosphate isomerase is specific for removal of the 1-pro-R hydrogen of fructose 6-phosphate (fructose-6-P). The conversion of [2-3H]glucosamine-6-P to fructose-6-P plus ammonia is accompanied by 99% exchange of tritium with water and 0.6% transfer to C-1 of fructose-6-P. The enzyme is active toward alpha-glucosamine-6-P and apparently inactive toward the beta anomer. The combination of the above results supports a cisenolamine intermediate for the reaction. The labeling of substrate and product pools in tritiated water shows that the two halves of the reaction are each freely reversible. No single step appears to be rate determining. 2-Amino-2-deoxyglucitol-6-P is an unusually strong competitive inhibitor (K1 = 2 X 10(-7) M, compared with the Km = 4 X 10(-4) M for glucosamine-6-P), suggesting the enzyme has a strong affinity for the open-chain form of glucosamine-6-P.


Purification and characterization of covalently closed replicative intermediates of ColEl DNA from Escherichia coli. Pulse-labeled ColEl DNA molecules, undergoing replication in Escherichia coli cells either in the absence or presence of chloramphenicol, were extracted and purified by neutral sucrose density gradient sedimentation and equilibrium centrifugation in an ethidium bromide-cesium chloride gradient. In the dye-buoyant density gradient, the replicating molecules were found in regions between the supercoiled and open-circular nonreplicating plasmid DNA, as well as in the open-circular region. In a neutral sucrose gradient, peaks of pulse label were found in the region of 26 to 38 S as well as at the 23 and 17 S positions corresponding to the positions of supercoiled and open-circular ColEl DNA. In alkaline sucrose gradient, nascent ColEl DNA was found to sediment as discrete peaks corresponding to 5-6, 7-9, and 14-16 S, indicating that at least one growing strand of the replicating molecule is produced discontinuously. In the electron microscope, many of the molecules appeared as partially supercoiled structures containing two open-circular branches of equal length, of less than 20% to more than 90% replicated. Branched open-circular molecules were not observed to any significant extent without prior treatment to induce single-strand scissions. The parental strands of the replicating molecules were determined to be covalently closed, but the superhelical density of the DNA was shown to be progressively decreased as replication proceeded.


Initial rate kinetic analysis of the mechanism of initiation complex formation and the role of initiation factor IF-3. Initial rate kinetics of the formation of ternary complexes of Escherichia coli 30S ribosomal subunits, poly(uridylic acid), and N-acetylphenylalanyl transfer ribonucleic acid in the presence and in the absence of IF-3 are consistent with the hypothesis that the ternary complex is formed through a random order of addition of polynucleotide and aminoacyl-tRNA to separate and independent binding sites on the 30S ribosomes. The transformation of an intermediate into a stable ternary complex which probably entails a rearrangement of the ribosome structure leading to a codon-anticodon interaction represents the rate-limiting step in the formation of the ternary complex. The rate constant of this transformation, as well as the association constants for the formation of the 30S-poly(U) and 30S-N-AcPhe-tRNA binary complexes, are enhanced by the presence of IF-3 which acts as a kinetic effector on reactions which are intrinsic properties of the 30S ribosome. The IF-3-induced modification of these kinetic parameters of the 30S ribosomal subunit can per se explain the effect of IF-3 on protein synthesis without invoking a specific action at the level of the mRNA-ribosome interaction. This seems to be confirmed by the finding that IF-3 can stimulate several-fold the formation of a ternary complex even if one by-passes the ribosome-template binding step by starting with a covalent 30S-polynucleotide binary complex. Furthermore, the above-mentioned changes induced by IF-3 appear to be compatible with the previously proposed idea that the binding of the factor modifies the conformation of the 30S subunit. The random order of addition of substrates determined for the 30S-N-AcPhe-tRNA-poly(U) model system was found to be valid also for the more physiological 30S initiation complex containing poly(A,U.G) and (fMet-tRNA formed at low Mg2+ concentration in the presence of GTP and all three initiation factors.


Hydrolytic action of aminoacyl-tRNA synthetases from baker's yeast. "Chemical proofreading" of Thr-tRNA Val by valyl-tRNA synthetase studied with modified tRNA Val and amino acid analogues. The properties of native and of two modified tRNA Val species in the correction of misactivated threonine by valyl-tRNA synthetase have been studied. Whereas Thr-tRNA Val-C-C-A could not be isolated in the valyl-tRNA synthetase catalyzed reaction, Thr-tRNA Val-C-C-3'dA is isolable in up to 50% yield in this system and tRNA Val-C-C-3'NH2A is fully aminoacylated with threonine by the same enzyme. The hydrolysis of preformed Thr-tRNA Val-C-C-A by free valyl-tRNA synthetase is 30 times faster than the corresponding breakdown of Val-tRNA Val-C-C-A. This hydrolytic activity is also observed with Thr-tRNA Val-C-C-3'dA although the rate is reduce to that of the reaction of Val-tRNA Val-C-C-A. Modification of the threonine to O-methylthreonine, which is also a substrate for valyl-tRNA synthetase, leads to stabilization of the O-methylthreonyl-tRNA esters. The AMP/PP independent hydrolysis under aminoacylating conditions, which is a measure of the correction process, indicates that O-MeThr-tRNA Val-C-C-A is only very slowly corrected while the tRNA Val-C-C-3'dA and tRNA Val-C-C-3'NH2A esters are completely stable. Removal of the methoxy group of O-methylthreonine as in alpha-amino-butyric acid increases the rate of the hydrolytic reaction and once again alpha-Abu-tRNA Val-C-C-A and alpha-Abu-tRNA Val-C-C-3'dA are unstable under aminoacylating conditions and not isolable.


Studies on gene control regions. 1. Chemical synthesis of lactose operator deoxyribonucleic acid segments. The chemical synthesis of lactose operator DNA segments is described. The 31-base-paired duplex contains the DNA recognized by lac repressor protein and twofold rotationally symmetric base pairs on either side of the tight binding region. The synthesis includes the deoxyoligonucleotides d(T-G-T-G-G), d(A-A-T-T-G-T-G-A-G), d(C-G-G-A-T-A-A-C-A-A-T-T), d(T-C-A-C-A), d(T-G-T-G-A-A-A-T-T-G-T), d(T-A-T-C-C-G-C-T-C-A-C), and d(A-A-T-T-C-C-A-C-A). These deoxyoligonucleotides were characterized by two-dimensional sequencing techniques, paper chromatography, and thin-layer chromatography.


