Normalization of the paradoxic secretion of glucagon in diabetics who were controlled by the artificial beta cell.

Since it is important to elucidate the precise significance of pancreatic A-cell hypersecretion in the pathogenesis of diabetes mellitus, the change in the immunoreactive glucagon (IRG) response to 100 g oral glucose challenges was studied in diabetics whose blood glucose responses and plasma immunoreactive insulin concentrations (IRI) simulated those in normal subjects with the aid of the artificial beta cell system that we developed originally. In six nonobese adult-onset and four insulin-dependent diabetics whose blood glucose responses and plasma insulin concentrations after 100 g oral glucose load were made equivalent to those seen in normal subjects by the artificial beta cell, the glucagon release was similar to the response in normal subjects. In one insulin-dependent diabetic with high anti-insulin-binding capacity, the blood glucose response after an oral glucose challenge was not normalized by the artificial beta cell and the glucagon secretion was paradoxically increased. This fact suggested that the paradoxic rise in glucagon, seen in response to an oral glucose load in some diabetics, is secondary to insulin deficiency.


Insulin and glucagon secretion from rat islets maintained in a tissue culture-perifusion system.

A tissue culture-perifusion system is described that allows for long-term culture of pancreatic islets and study of the dynamics of islet hormone secretion. Islets cultured in this system demonstrate brisk, reproducible biphasic insulin and glucagon release. Glucose-stimulated insulin release is similar after 1 or 14 days in culture. Freshly isolated islets are relatively insensitive to somatostatin, requiring 100 ng/ml to suppress partially the glucose-induced insulin secretion. After 24 h of culture, the same islets demonstrate a marked increase in sensitivity to this hormone. Glucagon secretion from islets maintained in this system occurred in a predictable fashion to arginine stimulation and glucose inhibition.


Antitumor activity of peritoneal exudate cells induced by cell-wall skeleton of Mycobacterium bovis BCG.

Peritoneal exudate cells (PEC) induced by oil-attached cell-wall skeleton of Mycobacterium bovis BCG (BCG-CWS) in ACI/N rats were tested for their effect on both in vivo and in vitro growth of syngeneic fibrosarcoma cells (AMC-60). Treatment of rats with intraperitoneal injections of BCG-CWS induced regression of syngeneic ascites tumor and increased the number of survivals. Whole PEC and adherent PEC from rats injected intraperitoneally with BCG-CWS inhibited the uptake of tritiated thymidine into the fibrosarcoma cells in an in vitro cytostasis test. This in vitro cytostatic effect was more marked as the ratio of effector to target cells increased. In addition, when tumor cells were inoculated subcutaneously with BCG-CWS activated PEC, tumor takes decreased markedly. Oil-stimulated PEC and normal peritoneal resident cells were inactive in inhibition of tumor growth in vivo and in vitro.


Effect of motilin on the rate of gastric emptying and gut hormone release during breakfast.

The effect of an infusion of natural motilin on the rate of gastric emptying of a standard breakfast was studied in 5 subjects using Caesium 129-tagged resin particles. Emptying rates were measured on 4 separate days.  On 2 days, the subjects receive only saline (controls), while on the other 2 days a motilin infusion of 0.34 pmol/kg/min for 60 min, followed by 0.68 pmol/kg/min for 60 min, was given. All infusions were blind and given in random order. At the end of the low-dose motilin infusion 31 +/- 4% of the meal had emptied, compared with 17 +/- 2% with saline infusion (P less than 0.01). A similar effect was also seen with the high-dose infusion. Plasma motilin concentrations rose from a basal of 68 +/- 13 pmol/liter to 126 +/- 10 pmol/liter during low-dose motilin infusion and to 170 +/- 11 pmol/liter during the high dose. No significant change in basal or postprandial levels of glucose, gastrin, pancreatic glucagon, pancreatic polypeptide, gastric inhibitory peptide, enteroglucagon, or vasoactive intestinal peptide was noted, but postprandial insulin release was greater during motilin infusion.


Treatment of chronic portal systemic encephalopathy with bromocriptine: a double-blind controlled trial.

A randomized double-blind clinical comparison of bromocriptine, a new dopamine agonist, and placebo was performed on 7 cirrhotic patients with chronic portal systemic encephalopathy (PSE). Before given either medication, patients were stabilized with a standard treatment (neomycin and cathartics). Serial semiquantitative assessments were done, including mental state, asterixis, number connection test, electroencephalogram, and ammonia blood levels. Three patients developed signs of precoma while ingesting both placebo and bromocriptine. Two patients experienced precoma only with placebo, and another patient only while taking bromocriptine. One patient remained awake throughout the study. All patients responded initially to neomycin and cathartics. Bromocriptine proved not to be significantly superior to placebo and was always inferior to standard treatment. During treatment with bromocriptine, 3 patients experienced constipation. This may be partially responsible for the ineffectiveness in the treatment of PSE.


Prophylactic doxycycline for travelers' diarrhea: results of a prospective double-blind study of Peace Corps volunteers in Morocco.

A second randomized double-blind study to determine the efficacy of doxycycline, 100 mg daily, for the prevention of travelers' diarrhea was carried out among 50 Peace Corps Volunteers during their first 10 wk in Morocco. The volunteers took either doxycycline or placebo for 3 wk, and were observed for an additional 7 wk. Eleven of 24 taking the placebo and 2 of 26 taking doxycycline had travelers' diarrhea during the treatment period (P less than 0.01). One week after cessation of the doxycycline, however, persons in that group developed an increase in frequency of travelers' diarrhea (P less than 0.05) so that by 3 wk after the drug was stopped, there were no differences between groups. Enterotoxigenic E. coli, most of which were sensitive to doxycycline, were the most frequently isolated pathogens during the entire study. This study corroborates the effectiveness of doxycycline prophylaxis for travelers' diarrhea.


An evaluation of some theories of the mechanism of aging.

Two theories of aging are considered in this review. Although there exists substantial experimental evidence in support of the somatic mutation and error catastrophe hypotheses, several experiments have been published which are extremely difficult to reconcile with these models, at least in their simplest forms. These include the observation that biochemical and morphological degenerative changes observed in fibroblasts aged in vitro do not resemble alterations observed in cells obtained from aged donors, and the fact that tissues transplanted serially through different hosts do not decline in vigor in the manner predicted by the somatic mutation theory. Although biochemical and mutational alterations appear to accumulate in fibroblasts aged in vitro (in support of the error catastrophe model), there are substantial problems with the interpretation of such experiments, and some observations (such as the lack of increase in translational error in hemoglboin synthesis as a function of age) seem to argue directly against the error catastrophe theory. Some alternative theoretical and experimental possibilities are discussed, including the concept of programmed aging as the cause of senescence.


Effect of age on the retraction of skin.

The retraction of skin flaps was measured. A constant decrease in retraction of 0.33% per year was found throughout life between the ages of 20 and 70.


Non-invasive techniques in the diagnosis of jaundice--ultrasound and computer.

This study describes the use of ultrasound in 84 consecutive patients to distinguish between medical (intrahepatic) and surgical (extrahepatic) types of jaundice. Accuracy rates in the 84 patients investigated were compared with those found in 169 patients using the computer-aided diagnostic model for jaundice. In 50 patients in whom both techniques were carried out direct comparison was possible. Ultrasound failed to give adequate visualisation for technical reasons in two patients, but 75 of the remaining 82 (91.5%) were correctly separated into medical and surgical categories. A similar percentage (90%, 152 of 169 patients) were correctly classified by the computer-assisted model, and in the 50 patients assessed by both techniques correct diagnostic separation was achieved in 43 (86%) by ultrasound and in 42 (84%) by computer. However, in this latter group two of the errors with computer diagnosis were falsely positive for a surgical jaundice (extrahepatic obstruction), and a surgical exploration performed on the basis of this test could have been unnecessary. This was not found with ultrasound in these same patients, all the errors being false negative. A high degree of confidence can therefore be attached to the demonstration by ultrasound of a dilated biliary tree.


Oral BCG vaccine in Crohn's disease.

In a randomised double-blind trial over one year oral BCG has been compared with a control preparation in the treatment of chronic Crohn's disease. Overall assessment scores deteriorated in nine of 22 patients taking BCG, and 11 of 26 in the control group (P = 0.25);  this deterioration was great enough to be regarded as a clinical relapse in three patients taking BCG and in seven taking the control preparation (P greater than 0.1). No significant benefit from oral BCG treatment has been demonstrated.


Effectiveness of etamsylate in intrauterine-device menorrhagia.

107 parous women wearing Szontágh-type IUDs were treated with either etamsylate or placebo tablets by a double-blind technique with random allocation. The length of the cycles, the duration of menstrual bleeding and the amount of bleeding were analyzed. Etamsylate treatment favourably influenced menstrual bleeding in IUD users. In the etamsylate group, the number of "bleeding days' and the amount of menstrual flow were significantly less than in the placebo group.


Radioimmunoassay of LH-RH. Standard curves and specificity.

As important discrepancies in blood concentrations of LH-RH remain, the basic problem of the specificity of the radioimmunoassay was reinvestigated using an antiserum raised against a new conjugate (33). Cross-reactions with related and unrelated peptides were studied in both charcoal-dextran and second antibody methods of separation. The possible relation between the nature of the conjugate and the specificity is discussed.


Radioimmunoassay of LH-RH. Application to human plasma.

Comparison of standard curves with different concentrations of human plasma in the medium of incubation is presented. It appears that Bo varies with the dilution although the ratio B/Bo is independent of the dilution. This last observation was used to research 'endogenous' LH-RH in four menstrual cycles, in postmenopausal women and in men. The validity of the dosage is discussed in a more general way in the light of the results published in the literature.


[Radioenzymatic assay for catecholamines (author's transl)].

Catecholamines are neurotransmitters produced and secreted by the central and autonomic nervous systems. In addition to being neurotransmitters, amines produced mainly by the adrenal medulla also act as hormones. Fluorometric measurements of amines yield variable results because of the poor sensitivity of the techniques, and the low concentration of these amines in tissues and biological fluids. The lack of specific and sensitive analytical methods has been an obstacle to resolving the mechanism of action of these neurotransmitters and hormones. The possibility of achieving qualitative and quantitative determination of picomole or femtomole amounts of these amines is a major need. Recently, radioenzymatic procedures for catecholamine assay have been developed and there has been a significant improvement in both sensitivity and accuracy of catecholamine assays. In this article, details of these radioenzymatic assay methods are reviewed.


[Effect of alpha-mercaptopropionylglycine (alpha-MPG) and sodium dipropylacetate (DPA) on antibody formation (IV). Tumor immunity (author's transl)].

