[Morphological ultrastructural and immunological studies of a rickettsia isolated from a parrot (author's transl)]. A rickettsia isolated from a parrot was found by use of electron microscopic and serological technic to be a Coxiella burneti. In cell culture one can see a dramatic polymorphism and a Gram negative-like structure of the cell wall. They divide by binary fission with no appearance of a complexe cycle of development like the one described for Chlamydia psittaci. Many myelinic fibrils and altered Coxiella suggest that the host cell react very deaply.


[Nutritional value and acceptibility of soy proteins in human diet]. These studies on textured soya protein (TSP) were directed towards the evaluation of its acceptability and nutritional value in man. ACCEPTABILITY. In the beginning, the reaction of adults to the incorporation of 40 g TSP in their daily diet was studied. After a three-week period, no intolerance or fatigue with the regim was shown. In a following study, observations made on young children's attitudes along with quantitative measurements of their food intake were noted during four one-wekk periods: -- period of "imposed diet", without TSP; -- period of imposed diet", with TSP; -- period of "free diet", without TSP; -- period of "free diet", with STP; During the periods of "imposed diets", children could eat only the proposed quantities of foodstuffs. During periods of "free diets", children were able to choose the quantity and sort of food they preferred from the proposed menu. Total protein ingested (differentiating between that of animal and vegetable origin) was calculated for each child from his daily food record. Results show that only one of the twenty-four children systematically refuse soya-containing preparations. Average consumption of soya protein during "free diet" periods was close to the desired level and not significantly different to levels achieved in the "imposed diet" period. The acceptance of TSP by the children was therefore evident. NUTRITIONAL VALUE. The following studies were designed to determine the effects of the replacement of animal protein by TSP (at the maximal levels of its tolerability) in a normal diet. In the first nitrogen balance study, with 10 convalescent subjects, the following protocol was developed: -- the habitual caloric and protein intakes of the subjects was established; -- following a four-day adaptation period, the subjects entered consecutively: a control period at established levels of calories and protein, and a test period in which 20 g of soya protein replaced an equivalent quantity of animal protein. The average nitrogen balances for control and test periods were + 4.49 g and 3.62 g. The difference not being significant at a probability of 5 p. 100. In a second study with 12 adults in good health (6 males and 6 females) soya protein replaced 34-39 p. 100 of the daily protein intake (according to the menu) and the average nitrogen balances in control and test periods were + 0.16 g and 0.91 g. These studies confirm the TSP replacement of animal protein in the normal diet does not affect nitrogen balance and that TSP can make a contribution to the fulfilment protein requierements in man.


Effect of intra-articular 90Y resin treatment on the chemical properties of synovial fluid in rheumatoid arthritis. Lyaluronic acid and total protein concentrations in synovial fluid of the knee joint obtained from 48 patients with classical or definite rheumatoid arthritis were determined before, and 1 week, 1 month and 6 months after intra-articular injection of 90Y resin colloid. The concentration ratio of total protein to hyaluronic acid, which is a sensitive index of the abnormality of the synovial fluid, was worse a week after the injection in each group as regards the clinical effect of the 90Y resin injection, but was better 1 month and 6 months after the injection only in the remission group.


Detection of homozygotes and heterozygotes for metachromatic leukodystrophy in lymphoid cell lines and peripheral leukocytes. Assays of arylsulphatase A have been performed on peripheral leukocytes and lymphoid cell lines from normal subjects and heterozygotes and homozygotes for metachromatic leukodystrophy (MLD) by utilizing an improved procedure based on the removal of excess protein from the final digest. No overlap of arylsulphatase A activity was observed between peripheral leukocytes obtained from 11 normal donors, 8 heterozygotes and 5 homozygotes for MLD. Absence of enzyme activity was demonstrated in the patients' leukocytes, when the modified assay procedure was employed. Arylsulphatase A activity (mean +/- S.D.) in 30 lymphoid lines and sublines established from 18 normal donors, 2 heterozygotes and 1 homozygote for MLD was 54-5 +/- 23-1, 24-1 +/- 3-2 and 0-9, respectively. The enzymic activity of arylsulphatase A and B remained stable over a period of several months. These findings indicate that long-term lymphoid lines express the genotype of the donor and maintain enzyme stability over long periods in culture. Moreover, it appears that peripheral leukocytes are a reliable system for the identification of the carrier state of MLD.


[Effects of sleep on ovary development during hibernation and aestivation in the dormouse (Eliomys quercinus L.)(author's transl)]. Hibernation does not correspond to a period of rest of the ovary, nor to the prolongation of a period of anoestrus. Hibernation is necessary for the normal course of the period of genital activity. Its suppression tends to modify more or less severely the physiological equilibrium implicated in the control of genital activities. However, the animal can find a more or less normal rythm even if is deprived of hibernation during the course of a second year. Prolongation of hibernation in hemicastrated animals during the entire period of genital activity suppresses the ovular discharge and the ovulating secretion of FSH and LH, without affecting the tonic secretions of FSH. It inhibits the phenomenon of compensatory hypertrophy. However, in hemicastrated animals deprived of hibernation, the reaction to hemicastration does not take place nor during anoestrus nor during mid-december or mid-march but only when the period of genital activity begins again.


Rupture of the spleen in erythroblastosis fetalis. A male infant of 36 weeks' gestation, weighing 3080 g, with erythroblastosis, ruptured spleen, and bilateral suprarenal haemorrhages is described. The infant survived after exchange transfusions and splenectomy.


The missing umbilical artery. II. Paediatric follow-up. Bryan, E. M., and Kohler, H. G. (1975). Archives of Disease in Childhood, 50, 714. The missing umbilical artery.  II. Paediatric follow-up. Of 143 infants with single umbilical artery detected by routine examination of the placenta, 25 had major malformations at birth; 3 of these survive. Another 6 were stillborn and 2 died during the first year of life. At follow-up 14 children could not be traced. 18 were assessed on the basis of reports by family doctors or parents ('report group'); 14 of these were considered normal. The remaining 78 infants and children were given a clinical examination ('examination group'); 64 were found to be normal. Malformations found in 10 children (6 from the examination group and 4 from the report group) are discussed. Most of the abnormalities detected were less severe and less conspicuous than those revealed at birth, and in a few instances only might have been diagnosed by a more thorough examination in the perinatal period. Failure to detect these 'less severe and less conspicuous' malformations is generally unlikely to be detrimental to the infant, with the exception of urinary tract anomalies which are known to predispose to infection. Included in the examination group were 16 children (out of an original 22) who had been 'normal' but small-for-dates at birth; 14 of these had now caught up. The remaining 2 were found to have abnormalities that had not been manifest at birth. The finding of single umbilical artery at birth commits the paediatrician to an intensive search for malformations which are not immediately apparent, but prolonged surveillance for this reason alone is not advocated.


An analysis of pancreatic enzymes used in epidermal separation. A comparative enzyme analysis was performed on 3 pancreatic extracts generally used for dermal-epidermal separation, namely, crude trypsin (Difco), crude trypsin (Sigma) and pancreatin. A fourth pancreatic extract, crude lipase, was subjected to a corresponding analysis. The 4 extracts were assayed for activities of: protease (total), trypsin, chymotrypsin, carboxypeptidase-A, amylase, elastase, lipase, esterase, arylesterase and ribonuclease. Relative activities of the different proteolytic enzymes were individualized by utilizing specific inhibitors. Insignificant differences were observed between the enzyme activities of crude trypsin (Difco) and pancreatin. Crude lipase displayed similar enzyme activities as these two extracts in addition to high lipolytic, esterolytic and arylesterolytic activities. Crude trypsin (Sigma) exhibited higher tryptic and chymotryptic activities than the other extracts but lacked all further enzyme activities. Epidermal separation was performed using similar incubation conditions for each extract and skin from the same donor. Ultrastructural examination of the detached epidermis revealed that a more effective separation could be achieved by crude lipase.


The distribution and binding of cadmium in oyster, soybean, and rat liver and kidney. Exposure to Cd and/or 109Cd has shown that the element was efficiently accumulated in oysters and soybeans as well as in rats. Differential centrifugation of oyster, soybean, rat liver, and rat kidney homogenates followed by analysis showed that Cd was associated primarily with proteins in the 105,000 X g supernatants. Separation of these proteins by Sephadex chromatography and subsequent analysis by atomic absorption spectroscopy or by radioactivity measurements revealed that Cd in oysters and rat organs was principally bound to proteins of 9,200 to 13,800 molecular weight. A significant amount of Cd in oysters was also associated with fractions of greater than 50,000 and less than 3,000 molecular weights. Almost all of the Cd in soybeans was found to be bound to molecules of greater than 50,000 molecular weight.