In vitro transcription of Moloney leukemia virus genes in infected cell nuclei and chromatin: elongation of chromatin associated ribonucleic acid by Escherichia coli ribonucleic acid polymerase. The in vitro transcription of viral specific DNA sequences in nuclei and chromatin isolated from mouse cells chronically infected with Moloney murine leukemia virus (Mo-MuLV) has been studied. The in vitro RNA synthesized by Escherichia coli RNA polymerase has been isolated by sulfhydryl affinity column following reaction in the presence of 5-mercuriuridine triphosphate. By comparison of the Crt curves of the in vitro RNA with that of 70S viral RNA, the content of viral sequences is found to be 1.3% in nuclei product and 0.24% in chromatin product which is lower than the 2.5% found in chromatin associated RNA. This latter value, however, is very close to the in vivo viral RNA content in pulse-labeled [3H]RNA of the infected cells. Unexpectedly, it is observed that over 20% of the chromatin associated RNA prelabeled in vivo with [5-3H]uridine is elongated and tagged with Hg atoms during RNA synthesis catalyzed by the exogenous E. coli RNA polymerase in the presence of Hg-UTP. The elongation reaction is dependent on the presence of all four nucleotide triphosphates and appears to be due to E. coli RNA polymerase per se. It is suggested that most of the viral specific sequences observed in the in vitro RNA products are very likely initiated and derived from the chromatin associated species. The implication of the present findings for in vitro RNA synthesis in nuclei and chromatin as related to regulation of gene expression is discussed.


Stimulation, by two Escherichia coli supernatant proteins, of the initiation of polypeptide synthesis. Two protein factors (A and B) have been partially purified from Escherichia coli supernatant which, in combination, are more effective than 0.5 M NH4Cl in stimulating ribosomes for AcPhe-tRNA and fMet-tRNA binding, for the puromycin reaction, and for incorporating acetylphenylalanine from AcPhe-tRNA into polypeptide. The factors appear to differ from the initiation factors, the elongation factor EF-T, and ribosomal proteins. Some uncertainty exists as to whether factor B is different from EF-G. To maximize the effect of the factors in initiator tRNA binding, we preincubated the ribosomes with the factors and carried out the binding assay for a short period at 15 degrees C. Maximal stimulation of binding occurred after about a 2-min preincubation at 37 degrees C. Longer preincubation times were required at 15 degrees C, and only slight stimulation was observed after preincubation at 0 degrees C. The extent of stimulation by the factors was not affected when the NH4Cl concentration was increased from 40 to 500 mM in the preincubation. The presence of both the 30S and 50S ribosomal subunits is required for the enhancement of AcPhe-tRNA binding. Polyphenylalanine synthesis carried out without AcPhe-tRNA is inhibited by the factors. It is suggested that the factors may act by inducing a structural rearrangement of the ribosomes.


Purification and structural analysis of the fourth component of human complement. The fourth component of human complement (C4) has been purified in 20% yield from fresh plasma using as starting material the 5-12% poly(ethylene glycol) precipitate which had been depleted of plasminogen by an affinity adsorbent. Sequential ion-exchange chromatography on diethylaminoethylcellulose, QAE-Sephadex, and DEAE-Bio-Gel A resulted in C4 homogeneous by immunological criteria and by polyacrylamide gel electrophoresis, the last chromatographic step achieving separation of native from inactivated C4. Reduction with 20 mM dithiothreitol for 2 h at 37 degrees C in 0.25 M 2-amino-2-hydroxymethyl-1,3-propanediol hydrochloride, pH 8.6, effected cleavage of the interchain disulfide bonds. A three-chain structure for C4 was confirmed, and molecular weight estimates of 93 000 +/- 9300, 75 000 +/- 7500, and 30 000 +/- 3000 determined for the alpha, beta, and gamma chains, respectively. The effects of known inactivators of C4 upon the chains of C4 were investigated, confirming that the inactivations by C1s and trypsin were accompanied by the fragmentation of the alpha chain. Inactivation of C4 by hydrazine, on the other hand, produced no detectable change in chain size. Separation of the chains was accomplished by gel filtration in the presence of 1 M acetic acid. Amino acid compositions of native C4 and the constitutive chains have been performed, and N-terminal sequences of the latter established by automated Edman degradation.


Biosynthesis and structure of lipopolysaccharide in an outer membrane-defective mutant of Escherichia coli J5. Membrane-defective mutants of Escherichia coli J5 were isolated on the basis of supersensitivity to the antibiotic novobiocin. These mutants display an increased sensitivity to a wide range of antibiotics and to several dyes and detergents. In addition, several mutants leak the periplasmic enzymes, alkyline phosphatase and ribonuclease. This evidence indicates an outer membrane defect in these mutants. The inner and outer membranes of one mutant were separated and subjected to compositional analysis. A deficiency in galactose containing lipopolysaccharide in the outer membrane of the mutant was observed. Two possible causes of this deficiency were examined and discounted: defective galactose uptake into the cell, and defective translocation of lipopolysaccharide from the inner membrane. Extraction and chemical analysis of mutant and wild type lipopolysaccharides suggests that the mutant is defective in the enzyme which transfers glucose to the growing lipopolysaccharide core, UDPglucose transferase. Thus, the mutant's deficiency in galactose-containing lipopolysaccharide can be ascribed to the fact that addition of glucose to the lipopolysaccharide core is a prerequisite for galactose addition. The physiological implications of this alteration are discussed.


The use of a phospholipase A-less Escherichia coli mutant to establish the action of granulocyte phospholipase A on bacterial phospholipids during killing by a highly purified granulocyte fraction. Phospholipase A2 present in a highly purified, potently bactericidal, fraction from rabbit graulocytes produces net bacterial phospholipid degradation during killing of a phospholipase A-less strain of Escherichia coli. In the wild-type parent strain phospholipid breakdown is caused not only by the action of phospholipase A2 but also by phospholipase A1, indicating activation of the most prominent phospholipase of E. coli. This activation occurs as soon as the bacteria are exposed to the granulocyte fraction. Phospholipid breakdown by both phospholipases A is dose dependent but reaches a plateau after 30-60 min and at higher concentrations of the fraction. Phospholipid degradation is accompanied in both strains by an increase in permeability to actinomycin D that is also dose dependent. Even though net hydrolysis of phospholipids is greater in the parent strain than in the mutant, the increase in permeability is the same in the two strains. The addition of 0.04 M Mg2+, after the effects on phospholipids and permeability have become manifest, initiates in both strains the restoration of insensitivity to actinomycin D, the net resynthesis of phospholipids, and the disappearance of monoacylphosphatides and the partial disappearance of free fatty acids that had accumulated. Loss of ability to multiply is not reversed by Mg2+ in either strain. Less than 5 micrograms of granulocyte fraction causes loss of viability of from 90 to 99% of 1 X 10(8) microorganisms of both strains. However, at lower concentrations the parent strain is considerably more sensitive to the bactericidal effect of the granulocyte fraction than the mutant strain.