E.L.4 lymphosarcoma (E.L.4) s.c. as a syngeneic tumor was transplanted into C57BL/6 mice, 4 weeks old. The tumor growth was more rapid in male mice. An i.p. injection of alpha-MPG or DPA caused a decrease of the tumor growth and prolongation of the survival time in male mice. In contrast, an acceleration of the growth and a reduction of the survival time were observed in females dosed with alpha-MPG or DPA. Such a sexual difference and the activity of drugs decreased with the progress of age. alpha-MPG and DPA had little effect on the number of recovered and living E.L.4 cells in vitro. However, with the inoculation of alpha-MPG-treated E.L.4 into mice, the tumor rapidly grew in a dose dependent manner. On the other hand, there was a tendency toward suppression of the tumor growth when DPA was given. Development and growth of tumor in mice induced by 20-methylcholanthrene showed a tendency toward suppression in male mice when alpha-MPG in a dose of 5 mg/kg was given every other day, and in both sexes with the same dose of DPA. Responses of spleen and lymph node cells to phytohemagglutinin-P (PHA) or lipopolysaccharide (LPS) were reduced by the transplantation of E.L.4 in 4 week old mice. With alpha-MPG and DPA a reduction in PHA response of both the cells tended to recover, and that in LPS response of lymph node cells was recovered. In these mice, the decrease in cytotoxicity of spleen and lymph node cells against E.L.4 was recovered by treatment with both drugs. There was also a complement-dependent cytotoxicity against E.L.4 in their sera, and the activity was further increased by the administration of alpha-MPG. These findings suggest that alpha-MPG and DPA are anti-tumor agents which act through immune mechanisms.


Gnotobiotic techniques.

A brief review is given of the development of equipment used for rearing germfree animals in various laboratories and of our experience with designing germfree isolators and with the technology of rearing germfree piglets is presented. The philosophy of isolation as applied to the animal as a biological unit, the elimination of contamination and the theoretical prerequisites of isolation were considered by Reyniers (1943, 1959) and Trexler (1960).


[Experiences with research on pharmacopsychological effects on older people].

During the last years we tested a number of cerebral vasoactive and cerebral metabolic active substances on aged people with the help of the double blind method against with placebo in order to measure the change of the "nutritive insufficiency" with its neurophysiological and vascular-psychological syndromes under pharmacological influence. We used psychological test instruments (groups of 60-80 patients between 50 and 70 years). Duration of the treatment was 3-4 months. Analysis of the dates by way of variance and covariance analysis. Significant results of successive treatments were achieved: improvements of the mental ability as far as the functions of the memory are concerned, higher capacity of attention and concentration (vigilance), fluency and adaptability of cognitive functions, increasing openess of mind and the ability of experiencing, improvement of the general mood and good health. Longer lasting effects in comparison of the drugs were noticed only after treatment with metabolic agents. Publication is planned.


[Urinary bladder plastic enlargement with the use of autologous, homologous and heterologous skin transplants].

1. Expanded urinary bladder resection and the following replacement of the defect with various skin grafts were well tolerated by 161 cats. 2. Grafts of various species were shown to be of significantly different biomechanical properties. This depended on basic substance, compound and texture of collagenous and elastic fibres and the methods of conservation. Preoperatively the transplants were sufficiently resistent against urine and bladder pressure. 3. Within of few weeks postoperatively there was an increase in bladder capacity. This was more evident and arose in an earlier period than in cases without bladder wall replacement (x-ray-control). 4. Autologous grafts were infiltrated by young new tissue. Homologous and heterologous patches served as "guide" of regeneration. Complete transitional cell epithelium was proven to be present after four weeks in the predominantly connective tissue regenerates. The development of smooth muscle fibres in the regenerates is discussed. 5. The microcytotoxicity test showed a host response, but without clinically manifest rejection of the foreign transplants. 6. Apart from clinical urological application the results of this experimental investigation could be useful in general plastic surgery.


[Seasonal variations of testicular and pituitary function in men].

In 21 young, healthy men typical parameters of pituitary and testicular function were recorded over a period of one year in intervals of two months. The mean sperm count in semen had a minimum in december, a maximum in june. Serum-FSH showed low levels in june, october and december, a peak in august. Levels of FSH following stimulation by LH-RH varied in the same manner, but the response to LH-RH was lower in april. A peak of serum-LH appeared in april, other values being rather constant. The response to LH-RH showed parallel variations. Testosterone levels exhibited a significant peak in june. The mean increase of testosterone following hCG depended on basal levels, but showed a distinct peak in december.


[The amnion epithelium in hydramnion. Light- and electron-microsocpy studies].

Human amniotic epithelium was examined in cases of pregnancy complicated with hydramnios; the light- and electronmicroscopical results were compared with those of epithelium in normal pregnancy. The cells of the hydramniotic epithelium show an increased number of microvilli in apical and lateral cell surfaces. There are also a greater number of desmosomes and enlarged intercellular spaces. In a single case where hydramnios was correlated with maternal diabetes mellitus the amniotic epithelium is characterized by local foci showing cellular dystrophy.


Effects of histamine, of histidine and of anti-histaminic agents on the release of glucagon and insulin from the rat pancreas.

We have studied the effect of histamine (HA) and histidine (HIS) on the release of immunoreactive glucagon (IRG) and insulin (IRI) by isolated rat islets and pieces of pancreas. In isolated islets, HA and HIS stimulated IRG release at a glucose concentration of 3.0 mg/ml and IRI release at a glucose concentration of 0.5 mg/ml. In pieces of pancreas incubated in the presence of glucose (3.0 mg/ml), HA at a 2mM concentration stimulated IRG release and had no effect on the release of IRI; however, when the concentration of HA was doubled (4 mM), an inhibition of IRI release could also be demonstrated. HIS was ineffective. Perphenazine and dexchlorpheniramine, two anti-histaminic agents, inhibited IRG and stimulated IRI release. These results indicate that histamine, whether endogenous or exogenous, directly stimulated IRG and inhibits IRI release and suggest that some of the in vivo effects of histamine, such as hyperglycemia and lipid mobilization, may be mediated, at least in part, by these endocrinologic effects.


Prolactin. I. Mechanisms of control, peripheral actions and modification by drugs.

The role of neurotransmitters in the release of prolactin (PRL) is reviewed. Special attention is paid to dopamine (DA) as the possible prolactin-inhibiting factor (PIF). Among other agents, estrogens alone can act directly on the pituitary galactotropes without involving hypothalamic factors. Peripherally, in addition to its stimulatory action on mammary tissue, PRL exerts a permissive role on ovarian steroidogenesis. A possible physiological action of this hormone on the regulation of adrenal function remains uncertain. The secretory rhythms of PRL are described and the mechanisms involved are discussed. A number of drugs can modify the secreting pattern of PRL mainly by acting on the dopaminergic control mechanisms. The usefulness of such agents in the clinical evaluation of galactotrope cell function is reviewed.


Enkephalins and pituitary hormone release: modification of responsiveness to LHRH.

An intraperitoneal injection of leucine-enkephalin into rats stimulates gonadotropin and prolactin release. To elucidate the mechanism of this releasing property of leucine-enkephalin, rat hemipituitaries were incubated with either enkephalin alone or enkephalin in combination with OHRH. Enkephalin alone had no effect on LH or prolactin release in vitro but potentiated the LH response to LHRH. Neither leucine-enkephalin nor LHRH alone had an effect on GH release; however, when combined, a GH response to LHRH occurred. These results suggest that leucine-enkephalin can modify the pituitary responsiveness to certain hypothalamic releasing hormones by a direct pituitary action.


Immunofluorescence studies indicate that the basic trypsin-kallikrein-inhibitor of bovine organs (Trasylol) originates from mast cells.

Using the indirect immunofluorescence technique, the basic kallikrein-trypsin inhibitor of bovine organs, Trasylol, could be localized in tissue mast cells of bovine lung, liver, pancreas and parotid gland. Identification of cells exhibiting specific fluorescence as tissue mast cells was achieved by combined light and electron microscopic diagnosis of bovine liver tissue sections. The presence of Trasylol in mast cells explains the widespread distribution of this inhibitor in functionally totally different organs or tissues of the bovine organism, as determined earlier by biochemical means. Identification of Trasylol as a mast cell constituent will facilitate the search for the biological function of this inhibitory protein in connection with a unique and highly specialized cell population.


Physical map of PM2 DNA.

The cleavage sites of the restriction nucleases HpaI, HpaII, HindII, HindIII, and PstI have been mapped on the DNA of the bacteriophage PM2. This map has been used for the localization of two strong binding sites of Escherichia coli RNA polymerase on PM2 DNA.


Health care delivery in Belgium.

Formerly Belgium's Catholic hospitals prospered within a system based on collectivized financing and individualistic service delivery patterns. Now faced with decreasing vocations, the religious congregations and the national Catholic hospital association are working toward a comprehensive health care delivery system in which laypersons' role is expanded and regional activities are coordinated.


Satellite DNA and heterochromatin variants: the case for unequal mitotic crossing over.

Variations of constitutive heterochromatin (heteromorphisms) appear to be a general feature of eucaryotes. A variety of molecular and cytogenetic evidence supports the hypothesis that heteromorphisms result from unequal double-strand exchanges during mitotic DNA replication. Constitutive heterochromatin consists of highly repeated DNA sequences that are not transcribed. Thus, heteromorphisms are tolerated without overt phenotypic effect. Several of the highly repeated DNAs that comprise constitutive heterochromatin have been shown to contain site-specific endonuclease recognition sequences interspersed at regular intervals dependent upon nucleosome structure. These interspersed short repeated sequences could mediate unequal crossovers, resulting in quantitative variability of constitutive heterochromatin and satellite DNA. De novo variations of constitutive heterochromatin may be useful as markers of exposure to mutagens and/or carcinogens.


In vitro studies of gonadal organogenesis in the presence and absence of H-Y antigen.

In a very strict sense, the primary (gonadal) sex of mammals is determined not so much by the presence or absence of the Y but the expression or nonexpression of the evolutionary extremely conserved plasma membrane H-Y antigen. The central somatic blastema of embryonic indifferent gonads contains one cell lineage characterized by the possession of S-F differentiation antigen that differentiates into testicular Sertoli cells in the presence of H-Y and into ovarian follicular (granulosa) cells in its absence. This cell lineage appears to play the most critical role in gonadal differentiation. Whether or not testicular Leydig cells and ovarian theca cells are similarly derived from the common cell lineage has not been determined. Nevertheless, if given H-Y antigen, presumptive theca-cell precursors of the fetal ovary acquire hCG (LH?)-receptors-the characteristic of fetal Leydig cells.


[In vitro investigations of microphages function in patients with chronic mucocutaneous candidosis (author's transl)].