Cell wall enzymatic lysis of the yeast form of Pullularia pullulans and wall regeneration by protoplasts. During the process of degradation of the cell wall of the yeast form of Pullularia pullulans by the lytic system of micromonospora chalcea samples were withdrawn at different times and observed under phase contrast and electron microscope. The progressive lysis of the walls reveals a fibrillar component inside the apparently amorphous wall. Freeze etched preparations of cells during the formation and regeneration of protoplasts show that the cellular membrane is split and this method allows the smooth external face of the membrane and other internal face covered by particles to be seen. The fact that the smooth face of the membrane is only visible during the preparation or the regeneration of protoplasts and very rarely when intact cells are fractured, suggests a strong adherence between cell wall and this external layer of the membrane. During the regeneration which takes place as in most of the yeasts and moulds, a special study of the extension of the cell wall is made and a possible mechanism for this extension of the regenerated cell wall is proposed.


Oxygen pressure, fatty acid composition and ergosterol level in Rhodotorula gracilis. Cells of Rhodotorula gracilis grown for 6 hrs at 2 mm Hg O2 pressure when compared with cells grown for 6 hrs at 140 mm Hg, show: a) a large decrease in the level of ergosterol, b) a significant increase in the level of oleic acid and a decrease in the levels of linoleic and linolenic acids, both in fatty acid fraction of the phospholipids and in the free fatty acids and neutral fat fractions. The results suggest that the dehydrogenation of oleic acid to linoleic acid is preferentially inhibited at low O2 pressure. The possibility is considered that these changes of lipid metabolism might be causally related with decrease of the growth rate observed at low O2 pressure.


Differentiation in Coprinus lagopus. III. Expansion of excised fruit-bodies. Fruit-body expansion was studied in Coprinus lagopus (sensu Buller) following surgical procedures. Elongation occurred after denuding mushroom caps of essentially all peripheral scales. Young primordia (1 - 5 mm) failed to develop after vertical bisection. Older primordia (e.g. 10 mm) expanded 3 - 4 fold after vertical bisection or quadrisection, underwent autolysis and basidiospore production. An amorphous brown gel in the stripe lumen disappeared during development of bisected primordia. Stripes isolated from primordia expanded autonomously and exhibited negative geotropism when incubated upside down or when the stripe apex was removed. Displacement of charcoal particles dusted on intact stripes revealed the most active zone of expansion to be the upper mid-region of the stripe. Segmented stripes likewise showed most active elongation in the mid-region. Vertically bisected stripes also expanded.


[Diurnal rhythm of the reproductive activity of cells and the functional morphology of the adenohypophysis of rats (histological and the autoradiographic study)]. The adenohypophysis of intact mongrel male rats was studied histologically and autoradiographically. The diurnal periodicity was revealed and the successive spikes of indices of the DNA synthesis, mitotic process and secretory activity of the adenohypophysis cells were established. The maximum amount of the dividing cells was found about 12 o'clock p. m.-at the period of the least secretory and synthetizing activity of the gland.


Electron microscopy of Japanaut and Tilligerry viruses: two proposed members of the orbivirus group. Japanaut and Tilligerry viruses were studied by thin-section and negative-contrast electron microscopy, and from their morphology and morphogenesis appear to be typical orbiviruses. Characteristic intracellular structures are associated with the development of the two viruses. These structures include coated tubules and one or more types of structure which appear to be composed of a number of parallel sheets of moderately electron-dense material.


Absence of a pyrimidine dimer repair mechanism for mitochondrial DNA in mouse and human cells. We have assayed the ability of mammalian cells to remove pyrimidine dimers from their mitochondrial DNA. The results indicate that pyrimidine dimers are not repaired for as long as 48 hr after UV irradiation. Furthermore, molecules containing pyrimidine dimers are unable to replicate and are simply diluted out in subsequent cell divisions.


Excision-repair in primary cultures of mouse embryo cells and its decline in progressive passages and established cell lines. Fibroblasts of mouse embryo cells from early subcultures excise pyrimidine dimers to an extent and at a rate comparable to those observed in human cells. The only apparent difference is that in primary mouse cells dimers are excised in an acid-insoluble form. Dimer excision in mouse embryo fibroblasts declines abruptly after the fourth to the sixth subculture and is not detectable in the permanent cell line 3T3. It is suggested that cessation of excision-repair may be due to genetic repression.


Use of enzymatic assay to evaluate UV-induced DNA repair in human and embryonic chick fibroblasts and multinucleate heterokaryons derived from both. A sensitive enzymatic assay has been utilized to monitor repair of UV-induced damage to DNA in primary human and embryonic chick cells and in multinucleate heterokaryons artificially derived from both. The assay exploits the unique ability of a purified repair endonuclease to attack UV-irradiated DNA at sites containing pyrimidine dimers. These nuclease-susceptible sites are subsequently observed as single-strand scissions by velocity sedimentation in alkaline sucrose gradients. Incubation of UV-damaged cultures followed by extraction and enzymatic analysis of the radioactively labeled DNA enables one to trace the disappearance of such sites in vivo and hence to monitor endogenous repair activity. When UV-irradiated human cells are incubated in the dark, the curve for site removal exhibits a two-phase exponetial decline; i.e. there exists a fast component responsible for elimination of 60% of the initial damage and a second one approximately 7 times slower in rate. The removal of sites is not further enhanced by exposing cells to blacklight during post-UV incubation. Conversely, UV-damaged chick cells rid their DNA of all nuclease-susceptible sites rapidly (i.e. at an exponential rate approximately 13 times faster than the fast component of site removal in human cells) when incubated under blacklight but not when kept in the dark. These data indicate the presence in human and embryonic chick cells of distinct enzymatic mechanisms for the elimination of dimer-containing sites. Wheneras human fibroblasts rely heavily on a light-independent process, excision-repair, chick fibroblasts possess a light-dependent mechanism, presumably photoenzymatic repair. Advantage has been taken of the contrasting repair properties of the human and embryonic chick fibroblasts to evaluate the extent to which each can assist the other in the removal of UV-induced damage from its DNA. The two cell types were fused to form giant human/chick heterokaryons containing a number of intact nuclei from both strains. Experimental conditions were selected so that UV-induced damage resided only in DNA foreign to the repair enzymes under study. Our results strongly suggest that repair enzyme(s) coded for by either fusion partner can remove dimer-containing sites from the DNA of the other with an efficiency comparable to that attained when acting on its own DNA in unfused, parental cells. Further, the light-requiring repair process supplied by the chick is more proficient at operating on these sites in human DNA than is excision-repair, the parallel mechanism available to human cells for this purpose.


The binding of 8-anilinonaphthalene-1-sulphonate to cytoplasmic aspartate aminotransferase from pig heart. Anilinonaphthalenesulphonate binds to cytoplasmic aspartate aminotransferase with high affinity (Kd about 10 muM) and with a stoicheiometry of one molecule per dimer. It is not displaced by aliphatic or aromatic dicarboxylate substrate analogues. The enzyme is believed to be a symmetrical dimer with identical subunits; it can evidently function asymmetrically in binding anilinonaphthalenesulphonate.


Purification and properties of rat brain pyruvate carboxylase. Rat brain pyruvate carboxylase was purified 2000-fold and some of its properties and kinetic parameters were investigated. The use of (NH4)2SO4 gradient solubilization on a Celite column and precipitation with polyethylene glycol permitted purification to an estimated 20% purity. Except for a few subtle kinetic differences this enzyme is indistinguishable from rat liver pyruvate carboxylase.


Activation and inhibition of rabbit muscle pyruvate kinase by transition-metal ions. The paper reports a comparative study of the effects of Mn2+, Ni2+ and Co2+ on the reaction of ADP with phosphoenolypyruvate when catalysed by K+-activated rabbit muscle pyruvate kinase. The activation and subsequent inhibition that occurs as the bivalent ion concentration is increased is taken as evidence that the substrates of the enzyme are phosphoenolypyruvate, uncomplexed ADP and free bivalent cation. Kinetic constants for the binding of the bivalent cation to the enzyme are reported.


Fixation of the first component of complement by immune complexes: effect of reduction and fragmentation of antibody. The integrity of both covalent and non-covalent bonds between the heavy chains of rabbit immunoglobulin gamma antibody is essential for maximal efficiency of binding of the first component of complement to immune complexes.