Relationship between phospholipid compositions and transport activities of amino acids in Escherichia coli membrane vesicles. Cells of Escherichia coli were incubated in broth medium in the presence of 5 mM of hydroxylamine which completely inhibited growth but did not affect viabilities. Hydroxylamine is known to inhibit phosphatidylserine decarboxylase. A large amount of phosphatidylserine (up to 20% of total phospholipids), which did not occur in normal cells, accumulated accompanied with a decrease in phosphatidylethanolamine. Higher uptake activities of serine and glutamate were observed with the hydroxylamine-treated cells than control cells. When membrane vesicles from hydroxylamine-treated cells were prepared, they also displayed higher uptake activities of serine, proline, glutamate, and threonine than those of normal membranes. When hydroxylamine-treated cells were incubated with chloramphenicol, at concentrations which almost completely inhibited protein synthesis, the composition of phosphatidylserine decreased with a concomitant increase in that of phosphatidylethanolamine. The phospholipid composition of these cells incubated for 5 h with chloramphenicol became almost normal. Membranes vesicles prepared from such cells displayed reduced uptake activities, which were close to those of normal vesicles. These results were interpreted as indicating the altered transport activities due to the altered phospholipid composition.


Cross-linking of the proteins in the outer membrane of Escherichia coli. 1. The organization of the proteins in the outer membrane of Escherichia coli was examined by the use of cross-linking agents and two-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Treatment of protein A-peptidoglycan complexes with dithiobis(succinimidyl propionate) or glutaraldehyde produced the dimer, trimer, and higher oligomers of protein A. Both forms of this protein, proteins A1 and A2, produced similar cross-linking products. No cross-linking of protein A to the peptidoglycan was detected. 2. The proteins of the isolated outer membrane varied in their ease of cross-linking. The heat-modifiable protein, protein B, was readily cross-linked to give high molecular weight oligomers, while protein A formed mainly the dimer and trimer under the same conditions. The pronase resistant fragment, protein Bp, derived from protein B was not readily cross-linked. No linkage of protein A to protein B was detected. 3. Cross-linking of cell wall preparations, consisting of the outer membrane and peptidoglycan, showed that protein B and the free form of the lipoprotein, protein F, could be linked to the peptidoglycan. A dimer of protein F, and protein F linked to protein B, were detected. 4. These results suggest that specific protein-protein interactions occur in the outer membrane.


Architecture of the outer membrane of Escherichia coli K12. I. Action of phospholipases A2 and C on wild type strains and outer membrane mutants. Phospholipids in whole cells of wild type Escherichia coli K12 are not degraded by exogenous phospholipases, whereas those of isolated outer membranes are completely degraded. It is concluded that the resistance of phospholipids in whole cells is due to shielding by one or more other outer membrane components. The nature of the shielding component(s) was investigated by testing the sensitivity of whole cells of a number of outer membrane mutants. Mutants lacking both major outer membrane proteins b and d or the heptose-bound glucose of their lipopolysaccharide, are sensitive to exogenous exogenous phospholipases. Moreover, cells of a mutant which lacks protein d can be sensitized by pretreatment of the cells with EDTA. From these results and from data on the chemical composition of the outer membranes, it is concluded that proteins b and d, the heptose-bound glucose of lipopolysaccharide and divalent cations are responsible for the inaccessibility of phospholipids to to exogenous phospholipases.


Phase transitions of phospholipid bilayers from an unsaturated fatty acid auxotroph of Escherichia coli. Total phospholipids were extracted from cells of temperature sensitive unsaturated fatty acid auxotrophs of Escherichia coli (K-12 UFAts) grown at 28degrees C (PL28), and at 42degrees C in the presence of 2% KCl as an osmotic stabilizer (PL42 (KCl)). From the analysis of fatty acids, it was shown that the content of unsaturated fatty acids of PL42 (KCl) is only 9% of the total fatty acids, while that of PL28 is 54%. The thermal phase transitions of the bilayers prepared from the phospholipid fractions were studied by proton magnetic resonance. The line widths of the methylene signals and the sums of the methylene and methyl signal intensities were plotted against reciprocal values of absolute temperature 1/T or temperature itself. From the plots phase transitions were detected at about 19degrees C for PL28 and at 43degrees C for PL42 (KCl). In spite of its complex composition of fatty acids a highly cooperative transition was observed in the case of PL42 (KCl). It was also suggested that the phospholipids bilayers in the biomembranes of this strain at the growth temperature (42 degrees C) are in the state where the gel and liquid crystalline phases coexist.


Ribonucleic acid from the higher plant Matthiola incana. Molecular weight measurements and DNA-RNA hybridisation studies. The percentage of DNA from the crucifer Matthiola incana coding for different types of RNA was measured by filter saturation hybridisation experiments using RNA labelled in vivo. In addition, the melting curves of the various DNA - RNA hybrids formed and the buoyant densities of the DNA sequences complementary to different types of RNA were measured. 1. The RNA preparations used were 25, 18, and 5 S rRNA and 4 S RNA, purified by gel electrophoresis, and poly(A)-containing RNA purified by oligo-(dT)-cellulose chromatography. The molecular weights of the 25 S and 18 S rRNAs, calculated from the mobility in formamide-acrylamide gels relative to Escherichia coli RNA, are 1.25 - 10(6) and 0.64 - 10(6). The rRNA precursor has a molecular weight of approx. 2.1 - 10(6) and the average molecular weight of the poly(A)-containing RNA from both cotyledons and roots is 4 - 10(5). 2. The percentage of the genome, calculated on the basis of double-stranded DNA, coding for these RNAs and the estimated number of genes per haploid DNA amount are approximately 0.46% and 1100 for 25 S plus 18 S rRNA, 0.032% and 3600 for 5 S rRNA and 0.072% and 13 000 for 4 S RNA. In filter hybridisation experiments very little hybridisation of poly(A)-containing RNA was found. A rapidly-hybridising component is attributed to small amounts of contaminating rRNA. 3. M. incana DNA has a main band at 1.697 g - ml-1 in CsCl and a satellite constituting approximately 3% of the DNA, at 1.708 g - ml-1 - 25 and 18 S rRNA hybridise to DNA with a buoyant density of 1.701--2 g - ml-1. The buoyant density of 5 S DNA is slightly less at 1.700--1 g - ml-1. 4. S RNA hybridises to at least two separate regions, one within the main-band DNA and a second lighter component. None of the RNAs tested hybridised to the satellite DNA. The Tm of the DNA - RNA hybrids in 1 X SSC is 89 degrees C for 25 S rRNA, 85 degrees C for 5 S rRNA and 82 degrees C for 4 S RNA. 4. 5 and 4 S RNA preparations contain fragments which hybridise to sequences complementary to high-molecular-weight rRNA. This spurious hybridisation can be eliminated by competition with unlabelled high-molecular-weight RNA.