In five patients with either familial or non-familial type of chronic mucocutaneous candidosis some properties of phagocytic function of the polymorphonuclear leucocytes (PMNL) have been studied in vitro. In each of the patients there were found: a) a decreased chemotactic activity of PMNL, b) a weakness of intake and of intracellular destruction of Candida albicans cells by PMNL, c) an impairment of phagocytosis and intracellular killing of Candida albicans as well of Staphylococcus aureus by PMNL. The rate of phagocytosis of heat-inactivated Candida albicans cells by PMNL was normal in each case. In the serum of two patients a phagocytosis inhibiting factor is supposed to exist. In PMNL of 3 patients a defective activity of NADH-dependent oxidase was found. The occurrence of hereditary CMCC in a father and his two daughters points to an autosomal dominant trait, whereas in most cases of familiar CMCC hitherto described an autosomal recessive mode of transmission was found.


Suppression of the humoral immune response by particulate material extracted from spleen cell subpoulations of MOPC-315 tumor-bearing mice.

Particulate extracts were prepared from MOPC-315 plasmacytoma cells, from whole spleen cells or from splenic macrophages of MOPC-315 tumor-bearing mice. These preparations, when injected into normal mice, inhibited their primary humoral immune response to sheep red blood cells (SRBC). At the same time, particulate extracts from whole spleens or splenic macrophages of normal mice and T cell extracts from spleens of MOPC-315 tumor-bearing mice had no effect on the primary immune response to SRBC. The results suggest that phagocytic spleen cells of tumor-bearing mice contain immunosuppressive particles which originate from the tumor cells.


Modification of kidney immunogenicity with standard Concanavalin A and its subfractions.

Pretreatment of kidneys with purified Concanavalin A (25 mg/L) allowed for a significant prolongation of survival after allotransplantation in minimally immunosuppressed mongrel dogs. Kidneys pretreated with standard Concanavalin A, impure fraction or the lipopolysaccharide had no prolongation of survival. In fact, the impure fraction and the lipopolysaccharide showed a diminished transplant kidney survival. These studied confirm the beneficial effect of Concanavalin A when the purified form is utilized in modifying kidney allograft survival.


The effect of dietary amino acids on immune reactivity.

The plaque-forming cell response and serum antibody titres to sheep red blood cells (SRBC) were enhanced in mice fed diets moderately deficient in the amino acids phenylalanine-tyrosine. These diets sustained normal growth during a 4 week study period. More severe limitation of phenylalanine-tyrosine or multiple essential amino acid restriction gave rise to a slight increas in the delayed hypersensitivity reaction to SRBC. These latter diets usually limit growth; nevertheless the humoral immune response to SRBC was not significantly depressed except in the mice fed a diet (diet 7) with severe restriction of seven essential amino acids. These results suggest that dietary restriction may suppress the production or function of an inhibitory cell, such as a suppressor T cell, whilst not affecting the effector cell to the same degree.


The differentiation of murine thymocytes in vivo and in vitro.

The differentiation of lymphoid cells in the early foetal mouse thymus was studied in vivo and in organ culture. The lymphoid precursors found in the 13 and 14 day foetal thymus constituted about 50% of the total thymus cell population. These T precursors were large blast-like cells which already expressed the Thy-1 antigen but were mostly TL negative. Both in vivo and in vitro the blasts were rapidly replaced by a population of typical small lymphocytes which were strongly Thy-1 and TL positive but Ig negative. In organ cultures grown under optimal conditions, large bimodal increases in cell numbers occurred. An initial population of about 2 x 10(4) T precursors per thymus lobe gave rise to nearly 10(6) predominantly Thy-1 and TL positive small lymphocytes by the 6--8th day in vitro. After this time lymphocyte numbers decreased until about the 10th day when they again increased to form a second peak of small lymphocytes on the 12--13th day. These cells were also predominantly Thy-1 positive but the majority were now TL negative. No Ig positive B lymphocytes were detected either by immunofluorescence or by autoradiography using polyvalent anti-MIg sera and no plasma cells were detected by electronmicroscopy. At all times however, minor subpopulations of Thy-1 negative small cells were present. The production of small lymphocytes during the 1st week of culture was critically dependent on culture conditions and particularly on the batch of FCS used. The population developing during the 2nd week required less stringent conditions and was less dependent on FCS. The culture systems described should prove useful in the study of T-lymphocyte differentiation.


Specific antibody-forming B-lymphocyte colonies. I. Distribution and nature of SRBC antibody-forming B-lymphocyte colonies in mouse lyphomyeloid organs.

Wehn normal mouse spleen or lymph node cells are cultured for 7 days in agar-medium containing 2-mercaptoethanol and sheep red blood cells (SRBC), approximately one B lymphocyte colony (BLC) develops per 50-100 cells seeded. Incubation of cultures for 3 h with guinea-pig complement at day 7, demonstrated that 0.05-0.25% of all BLC form specific antibody against SRBC (SRBC-AF-BLC). The SRBC specific colonies appear centrally in lytic plaques of 2-5 mm in diameter and cells recovered from individual SRBC-AF-BLC were shown to produce antibodies of the IgM class against SRBC. Cells forming SRBC-AF-BLC are absent in the new-born and infantile spleen but appear in adult mice with a frequency of 5, 10-25 and 25-70 per 10(6) bone marrow, spleen and lymph node cells respectively. specific immunization in vivo or in vitro does not greatly affect the number of SRBC-AF-BLC-forming cells. Cytolysis of spleen cells with anti-Ig serum plus complement prior to culture reduced the number of total BLC and that of specific SRBC-AF-BLC by 93% and 94% respectively. The peak sedimentation velocity of both SRBC-AF-BLC-forming cells and total BLC-forming cells was 3.5 mm/h. Spleen cells enriched 200-300 times for cells that bind specifically to the hapten NIP were not enriched for cells forming colonies with specific antibody production against NIP. The data indicate that the cells that give rise to specific antibody-forming colonies belong to a mature virgin B cell group of small Ig-positive B lymphocytes.


Macrophage fatty acid composition and phagocytosis: effect of unsaturation on cellular phagocytic activity.

In order to manipulate the physical properties of the macrophages membrane, methods were developed which potentiated the incorporation of exogenously supplied fatty acids into membrane lipids. Chromatograms of macrophages which were grown in the presence of a variety of fatty acids demonstrated that exogenously supplied unsaturated fatty acids (palmitoleic, oleic, elaidic, linoleic, linolenic and arachidonic acids) were readily incorporated into the cells and selectively altered the fatty acyl composition of macrophage phospholipids. Up to 38% of the total cellular phospholipids were found to be derived from the exogenously added fatty acid supplements. The incorporation of the different fatty acids into cellular phospholipids had striking effects on cellular phagocytic activity. These effects were found to correlate with the degree of unsaturation, and the cis- or trans-double bond configuration. Thus, macrophage phagocytic ingestion rates of 125I-labelled Shigella flexneri were found to alter by more than 2-fold after the cells were cultivated in the presence of cis unsaturated fatty acids.


Time-dependence and selectivity of immunosuppressive agents.

The effects of a variety of agents on the mouse antibody response to sheep red cells (SRC) and Escherichia coli lipopolysaccharide (LPS) were investigated. Agents formed two broad groups according to the time-dependence of their effects. Class I agents inhibited responses if given at any time over a broad period from before administration of antigen to 1-2 days afterwards. They included ionizing radiation, cortisone acetate, cholera toxin and several alkylating agents. Class II agents inhibited responses substantially only if given 1-2 days after the antigen. These agents were all either antimetabolites or vinca alkaloids. Class II agents had only moderate selectivity. When given at the time of maximal effectiveness they usually suppressed the response to SRC more than that to LPS. Some class I agents had striking selectivity. Radiation, cortisone acetate, and cholera toxin markedly suppressed the SRC response but had much less effect on the LPS response. In contrast, mitoclomine and its analogues, which are lipid-soluble alkylating agents, suppressed the response to LPS far more than that to SRC. Possible reasons for these differences in time-dependence and selectivity are discussed.


Mitogenic activity of Mycoplasma pulmonis. I. Stimulation of rat B and T lymphocytes.

The mitogenic activity of Mycoplasma pulmonis towards both rat B and T lymphocytes has been demonstrated. The data summarized in this report show that spleen cells obtained from T X BM rats were extensively activated by M. pulmonis. Furthermore, M. pulmonis has been demonstrated to induce the development of antibody producing cells, as attested by the appearance of direct plaque forming cells against SRBC and TNP-SRBC in spleen cultures exposed to this mitogen. It was also demonstrated that rat thymus cells and a part of the lymph node T-cell population responding to either Con A or PWM, were stimulated by M. pulmonis, the response being weaker than that of rat B-cell populations. It was thus concluded that M. pulmonis activates both rat B and T lymphocytes. This mitogenic stimulation, however, is not equally exerted on both these populations, being strongly effective upon B cells and less so on T cells.


Reduction of phagocytosis, surface hydrophobicity and charge of Salmonella typhimurium 395 MR10 by reaction with secretory IgA (SIgA).

Binding of human colostral secretory IgA (SIgA) to Salmonella typhimurium 395 MR10 decreased the liability to hydrophobic interaction of the bacteria, as analysed by hydrophobic interaction chromatography on Octyl-Sepharose and partition in an aqueous polymer two-phase system consisting of dextran, poly(ethyleneglycol) (PEG) and poly-(ethyleneglycol)-palmitate (P--PEG). SIgA also reduced the negative charge of the bacteria. Treatment of the bacteria with centrifuged but not further fractionated colostrum added positive charge to the bacteria which was removed by treatment with pepsin. Colostral SIgA reduced the in vitro phagocytosis of S. typhimurium MR10 by polymorphonuclear leucocytes. The adhesion of the bacteria to cellulose membrane filters in the absence of phagocytes was also reduced after the interaction with SIgA. It is proposed that the binding of SIgA to bacterial surfaces has hydrophilic and anti-adhesive effects, which may serve to exclude antigen from mucosal surfaces.


Lymphocyte binding of fluid phase mouse C3b.

Isolated fluid phase mouse C3b adhered to human and to mouse lymphocytes. On human, but not on mouse cells it could be stained by fluoresceinated F(ab')2 anti-mouse C3. Binding to mouse cells was, however, shown by inhibition of C3-dependent rosette formation and by direct staining with fluorescein-conjugated C3b (FITC-mouse C3b). Optimal staining of lymphocytes by FITC-mouse C3b depended on a sufficient intensity of fluorochrome conjugation and on a degree of aggregation of the C3b. FITC-mouse C3b preparations which initially stained weakly, stained strongly after being aggregated with glutaraldehyde. The failure of immunofluorescent techniques to demonstrate binding by mouse lymphocytes of mouse C3b which had not been so aggregated was apparently due to inaccessibility to anti-C3 of the receptor-bound C3b. Aggregated mouse C3b-FITC induced sequential patching and capping of lymphocyte complement receptors followed by endocytosis, all of which was inhibited completely by cold (4 degrees), sodium azide (2 x 10(-3) m), cytochalasin B (28 microgram/ml) and chlorpromazine (10(-4) m), and partially by lignocaine (2 x 10(-3) m), and colchicine (10(-4) m). That mouse C3 which has interacted with mouse complement receptors may not be demonstrable by anti-C3 antibody may have important implications for immunohistochemical localization of C3 bound in vivo to lymphoid cells.