Modification of pig M4 lactate dehydrogenase by pyridoxal 5'-phosphate. Demonstration of an essential lysine residue. 1. Pig M4 lactate dehydrogenase treated in the dark with pyridoxal 5'-phosphate at pH8.5 and 25 degrees C loses activity gradually. The maximum inactivation was 66%, and this did not increase with concentrations of pyridoxal 5'-phosphate above 1 mM. 2. Inactivation may be reversed by dialysis or made permanent by reducing the enzyme with NaBH4. 3. Spectral evidence indicates modification of lysine residues, and 6-N-pyridoxyl-lysine is present in the hydrolsate of inactivated, reduced enzyme. 4. A second cycle of treatment with pyridoxal 5'-phosphate and NaBH4 further decreases activity. After three cycles only 9% of the original activity remains. 5. Apparent Km values for lactate and NAD+ are unaltered in the partially inactivated enzyme. 6. These results suggest that the covalently modified enzyme is inactive; failure to achieve complete inactivation in a single treatment is due to the reversibility of Schiff-base formation and to the consequent presence of active non-covalently bonded enzyme-modifier complex in the equilibrium mixture. 7. Although several lysine residues per subunit are modified, only one appears to be essential for activity: pyruvate and NAD+ together (both 5mM) completely protect against inactivation, and there is a one-to-one relationship between enzyme protection and decreased lysine modification. 8. NAD+ or NADH alone gives only partial protection. Substrates give virtually none. 9. Pig H4 lactate dehydrogenase is also inactivated by pyridoxal 5'-phosphate. 10. The possible role of the essential lysine residue is discussed.


Continuous assessment of left ventricular shape in man. Frame by frame analysis of left ventriculograms has been performed in 10 normal subjects and 40 patients with heart disease. Left ventricular shape index was derived as 4 pi (cavity area)/(perimeter)2, which has a maximum value of 1 when the outline is circular. In normal subjects systole was always associated with progressive reduction in shape index, indicating that the cavity projection had become less circular. This change was smaller in patients with low ejection fraction and also when inferior or anterior hypokinesia was present, even though ejection fraction was normal. During early diastole shape index rose rapidly due to an increase in minor diameter occurring throughout the period of rapid filling. In some cases this preceded any change in long axis, which was due to upward movement of the aortic root as well as outward movement of the apex. These results have functional implications, suggesting in particular that wall movement during filling may be non-uniform and that assumptions about cavity shape used in the derivation of wall properties from estimates of ventricular volume may require modification.


The immunological localization of factor V in human tissue. Antibody to factor V was produced by immunizing rabbits with purified factor V from human plasma. Various tissues were examined for the presence of factor-V antigen using this antiserum. It was consistently demonstrated in homogenates of liver and spleen by means of an antibody (coagulation inhibitor) neutralization technique. The antigen was further localized histologically by the indirect fluorescent antiglobulin technique. It was present on the endothelium of normal blood vessels in all organs examined. In the liver it was detected in hepatic parenchymal cells and a distinctive pattern of fluorescence in the spleen suggested that it was being detected on platelets. Results were negative in all other tissues examined. The findings confirm the presence of factor V in hepatic parenchymal cells and support the suggestion that endothelial coagulation factors may play a role in haemostasis and thrombosis.


Haemoglobin Bart's in Saudi Arabia.

Haemoglobin Bart's in Saudi Arabia. The haemoglobin (Hb) patterns of 345 Shiite Saudi Arab cord bloods were examined by alkaline starch-gel electrophoresis. A fast-moving component, identified by structural analysis as Hb Bart's, was found in 52% of cases, the highest incidence of this variant yet recorded. The levels of Hb Bart's ranged from 0.5 to 16% of the total haemoglobin. The relative rates of synthesis of the alpha, beta and gamma-chains, measured by [3H]leucine incorporation, were estimated in 12 newborn Arab infants. There was an excellent correlation between the amount of Hb Bart's and the alpha/non-alpha-globin-chain production ratio. Furthermore there was a significant correlation between the level of Hb Bart's and morphological abnormalities of the red cells and the mean cell haemoglobin (MCH). These findings indicate that elevated levels of Hb Bart's in this population are due to the presence of alpha thalassaemia. The absence of hydrops fetalis and the rarity of Hb-H disease despite the intense inbreeding in this population, points to an alpha-thalassaemia genotype that is, in terms of phenotypic expression, intermediate between the heterozygous state for alpha-thalassaemia I and Hb-H disease. A possible molecular basis for this genotype is suggested.


Platelet kinetics in idiopathic thrombocytopenic purpura (ITP) before and at different times after splenectomy. Platelet kinetics were studied in 18 patients with idiopathic thrombocytopenic purpura (ITP) before splenectomy, 1 month and 8 months after splenectomy. A gamma function was used for the calculation of platelet mean life span (MLS). Before splenectomy platelet MLS was greatly reduced and platelet production was 2.8 times normal. The peripheral platelet count and platelet recovery were significantly related to platelet MLS. One month after splenectomy platelet MLS increased appreciably while platelet production decreased but continued to be significantly above normal. These changes together with the removal of the splenic platelet pool resulted in a peripheral platelet count significantly higher than normal and a total platelet mass not differing from normal. Eight months after splenectomy platelet MLS increased slightly but was not significantly different from that found 1 month after splenectomy. Platelet production, however, decreased significantly and became normal. This resulted in a normal peripheral platelet count and a decreased total platelet mass which was significantly lower than in non-splenectomized control subjects. The results suggest that platelet production is regulated by a feed-back mechanism that is governed by the peripheral platelet count. The high platelet count encountered 1 month after splenectomy is due to a sluggishness of this regulatory mechanism.


Foetal erythropoiesis in steel mutant mice. II. Haemopoietic stem cells in foetal livers during development. A rapid and reliable method of definitively identifying mutant (Sl/Sld) and wild type (+/+) mouse embryos in segregating litters is described, based on the mean cell volume determination of circulating foetal erythrocytes by an electronic particle counter. The mean cell volumes of yolk sac derived nucleated erythrocytes from +/+ and Sl/Sld embryos are similar, whereas the foetal liver derived non-nucleated red blood cells are much larger in Sl/Sld than +/+ embryos. There is no significant retardation in the growth of mutant embryos in spite of the severe anaemia which is macrocytic and normochromic. Evidence is also presented that the proportion of haemopoietic stem cells among hepatic erythroblasts, assayed by the macroscopic spleen colony technique, is higher in the mutant embryos, even though the total number of these progenitor cells present in each mutant foetal liver is less than the normal. Furthermore, these stem cells undergo active proliferation in Sl/Sld foetuses during development. The data indicate that the mutant Sl genes do not affect the primitive erythroid cell lineage derived from the yolk sac blood islands, but seriously interfere with the differentiation of the definitive erythroid cell lineage of foetal liver origin. It is further suggested that the mutant foetal liver fails to support or interferes with the normal rate of differentiation from immature precursor cells in to haemoglobinized erythroblasts. The reduction of total haemopoietic stem cells in mutant embryos may be secondary to this defect in cellular differentiation.


Interaction of thrombin with human platelets. The first step in thrombin-induced aggregation of blood platelets is binding of thrombin to specific receptors on the platelet membrane. Elucidation of the nature of this receptor in human platelets was attempted using radioactively labelled thrombin. In disc gel electrophoresis an extract of thrombin-treated platelets showed one peak of radioactivity near the origin. Gel filtration of the platelet extract through Sephadex G-200 showed mainly one protein peak at the void volume which contained radioactivity. After column fractionation the final isolate reacted with antiserum to thrombosthenin but did not react with antiserum to serum, fibrinogen or soluble platelet proteins. Gel electrophoresis of the reduced isolate in the presence of sodium dodecyl sulphate showed a pattern similar to thrombosthenin. To explore the possibility that thrombosthenin might be the receptor of thrombin, attempts were made to complex the receptro sites with thrombosthenin antibody or its univalent fragment. It was observed that complexing these receptors potentiates, rather than inhibits, platelet aggregation by thrombin or by adenosine diphosphate. Univalent fragment of antibody to albumin failed to cause this potentiation. Thus, blocking of the thrombosthenin sites is necessary for sensitization of platelets.