Release of template restriction in chromatin by nuclear 4.5s RNA. The stimulatory mechanism of RNA synthesis of calf-thymus chromatin by nuclear 4.5 S RNA from the homologous tissue was investigated by using exogenously added Escherichia coli RNA polymerase. The RNA synthesis was initiated at low concentration of salt, and then the chain elongation was achieved at high concentration of ammonium sulfate in the presence of polyvinyl sulfate. Under these conditions the number of binding sites of RNA polymerase on chromatin which were capable of initiating RNA chain was increased by the addition of the 4.5 S RNA. This stimulation was presumed to result from the release of template restriction in chromatin. The polyvinyl salt minimized ribonuclease activity without changing the RNA polymerase activity bound to the template. Neither rearrangement nor release of chromatin proteins affected the amount or size of RNA produced. Preliminary analysis suggested that the molecular species of RNA produced upon the addition of the 4.5 S RNA from various tissues seemed to be heterologous.


Invertase messenger ribonucleic acid in Saccharomyces cerevisiae. Kinetics of formation and decay. Saccharomyces cerevisiae -136ts (Hutchison, H.T., Hartwell, L.H. and McLaughlin, C.S. (1969) J. Bacteriol. 99, 807-814) incubated in the presence of maltose at 23 degrees C (permissive temperature) synthesized the RNA messengers which codify derepressed invertase (an external mannoprotein) and induced alpha-glucosidase (a non-glycosylated internal enzyme). The enzymes were not synthesized if the mutant was transferred to the maltose-containing medium at the moment of incubation at 37 degrees C indicating that the cells had no pools of the specific RNA messengers and that transcription of the DNA was a prerequisite to enzyme synthesis. Cycloheximide inhibited syntheses of the enzymes both at 37 and at 23 degrees C suggesting that the enzymic activities were the result of "de novo" synthesis of the proteins and did not result from the activation of proenzymes. In derepressed cells the number of invertase mRNA molecules is probably larger than that actually being translated. The half-life of the derepressed invertase mRNA was calculated from the moment that the molecules of RNA messenger were limiting the enzyme synthesis and a value of 30-35 min was estimated. The value found for the basal (repression independent) invertase mRNA was of 45-50 min. The half-life of alpha-glucosidase mRNA was computed following the mathematical procedure described in the Appendix, and a value of 23 min was obtained. These results are consistent with the existence of relatively long-lived RNA messengers involved in the synthesis of extracellular macromolecules.


Discrimination between bromouracil and thymine for uptake into DNA in drm- and dra- mutants of Escherichia coli K12. The relative efficiency of bromouracil and thymine for uptake into DNA was measured in various thymine-requiring strains of Escherichia coli K12. It was found that: 1. Mutants with genotype thyA- dra- discriminate against bromouracil to much greater extent than do mutants with genotype thyA- drm-. 2. The discrimination in dra-mutants is dependent on thymine concentration, whereas discrimination in drm- mutants is almost independent of thymine concentration. It is suggested that the intracellular level of deoxyribose 5-phosphate affects the efficiency of uptake into DNA of bromouracil relative to thymine.


Comparative inactivation of yeast fatty acid synthetase component enzymes by 100 atmospheres of oxygen. Yeast fatty acid synthetase at 4 degrees C was stable during 1- and 2-h exposures to oxygen at 100 atm, but was 48% and 90% inactivated after 20 h and 40 h, respectively, with fatty acid synthesis measured by both radioactive and optical assays. Incubation with dithiothreitol did not restore activity. Component enzyme activities were compared before and after 40 h in 100 atm of oxygen. Ketoacyl reductase activity was most reduced followed by ketoacyl synthetase and then acetyl transferase while malonyl transferase, enoyl reductase and palmitoyl transferase were not significantly inactivated.


Quantitative effects of unsaturated fatty acids in microbial mutants. VII. Influence of the acetylenic bond location on the effectiveness of acyl chains. The ability of a series of 18 carbon acetylenic fatty acids to fulfill the unsaturated fatty acid requirements of Escherichia coli and Saccharomyces cerevisiae was investigated. Despite their high melting points (greater than 40 degrees C), several isomers of the acetylenic fatty acids were as efficient or more efficient in supporting growth than the analogous fatty acid having a cis-double bond. The efficiencies of the different positional isomers in supporting cell proliferation varied from essentially 0 cells per fmol for the 2-5 and 13-17 isomers to high values when the acetylenic bond was near the center of the chain: e.g. 45 E. coli and 5.5 S. cerevisiae cells/fmol for the 10 isomer. A striking ineffectiveness of the 9 isomer was observed with E. coli. The 7, 8 and 10 isomers were at least 10-fold more efficient than any of the other positional isomers in supporting the growth of E. coli. In contrast, the 9 isomer was among the most effective acetylenic fatty acids tested with the yeast mutant. Chromatographic analysis of the extracted lipids indicated that each of the acetylenic isomers tested (except delta2 and delta3) could be esterified by the prokaryotic and eukaryotic microorganisms. The content of unsaturated plus cyclopropane acids observed when growth ceased in E. coli cultures supplemented with growth-limiting concentrations of the acetylenic fatty acids ranged from approx. 15 mol% for the 8 isomer to approx. 35 mol% for the 14 and 17 isomers. The 8-11 isomers were observed to be esterified predominantly at the two position in phosphatidylethanolamine of E. coli and in phosphatidylcholine of S. cerevisiae.


Glycolipid intermediates involved in the transfer of N-acetylglucosamine to endogenous proteins in a yeast membrane preparation. A particulate membrane fraction from Saccharomyces cerevisiae contains transferases which catalyze the incorporation of N-acetylglucosamine from UDP-N-acetylglucosamine into a lipid fraction as well as into a protein fraction. The lipid fraction contains two alkali-stable lipids which can be separated on a silica G-60 column. The sugar moieties of these polyprenoid lipids are: N-acetylglucosamine and di-N-acetylchitobiose. The transfer of carbohydrate from isolated glycolipids to endogenous protein has been examined. After separation of protein and saccharide by hydrazinolysis and reacetylation only di-N-acetylchitobiose is found, and also when glycolipid containing only one N-acetylglucosamine is used as substrate. Maximum transfer of saccharides from glycolipids to protein is obtained at a Triton X-100 concentration of 1%. At this Triton X-100 concentration there is practically no transfer of N-acetylglucosamine from UDP-N-acetylglucosamine to the phosphorylated lipid. Therefore, when polyprenyl diphosphate N-acetyl[3H]-glucosamine is incubated together with UDP-N-acetyl[14C]glucosamine with the membrane fraction in the presence of 1% Triton X-100, a doubly labelled di-N-acetylchitobiose linked to lipid is formed with N-acetyl[14C]glucosamine at the non-reducing end of the chain.