Comparison of the direct antiglobulin rosetting reaction (DARR) and direct immunofluorescence (DIF) for demonstration of sIg-bearing lymphocytes in pigs, sheep and cattle.

Tests with untreated and trypsin-treated red cells (rbc) from a variety of species showed that anti-Ig-coupled pig RBC are good indicator cells for the study of ruminant blood sIg + lymphocytes by the DARR test; coupled donkey and rabbit RBC are suitable for investigating pig lymphocytes. The different species showed the following percentages of sIg + lymphocytes (M +/- SE) by direct immunofluorescence (DIF) and the direct antiglobulin rosetting reaction (DARR) respectively:pigs 9.2 +/- 0.7% and 16.3 +/- 1.2%; sheep 20.2 +/- 1.2% and 33.1 +/- 1.6%; Cattle 13.5 +/- 1.4% and 28.9 +/- 3.5%. The mean ratio of sIg + lymphocytes shown by the two tests (DARR/DIF) for each species was 1.80 +/- 0.08 for pigs, 1.73 +/- 0.7 for sheep and 2.15 +/- 0.18 for cattle. Preincubation of pig and sheep lymphocytes at 37 degrees for 1 h did not alter the proportion of sIg + lymphocytes detected by either test. Thus the DARR test reveals a further population of sIg + lymphocytes in addition to that detected by immunofluorescence, whose number is proportional to the B population as measured by DIF and whose sIg is intimately associated with the membrane.


Development of delayed hypersensitivity responses in Mycobacterium lepraemurium infections in resistant and susceptible strains of mice.

C57Bl mice are relatively resistant to a moderate subcutaneous infection with Mycobacterium lepraemurium while BALB/c mice are much more susceptible. Cutaneous delayed hypersensitivity reactions which develop in the first 3 weeks of infection were compared in these two strains of mice. Both strains gave a peak of delayed hypersensitivity between 6 and 10 days after infection which was followed by a period of low reactivity before the development, in the third week, of a stable persistent delayed hypersensitivity reaction. There was no difference between the strains in the size at 24 h of the delayed hypersensitivity reaction but the reactions differed in their kinetics. The low resistance strain, BALB/c, gave a Jones-Mote-type of response while the high resistance strain gave a response which could be described as a tuberculin-type reaction.


A sensitive technique for measuring specific macrophage aggregation: a comparison with macrophage migration inhibition, for the detection of lymphokine activity.

A new quantitative method for the measurement of macrophage aggregation, induced by lymphokine preparations, is described. This involves the continuous measurement of the light absorbance of stirred macrophage suspensions. The results have been compared with those obtained by measuring macrophage migration inhibition activity for the detection of lymphokine activity. Separation of crude lymphocyte culture supernatants by Sephadex G-200 shows that the material with aggregating activity is heterogeneous. The greater amount of this activity is not separated from macrophage migration inhibition activity, and has a molecular weight of 35,000--70,000. A comparison has been made with other published methods for measuring macrophage aggregation.


Immunosuppression in murine malaria. III. Induction of tolerance and of immunological memory by soluble bovine serum albumin.

The primary antibody response to alumadsorbed BSA was depressed when initiated during low-grade chronic Plasmodium berghei malaria in mice, as previously reported during acute P.y. yoelii infection. Induction of immunological memory by soluble polymerized BSA was abolished in both infections; in infected hosts this normally immunogenic stimulus resulted in partial tolerance. In contrast to the depression of immune response, neither infection interfered with the induction of low-zone tolerance by monomeric BSA. The rate of non-immune elimination of BSA was found to be normal during acute malaria, and only slightly reduced in chronic infection. These results may be explained in terms of abnormal antigen handling in infected mice, due to some functional defect in macrophages, although this does not seem to be a sufficient explanation for all the phenomena of malaria-associated immunosuppression.


Peritoneal exudate T lymphocytes with specificity to sheep red blood cells. I. Production and characterization as to function and phenotype.

T lymphocytes which mediate DTH reactions to sheep red blood cells (SRBC) in mice enter casein-induced peritoneal exudates from which they can be recovered and assayed in a passive transfer system. Peritoneal exudates need not contain specific antigen for inducement of T-cell immigration. The amount (or biological activity) of DTH-transferring peritoneal exudate lymphocytes is enhanced by the previous use of immune modulating agents, such as cyclophosphamide (Cy) (200 mg/kg 2 days prior to sensitization), or BCG (10(7) live organisms i.v. 14 days prior to sensitization). SRBC-specific peritoneal exudate lymphocytes phenotypically are Thy 1+ and Ly 1+, 2-. In vivo, peritoneal exudate T cells from Cymodulated donors persist in circulation for a short period only and are subject to the suppressive mechanisms acting in anergic mice. Cells from BCG-plus-Cy-modulated donors, on the other hand, persist in circulation for a longer period and appear to be less susceptible to immune suppression.


Subpopulations of Ig-secreting cells induced by peroxidase immunization: discrimination according to antibody storage and secretion.

Mice were injected in their hind footpads with peroxidase (PO) emulsified in Freund's complete adjuvant. The development of cells secreting anti-peroxidase antibody (Ab) and cells secreting immunoglobulins (Ig) were detected in the draining popliteal lymph nodes in the subsequent 35 days, using local haemolysis plaque assay with sheep red cell blood cells coated with either PO or anti-mouse Ig antibody. Plaque-forming cells (PFC) were isolated from the centre of plaques by micromanipulation and after appropriate treatment, were examined by electron microscopy for their intracellular Ab content and in corporation of [3H]-thymidine. Four subpopulations of Ig secreting cells were distinguished: (1) cells secreting Ig without Ab function and not containing intracellular Ab detectable between days 5 and 20; (2) cells secreting Ig without Ab function but containing Ab appearing on day 6 and present throughout the immune response; (3) cells secreting Ab and containing Ab; (4) cells secreting Ab, but without detectable intracellular Ab. These last subpopulations appeared on day 7 and were found in all subsequent assays. The analysis of the kinetics of these subpopulations suggest that cells secreting Ig without Ab function might be precursors of Ab secreting cells.


Induction of autoimmunity to antigens of the glomerular basement membrane in inbred Brown-Norway rats.

Induction of autoimmune antibodies against antigens of glomerular basement membrane (GBM) was studied in nine inbred strains of rats each with a different major histocompatibility complex H-1. Brown-Norway (BN) (H-1n), Lewis (H-1(1)), PVG/c (H-1c), AS2 (H-1f), AVN (H-1a), BD V (H-1d), DA (H-1a) and F344 (H-1(1)) rats were immunized with bovine GBM and Freund's complete adjuvent (CFA). A pronounced linear deposition of host IgG (IgG1 and IgG2a) along the GBM was found in BN rats. No deposition of C3 could be detected in the glomeruli nor did the animals develop proteinuria. The quantity of autoimmune antibodies fixed to the GBM was low (48 microgram +/- 14) which could explain the absence of C3 deposition and proteinuria. The antigenic specificity of the antibodies deposited along the GBM in BN rats was shown by the fixation in vitro of the eluted antibodies to the GBM and tubular basement membrane (TBM) of normal kidneys. A much weaker and irregular deposition of host IgG along the GBM was observed in PVG/c, AS2. AVN, BD V, DA and F344 rats. Of these strains, eluates from the glomeruli of PVG/c, AVN, BD V and DA rats fixed very weakly to the GBM of normal kidneys whereas eluates from AS2 and F344 rats did not fix to GBM or TBM. No deposition of host IgG was found in Lewis rats, and the eluates did not fix to normal kidneys. Congenic L.BN rats with the BN H-1n haplotype and a Lewis background did not respond. This study shows a genetic predisposition in rats to an autoimmune anti-GBM response which is not, or not exclusively, controlled by genes linked to the H-1 histo-compatibility complex.


Development of antigen-induced proliferative responsiveness by murine lymph node cells. I. Identification of differences in the in vitro proliferative responses during a first and a second period of responsiveness.

The development and course of antigen-induced proliferative responsiveness by murine lymph node cells (LNC) was observed for 16 weeks post-immunization. The initial phase of responsiveness was characterized by antigen-induced proliferative responsiveness in vitro which reached a maximum 3-5 weeks post-immunization and then declined to low levels by 6-8 weeks. Without injection of additional antigen, the initial phase of responsiveness was followed by the development of a second phase of antigen-induced proliferative responsiveness 10-12 weeks post-immunization. These findings suggest that the in vivo development of lymph node lymphocytes capable of a proliferative response to antigen is under some type of modulation which is maximal 6-8 weeks post-immunization. Early in the first phase the proliferative responses to higher concentrations of antigen peaked early in the culture period (days 3-4), whereas responses to the lower concentrations of antigen were optimal after 5-6 days of culture. During the latter half of the first phase, however, peak proliferative responses were made to all the concentrations of antigen on the same day of culture (day 6). In contrast, the responses detected at the beginning and throughout the second phase of responsiveness were characterized by maximum proliferation to all the concentrations of antigen late in the culture period (day 7). These results delineate the temporal requirements for maturation of antigen-induced proliferative responsiveness of murine LNC post-immunization and indicate the time interval when optimal responses may be detected.


The distribution of 2,4-dinitrophenyl groups in lymphoid tissue of guinea-pigs following skin painting with 2,4-dinitrochlorobenzene.

Cellular localization of 2,4-dinitrophenyl (DNP) groups in the peripheral lymphoid system of guinea-pigs was studied at various times after painting the skin with 2,4-dinitrochlorobenzene (DNCB) by the immunofluorescent method using anti-DNP antibody. The cells taking up the stain (DNP cells) were shown to be mainly lymphocytes. At 1-6 h after painting the majority of DNP cells were found in the peripheral blood and the spleen but the maximum number was reached in the lymph node draining the site of DNCB application 12 h after painting. Injecting cyclophosphamide (CY) 3 days before painting with DNCB, heightened the number of DNP cells residing in the draining node. The animals treated with the tolerogen, 2,4-dinitrobenzene sulphonic acid sodium salt (DNBSO3Na), and then painted with DNCB, had fewer DNP cells than those animals which had simply been painted once with DNCB. The culture supernatants prepared from the draining nodes of both normal and tolerant animals partially blocked the anti-DNP antibody binding with DNP cells. It is suggested that the cells associated with DNP groups residing in the draining lymph node act as immunogens in the immunizing process of contact sensitivity.


In vitro production and cellular origin of murine type II interferon.