Ultrastructure of lymphocyte-mediated cytotoxicity. I. Interaction between phytohaemagglutinin-stimulated human lymphocytes and chicken erythrocytes. The ultrastructural appearances of the interaction between normal phytohaemagglutinin (PHA)-stimulated human lymphocytes and chicken red cells (ChRBC) were studied. Close contact between ChRBC and control non-stimulated lymphocytes was often observed indicating that this is a non-specific phenomenon. In mixtures of PHA-stimulated lymphocytes and ChRBC, time-dependent development of interdigitating pseudopods between the two cell types was observed which was not seen in controls. Their development followed the same time-course as release of 51Cr when the ChRBC were pre-labelled with this marker. The findings suggest a relationship between the morphorlogical changes observed and the destruction of the target cells (ChRBC).


Beta-thalassaemia of clinical significance in adult Jamaican Negroes. Over a 9-year period, three adult Negro patients with beta-thalassaemia of clinical significance were recognized out of approximately 185 000 new adult patients attending the University Hospital. These patients, aged 15-58 years, have clinical and haematological characteristics within the spectrum of beta-thalassaemia intermedia; which in this paper refers to phenotypes resulting from defects in beta-chain synthesis clinically intermediate between classical Cooley's anaemia and beta-thalassaemia trait, genetic classification being dependent on family study. Family studies established the presence of two beta-thalassaemia genes conclusively in one case (proposita, family A); presumptively in another (propositus, family C); while in the remaining subject (proposita, family B), who has two similarly affected siblings, homozygosity is suspected, but not proven by family study. In simultaneous 59Fe and 51 Cr studies, estimates of effective erythropoiesis are in reasonable agreement with measurements of red cell destruction.


Behaviour of rats exposed to trichloroethylene vapour. Rats were exposed to trichloroethylene (TCE) vapour for about five five-day weeks at concentrations from 100 to 1 000 ppm, and at 100 ppm for 12 1/2 weeks. The social behaviour of paired male rats was observed periodically in the home cage for five minutes after they had been exposed to TCE. The principal finding was a consistent reduction of up to 24% in the total acitivity. A single day's exposure to TCE was sufficient at the highest concentration. At 100 ppm, a similar decline in activity was significant after 1 1/2 weeks' exposure in one experiment and 8 1/2 weeks' in another. The decline in activity was fairly uniform and not usually because of specific reductions in particular kinds of behaviour. However, exploration of the cage and submission to, or escape from, the other rat were sometimes specifically reduced. In an 'exploration-thirst' test, rats were deprived of water overnight and placed on two or three occasions in a previously unfamiliar cage. Rats exposed to 100, 200, or 1 000 ppm TCE found water and began drinking sooner than their controls without altering the rate of movement about the cage. These results suggest lowered performance in a familiar situation where rats are usually very active and some loss of inhibitory control in an unfamiliar one. At the present threshold limit value, repeated exposure to TCE eventually had effects similar to those of one day's exposure to a higher concentration, but only after a widely variable delay.


An epidemiological study of recent trends in ectopic pregnancy. Data from two independent sources in England and Wales, the first based upon persons discharged from hospital and the second upon general practitioner consultations, indicate that the incidence of ectopic pregnancy has been increasing since the late 1950s, with a particularly sharp increase since 1970. Time trends in the age and regional distribution of ectopic pregnancy suggest that the increasing use of intrauterine contraceptive devices may be a major factor contributing to this recent increase in extrauterine gestations. In contrast, recent age and regional trends in tubal infection appear to be unrelated to the trends in ectopic pregnancy. The possible contribution of progestogen-only contraceptives, induced abortion and tubal surgery to the recent increase in extrauterine pregnancy cannot be assessed from the available data.


Thresholds of response to electric shock in previously undernourished rats. 1. Rats were undernourished in early life by feeding their mothers a restricted quantity of a good-quality diet during pregnancy and lactation. All offspring were fed ad lib. from weaning. Behavioural tests were done using adult males. 2. Thresholds of aversion to electric shock were determined in a rectangular box, one side of which was designated 'live' and the other 'safe' (o V side) for any particular shock level. Three 15 min sessions were given at each of five shock levels. Previously undernourished rats had a lower aversion threshold than controls. They spent more time than controls on the o V side at two intermediate levels of shock. 3. Other male rats were subjected to six graded series of brief, unavoidable shocks. Previously undernourished rats responded to a greater total number of shock than controls. 4. These results support the hypothesis that previously undernourished rats have a lowered threshold of arousal.


Catalysis by serine proteases and their zymogens. A study of acyl intermediates by circular dichroism. p-Nitrophenyl p'-guanidinobenzoate and methylumbelliferyl p'-guanidinobenzoate, which are active site titrants for trypsin, and p-nitrophenyl p'-dimethylsulfonioacetamidobenzoate and methylumbelliferyl p'-trimethylammoniocinnamate, which are active site titrants for chymotrypsin, are also hydrolyzed by the respective zymogens. Hydrolysis in each case proceeds via the formation of acyl-zymogens. The acylation rates for the zymogens are 10(3)-10(7) times slower than for the enzymes whereas the deacylation rates of acyl-enzymes and acyl-zymogens are comparable. These findings are consistent with the idea that the diminished catalytic activity of these zymogens is due primarily to their distorted substrate binding sites. The circular dichroic spectra of the acyl-enzymes show induced negative ellipticities in the region of absorption of the acyl group, due to binding of the group in an asymmetric environment. The circular dichroic spectra of the acyl-zymogens do not, but conversion of the acyl-zymogens to acyl-enzymes changes the circular dichroic spectra to those characteristic of the acyl-enzymes. alpha-Carbamyl-epsilon-guanidinated trypsin is a derivative which resembles trypsinogen in lacking activity toward specific ester substrates but possessing low activity toward p-nitrophenyl p'-guanidinobenzoate. The circular dichroic spectrum of the acyl-enzyme formed during hydrolysis of p-nitrophenyl p'-guanidinobenzoate by this derivative resembles that of guanidinobenzoyltrypsinogen, and not that of guanidinobenzoyltrypsin. These circular dichroism studies confirm that the same serine residue is involved in catalysis by both enzymes and zymogens. They demonstrate directly that the acylating group is in a different environment in each and indicate that this specific environment is a determinant in the catalytic activity of each. Thus the circular dichroic spectra of these acyl intermediates provide a sensitive probe of the subtle conformational changes which occur on zymogen activation. The results support the previous conclusion that the major feature of the activation of trypsinogen and chymotrypsinogen is the rearrangement of the substrate binding site and that the appearance of a new amino terminus causes this rearrangement.


Circular dichroism studies of myoglobin and leghemoglobin. The circular dichroism spectra of leghemoglobin a from the root nodules of soybean have been compared with those for sperm whale myoglobin in the fat- and near-ultraviolet and the Soret and visible regions of the spectrum. Circular dichroism spectra in the far-ultraviolet show that the leghemoglobins all have a high alpha-helix content (soybean leghemoglobin a, 55%) regardless of the nature of bound ligands and oxidation or spin state of the heme iron. The known sequence homologies with mammalian hemoglobins may therefore be reflected in conformational homologies as suggested by the x-ray studies of Vainshtein et al. ((1975) Nature (London) 254, 163-164) on lupin leghemoglobin. Removal of the heme moiety decreases helicity by only 9% for leghemoglobins, compared with 23% for myoglobin. This, the much smaller heme contribution to the near-ultraviolet circular dichroism than in myoglobin, and the greater accessibility of the heme moiety to aqueous solvent (Nicola et al. (1974), Proc. Aust. Biochem. Soc. 7, 21) suggest that the association between heme and protein is much weaker in leghemoglobins than in myoglobin. The aromatic Soret and visible circular dichroism spectra for all derivatives of leghemoglobin are opposite in sense to those for myoglobin, showing that the patterns of protein side chain contacts with the heme are different in the two classes of heme proteins. There is strong evidence that one of the two tryptophans whose identity and structural role in myoglobin is known, is present also in plant leghemoglobins, hydrogen-bonded and in a similar nonpolar environment whether heme is present or not. The above findings help to explain the remarkably high oxygen affinity and some other ligand-binding properties of leghemoglobins which differ from those of myoglobin.


Intermediates in the limited proteolytic conversion of procollagen to collagen. The conversion of chick bone procollagen to collagen proceeds in a stepwise fashion to produce a limited number of intermediates. Initial proteolytic cleavages remove NH2-terminal nonhelical extensions and yield an intermediate which remains disulfide-bonded via COOH-terminal extensions. Subsequent stepwise scission of one or two chains of the triple-stranded molecule in its COOH-terminal domain produces intermediates which can only be distinguished after dissociation of the noncovalently bonded alpha chains. A final cleavage in this region produces the collagen molecule and a disulfide-bonded triple-stranded fragment which represents the COOH-terminal domain. In all likelihood the endopeptidases which effect cleavage in the NH2- and COOH-terminal regions differ. More than two enzymes may be required for conversion of procollagen to collagen if the nonhelical domains are not released in an en bloc fashion.