Membrane lipid metabolism of Bacillus Calmette-Guerin-induced rabbit alveolar macrophages. We examined the uptake of radiolabeled lysophospholipids and oleic acid by Bacillus Calmette-Guerin-induced rabbit alveolar macrophages either in the presence or absence of challenge particles. There was no difference in the uptake and metabolism of lysophospholipids by control or challenged cells for incubation periods up to 5 h. When incubated with [3H]oleic acid, challenged cells consistently exhibited a slightly greater uptake of radioactivity. Extraction of the whole cells revealed that the greater amount of radioactivity found in the challenged cells primarily was in triacylglycerol. There was no marked difference in the amount of radioactivity associated with the phospholipids in the whole cell extracts from control and challenged cells. When the macrophages were pre-labeled for 15 min with [3H]oleic acid and then reincubated in fresh medium in the presence or absence of autoclaved Escherichia coli B, more radioactivity was retained by the challenged cells, again in the form of triacylglycerol. Only in isolated plasma membrane fractions did we observe a difference in the amount of radioactivity associated with phospholipids from control and challenged cells. Plasma membranes isolated from Bacillus Calmette-Guerin-induced rabbit alveolar macrophages that had been incubated for 6 h with [3]oleic acid in the presence of E. coli B contained significantly higher level of radioactivity in all lipids than plasma membranes from control cells. Since the greatest and the most consistent difference between control and challenged cells is associated with the triacylglycerol molecule, it is postulated that this molecule may serve as a precursor in the synthesis of alveolar macrophage phospholipids, both by the reacylation pathway and the de novo pathway. It is possible that the high level of radiolabeled phospholipid found in the plasma membrane arose via the de novo pathway following the cleavage of an acyl group as we have found cytidine diphosphocholine phosphotransferase in the plasma membrane fraction (Wang, P., DeChatelet, L.R., and Waite, M. (1977) Biochim. Biophys. Acta 450, 311--321).


Electrophysiological evidence for the autoregulation of beta-cell secretion by insulin. Electrophysiological studies of cultured rat pancreatic beta-cells using intracellular microelectrodes show that exogenous insulin over the range of 0.1 -- 10.0 microng/ml inhibits the electrical activity due to 27.8 mM glucose in a dose-related manner. This inhibitory effect is manifested by a mean increase of the membrane potential from about --20 to --30 mV and inhibition of the number of cells impaled showing spike activity from 60 to less than 10%. The inhibitory influence of insulin is rapid occurring within 5 min for the highest level used. The results provide evidence for a negative feedback role of insulin in regulating its own release.


Stability of 70S ribosomes in relation to misreading and antibacterial activity of aminoglycosides. The anomeric aminoglycosides, RU 21886 and RU 23468, which both have a 2-deoxystreptamine residue, stabilize 70S ribosomes to similar extents at low magnesium ion concentrations. Only RU 21886, however, has marked antibacterial and bactericidal activity and gives rise to a high level of misreading in cell-free protein synthesizing systems. It would thus appear that the ability to stabilize the association of the two ribosomal subunits does not necessarily lead to errors in translation.


[Gratuitous induction of beta-glucuronidase of Escherichia coli K 12 and the double repression mechanism]. Using natural inducers of beta-glucuronidase, methyl-glucuronide and fructuronate, under gratuitous conditions (without metabolic conversion of these two compounds), we corroborate the fact that both molecules are required simultaneously in order to derepress the enzyme synthesis to a maximum level. Structurally related analogs of the natural inducers, thiophenyl-glucuronide and mannonic amide respectively, were assayed in the wild type and suitable mutant strains of E. coli. The results are in agreement with the model where the dual negative regulation of the enzyme synthesis is exerted by two regulatory genes uidR and uxuR. The concerted action of mannonic amide and thiophenyl-glucuronide, which alone fail to induce significantly beta-glucuronidase synthesis, reveals that a cooperative effect of the two repressor molecules responsible for the complete blocking of the enzyme synthesis is occuring.


Single-stranded bacteriophage T5 stO DNA as template for in vitro fMet-tRNA binding to ribosomes and protein synthesis. Good evidence is provided that fMet-tRNA binding and aminoacid incorporation, when single-stranded DNA is used instead of mRNA in an E. coli cell-free system, are strictly dependent on the magnesium concentration. Ten sites homologous to the initiation sites of translation can be detected on denatured T5 stO DNA when using ribosomes and initiation factors from uninfected E. coli F. In S-30 extracts, at high magnesium concentrations and in the presence of neomycin, initiation of the translation of denatured T5 stO DNA begins anywhere on the molecule, and yet high molecular weight polypeptides are synthesized. The template potentiality of the denatured T5 stO DNA decreased when using ribosomes plus initiation factors and crude extracts from T5 stO-infected bacteria. By in vitro formation of initiation complexes sites analogous to initiation sites of translation were localized on T5 stO DNA molecules using single-stranded fragments separated by sedimentation in alkaline sucrose gradient.


[Structural lability of membranes].

[Structural lability of membranes]. The present-day state of the problem of membrane structural lability is reviewed. The ways of initiation and the nature of structural rearrangements, the mechanisms of generalization of local structural perturbations in membranes and the effects of rearrangements on the functional activity of organoids and cells are discussed.


[Effect of polyenic antibiotics on the activity of alkaline phosphatase from Candida albicans]. Polyenic antibiotics (levorin, amphotericin B, nistatin) inhibit in vivo and in vitro the activity of membrane alkaline phosphatase from sensitive Candida albicans strain, and their inhibitory effect is twice lower on the enzyme from the resistant strain. A correlation is observed between the antibiotic concentration and the inhibitory effect on alkaline phosphatase activity. Nistatin is found to be the least efficient inhibitor (among the antibiotics studied) of alkaline phosphatase. The treatment of membranes with polyenic antibiotics does not result in solubilization of membrane proteins nad alkaline phosphatase. The data obtained are considered with respect to the effect of polyenic antibiotics on cell membrane structure.


[Fractionation and purification of DNA methylases and specific endonucleases from cells of Escherichia coli SK]. Fractionation and purification of DNA methylases and specific endonucleases from E. coli SK responsible for DNA specificity to host prokaryotic cells were studied. The most efficient purification was achieved by precipitation of proteins by 0.6 saturated ammonium sulfate with subsequent chromatography on KM-cellulose and concentration of fractions by dialysis against glycerol. Under these conditions the methylase activity produced 4 discrete fractions. Due to purification the specific activity of methylases increased 11--20-fold in various fractions. Methylase from the first (A) and fourth (BII) peaks catalyzed the methylation of cytosine to produce 5-methylcytosine; methylase from the third peak (BI) methylated adenine to produce 6-methylaminopurine. The chemical specificity of the second peak (B) methylase could not be established due to very high lability of the enzyme in this fraction. Specific endonuclease was found in the gradient zones eluted by 0.1--0.2 M and 0.65--0.75 M NaCl. It is assumed that those enzymes providing for DNA hydrolysis up to the formation of high--molecular discrete fragments, are restricting endonucleases of the SK system. The results obtained strongly suggest the existence of several types of methylases and restricting endonucleases in E. coli SK cells.