Antigen-specific type II interferon was produced in vitro by harvesting supernatants of spleen cell cultures from Swiss-Webster mice sensitized with Mycobacterium bovis strain BCG and challenged with old tuberculin. Treatment of C3H mouse spleen cell cultures with appropriate anti-Ia, anti-IgG, anti-Thy-1 or anti-Ly-2,3 sera resulted in a significant decrease in production of type II interferon. Removal of nylon wool adherent cells or cells with histamine receptors by column chromatography similarly caused reduced production of type II interferon. Recombination of spleen cell cultures treated with anti-Ia and anti-Thy-1 sera or of cells treated with anti-IgG and anti-Thy-1 resulted in restored production of type II interferon. Interferon production was also restored by combination of cells passed through histamine columns with anti-Ia treated cells, or those passed through nylon wool columns with anti-Thy-1 treated cells. Anti-Ly-1 serum treatment had no effect on interferon production. Removal of plastic-adherent cells or cells that had phagocytosed carbonyl iron also decreased interferon production, suggesting that macrophages were also involved in type II interferon production. Recombination of non-adherent spleen cells with anti-Ia and anti-Thy-1 sera treated spleen cells, however, did not restore interferon production, suggesting that other cells in addition to macrophages are depleted by the adherence procedure. These findings indicate that type II interferon is produced by suppressor or cytotoxic (Ly-2,3+) T lymphocytes in co-operation with one or two additional cell types: (i) B lymphocytes, and (ii) macrophages.


Ontogeny of B-lymphocyte function. IX. Difference in the time of maturation of the capacity of B lymphocytes from foetal and neonatal mice to produce a heterogeneous antibody response to thymic-dependent and thymic-independent antigens.

The ontogeny of the capacity of the B-lymphocyte population to produce a response which is heterogeneous with respect to antibody affinity was studied in a cell transfer system. Lethally irradiated mice were reconstituted with B cells from donors of various ages, together with adult thymus cells when the response to T-dependent antigens was studied. The animals were immunized with one of a variety of antigens one day after cell transfer and the distribution of their splenic plaque-forming cells (PFC) with respect to affinity was assayed, by hapten inhibition of plaque formation, 2 to 3 weeks after immunization. Mice reconstituted with B cells from neonatal donors produced a response of low affinity and restricted heterogeneity. With four different thymic-dependent antigens (DNP-BGG, F-BGG, DNP-KLH and Dan-KLH) the splenic B-cell population acquired the capacity to reconstitute irradiated mice to produce a normal adult-like, highly heterogeneous, high affinity PFC response between 7 and 10 days after birth. The capacity to produce a heterogeneous response to the thymic-dependent protein antigen BGG matured just slightly later between 10 and 14 days of age. The bone marrow matures with regard to the capacity to reconstitute irradiated mice to give a heterogeneous response several days after the spleen, possibly as a consequence of the redistribution of peripheral B cells to the bone marrow. In contrast, maturation of the capacity of the splenic B-cell population to reconstitute irradiated recipients to give a heterogeneous, adult-like PFC response to three 'thymic-independent' antigens (TNP-PA, DNP-Ficoll and TNP-BA) takes place considerably later (between 3 and 4 weeks of age). These results suggest that the population of B-cell precursors which responds to thymic-dependent antigens may represent a different subpopulation of B cells from the population that responds to thymic independent antigens. Furthermore, the results suggest that these B-cell subsets mature at different times, presumably under independent controls.


Effect of in vitro treatment with reducing drugs on structure and function of human secretory immunoglobulin A.

Samples of unstimulated whole saliva from 15 healthy children with 0.5--6 mg/100 ml of secretory IgA and from 10 healthy adults with 4--18 mg/100 ml of secretory IgA were pooled and treated in vitro with dithiothreitol and alpha-mercaptopropionylglycine. The effect of these reducing drugs on the immunochemical properties of secretory IgA was evaluated. Dithiothreitol induced depolymerization of secretory IgA and splitting of the secretory piece from the IgA molecule; furthermore it strongly reduced the titer of secretory antibodies to Escherichia coli antigens. The drug alpha-mercaptopropionylglycine apparently did not affect either the polymeric structure of secretory IgA or the titer of secretory anti-E. coli antibodies; however it induced splitting of the secretory piece. On the whole it appers that drugs with reducing properties, currently employed for liquifying mucous secretions in clinical practice, should be carefully evaluated for possible depressive side-effects on local immunity.


Antibacterial activity of co-trimazine in vitro and in vivo.

Co-trimazine is a new drug combination especially designed for the treatment of urinary tract infections. It consists of trimethoprim (90 mg) and sulphadiazine (410 mg). When combined in vitro, the components show high activity and a high frequency of synergy against urinary tract pathogens. After oral absorption sulphadiazine has a serum half-life similar to that of trimethoprim and is excreted in active form into the urine to a much higher degree than sulphamethoxazole. The ratio of the concentrations of trimethoprim and sulphadiazine in the urine following co-trimazine is favourable for a strong synergistic action between the compounds. In cross-over studies in volunteers receiving repeated daily doses of co-trimazine, either 500 mg twice daily or 1000 mg once daily, it was found that antibacterial activity in the urine was at least as high as that provided by co-trimoxazole (2 x 960 mg) and considerably higher and more uniform than that given by nitrofurantion (3 x 50 mg).


[Clinical efficacy, nephrotoxicity and ototoxicity of amikacin].

The clinical efficacy and toleration of amikacin was investigated in 22 patients most of whom had chronic urinary tract infections that had already been treated unsuccessfully on several occasions with other antibiotics. Amikacin was administered i.m. in a dosage of 7.5 mg per kg twice daily for an average of 11.4 days. This new aminoglycoside antibiotic proved highly effective in the treatment of chronic pyelonephritis and cystitis, as well as in septicaemia caused by gentamicin-resistant Pseudomonas. The pharmacokinetic studies did not show any retention after a ten day treatment with amikacin. Thorough nephrologic and otologic investigations for side-effects did not show any permanent nephrotoxic or ototoxic damage or delayed damage. The results were compared with the clinical and experimental data in the literature and with the results of experimental studies which have not yet been published.


[Typing of Escherichia coli O and K antigens in patients with urinary tract infections (author's transl)].

In a long-term investigation in children and adult patients with urinary tract infection, 1403 Escherichia coli strains were serotyped and 266 strains were subjected to serological investigation. The most frequently occurring O groups were O6, O8, O2, O18, O4, O75, O1, O22, O7 and O9. L type antigens were predominant among the K antigens. Infants in whom pyelonephritis manifested itself for the first time exhibited an above average rate of infection with O2, O4 and O75. A reinfection rate of 84 to 91% was observed in children in follow-up investigations.


[Erythromycin determination in organ tissues (author's transl)].

The recovery rates of erythromycin after in vitro and in vivo administration were studied comparatively in liver, lung, and kidney. Using buffer standards the recovery rate in homogenates of in vitro administered erythromycin decreased with increasing protein concentration. At constant protein concentration the erythromycin administered recovered correlated linearly. Mincing the tissues with scissors was not found to be adequate, even when followed by a diffusion period of up to 24 hours. Similarly, shock freezing resulted in lower values. Even and optimal breaking down of tissues was found after homogenisation or ultrasonics. The recovery rates were not raised by treating the tissue with trypsin. There was no evidence of enzymatical erythromycin degradation in liver homogenates under the assay conditions.


Decomposition of a mixture into normal components: a review.

The problem of estimating the parameters of a finite mixture of normal distributions has been considered by several researchers in different fields of application. This paper consists of a brief historical background and a comprehensive account of the theoretical aspects behind the principal lines of approach of different techniques of estimation. These include moment- and maximum likelihood estimation, estimation by minimum variance decomposition and graphical techniques. Further, three different methods of estimation are compared on medical data.


The metabolic activation of benzo(a)pyrene and 9-hydroxybenzo(a)pyrene by liver microsomal fractions.

A rat liver microsome-mediated bacterial mutagenicity test showed 9-hyroxybenzo(a)pyrene to be significantly more effective as a pre-mutagen than benzo(a)pyrene. Experiments measuring the ability of these compounds to be metabolically activated to moieties that alkylate exogenous DNA demonstrated that 9-hydroxybenzo(a)pyrene was almost six times more effective than benzo(a)pyrene itself. Addition of trichloropropene-2,3-oxide to the reaction mixture enhanced the mutagenicity and DNA alkylation by benzo(a)pyrene but had little or no effect on the 9-hydroxybenzo(a)pyrene-mediated mutagenicity and alkylation. On the other hand, 7,8-benzoflavone inhibited the microsome-mediated mutagenicity and DNA alkylating activity of both hydrocarbons.


An immunoregulatory factor associated with spleen cells from tumor-bearing animals. III. Characterization of the factor's target cells.

An immunoregulatory factor associated with spleen cells from tumor-bearing mice was found to potentiate the generation of antibody-producing cells (APC). In an attempt to characterize the target cell of this enhancing factor (EF), its activity in mice devoid of mature T lymphocytes was tested. Levels of anti-SRBC APC were augmented when EF was injected together with SRBC to nude mice and "B" mice. In addition, EF potentiated the antibody response against the IgM-inducing T-independent pneumococcal plysacchardide SIII antigen. These results suggest that EF most probably exerts its enhancing influence directly on B lymphocytes. In a different line of experiments adoptive secondary responses were performed. Mice were immunized with SRBC as carrier or with the NIP-chicken erythrocytes (as a source of NIP-specific primed B cells) in the presence or absence of EF. Various combinations of spleen cells from the donor immunized mice were transferred in a mixture with NIP-SRBC to lethally irradiated recipient mice, EF did not exert any potentiation effect on helper function, except when primed B cells were used. In contrast, a clear activation or clone expansion of hapten-specific B cells was observed. These findings indicate that the enhancing factor from tumor-bearing animals directly affected the antigenic triggering of B lymphocytes and their subsequent proliferation and differentiation to antibody-producing cells.


Expression of Fv-4r allele in hematopoietic cells from G mice resistant to Friend leukemia virus.

G mice carrying the Fv-4r resistant allele supported virus growth neither at an early nor a later (Kai et al., 1976) stage of infection with NB-tropic FLV. This resistance could not be abolished by treatment of G mice with cyclophosphamide or cortisone acetate. By bone-marrow or spleen-cell transplantation into irradiated mice, the resistan-e of G mice could be transferred to Fv-4-susceptible mice. Conversely, transfer of bone-marrow or spleen cells of Fv-4-susceptible mice rendered G mice susceptible. It could be concluded that, as assessed by the virus content in the spleen, helper LLV grows mainly in radiosensitive, bone-marrow-derived cells, and the Fv-4 gene is expressed in these cells.


Preserved foods as possible cancer hazards: WA rats fed salted fish have mutagenic urine.