Structural and thermodynamic basis of affinity in anti-dinitrophenyl antibody. The thermodynamic quantities of the anti-dinitrophenyl antibody-hapten interaction are reported for rabbit, goat, and guinea pig antibodies. Rabbit and goat antibodies had similar exothermic enthalpy changes for their reaction with 2,4-dinitrophenyl-L-lysine (-13.9 and -14.8 kcal/mol, respectively). The enthalpy change with guinea pig antibody was much less exothermic (-8.7 kcal/mol), and this value was the same for two guinea pig antibody preparations that differed in affinity by almost two orders of magnitude. A heterogeneous goat anti-dinitrophenyl antibody preparation was fractionated on a molecular sieve column in the presence of a bivalent ligand, a procedure that has been reported to separate antibodies according to differences in the depth of interaction with the ligand. The relationship of these differences in apparent site depth to changes in interactions with the hapten tail was examined by comparing the affinities of various fractions for two haptens. The results show that the presumed deeper sites have stronger interactions with the hapten tail. These studies suggest that the heterogeneity of anti-dinitrophenyl antibodies with respect to affinity results from differences in entropy driven lysyl side-chain interactions which arise from a heterogeneity in antigen binding site depth.


The mechanism of action of dipeptidyl aminopeptidase. Inhibition by amino acid derivatives and amines; activation by aromatic compounds. A variety of amino acid and peptide amides have been shown to be inhibitors of dipeptidyl aminopeptidase. Among these compounds derivatives of strongly hydrophobic amino acids are the strongest inhibitors (Phe-NH2, Ki = 1.0 +/- 0.2 mM), while amides of basic amino acids were somewhat less effective (Lys-NH2, Ki = 36 +/- 3 mM). Short chain amino acid amides are notably weaker inhibitors (Gly-NH2, Ki = 293 +/- 50 mM). The interaction of the side chains of compounds with the enzyme appears to be at a site other than that at which the side chain of the amino-penultimate residue of the substrate interacts since the specificity of binding is different. Primary amines have been shown to inhibit, e.g., butylamine, Ki = 340 +/- 40 mM, and aromatic compounds have been shown to stimulate activity toward Gly-Gly-NH2 and Gly-Gly-OEt (phenol, 35% stimulation of activity at a 1:1 molar ratio with the substrate). The data suggest that inhibition involves binding at the site occupied by the free alpha-amino group and the N-terminal amino acid.


Specific transformations at the N-terminal region of phospholipase A2. Treatment of porcine pancreatic prophospholipase A2 with methyl acetimidate converted all lysine residues into epsilon-acetimidolysine residues. Enzymatically active epsilon-amidinated phospholipase A2 (AMPA) was obtained from the epsilon-amidinated zymogen by limited tryptic proteolysis cleaving the Arg7-Ala8 bond. AMPA was used to prepare des-Ala8-, des-(Ala8,Leu9)- and des-(ALa8),Leu9,Trp10)-AMP by successive Edman degradations, and des-(A la 8-Arg13)-AMPA by selective splitting of the Arg13-Ser14 bond by trypsin. Structural analogues of AMPA with different N-terminal amino acid residues, viz., D-Ala, beta-Ala, and Gly, have been prepared by reacting des-Ala8-AMPA with the corresponding N-t-Boc-N-hydroxysuccinimide esters of these amino acids. Similarly, the only Trp10 residue has been substituted for Phe by coupling of des-(Ala8-,Leu9,Trp10)-AMPA with N-t-Boc-L-Ala-L-Leu-L-Phe-N-hydroxysuccinimide ester. The feasibility of these substitutions has been proven unambiguously by the retroconversion of des-Ala8-AMPA and of [Ala7]AMPA into AMPA having identical enzymatic activity as the starting AMPA. The single Trp10 residue in native phospholipase A2 and its zymogen was specifically sulfenylated using 0-nitrophenyl-sulfenyl chloride. The homogenous proteins were kinetically analyzed using short-chain lecithins in the monomeric and micellar region. All modified AMPA analogues, except those in which two or more of the N-terminal amino acid residues are removed, show enzymatic activities toward monermic substrate comparable to that of AMPA, indicating that the active site region is still intact. Only [Gly8]-, [beta-Ala8]-, and [Ala8,Leu9,Phe10]AMPA exhibit a dramatic increase in enzymatic activity similar to that of AMPA upon passing the critical micellar concentration (cmc) of the substrate. From these results it can be concluded that the N-terminal region of the enzyme requires a very precise architecture in order to interact with lipid-water interfaces and consequently to display its full enzymatic activity.


13C Fourier transform nuclear magnetic resonance studies of fractionated Candida utilis membranes. 13C Fourier transform nuclear magnetic resonance has been used to study the lipid structure and dynamics of fractionated Candida utilis cell membranes. Measurements of the spin-lattice relaxation times indicate the existence of mobility gradients in the direction of increased mobility from the glycerol backbone toward the terminal methyl group of the fatty acid and toward the choline methyls. The temperature dependence of the relaxation times gives activation energies of approximately 4-6 kcal/mol for the rotations about various carbon-carbon bonds which determine the relaxation rates. In general, comparison with data which have been reported for artificial membrane systems indicates that the contributions of protein-lipid interactions to the T1 gradient are of negligible importance in the yeast membrane system. A dynamical model for the motion about bonds near unsaturated bonds which determined the relaxation of the unsaturated carbons is also proposed. Measurements of chemical shifts with temperature also exhibit a correlation with chain position. On the basis of these data a correlation of deltaE, the energy difference between gauche and anti conformations for gamma carboms, with chain position is inferred. In addition, an estimate of 1.2 kcal/mol can be obtained for deltaE for carbons near the end of the fatty acid chain. This value indicates that intermolecular interactions contribute substantially to deltaE since a value of approximately 0.5 kcal/mol can be ascribed to intramolecular interactions.


Isolation and partial characterization of the major glycoproteins of horse and swine erythrocyte membranes. The major glycoproteins of horse and swine erythrocyte membranes were isolated and examined chemically and immunologically. The major glycoprotein of horse erythrocyte membranes had a molecular weight of 33 000 and consisted of 46.2% protein and 53.8% carbohydrate, of which 9.4% was hexose, 10.1% hexosamine and 33.7% sialic acid. This glycoprotein was associated with activity for the infectious mononucleosis heterophile antigen. There were two different major glycoproteins in swine erythrocyte membranes. One major glycoprotein had a molecular weight of 46 200 and consisted of 34.2% protein and 65.8% carbohydrate, of which 18% was hexose, 19% hexosamine and 27.2% sialic acid. This glycoprotein had phytohemagglutinin (Phaseolus vulgaris) binding activity. The other glycoprotein had a molecular weight of 29 000 and consisted of 50.4% protein and 49.6% carbohydrate, of which 6.4% was hexose, 7.0% hexosamine and 36.3% sialic acid. This glycoprotein had weak or absent phytohemagglutinin binding activity.


Adaptive regulation of amino acid transport across the cell membrane in avian and mammalian tissues. The regulation of amino acid transport across the cell membrane by adaptive mechanisms has been studied in a variety of mesenchymal and epithelial cells and tissues of avian and mammalian origin. Changes in transport activity as a function of time under various in vitro conditions (amino acid dependence, active and inhibited protein synthesis) have been evaluated by measurements of initial entry rates with representative amino acids. Results and conclusions based on the adopted experimental approach include the following. (1) An adaptive control mechanism for the transport of neutral amino acids corresponding to the typical substrates of the A mediation is operative in (a) mesenchymal cells (fibroblasts, chondroblasts, osteoblasts and myoblasts) from embryonic tissues of avian (chick embryo) origin and (b) mesenchymal cells from immature rat uterus (fibroblasts and smooth muscle cells) and other mammalian tissues (cardiac cells from newborn mouse and rat heart). (2) Adaptive regulation is restricted to a discrete subgroup of amino acids (L-proline, glycine and the analogue alpha-aminoisobutyric acid) in rat peritoneal macrophages and thymic lymphocytes. (3) Adaptive regulation is absent in erythroid cells (human erythrocytes, rabbit erythrocytes and reticulocytes, avian erythrocytes) which lack the A mediation and are incapable of active gene transcription. (4) Adaptive regulation is absent in the epithelial kidney cortex tissue and possibly absent in the epithelial component of liver tissue from adult rats; it is fully operative in the chick embryo crystalline lens, i.e. an epithelial preparation of embryonic origin. (5) These observations indicate that adaptive control mechanisms of amino acid transport across the cell membrane are quite common among tissues and species and emphasize their broad biological significance in eukaryotes.