A mechanistic model of the aerobic growth of Saccharomyces cerevisiae. A two-stage deterministic model of the growth of Saccharomyces cerevisiae is presented. The cell cycle of this organism was used to suggest the basic model structure. The model represents the preparatory processes of substrate uptake and conversion separately from replication and division. The regulation of the fraction of the culture devoted to each of these broad areas of metabolism, and the overall growth rate, is related to the nature and availability of the energy substrate. The simulation of respiration and glycolysis is achieved by including two alternative energy producing pathways. The regulation of these pathways is described in terms of the postulated primary regulation of the proportion of the culture required for substrate uptake and conversion, and the overall kinetic constants for each pathway. This regulation is dictated primarily by the growth rate rather than the nature or concentration of the energy substrate. The model successfully describes both batch and continuous growth of S. cerevisiae under conditons of glucose limitation and oxygen excess. A preliminary assessment indicates that adjustment of the relevant parameters will allow the model to describe the growth of S. cerevisiae on other sugars and under oxygen limitation. Similarly the model could be expected to describe the growth characteristics of other yeast species.


Assimilation of liquid hydrocarbon by microorganisms. I. Mechanism of hydrocarbon uptake. The uptake mechanism of liquid hydrocarbons of low solubility in water was investigated, using microorganisms with different affinities for liquid hydrocarbon. Microorganisms which could utilize hydrocarbon were much more adherent to hydrocarbon than those which could not. The adhesive force between Candida intermedia IFO 0761 and hydrocarbon was higher than that of Candida tropicalis ATCC 20336, though both could utilize hydrocarbon; The total hydrocarbon uptake from the drop and accommodation forms of hydrocarbons was much higher than that from dissolved hydrocarbon. The uptake rate of drop-form hydrocarbon was nearly equal to that of accommodation-form hydrocarbon for C. intermedia, but was lower for C. tropicalis which shows lower adhesion to hydrocarbon.


Limb anomalies in chromosomal aberrations. This survey shows that there are at least 6 autosomal and 2 gonosomal aberrations which may produce specific types of limb anomaly in 30%-80% of cases. The "expressivity" of these anomalies covers a wide range within the morphogenetic pattern. No entirely specific malformation type is seen. The most unusual malformation, aplasia of the thumbs with proximal synostosis of the 4th aand 5th metacarpals, is seen in 13q- (13r) but the precise cytogenetic basis is not clear. Aplasia of the thumb associated with synostosis of the 4th and 5th metacarpals was occasionally described before (unilateral [105], bilateral [106] while synostosis only (V or Y shaped) may be due to a dominant [107] or an X-linked recessive gene [108]. Reduction malformations limited to radial heminelia have been noted in 4q- (4r) and in trisomy 18. Although the number of cases is still small the pattern is similar to that of thalidomide embryopathy, radial hemimelia (AD, 17910), cardiodigital syndromes (AD, 14290), and even Fanconi panmyelopathy (AR, 22790). It ranges from hypoplasia of the thenar muscles and thumb to pseudophocomelia which should be clearly distinguished from phocomelia because of the absence of the thumb and frequently of the 2nd and 3rd fingers. Nothing has to be added to the teratologic series published by Müller [58] and, more recently, Willert and Henkel [109], but the distribution of the various manifestations may diverge. Asymmetry in 4q- (4r) is noteworthy. Postaxial polydactyly which is noted in trisomy 13 and trisomy of the terminal portions of the long arm of No. 13 is as variable in distribution and morphology as is observed in families in which the gene (AD, 17420) is transmitted. The question cannot yet be answered whether infrequent anomalies of the limbs which do not fit into the morphologic pattern of these types, eg postaxial polydactyly in + 18 or absence of the radius in + 13 are random. Syndactyly of the 3rd and 4th but also of other digits is a frequent but variable anomaly in triploidy. It is very similar to hereditary zygodactyly (AD, 18590). Peripheral hypoplasia of several digits accompanied by onychodysplasia seems to be a frequent anomaly in 9p+ syndrome. It is similar to that seen in a syndrome with mental deficiency, peculiar facies, and stunted growth [110] in which no chromosomal aberration has been found up to the present. Dysostoses have been frequently noted in gonosomal aberations. Brachymetopodia in XO females maybe confused with pseudo-pseudohypoparathyroidism (XR, 30080) or brachydactyly type E (AD, 11330) when only the lateral metacarpals and/or metatarsals are shortened. However, further studies are needed in order to delineate these syndromes on the basis of different frequencies and radiologic patterns. The radioulnar synostosis noted in cases with supernumerary X chromosomes cannot be distinguished from the inherited anomaly (AD, 17930), but associated anomalies of the hand are uncommon.


[Role of substrate inductors in the mechanism of cortisol action on beta-galactosidase activity in different strains of E. coli k-12 and in rat liver]. A comparative study of the effect of cortisone on the beta-galactosidase synthesis in E. coli K-12 strains with an induced (E. coli 200 PS/Iac), constitutive (E. coli ML-308), and superrepressed (E. coli 200is) type of the enzyme synthesis and in the cells of rat liver demonstrated that the hormone proper had no derepressive effect. An increase of the beta-galactosidase synthesis occurred only in the presence of specific substrate inductors. It is supposed that the principal link in the action mechanism of cortisone on the laclose operon of E. coli and the enzyme production in the cells of the rat liver is preliminary derepression of the genome areas by means of the substrate inductors.


[Classes of rat lymphocyte surface immunoglobulins detected by enzymatic radioiodination]. By means of enzymatic radioiodination, immunopre-cipitation and acrylamide gel electrophoresis in sodium dodecyl sulfate it was found that on the surface of normal rat splenocytes there were two main immunoglobulin classes--monomeric IgM and Ig (H2L2) which had a heavy chain larger than they phi, yet smallerthan the micron chain, differing from them by the antigenic properties. It is possible that this Ig class corresponds to the human IgD and the mouse IgD-like cell surface molecules. Small amounts of thedetected cell surface IgG may indicate both its cytophilic properties and the immunological status of the experimental animals.


[Comparative analysis of the adjuvant action of fab-fragments of normal rabbit IgG obtained using pepsin and papain]. The capacity of Fab fragments of normal rabbit IgG to enhance the immune response to sheep erythrocytes in the homologous recipients was determined by the structure of the C-terminal part of the heavy chain Fd fragment. This followed from the fact that pepsin F(a')2 and Fab' fragments enhanced considerably the hemagglutinin production and proliferation of the antibody-forming cells in the spleen, whereas papain Fab fragments, used in the same dose as the pepsin fragments, possessed only negligible adjuvant activity. As shown the adjuvant activity of pepsin and papain fragments displayed an inverse correlation with the titres and papain homoreactants in the sera of the homologous recipients. The data obtained suggested that the target cells for Fab fragments were lymphocytes carrying cytophilic homoreactants as receptors for the fragments.