Six batches of food traditionally and commonly consumed by southern Chinese, including two samples of dried shrimps and four samples of different species of salted fish, were tested for mutagenic properties using Salmonella typhimurium TA98 and TA1000. Mutagenic activities toward both tester strains were found in all preparations. In most cases, these activities were enhanced by liver microsomal activation. Urine collected from experimental rats regularly fed salted fish also showed mutagenic activity. The level of this activity decreased markedly when the experimental rats were transferred from a salted fish diet to Purina rat chow. Our data suggested the presence of mutagenic/carcinogenic substances in some local preserved foods and at least one of them, salted fish, has been suspected on epidemiological and experimental evidence to be a possible co-carcinogenic factor in the development of nasopharyngeal carcinoma in southern Chinese. Our finding is compatible with this hypothesis.


Untangling the logics of urban service bureaucracies: the strange case of the San Francisco Municipal Railway.

Marxist analysts have developed theoretical propositions concerning the conflicting functions of government in advanced capitalist countries, while public policy analysts have examined the logic of public resource allocation. Through an historical analysis of a typical urban bureaucracy, the San Francisco Municipal Railway, this paper describes how bureaucratic structure and functioning relate to broader conflicts over whom government actions should benefit. It concludes that the apparent shortcomings of the organization can be traced to, and were caused by, this conflict. The argument thus draws from each of these two quite different approaches to the question of what logic motivates the public sector.


What's inside the black box: a case study of allocative politics in the Hill-Burton program.

This paper explores the political and bureaucratic determinants of grant allocations within the Hospital Survey and Construction Act of 1946 (the Hill-Burton Act), which established the major hospital construction subsidy program in the United States. Resource allocation within the Hill-Burton Program was neither purely a function of external political forces impinging on the state nor purely a result of internal organizational factors. Instead, as shown in the analysis which follows, external political forces shaped program structure, operating routines, and rules and regulations at the time of the Program's formation. Rules and regulations depoliticized the grant allocation process while institutionalizing the interests of hospitals and other producers into the program structure. Consumer interests and influence are excluded from program decision making. Once established, the rules and regulations and a set of bureaucratic behaviors play a critical role in determining resource allocation. Rules and regulations facilitate distributing divisible benefits to producer interests in a politically predi-table way. Yet rules and regulations must also regulate the supply of hospital beds in accordance with local and national market conditions. The mode of operations required for distributing benefits in a politically predictable way limited the Program's ability to regulate hospital bed supply. This was a critical factor which led the Hill-Burton Program to subsidize excess capacity in the U.S. hospital system and is a useful way of explaning many similar dysfunctions within allocative programs such as Hill-Burton.


Peyronie's disease revisited.

Various treatment modalities have been suggested for Peyronie's disease. Varying degrees of success have been reported. Certain numbers of cases will have spontaneous remission of the disease process. The treatment, however, should begin as early as possible and recognition should be achieved earlier. The injection technique utilizing a high-pressure metal syringe or syringe jacket with screw-type barrell is still the most successful for the "soft fibrosis" lesions. This was reported about twenty-five years ago. A small number of patients treated by plaque excision and full thickness skin graft replacement have also shown good results. The type of lesion present best dictates the proper treatment.


A new technique for isolation of Descemet's membrane: preliminary studies.

The structure and function of basement membranes have been the subject of extensive investigation. The present study takes advantages of a new experimental procedure to yield ultrastructurally pure basement membranes and applies this methodology to Descemet's membrane, a highly specialized ocular basement membrane. Rabbit Descemet's membranes and associated endothelial cells were mechanically isolated without contaminating stromal elements. The endothelial cells were then solubilized and removed by treatment with detergents as verified by light microscopy and scanning and transmission electron microscopy. The Descemet's membrane remained intact and retained its fibrillar fine structure. Therefore purity of starting material for ongoing morphological and biochemical studies of isolated Descemet's membranes is demonstrated. These investigations will provide a valuable data base for comparison with disease-altered Descemet's membranes.


[Treatment of psoriasis using vitamin A, vitamin A acid and oral retinoids].

Vitamin A, Vitamin A acid (retinoic acid) and their synthetic derivatives were variously applied in the management of psoriasis with different therapeutic results. Obviously, they influence the proliferation rate and the differentiation of human keratinizing epithelia and have beneficial effects on skin diseases with disturbances of keratinization, including psoriasis. Systemic application of Vitamin A has been yet largely abandonned since high dosages leading to clearing develop evident systemic toxicity. The anti-psoriatic effect of Vitamin A acid is either moderate or restricted, because of side effects. Only its combined local application with topical corticosteroids may be considered. Oral application of newly synthesized retinoids, however, was beneficial in psoriasis, particularly in erythrodermic or pustular types. With this group of retinoids new pathways were opened in dermatotherapy which may help to replace cytostatic drugs in these cases. Additionally, oral retinoid treatment may be introduced as an adjuvans in the management of widespread psoriasis, in order to enhance the effect of anthralin, PUVA or UVB treatments.


[Swellings of the cheek from sialosis (author's transl)].

The term sialosis was introduced by Rauch (1956) and was associated with a symmetrical, painless, recurrent enlargement of the salivary glands. Included among the sialoses were non-inflammatory and inflammatory diseases of the salivary glands. A more recent definition still in use today was made by Seifert (1960) and includes disturbances in metabolism and secretion of the parotid gland. The term sialosis was also recommended by the World Health Organization (Thackray and Sobin, 1972) to replace the descriptive term "asymptomatic salivary enlargement." In addition to inflammatory diseases and tumors of the salivary glands, sialosis is the most common disease of salivary tissue recorded at the Institute for Pathology, University of Hamburg. In 50% of patients with sialosis, a syntropic occurrence of other diseases can be found which can be further classified as endocrine, dystrophic-metabolic, and neurogenic sialoses. All clinical types of sialosis are represented by only one morphological pattern: enlarged acinar cells with either a granular pattern, a numerical increase in secretory granules or a vacuolar transformation of the cytoplasm; degenerative changes are seen in myoepithelial cells and the autonomic nervous system. Three types of sialosis can be further distinguished by the electron density of the acinar granules. These are granular, vesicular or mixed. However, there is no correlation between the clinical and morphological forms of sialosis. Experimental destruction of the sympathetic nervous system results in tissue changes which are similar to sialosis. These ultrastructural alterations are interpreted as a disturbance of secretion, and suggest that degeneration of the autonomic nervous system is the common pathogenic principle in all types of sialosis.


[Differential diagnosis of facial skin swellings (author's transl)].

Enlargement of the cheeks may be due to a multitude of disorders, congenital, neoplastic, and in particular inflammatory. Congenital facial anomalies include cutaneous (and osseous) hemihypertrophy of the face and unilateral angiomatous malformations (e.g. Sturge-Weber-Krabbe Syndrome). Buccal enlargement due to dermal tumours include localized haemangiomas and lymphangiomas, lipomas and other benign connective tissue neoplasms, generalized disorders of the lymphatic or reticuloendothelial system including mycosis fungoides, reticulum cell sarcoma and other soft tissue malignancies, and cutaneous manifestations of malignant haemoblastoses, in particular chronic lymphatic leukaemia. Within the very large group of inflammatory skin swellings of the face a review is made of some bacterial pyodermias, severe forms of acne vulgaris, herpes zoster, lupus vulgaris, erysipelas, rosacea, steroid dermatitis, lupus erythematosus (discoid and systemic), toxic dermatitis, allergic eczema, urticaria, Quincke's oedema, and the Melkersson-Rosenthal syndrome. The importance of prevention and early detection of steroid-induced dermatitis is emphasized. This disorder, which is a pseudo-inflammatory disfiguring complication of prolonged topical steroid abuse, ranks in frequency with the skin problems most often seen in dermatological practice.


[Differential diagnosis of salivary gland swellings (excluding neoplasms)--the ORL view (author's transl)].

Important investigatory methods for the diagnosis of non-neoplastic salivary gland disease are discussed. The difficulties found with these different methods are emphasized, and biopsy is recommended as the most reliable means for diagnosis. Some salivary gland disorders are caused by hypotensive drugs. Chronic recurrent parotitis is presented as an example of the difficulties in differential diagnosis and treatment.


Clinical response of dairy cattle with ovarian cysts to single or repeated treatments of gonadotropin-releasing hormone.

Two hundred twenty-five dairy cows with ovarian cysts were treated with 100 microgram of gonadotropin-releasing hormone (GnRH). Cows failing to recover were retreated with a second or third dose of GnRH. Clinical recovery following 1st, 2nd, and 3rd treatments was 76% (170/225), 78% (43/55), and 66% (8/12), respectively. Of the 221 cows that recovered, 184 eventually became pregnant. The remaining 41 cows were culled primarily as problem breeders. First-service conception rate was 49%. More ovarian cysts were detected in herds examined at 2-week intervals than in herds examined at 4-week intervals. The increased diagnosis of ovarian cysts may have been related to detection of cysts prior to spontaneous recovery that occurs in some cases. Clinical recovery from ovarian cysts was lower in herds examined at 2-week intervals following treatment, compared with those examined at 4-week intervals. The GnRH-induced luteinization of ovarian cysts and response to treatment was more difficult to judge at 2 weeks than at 4 weeks.


Tadini, the man who invented the artificial lens.

Tadini, an Italian by birth, was an itinerant ophthalmologist living in the second half of the eighteenth century. The eye operations which he performed at many places throughout Europe are documented by his advertisements in newspapers of the day. It is likely that Tadini was the first to conceive the possibility for intraocular correction of aphakia. At any rate, Casanova's memoirs record a meeting with Tadini and make mention of the oculist's box of artificial glass lenses for placement in the eye. Presumably, Casanova conveyed Tadini's idea to Casaamata, an ophthalmic surgeon in Dresden who was the first to actually attempt correction of aphakia with artificial lens implantation.


Complications of implant surgery in children.

Rapid recognition and appropriate therapy will prevent or remediate most of the complications occurring in children after cataract aspiration and IOL implantation. Some of the complications of pediatric IOL implantations are related to the increased scleral pliability and decreased rigidity which predispose scleral collapse, vitreous loss, flat anterior chamber and corneal endothelial damage. A second group of complications is associated with the enhanced inflammatory and fibrotic responses peculiar to a child's eye. This group includes operative striate keratitis and iridocyclitis, late IOL precipitates, secondary and postpseudophakos membranes, iris erosion and synechiae formation, and IOL tilt and displacement. Although the intraocular lens is a possible means of visual rehabilitation for children with traumatic cataracts or unilateral infantile cataracts, its long-term risk/benefit ratio must still stand the test of time.


Isolation and characterization of antibiotic X-14547A, a novel monocarboxylic acid ionophore produced by Streptomyces antibioticus NRRL 8167.