Indophenyl acetate and acetylcholinesterase: binding of a non-specific substrate on the margin of the active center. 1. Indophenyl acetate is a very poor substrate of eel or bovine acetylcholinesterase (acetylcholine hydrolase, EC 3.1.1.7), with a V less than 5% of that of phenyl acetate, but it is a labile ester and in imidazole buffer is hydrolyzed, non-enzymically, even faster than phenyl acetate. 2. Indophenyl acetate completely protects the enzymes against inhibition by diisopropylphosphorofluoridate but promotes inhibition by methanesulfonyl fluoride. 3. With either of these inhibitors the measured rate of inactivation of eel acetylcholinesterase is the same whether activity is determined with this poor substrate or with a good substrate, acetylthiocholine. With bovine enzyme the inactivation rate is 25% lower when assayed with the former substrate. However this preparation contains a minor enzyme component which is involved in hydrolysis of indophenyl acetate but not good substrates, and which is not readily inhibited. When this is taken into account the inactivation rates for bovine acetylcholinesterase, too, are found to be the same in either assay. These and other observations in the literature can be explained if indophenyl acetate, because of its size, cannot fully penetrate into the active center and is bound in adjoining non-polar regions of the protein. From this external position it makes only intermittent contact with the esteratic site. Hence it is slowly hydrolyzed and fails to protect the enzyme against methanesulfonyl fluoride, though it does protect, possibly sterically, against the larger inhibitor diisopropylphosphorofluoridate.


Purine and pyrimidine inhibitors of arginase. 1. Adenosine, inosine, adenine and uric acid are competitive inhibitors and cytidine and cytosine noncompetitive inhibitors of bovine liver arginase (L-arginine amidinohydrolase, EC 3.5.3.1). 2. The affinity of the enzyme for these inhibitors was 10--100 times as great as for substrate in terms of Ki versus Km. 3. These nucleic acid metabolites may thus function in vivo to regulate the urea cycle. 4. Several naturally occuring competitive and noncompetitive inhibitors of arginase of unknown structure have been isolated from plant and animal tissue. From their properties and methods of isolation, they may be the purines and pyrimidines herein described. 5. These purines and pyrimidines have no effect on tryptic hydrolysis.


Pallidin. Purification and characterization of a carbohydrate-binding protein from Polysphondylium pallidum implicated in intercellular adhesion. A carbohydrate-binding protein from Polysphondylium pallidum, a species of cellular slime mold, was purified to homogeneity by adsorption to formalinized erythrocytes and elution with D-galactose. The protein, for which we propose the name PALLIDIN, is assayed by its activity as an agglutinin of erythrocytes. It was previously shown to have different carbohydrate-binding specificities than discoidin, a carbohydrate-binding protein from Dictyostelium discoideum, another species of slime mold. Evidence has been presented previously that each of these proteins is detectable on the cell surface. In the present report we show that the physico-chemical properties of pallidin are different from discoidin. Pallidin has a subunit molecular weight of 24 800 +/- 1100 determined by polyacrylamide electrophoresis in the presence of dodecyl sulfate and 2-mercaptoethanol, compared to 26 100 +/- 1000 for discoidin. The weight-average molecular weight of pallidin is 250 000 +/- 50 000 determined by equilibrium sedimentation in the presence of D-galactose compared to 100 000 +/- 2000 for discoidin. In equilibrium sedimentation studies, pallidin exhibited some heterogeneity at equilibrium while discoidin was homogeneous. The amino acid composition of pallidin is generally similar but clearly different from the composition of discoidin. The isoelectric point of pallidin is 7.0 compared to 6.1 for discoidin. Like discoidin, pallidin contains no detectable hexosamine or neutral sugar. These results establish that agglutinins from two species of cellular slime molds are distinct. The different properties of the cell-surface agglutinins, pallidin and discoidin, are consistent with their suggested role in species-specific cellular recognition and adhesion in the species of slime mold from which they are derived.


The effect of calcium on the tryptic digestion of bovine intestinal calcium-binding protein. The tryptic hydrolysis of bivine intestinal calcium-binding protein in the presence and absence of excess calcium has been investigated. Calcium-binding activity and immunological reactivity of the protein were not significantly affected in the presence of 1.0 mM CaCl2 following 24 h incubation at 38 degrees C with trypsin at ratios of 1:9 of enzyme to calcium-binding protein. Some modification of the protein did occur under these conditions, however, since analysis by analytical acrylamide gel electrophoresis indicated the formation of a more rapidly-migrating species from the slower-moving original protein band. Omission of added calcium from the incubation medium resulted in rapid and essentially complete destruction of calcium-binding activity and immunological reactivity, and the formation of peptides of low molecular weight. This provides evidence that the conformation of the calcium-binding protein in the presence of calcium differs from that in its absence.


The structure, physical and chemical properties of the soy bean protein glycinin. The major storage protein of the soybean, glycinin, has been prepared in a homogeneous form and examined by a variety of techniques. It has been found that the protein has a molecular weight of 320000 and contains two sizes of subunits with different isoelectric points. There are six acidic subunits of approximately 35000 and six basic of approximately 20000. Analysis revealed three different kinds of acidic subunits and probably three kinds of basic ones also. These twelve subunits are packed in two identical hexagons, placed one on the other, yielding a hollow oblate cylinder of 110 X 110 X 75 A. Some or all of the subunits are non-spherical resulting in a partial blocking of the central hole. Information about the forces stabilzing the native structure is also discussed.


Surface distribution of the fatty acid side chains of phosphatidylethanolamine in mixed phospholipid vesicles. A method has been developed for the selective determination of the fatty acid side chain distribution associated with the amino containing phospholipids located in the inner and outer surfaces of membranes. Using sonicated phosphatidylethanolamine/phosphatidylcholine vesicles as a model, the analysis consists of selective labeling of the outer surface amino groups with the membrane impermeable reagent 2,4,6-trinitrobenzenesulfonic acid. Outer and inner surface phosphatidylethanolamine fractions are separated by thin-layer chromatography. Analysis of methyl esters derived from these two fractions, by gas-liquid chromatography, yields the fatty acid side chain distribution. Our results show that there is no mol fraction dependence of the incorporation of any specific fatty acid side chains of egg yolk phosphatidylethanolamine into the vesicle or any preferential distribution of these side chains in the inner or outer vesicle surface. The surface distribution of the egg yolk phosphatidylethanolamine molecules in these vesicles appears to be determined by the head group packing requirements and not the fatty acid side chain composition.


Step conductance increases in bilayer membranes induced by antibody-antigen-complement action. A sharp rise in the electrical conductance of lipid bilayer membranes was observed following the addition of antigen (bovine serum), antibody (rabbit anti-bovine serum), and complement to the neighboring aqueous phases. At low concentrations, step increases in the conductivity occurred which are consistent with the appearance of about 2.2 nm holes in the membrane. Probably attack or lysis of the lipid bilayer by complement is responsible.


Structural studies of biological membranes and related model systems by Raman spectroscopy. Sphingomyelin and 1,2-dilauroyl phosphatidylethanolamine. Raman spectra are reported at relatively high resolutions (1.5 cm-1) for sphingomyelin and for 1,2-dilauroyl phosphatidylethanolamine above and below their melting temperatures. The spectra of 1,2-dilauroyl phosphatidylethanolamine below Tm show the hydrocarbon chains to be less ordered in the solid phase than the fatty acid of the same chain length, without the significant occurrence of guache isomers. The spectra of sphingomyelin show significant formation of gauche isomers below Tm, indicating a less rigid environment for this molecule in the solid state.


Cell cycle dependent exposure of a high molecular weight protein on the surface of mouse L cells. The non-penetrating lactoperoxidase iodination probe has been employed in conjunction with synchronously dividing populations of mouse L cells to identify a high molecular weight protein is preferentially exposed on the L cell surface during G1 phase of the cell cycle. Progression of cells from G1 to S is accompanied by a marked decrease in tha availability of this structure, called band 1, to lactoperoxidase-catalyzed iodination and it remains unavailable until cells re-enter G1. It is suggested that the band 1 polypeptide may be functionally involved in the regulation of L cell growth.