[Characteristics of cytotoxic t-lymphocytes eluted from allogenic target cells]. Gain in the specific cytotoxic effect of immune lymphocytes after their absorption on the corresponding target cells (TC) and subsequent elution with pronase was caused not by the increase of the cytotoxic activity of individual cells, but by the quantitative enrichment of the population with the effector T-cells. Eluted and the initial immune lymphocytes failed to differ by the kinetics of absorption on the TC. The eluted lymphocyte fraction was characterized by a two fold increase in the T-cell content and a 4-5-fold increase in the number of DNA-synthesizing cells on account of an increase in the content of medium and large lymphocytes.


[Preparation of liposomes possessing immunologic specificity]. The authors obtained artificial lipid vesicles--liposomes containing immunoglobulins. IgG in the complexes with liposomes proved to retain their immunological activity: the liposomes containing rabbit anti-mouse IgG agglutinated in the presence of donkey anti-rabbit IgG or mouse serum. As shown by the use of liposomes containing H3-inulin and immunoglobulins against the cell surface determinants, these immuno-liposomes selectively bound the target, but not the control cells. Specific binding with the antigenic cell surface determinants was also demonstrated in the case of liposomes bearing the nonimmune globulins besides the immunoglobulins. By the indirect immunofluorescence method it was shown that the nonimmune globulins in complex with the immune liposomes were selectively bound by target cells. A possible use of the immuno-liposomes to deliver various substances selectively to the cells of particular types, and to incorporate new antigens into the cell membrane is discussed.


[Antibodies to an antigen of human thymus epithelial tissue common to the epidermis of the skin in malignant myasthenia]. It has been demonstrated by the indirect immunofluorecent technique that many of the sera of patients with myasthenia gravis react with the anticells of the human thymus epithelial tissue. Sorption of the sera with the suspension of the epidermis cells and the homogenates of the tissues of other human organs showed that the epithelial cell antigen with which the sera of patients with myasthenia reacted were epidermal heteroorganic thymus antigens, i.e. common for the thymus epithelium and skin epidermis. The presence of antibodies to the cells of the epithelial tissue of the thymus in the sera of patients suffering from myasthenia gravis permits to suppose the existence of an immunopathological process against the thymus tissue antigens (including the heteroorganic structures of its epithelium) in this disease.


[2 populations of bone marrow cells transfering the hematopoietic environment]. A bone marrow fragment transplanted under the kidney capsule created a focus of ectopic hemopoiesis, whose isze, measured by the number of hemopoietic cells, was proportional to the implant size. Dimensions of the focus proved to be 11/2--21/2 greater in the irradiated than in the intact recipients. Cells building up the focus of heterotopic hemopoiesis had a different radiosensitivity in the intact and irradiated recipients--their Do constituted about 160 and 350 rad, respectively. In this connection it is supposed that two cell populations of precursors took part in the creation of the focus. Their possible relations with the determined and inducible osteogenic precursor cells are discussed.


[Analysis of different reactive changes in nerve fibers]. A study was made of the morphological peculiarities of the varicose corpuscles revealed in man and animals at the terminal stages of ontogenesis along the peripheral nerve fibers. The properties of these varicose bodies were compared with the reactive varicose deformities experimentally observed under conditions of disturbed blood supply. Similar features of the two formations compared and the presence of transitional forms permit to regard the former ones as the reactive structures. However, more time and participation of the surrounding tissue is required for their formation.


[Effect of antrectomy on gastric secretion induced by insulin hypoglycemia in dogs]. Antrectomy was shown to cause a sharp and stable suppression of the secretory reaction of the fundal glands of the stomach in response to insulin hypoglycemia. Acid and pepsin secretion was reduced two-fold (on the average) as compared to that after histamine stimulation. A tendency to restoration of the secretory parameters was seen in 3 to 5 months; the next 7 months they persisted on a constant level.


[Suppression of antibody formation by bone marrow T-cells activated by histocompatibility antigens]. The suppressive effect of the bone marrow T-cells activated by histocompatibility antigens on antibody formation was studied. The bone marrow of CBA mice was found to contain the thymus-dependent lymphocytes which underwent hyperactivation after the repeated transfers to F1 recipients and acquired the capacity ot inhibit the development of cooperative immune response to sheep erythrocytes. Pretreatment of the bone marrow cells with antithymocyte globulin and complement prevented the suppression.


[Comparative assessment of the properties of a factor inhibiting macrophage migration and interferon]. The authors investigated some functional properties of interferon and the macrophages migration inhibitory factor (MIF) obtained by stimulation of human tonsil lymphocytes with the virus of Newcastle disease (NDV) or O-streptolysin. Both the interferon and MIF inhibited actively the migration of human tonsil cells, but differed by the anti-viral activity, and sensitivity to heating at 56 degrees C for 30 min. Stimulation of human tonsil lymphocytes with NDV leads to production of a more wide range of delayed hypersensitivity mediators than stimulation with O-streptolysin.


[Effect of somatotropic hormone on autoimmune responses induced in mice of different genotypes by syngenous heated erythrocytes]. A single administration of 1 X 10(9) heated erythrocytes to C57BL and BALB/c mice caused on the 13th day the appearance of antierythrocytic autoantibodies, an increase in the weight of the lymphoid organs, and lymphoreticular hyperplasia. These changes were more pronounced in BALB/c mice. During the development of autoimmune reactions the changes in the number of E- and EAC-rosette-forming cells in the thymus and spleen and in the immune response to the sheep erythrocyte immunization and E. coli endotoxin were revealed; distinct strain differences were observed. Daily somatotropic hormone administration (5 mg/kg of body weight) for 10 days decreased the degree of the autoimmune reactions development in mice of both strains. Its action was more expressed in BALB/c mice.


[Release of suppressor factors by the lymphocytes of mice in contact with syngeneic and xenogeneic erythrocytes]. Spleen cells from immune mice incubated in vitro with syngeneic and xenogeneic red blood cells release the factor(s) which possess the immunosuppressive activity. Release of suppressive factor(s) by spleen cells from the non-immunized mice occurs only after the contact with xenogeneic erythrocytes.


[Hybrid resistance to precursor cells of bone marrow stroma]. A focusl of hemopoiesis appearing after the transplantation of a bone marrow fragment of C57BL mice to syngeneic mice (under the kidney capsule) contained more hemopoietic cells than in transplantation to the semisyngeneic (CBA X C57BL) FI recipient. Experiments were conducted with a secondary seeding by intravenous injection of hemopoietic cells of the C57BL transplant genotype into the transplant depopulated by irradiation; it was shown that these differences were caused by lesser dimensions of the hemopoietic microenvironment in the focus in the hybrid organism in comparison with such in the syngeneic system. Thus, the hybrid resistance was expressed not only to the hemopoietic cells, but also to the stromal precursors transferring the hemopoietic microenvironment.


[Proliferative capability of nuclested rat bone marrow cells following freeze--thawing]. A study was made of the proliferative capacity of myelokariocytes of the rat bone marrow after freezing and thawing under protection of the oxyetyl derivative of the tetratomic alcohol; experiments were conducted on mouse-rat radiation chimerae. The rat bone marrow cells proved to retain their proliferative capacity.