A novel carboxylic acid ionophore, antibiotic X-14547A, closely related to the polyether antibiotics has been isolated along with four other metabolites from fermented cultures of a new strain of Streptomyces antibioticus. The structure, determined by X-ray analysis of the R(+)-1-amino-1-(4-bromophenyl)-ethane salt contained pyrrole carbonyl and trans-butadienyl chromophores in addition to the unusual tetrahydroindane bicyclic ring system. A second novel metabolite was identified as 3-ethyl-1,3-dihydro-3-methoxy-2H-indol-2-one.


X-14547A, a new ionophorous antibiotic produced by Streptomyces antibioticus NRRL 8167. Discovery, fermentation, biological properties and taxonomy of the producing culture.

X-14547A is a novel antibiotic produced by a new strain of Streptomyces antibioticus (NRRL 8167). The antibiotic is active in vitro against Gram-positive bacteria and is capable of complexing and transporting divalent as well as monovalent metal cations.


Occurrence of 1-methyladenosine and absence of ribothymidine in transfer ribonucleic acid of Mycobacterium smegmatis.

The minor base composition of Mycobacterium smegmatis tRNA has been studied. Thin-layer chromatographic patterns of a ribonuclease T2 digest of mycobacterial tRNA indicated the presence of appreciable amounts of 1-methyladenosine (which is commonly present only in eucaryotic tRNA), dihydrouridine, and 7-methylguanosine. Ribothymidine was absent. The S-adenosylmethionine-dependent tRNA methylases of M. smegmatis catalyzed the formation of 1-methyladenosine when Escherichia coli tRNA was used as acceptor. Similarly, E. coli extracts methylated the tRNA of M. smegmatis, forming ribothymidine.


Identification of the ftsA gene product.

A nonsense mutation was identified in the essential cell division gene ftsA of Escherichia coli. A gamma-transducing phage was isolated which complemented this mutation. This phage programmed the synthesis of four bacterial proteins in UV-irradiated cells. By substituting the nonsense mutation for the ftsA+ allele in this transducing phage and comparing the proteins programmed by it in UV-treated Su+ and Su- cells, the product of the ftsA gene was identified as a protein with a molecular weight of 50,000.


Effect of dna mutations on the replication of plasmid pSC101 in Escherichia coli K-12.

Escherichia coli strains with mutations in genes dnaB, dnaC, and dnaG were tested for their capacity to replicate pSC101 deoxyribonucleic acid (DNA) at a nonpermissive temperature. Only a small amount of radioactive thymine was incorporated into pSC101 DNA in the dna mutants at 42 degrees C, whereas active incorporation into plasmid DNA took place in wild-type strains under the same conditions. The effects of the dnaB and dnaC mutations were greater on plasmid DNA synthesis than on host chromosomal DNA synthesis, suggesting that these gene products are directly involved in the process of pSC101 DNA replication. In dnaG mutants, both plasmid and chromosomal DNA synthesis were blocked soon after the shift to high temperature; although the extent of inhibition of the plasmid DNA synthesis was greater during the early period of temperature shift to 42 degrees C as compared with that of the host DNA synthesis, during the later period it was less. It was found that the number of copies of pSC101 per chromosome in dnaA and dnaC strains, grown at 30 degrees C, was considerably lower than that in wildtype strains, suggesting that the replication of pSC101 in these mutant strains was partially suppressed even under the permissive conditions. No correlation was found between the number of plasmid copies and the tetracycline resistance level of the host bacterium.


Cloning the spoT gene of Escherichia coli: identification of the spoT gene product.

We have isolated five specialized transducing lambda bacteriophages (lambda dpyrE spoT) carrying the pyrE and spoT genes of Escherichia coli. A fragment from one of these phages was used as the source of DNA to clone the spoT and pyrE genes on a multicopy plasmid, pBR322. Insertions and deletions in this plasmid were obtained. These plasmids were used to transform a minicell-producing strain, and the gene products synthesized were determined. Our experiments demonstrate that the spoT and pyrE genes are separated by about 4 magadaltons and suggest that the spoT gene product is a protein whose molecular weight is 80,000. The strain in which the spoT+ allele is carried on a plasmid produced nine times more spoT gene activity than a normal spoT+ strain when assayed in crude extracts. This strain was used to prepare partially purified gene product, guanosine 5'-diphosphate, 3'-diphosphate pyrophosphatase. The enzyme has the following characteristics. (i) It hydrolyzes pyrophosphate from the 5'-pyrophosphate of guanosine 5'-diphosphate, 3'-diphosphate, yielding GDP and pyrophosphate. (ii) Its activity is strongly stimulated by Mn2+ and slightly stimulated by salt. (iii) Its activity is inhibited by uncharged tRNA. There are also two additional activities in the cell extract which degrade guanosine in 5'-diphosphate, 3'-diphosphate in vitro but which are not specified by the spoT gene.


Isolation and properties of Escherichia coli K-12 mutants impaired in the utilization of gamma-aminobutyrate.

We have isolated mutants of Escherichia coli K-12 CS101B that have lost the ability to utilize gamma-aminobutyrate as a source of nitrogen. One class of mutants, which were not affected in the utilization of other nitrogen sources (proline, arginine, glycine), included many isolates with lesions in gamma-aminobutyrate transport or in its transamination and one mutant completely devoid of succinic semialdehyde dehydrogenase activity and exhibiting low gamma-aminobutyrate transport and transamination. gamma-Aminobutyrate-utilizing revertants of the latter recovered full transport and transamination capacities but remained dehydrogenaseless. Another class of mutants showed pleiotropic defects in nitrogen metabolism. One such mutant was lacking glutamate synthase activity. The genes specifying the synthesis of gamma-aminobutyrate permease, gabP, gamma-aminobutyrate transaminase, gabT, and succinic semialdehyde dehydrogenase, gabD, and the control gene, gabC, that coordinately regulates their expression all form a cluster on the E. coli chromosome, linked to the srl and recA loci (at 57.5 min). The mutations with pleiotropic effects on the metabolism of nitrogenous compounds are not linked to the gab cluster.


Chromosome replication and cell division in plasmid-containing Escherichia coli B/r.

The kinetics of chromosome replication and cell division have been examined in recA mutants of Escherichia coli B/r containing F' plasmids of various sizes. Plasmid-mediated alterations in growth properties were detected only with the presence of the larger F' plasmids, and were reflected in decreased mean cell sizes and growth rates. The lengths of C and D in all plasmid-containing strains were in accord with the values for plasmid-free parental strains growing with similar generations times. The findings were consistent with an absence of competition between the chromosomal and extrachromosomal replicons for rate-limiting components involved in the initiation of deoxyribonucleic acid synthesis or in the elongation of deoxyribonucleic acid chains.


Synthesis and modification of proteins during the cell cycle of the yeast Saccharomyces cerevisiae.

We have used a novel technique to study the synthesis, modification and degradation of proteins during the cell cycle in Saccharomyces cerevisiae. Logarithmically growing cells were pulse-labeled twice, with the pulses separated in time by more than one generation. Subsequently, the cells were fractionated as to their position in the cell cycle by centrifugal elutriation, and for different proteins the ratio of radioactive material from the two pulses was then determined. Periodic degradation, synthesis, or modification would produce periodic variations in the ratio of counts. Two-dimensional gel electrophoresis was used to examine 110 different proteins at different times of the cell cycle. All but two proteins had a constant ratio of counts through the cell cycle. This indicates that the rate of synthesis of individual proteins increases exponentially during the cell cycle and that periodic degradation or modification of proteins is not a general feature of the cell cycle in S. cerevisiae.


Gene dosage effects on the synthesis of maltase in yeast.

Inbred strains of Saccharomyces cerevisiae carrying MAL1, MAL2, or MAL6 in a common background were used to construct (i) homo- or heterozygous diploids carrying one or two active alleles of a single MAL locus (MAL1, MAL2, or MAL6) and (ii) triploids carrying one, two, or three active alleles of MAL2. The diploid and triploid strains were used to investigate gene dosage effects of the differential rate of maltase synthesis (delta enzyme activity/delta growth) and the kinetics of induction (for MAL2). All three MAL loci exhibited a gene dosage effect on the differential rate of maltase synthesis; MAL2 also exhibited a gene dosage effect on the kinetics of induction. The dosage effects of the MAL1 and MAL6 loci were additive, but the effects of the MAL2 locus were not; the magnitude of the MAL2 gene dosage effect decreased with increasing dosage. These results are compatible with the current genetic evidence that the MAL genes are regulatory loci if the product(s) of the MAL1 and MAL6 locus is produced in limiting amounts but the product(s) of the MAL2 locus is produced in excess, except at very low genes dosages.


Substrate binding site for nitrate reductase of Escherichia coli is on the inner aspect of the membrane.

Escherichia coli grown anaerobically on nitrate exhibited the same transport barrier to reduction of chlorate, relative to nitrate, as that exhibited by Paracoccus denitrificans. This establishes that the nitrate binding site of nitrate reductase (EC 1.7.99.4) in E. coli must also lie on the cell side of the nitrate transporter which is associated with the plasma membrane. Because nitrate reductase is membrane bound, the nitrate binding site is thus located on the inner aspect of the membrane. Nitrate pulse studies on E. coli in the absence of valinomycin showed a small transient alkalinization (leads to H+/NO3- congruent to --0.07) which did not occur with oxygen pulses. By analogy with P. denitrificans, the alkaline transient is interpreted to arise from proton-linked nitrate uptake which is closely followed by nitrite efflux. The result is consistent with internal reduction of nitrate, whereas external reduction would be expected to give leads to H+/NO3-ratios approaching --2.


Mapping of ilvO loci of Escherichia coli K-12 with bacteriophage lambda dilv.

A set of lambda dilv phage have been used in a deletion mapping procedure to determine the location of two previously characterized ilvO alleles. In contrast to earlier conclusions derived from three-factor crosses and episome-shortening techniques with phage P1, the order found is ilvG-ilvO-ilvEDA. A three-factor cross with phage P1 is described that is not consistent with this location for an ilvO allele. Further analysis of this particular three-factor cross revealed than an artifact attributable to a mutual syntrophism had skewed the apparent frequency of inheritance of the ilvO locus. The role of mutual syntrophism is discussed as a source of mapping errors for the ilvO locus. The value of this set of lambda dilv phage and this mapping procedure for obtaining comparatively unambiguous data on the locations of the ilv structural and regulatory genes is demonstrated.


Enzymatic degradation of uracil-containing deoxyribonucleic acid. V. Survival of Escherichia coli and coliphages treated with sodium bisulfite.