Fluctuations in deoxyribo- and ribonucleoside triphosphate pools during the mitotic cycle of Physarum polycephalum. Fluctuations in the pools of deoxyribo- and ribonucleoside triphosphates have been measured during the synchronous mitotic cycle of the slime mould Physarum polycephalum. The most pronounced fluctuation of the deoxyribonucleoside triphosphates was seen shortly before and after initiation of DNA synthesis. The pools of dTTP, dATP, dCTP and dGTP expanded before initiation of DNA synthesis and decreased again in early S phase. The pools of ribonucleoside triphosphates increased during mitosis and again 1 h after mitosis and 5 h after mitosis.


Theory of hapten binding to IgM: the question of repulsive interactions between binding sites. Various workers in their studies of the binding of haptens to IgM have observed that at low hapten concentration IgM has an apparent valence of five or near five, while at high hapten concentration IgM has a valence of ten. A possible explanation for this is that there is an interaction between binding sites on the same F(ab')2 region of the IgM molecule. In this paper the theory for such an interaction is presented and an expression for the apparent valence is derived. It is shown that the apparent valence depends on both the interaction between binding sites on the IgM molecule and on the width of the affinity distribution which characterizes the antiserum. A broad affinity distribution can give an apparent valence of five even when there is no interaction between sites, i.e., even when the ten binding sites on the IgM molecule are identical and independent. The general properties of a Scatchard plot are also discussed. When there is no interaction between sites it is shown that the average affinity and the variance of the affinity distribution can be obtained from a Scatchard plot. To illustrate the theory, an antiserum with affinities characterized by a normal distribution is considered and a simple method is presented for determining alpha, the parameter which measures the width of the normal distribution.


Sugar production from agricultural woody wastes by saccharification with Trichoderma viride cellulase. The saccharification of agricultural woody wastes was studied using a commercial enzyme preparation, Cellulase onozuka, derived from Trichoderma viride or the solid culture extracts of the fungus. With the intention of producing sugar at low cost, a simple procedure of enzymatic saccharification of rice straw, bagasse, and sawdust was studied. Delignifying methods of these wastes were investigated using dilute sodium hydroxide solution and dilute peracetic acid. Rice straw and bagasse were effectively delignified by boiling in a 1% sodium hydroxide solution for 3 hr or by autoclaving at 120 degrees C in a 1% sodium hydroxide solution for 1 hr. The sawdust from a broad leaved tree (Machilus thunbergii) was delignified by autoclaving at 120 degrees C in a 1% sodium hydroxide solution for 1 hr and by subsequent boiling in diluted 1/5 peracetic acid for 1 hr. This type of sawdust was also delignified by boiling in 1/5 peracetic acid for 1 hr and by subsequent autoclaving at 120 degrees C in a 1% sodium hydroxide solution for 1 hr. The sawdust from a coniferous tree (Cryptomeria japonica) was delignified by boiling in 1/5 peracetic acid for 1 hr and by subsequent autoclaving at 120 degrees C in a 1% sodium hydroxide solution for 1 hr; however, the successive treatment by autoclaving with alkali solution and subsequent boiling with diluted peracetic acid failed to bring about the desired effect. The saccharification of delignified rice straw, bagasse, and sawdust was examined using Cellulase onozuka, wheat bran or rice straw solid culture at various substrate concentrations, resulting in the formation of 5 to 10% sugar solutions after incubation at pH 5.0, 45 degrees C for 48 hr. The optimum substrate concentration existed at around 10%. Reuse of cellulase solution and resaccharification of residual sawdust were considered to be inadequate.


[Effect of "immune" RNA on various types of cells interacting in the immune response]. Simultaneous injection of marrow cells previously incubated with "immune" RNA, thymus cells and sheep red cells to X-ray treated mice resulted in a greater accumulation of antibody-forming cells in the spleen, as compared to mice inoculated with intact cells or with those incubated with normal RNA. Preliminary incubation of thymus cells with "immune" RNA did not affect the accumulation of antibody-forming cells. Incubation of peritoneal exudate cells with "immune" or normal RNA failed to influence the mode of accumulation of the antibody-forming cells either.


Infantile autism in twins.

Infantile autism in twins. A pair of MZ male twins concordant for early infantile autism (E.I.A.) is presented. A distinction is drawn between E.I.A. and infantile psychosis (I.P.) with associated mental retardation as a guide to prognosis and the necessity to provide suitable social training and education. The relevance of aetiology is discussed, and the importance of abnormal ante-natal history and delivery status, even in cases with strong genetic predisposition, is noted.


Bilateral vesico-ureteral reflux with pyelonephritis in identical twins. A case of identical twins with bilateral vesico-ureteral reflux is presented. This case supports the hypothesis that primary vesico-ureteral incompetence may be an inherited lesion. The importance of investigating close relatives, especially identical twins, is stressed.


Mastalgia and total body water.

Mastalgia and total body water. Total body water (TBW) was measured early and late in a menstrual cycle in 56 women, 39 of whom had breast pain. The remainder were asymphtomatic controls. Most women did not conform to the traditional view that there is a premenstrual increase in TBW. In some TBW decreased, while in others there was no change from the early cycle measurement. No TBW pattern correlated with any syndromes of breast pain or with any psychoneurotic profile.


Apomorphine-induced stereotyped biting in the tortoise in relation to dopaminergic mechanisms. A biting behavior in the Moroccan pond tortoise (Clemmys caspica leprósa) after apomorphine administration is described. The biting behavior was antagonized by the specific anti-dopaminergic drug haloperidol (10 mg/kg) and trifluperazine (15 mg/kg). The behavior was compared to similar behaviors in birds and mammals. The dopamine metabolites, homovanillic acid and 3,4-dihydroxyphenylacetic acid, were measured by a gas chromatographic method in the tortoise brain. From the effects on behavior and the changes in the level of these metabolites upon drug administration it was concluded that the dopamine system in the tortoise is qualitatively similar to that in birds and mammals but less sensitive to blockade.


Effect of Nepsilon-alkylation of the substrate on the hydrolysis of benzoylglycyl-L-lysine by carboxypeptidase B1. The action of carboxypeptidase B (EC 3.4.12.3) on the benzoylglycyl dipeptides Bz-Gly-Lys(X) where X = methyl, ethyl, propyl, formyl, dimethyl, isopropyl, trimethyl, and benzyl has been investigated. All were hydrolyzed, at a rate decreasing in the order indicated, except where X = trimethyl and benzyl. Compounds where X = dimethyl, formyl, and isopropyl were hydrolyzed very slowly, and did not inhibit the hydrolysis of Bz-Glys-Lys by the enzyme. The kinetic parameters kcat and Km were determined for compounds with X = methyl and ethyl. The observed decrease in rate of hydrolysis of substrates with increasing size of X is consistent with increasing steric hindrance effects arising from the interaction of the Nepsilon-alkyl group with residues of the protein in the cleft which accommodates the substrate side-chain, and resulting in weaker binding of the substrate. Bz-Gly-Lys(Me2) was prepared by reductive methylation of Bz-Gly-Lys. Bz-Gly-Lys(Me3) was prepared by the reaction of Bz-Gly-Lys(Me2) with diazomethane in aqueous solution. Bz-Gly-Lys(Me) and Bz-Gly-Lys(Et) were synthesized by classical coupling procedures from the appropriately protected lysine derivatives.


54Mn uptake by the ovaries and reproductive tract of cycling and anestrous ewes. The uptake of 54Mn by the ovaries and reproductive tract of cycling and anestrous ewes has been investigated following intravenous injection of a single dose of 54MnCl2 and sacrifice of the ewes 6 h later. The uptake of 54Mn was greater in the Graafian follicle and the corpus luteum (CL) of the cycle than in the other components of the ovary. An increased uptake of radioactivity was recorded in the CL of the 11th day of the cycle when compared with that of the 4th day. The uptake of 54Mn was lower in the corpus albicans and follicles. A low uptake of radiomanganese was found also in the various tissues of the reproductive tract. These findings indicate that manganese may play a role in the normal functioning of ovarian activity in the ewe.


Ultrastructural studies of Chlamydia psittaci 6BC in situ in yolk sac explants and L cells: a comparison with gram-negative bacteria. Chlamydia psittaci (6BC) was grown in yolk sac explants and in L cells and fixed by perfusion in situ to provide undamaged material for comparison with gram-negative bacteria. Reticulate, intermediate, and elementary bodies were all seen to lack a well-defined periplasmic space; intermediate and elementary bodies showed condensations of the nucleoid which differ from common bacterial configurations; and the cytoplasm of highly condensed elementary bodies was much more electron dense than that of the gram-negative bacteria, while retaining its basically particulate nature. These important morphological distinctions are interpreted as reflections of a significantly different cellular level of organization in these two groups of organisms. No important morphological differences were noted in comparisons of the chlamydial particles grown in the two different host systems.