Pretreatment cytokinetic studies in 94 children with acute leukemia. Relationship to other variables at diagnosis and to outcome of standard treatment. The relationship of the pretreatment bone marrow mitotic index (MI) and in vitro 3H-thymidine labeling index (LI) to other variables present at diagnosis and to the outcome of standard therapy was examined in a series of 94 children with acute leukemia (71 ALL, 23 AML). The range of observed values at diagnosis was extremely broad, from less than 1% labeled cells to more than 25%. The median LI% for all patients studied was 5.2%; the median MI% was 0.3%. The values for AML and ALL did not differ significantly despite older age, higher initial white blood cell (WBC) counts, and poorer response to therapy of AML patients. The initial MI and LI were positively correlated with each other, but unrelated to age or initial WBC count in either ALL or AML. However, the LI and MI were significantly higher (p less than 0.01) in the group of 10 children of 54 studied with ALL whose blasts formed spontaneous rosettes with sheep red blood cells at 37 degrees C. The initial LI and MI were not shown to be related to the likelihood of attaining a complete response or to the length of remission in either ALL or AML. There was thus no evidence that either the initial MI or LI% of marrow blasts was of any prognostic significance in children with acute leukemia.


Depressed in vitro lymphocyte responses to PHA in patients with Hodgkin disease in continuous long remissions. Twenty consecutive patients with Hodgkin disease in continuous complete remission and off treatment for at least 5 yr (range 5-25 yr, median 9 yr) were studied with a battery of immunologic parameters. Skin test reactivity to four common antigens, sensitization to 2,4-dinitrochlorobenzene, absolute lymphocyte count, relative percentage of T cells (as measured by spontaneous rosette formation with sheep erythrocytes) and B cells (as measured by immunofluorescence with polyvalent antiserum), and absolute number of T and B cells were normal when compared with controls. However, the mean value of lymphocyte response in vitro to the mitogen phytohemagglutinin for the study population was significantly decreased (p less than 0.001) when compared with the controls. This abnormality in response to mitogen could not be correlated with age, sex, stage, symptoms, histologic subclassification, or previous treatment. The data suggest the existence of a persisting cell-mediated immune defect in the circulating lymphocytes in patients with long-standing Hodgkin disease that might otherwise be considered "cured."


[Long-term therapy using horse anti-dog lymphocyte globulin without sensitization against horse protein]. Eight mongrel dogs received a standard daily i.v. infusion of 20 mg/kg b.w. deaggregated horse-anti-dog-lymphocyte-globulin (ALG) and additional prednisolone (1 mg/kg b.w. daily i.v.) over a maximum period of 82 days following pretreatment with deaggregated normal horse IgG. No sensitization against horse protein was observed during therapy of afterwards as proved by lack of humoral antibodies against horse antigens, maintained lymphopenia, good compatibility, longterm prolongation of xenogeneic skin graft survival (85.6+/-20.6 days, n=8' untreated controls 12.5+/-1.3 days, n=4) and longterm suppression of cytotoxic antibodies against donor lymphocytes. The level of preformed agglutinating antibodies against horse erythrocytes was significantly reduced, while preformed antibodies against other species remained normal. The immune response to a challenge injection of anti-lymphocyte-serum (ALS) 6-11 weeks after termination of treatment was significantly lower in the ALG treated animals as compared to the control group. These results suggest the involvement of a specific mechanism of unresponsiveness against ALG other than immunosuppression only. It is concluded, that by the described method sensitization against ALG can be prevented during longterm treatment.


The fate of biogenic monoamines in perfused rabbit lung. 1. Inactivation of beta-phenylethylamine and several of its derivatives was studied in a preparation of rabbit lung perfused with Krebs physiological medium at 37 degrees C. Inactivation or removal of these compounds was calculated as the difference between the concentration of each amine in the perfusion medium and the effluent, collected separately from each lung. The extent of amine metabolic degradation was also measured, by column chromatography, in lung effluent. 2. With this technique the magnitude of amine removal as a function of concentration was determined and an apparent Km and Vmax of removal were calculated for each amine. 3. Percentage removal was highest with phenylethylamine (95%), and decreased, apparently in relation to increasing phenyl- and side chain-hydroxylation (and therefore likely increased hydrophilicity), and 5-hydroxytryptamine (64%), tyramine (53%), octopamine (35%), dopamine (32%) and noradrenaline (23%). 4. Inactivation of each amine could be accounted for by metabolic degradation to deaminated products, which appeared in lung effluent within 90 s of beginning amine perfusion. 5. When intrapulmonary metabolism of phenylethylamine was inhibited by simultaneous perfusion with semicarbazide (10 mM) and pargyline (10 micronM), the removal rate was unaltered, establishing that uptake of the amine from the vascular space is not dependent on metabolism at least for 4 min infusions.


Haloperiodl in the treatment of stuttering. Haloperidol treatment for stuttering was examined in a double-blind cross-over study of 26 adult volunteers with long-standing stuttering. Most had had unsuccessful speech and drug therapy. During the three-month study, their speech and stuttering patterns were repeatedly evaluated from videotaped readings of a standard passage and during spontaneous speaking. Of the 18 patients completing the trial 11 were significantly more improved on haloperidol than on placebo, three benefited equally from both, and four were unchanged. Improvement over placebo was indicated by fewer dysfluencies, increased speed of speaking, and reduced secondary "struggle" phenomena. Side effects were common on a dosage of 3 mg haloperidol daily. Poor concentration, akathisia and dystonic movements caused 8 patients to discontinue the trial despite significant improvement in 5 of them. Although "statistically significant" improvement occurred in most patients on haloperidol, the "clinical significance" of this form of therapy will be limited by the partial response, the need for continuous medication, the side effects of haloperidol and the attitude of stutters to this type of therapy.


Impairment of a motor skill in children with spina bifida cystica and hydrocephalus: an exploratory study. Twenty 7-10 year old children with spina bifida cystica and hydrocephalus and 20 normals matched for age, sex and IQ were compared on a 12-trial target task, first used by Connolly, Brown & Bassett (1968). Analysis of the results, in which particular attention was paid to statistical method and to ways of analysing individual differences, showed a significant impairment in dotting speed in the spina bifida group, although both groups improved with practice. In a second experiment immediately following the first, visual monitoring of this task was restricted. The spina bifida children were initially more affected than the controls but able to recover. The findings are discussed in relation to neurological abnormalities in the spina bifida group.


The effect of frusemide used for post-prostatectomy irrigation on total body potassium. The effects of 2 techniques of bladder irrigation on total body potassium have been compared. The results show that a net decrease was associated with both methods. This was marginally greater (0.1 greater than P greater than 0.05) in the group which underwent a forced diuresis using frusemide with a fluid load. This group received some potassium supplements without which it is possible that the decrease would have been greater. It is concluded that this regime cannot be overtly criticised for its potential loss of potassium.