A number of mutants of Escherichia coli defective in the ung gene (structural gene for uracil-deoxyribonucleic acid [ura-DNA] glycosylase) are shown to be abnormally sensitive to treatment with sodium bisulfite when compared with congenic ung+ strains. These results provide further evidence that sodium bisulfite causes the deamination of cytosine to uracil in DNA and that ura-DNA glycosylase is required for the repair of U-G mispairs. The effect of the chemical is apparently selective with respect to base damage; coliphages containing cytosine in their DNA are inactivated by treatment with sodium bisulfite, whereas those containing hydroxymethylcytosine are not. ura-DNA glycosylase and the major apurinic-apyrimidinic endonuclease of E. coli may function in the same repair pathway, since the extent of inactivation of a congenic set of strains which are ung xth (structural gene for the major apurinic-apyrimidinic endonuclease of E. coli) or ung xth+ is the same.


Salmonella typhimurium newD and Escherichia coli leuC genes code for a functional isopropylmalate isomerase in Salmonella typhimurium-Escherichia coli hybrids.

The supQ newD gene substitution system in Salmonella typhimurium restores leucine prototrophy to leuD mutants by providing the newD gene product which is capable of replacing the missing leuD polypeptide in the isopropylmalate isomerase, a complex of the leuC and leuD gene product. Mutations in the supQ gene are required to make the newD protein available. An Escherichia coli F' factor was constructed which carried supQ- newD+ from S. typhimurium on a P22-specialized transducing genome. This F' pro lac (P22dsupQ394newD) episome was transferred into S. typhimurium strains containing th leuD798-ara deletion; the resulting merodiploid strains had a Leu+ phenotype, indicating that supQ- newD+ is dominant over supQ+ newD+, and eliminating the possibility that the supQ gene codes for a repressor of the newD gene. Furthermore, transfer of the F' pro lac (P22dsupQ39newD) into E. coli leuD deletion strains restored leucine prototrophy, showing that the S. typhimurium newD gene can complment the E. coli leuC gene. Growth rates of the S. typhimurium-E coli hybrid strains indicated that the mutant isopropylmalate isomerase in these strains does not induce a leucine limitation, as it does in S. typhimurium leuD supQ mutants. In vitro activity of the mutant isopropylmalate isomerase was demonstrated; the Km values for alpha-isopropylmalate of both the S. typhimurium leuC-newD isomerase and the S. typhimurium-E. coli hybrid isomerase were as much as 100 times higher than the Km values for alpha-isopropylmalate of the wild-type enzyme, which was 3 x 10(-4) M. Mutagenesis of E. coli leuD deletion strains failed to restore leucine prototrophy, indicating that E. coli does not have genes analogous to the S. typhimurium supQ newD genes, of that, if present, activation of a newD is a rare event or is lethal to the cell.


Positive selection of mutants with cell envelope defects of a Salmonells typhimurium strain hypersensitive to the products of genes hisF and hisH.

Strain SB564 and its derivative DA78 are hypersensitive to the inhibitory action of the proteins coded for by genes hisF and hisH on cell division. Transduction of hisO1243, a regulatory mutation that elicits a very high level of expression of the histidine operon, into these strains resulted in the production of long filamentous cells carrying large "balloons" and in growth failure. Forty-one hisO1242 derivatives that escaped inhibition were isolated. These strains showed a large variety of alterations, many of which were related to the cell envelope. The more-frequent alterations included: changes in cell shape, increased sensitivity to one or more of several drugs (deoxycholate, cycloserine, penicillin, novobiocin, acridine orange), increased autolytic activity in alkaline buffer, anomalous fermentation of maltose on eosin--methylene blue plates, and temperature-conditional cell division. The alterations are produced, in some of the strains, by pleiotropic mutations in gene envB (Antón, Mol, Gen. Genet. 160:277--286, 1978) or envD (Antón and Orce, Mol. Gen. Genet. 144:97--105, 1976). Strains affected in divC, divD, and rodA loci have also been identified. Genetic analysis has shown that several strains carry more than one envelope mutation. It is assumed that envelope mutations are positively selected because they somehow alleviate the particularly severe inhibition of cell division caused, in strains SB564 and DA78, by the excessive synthesis of hisF and hisH gene products.


Chloramphenicol resistance mutation in Escherichia coli which maps in the major ribosomal protein gene cluster.

Localized mutagenesis and selection for streptomycin resistance were utilized to isolate a chloramphenicol resistance mutation in Escherichia coli K-12 linked to the strA (rpsL) locus. Bacteriophage P1 transduction verified the map position of the new resistance mutation at 72 min, placing it within a dense cluster of ribosomal protein genes. The map position differs from that of known cmlA and cmlB mutations, which map at 18 and 21 min, respectively. Ribosomes prepared from chloramphenicol-resistant and -sensitive isogenic transductants were analyzed in vitro for activity in formation of N-formylmethionyl-puromycin, polyphenylalanine, and polylysine in the presence of inhibitory concentrations of chloramphenicol. Comparisons were also made of 14C-chloramphenicol binding to 70S ribosomes and of the two-dimensional polyacrylamide gel electrophoresis pattern of ribosomal proteins from each strain. There was no detectable difference between ribosomes from sensitive and resistant strains as measured by these assays. Enzymatic modification by chloramphenicol acetyltransferase is not responsible for the observed phenotype.


Enumeration and identification of IS3 elements in Escherichia coli strains.

Escherichia coli K-12 strains ordinarily contain five IS3 elements. Three of these correspond to previously mapped IS3 elements (R. C. Deonier, G. R. Oh, and M. Hu, J. Bacteriol. 129:1129--1140, 1977; S. Hu, E. Ohtsubo, and N. Davidson, J. Bacteriol. 122:749--763, 1975), and two additional IS3 elements are identified. The distribution of IS3 elements among deoxyribonucleic acid fragments generated by digestion with EcoRI indicates a basic pattern from which deviation is detected.


Nucleoside diphosphate sugar hydrolase gene of Salmonella typhimurium: chromosomal location determined by intergeneric crosses.

The gene specifying a membrane-bound nucleoside diphosphate sugar hydrolase of Salmonella typhimurium was mapped near the metA locus by using intergeneric crosses between this bacterium and Escherichia coli.


Association of tellurium resistance and bacteriophage inhibition conferred by R plasmids.

Concomitant resistance to tellurium compounds (Ter) and inhibition of coli-phage development (Phi) are properties mediated by many H2 incompatibility group R plasmids which have been isolated from diverse bacterial and geographic sources. Ter plasmids from tellurium-resistant bacteria that were isolated from sewage and industrial wastes also mediated phage inhibition. Of these Ter plasmids, three from Citrobacter freundii belonged to the H incompatibility group, whereas three from Klebsiella pneumoniae did not.


Screening for amino acid pool mutants of Neurospora and yeasts: replica-printing technique.

A replica-printing technique has been developed for permeabilized colonies of Neurospora and yeast by which endogenous amino acid pools are visualized. By the use of cation-exchange paper, the method has detected Neurospora mutants unable to store the large pools of basic amino acids characteristic of wild type. Mutants of this sort were sought to study the function of the fungal vacuole.


Enhancement of deoxyribonucleic acid polymerase I-directed repair synthesis in toluene-treated Escherichia coli after growth in the presence of low levels of N-methyl-N'-nitro-N-nitrosoguanidine.

Deoxyribonucleic acid polymerase I-directed repair synthesis can be selectively measured in toluene-treated Escherichia coli cells exposed to alkylating chemicals such as N-methyl-N'-nitro-N-nitrosoguanidine. Prior growth of the cells in the presence of a low dose of N-methyl-N'-nitro-N-nitrosoguanidine results in an enhanced deoxyribonucleic acid polymerase I-directed repair synthesis and an increase in single-strand breaks.


Incorporation of synthetic fatty acid analogs into phospholipids of Escherichia coli.

Escherichia coli K1060-B5 (fadE fabB thi) incorporates into phospholipids several new synthetic fatty acid analogs including those containing phenyl, phenoxy, and azidophenoxy groups at the omega position.


Addition of basic amino acids prevents G-1 arrest of nitrogen-starved cultures of Saccharomyces cerevisiae.

Arginase-minus mutants of Saccharomyces cerevisiae were arrested in growth and accumulated at the unbudded G-1 stage of the cell cycle when starved for nitrogen. If, however, arginine was added to the culture medium at the time of starvation, growth ceased but the cells did not collect at the unbudded G-1 stage. We suggest that arginine addition prevented the cells from collecting at the G-1 stage by starving them for histidine and lysine, thereby inhibiting synthesis of proteins needed to complete the cell cycle.


Thermoresistant revertants of an Escherichia coli strain carrying tif-1 and ruv mutations: non-suppressibility of ruv by sfi.

Spontaneous thermoresistant revertants were isolated from Tif1 Ruv- and Tif1 Ruv+ strains of Escherichia coli K-12. They were divided into five groups; backmutants to tif+ and recA structural gene mutants accounted for at least two of these groups. Mutations with an unconditional RecA- phenyotype were detected at a higher frequency in the Tif1 Ruv- strains (65%) than in the Tif1 Ruv+ strains (25%). A third group consisted of revertants exhibiting a RecA- phenotype at low temperature. Revertants with normal recombination ability and UV resistance, but with a thermosensitive defect in propagating lambda bio11 phage, were also isolated (group 4). The alleles responsible for this property were cotransducible with the srl gene, suggesting that they are located at the recA locus. Other revertants, which might carry lex, LEXB, or zab mutations, were UV sensitive and were able to propagate lambda bio11 phage (group 5). The sfi mutation, which suppresses filamentation in the Tif1 and UV-sensitive Lon- strains, does not restore UV resistance of the Ruv- mutant.


Repair of ultraviolet light-damaged deoxyribonucleic acid in sbc-A strains of Escherichia coli K-12.

An Escherichia coli strain carrying both rec+ and sbcA has been constructed. Repair of ultraviolet light-induced deoxyribonucleic acid damage was examined by measuring survival and thymine-dimer excision in the rec+ sbcA strain as well as rec+ sbcA+ and recB recC sbcA strains. The sbcA mutation restores normal survival in both recB recC uvrB and recB recC uvr+ strains. Excision of thymine-containing dimers does not occur in uvrB mutants, regardless of the rec or sbcA genotype. Survival, after ultraviolet-light damage, of a rec+ sbcA strain is quantitatively similar to rec+ sbcA+ and recB recC sbcA strains.


Properties and biosynthesis of cyclopropane fatty acids in Escherichia coli.

The lipid phase transition of Escherichia coli phospholipids containing cyclopropane fatty acids was compared with the otherwise homologous phospholipids lacking cyclopropane fatty acids. The phase transitions (determined by scanning calorimetry) of the two preparations were essentially identical. Infection of E. coli with phage T3 inhibited cyclopropane fatty acid formation over 98%, whereas infection with mutants which lack the phage coded S-adenosylmethionine cleavage enzyme had no effect on cyclopropane fatty acid synthesis. These data indicate that S-adenosylmethionine is the methylene in cyclopropane fatty acid synthesis.