The effect of purification on the ultrastructure and infectivity of egg-attenuated Chlamydia psittaci (6BC). A procedure is described for the purification of mixed populations of the three different morphological forms of Chlamydia psittaci (6BC) from infected yolk sac membranes. Elementary bodies and small intermediate bodies are not perceptibly damaged during purification which involves homogenization of the host cells, differential centrifugation, sedimentation through 20% sucrose, and treatment with trypsin. The observation that elementary bodies undergo plasmolysis in 20% sucrose is interpreted as indicating that the cytoplasmic membrane of these particles is intact at that stage in the purification. Reticulate bodies and large intermediate bodies are damaged, to a degree, by the homogenization step. This damage takes the form of discontinuities of the outer envelope membrane, and results in the loss of the regular coccobacillary shape of these particles and in an increase in their size. Treatment with a combination of RNase and DNase was found to cause profound damage to all three morphological forms of the chlamydiae.


Specific inhibition of sclerotium formation by 2-mercaptoethanol and related sulfhydryl compounds in Sclerotium rolfsii. Sclerotium formation in Sclerotium rolfsii was completely inhibited by 2-mercaptoethanol at a concentration of 2-4 mM without any adverse effect on mycelial growth. Concentrations lower than 2 mM had no effect on mycelial growth and sclerotium formation, whereas both were inhibited at concentrations higher than 4 mM. Complete inhibition of sclerotium formation with no effect on mycelial growth was also obtained by propyl mercaptan, 1-butyl mercaptan and 2-butyl mercaptan at a concentration of 0.10 mM. Sclerotium formation was also inhibited by benzyl mercaptan and thioglycolic acid at 0.15 mM and 2-4 mM concentration respectively, whereas it was only partially inhibited by L-cysteine and glutathione at 20 mM. Mycelium grown for 21 days in nutrient medium supplemented with mercaptoethanol at a concentration of 3 mM, when transferred into fresh medium without the chemical, grew normally and produced abundant mature sclerotia. Mercaptoethanol inhibited the initiation as well as the further development of young, unpigmented sclerotia. The mechanism of sclerotium formation was arrested completely when mercaptoethanol was added to the growth medium at any time between inoculation and the appearance of sclerotia of the "development" stage. It is suggested that the specific inhibitory action of mercaptoethanol could be used to study the mechanism of sclerotium formation


Osmophilous fungi in the salt marshes of Kuwait. During an investigation of the mycoflora inhabiting saline soils in Kuwait, special attention was focused on osmophilous fungi. A total of 101 species belonging to 46 genera were encountered from 40 soil samples collected from salt marshes using Czapek's agar supplemented with 40% sucrose. Soil samples were collected from different habitats at different distances from the water edge. Soils near to the water edge were poor in their fungal content, while those taken from areas covered by Juncus arabicus contained highest fungal populations. The recorded genera were classified as follows: 7 were of high frequency of occurrence, 8 moderate, 17 low, and 13 were rare. The order of dominance was Aspergillus, Alternaria, Penicillium, Cephalosporium, Fusarium, Stachybotrys, and Drechslera. Comparison between our results and those in other studies showed that there is no fungal flora characteristic of saline soils.


Evidence for a relationship between chloroquine and complement from studies with lymphocyte mitogens: possible implications for the mechanism of action of chloroquine in disease. Chloroquine increases the inhibition of cultured lymphocytes by high concentrations of phytohemagglutinin (PHA) or concanavalin A (Con A). The inhibition is also increased by complement. Thus chloroquine and complement have similar effects. Time-course studies show that chloroquine increases the rate of onset of the complement-dependent inhibition. In serum preheated to inactivate complement, chloroquine can partially simulate the effect of complement. It is suggested that at certain stages in malaria or autoimmune disease the rate of clearance of parasitized erythrocytes or autoreactive lymphocytes is limited by the concentration of complement. Under these conditions a drug such as chloroquine, which could enhance or simulate the action of complement, might be of therapeutic value.


Concanavalin A as a selective agent in tissue culture for temperature-sensitive hamster cell lines. A novel method for isolating Chinese hamster cell cultures with temperature-sensitive growth properties is described. Concanavalin A can be used as a selective agent in tissue culture to isolate lectin-resistant cell lines which exhibit colony-forming abilities at the nonpermissive temperature relative to the permissive temperature as low as 10(-4) to 10(-6). A general correlation exists between resistance to the lectin and temperature sensitivity.


The function of slime from Physarum flavicomum in the control of cell division. A haploid cell of the myxomycete Physarum flavicomum undergoes cytokinesis, producing a large population of cells. However, after syngamy, cytokinesis no longer occurs but karyokinesis does and subsequent growth results in the formation of a diploid syncytial plasmodium. Slime, which is produced by the plasmodium but not the haploid cells, was aseptically isolated and purified, and tested for its effect as a cytokinetic regulator. Slime (a viscous, high molecular weight, acidic glycoprotein) affected cytokinesis of the haploid myxamoebae growing in pure culture in soluble media, and the effect was concentration dependent. In simple media, a slime concentration of about 6  10(-5) mug protein per cell suppressed cytokinesis about 50%, unequally inhibited the synthesis of protein, RNA, and DNA, but stimulated respiration. The biological activity of slime was not species specific and it also affected the bacterium Bacillus subtilis by inhibiting cytokinesis, stimulating oxygen uptake, and producing an aberrant cell morphology. Slime was inactivated by heat, fragmentation, and incubation with dithiothreitol, mercaptoethanol, and the proteolytic enzyme papain (EC 3.4.22.2). The inhibitory effect of slime on cell division of haploid cells could not be achieved using mucin or various polyanions. The possible role of slime in the production of the diploid syncytium is discussed.


The lipids of four unusual non-pathogenic host-associated spirochetes. The lipid compositions of two spirochetes isolated from the human oral cavity and two isolated from pig feces were examined. These isolates were unusual in that they did not require long-chain fatty acids for growth, as do the other host-associated spirochetes, but rather required isobutyric and valeric acids. Therefore, they could be cultured in a medium free of serum or fatty acid - albumin supplements. The major fatty acids synthesized were normal and iso fatty acids with 14 and 16 carbons. No unsaturated fatty acids were detected, nor were chain lengths longer than 16 carbons. The major complex lipids found were monogalactosyl diglyceride, phosphatidyl glycerol, and bis-phosphatidyl glycerol. Nitrogenous phospholipids, present in Treponema and Leptospira, were not synthesized by these novel strains. The data indicate an intermediate position of these isolates between Treponema and free-living Spriochaeta.


Oxygenated mixed venous PCO2 in healthy subjects. Despite the large number of blood gas measurements made over the years, precise information is lacking on intrasubject and intersubject variability of carbon dioxide tension. Accordingly, the oxygenated mixed venous CO2 tension was measured in 202 healthy volunteers and the influence of potentially disturbing factors such as intake of coffee and food, exercise and menstruation on the stability of this tension was studied in selected subjects. In the 102 men the mean value was 5.1 mm Hg (SD, 2.9 mm Hg), significantly higher (P less than 0.001) than that of the 100 women, 46.03 mm Hg (SD, 2.48 mm Hg). Among the women the value was significantly lower (P less than 0.01) in those taking oral contraceptives (mean, 44.9 mm Hg; SD, 2.3 mm Hg) than in those not taking these agents (mean, 46.5 mm Hg; SD, 2.4 mm Hg). Intrasubject variability of values was greatest in women taking oral contraceptives, mean day-to-day variation over two mentrual cycles being +/- 3.26 mm Hg. After moderate exercise the tension usually returned to +/- 1 mm Hg of resting values within 5 minutes. Intake of food and coffee had negligible effects on the tension.


Cancer incidence in the United States and Puerto Rico. This study compares cancer incidence in the United States National Survey areas and Puerto Rico for the period of 1969 to 1974. There are several differences between the 2 areas. Puerto Rico has a higher incidence of cancer of the mouth, pharynx, esophagus, stomach, cervix uteri, vulva, and penis. The Survey areas have higher incidences of cancer of the colon, rectum, lung, corpus uteri, female breast, and ovary. The differences between sexes are more or less the same as in other countries.


Unscheduled DNA synthesis in cells from N-2-fluorenylacetamide-induced hyperplastic nodules of rat liver maintained in a primary culture system. The level of unscheduled DNA synthesis in the parenchymal cells from hyperplastic nodules and from the entire liver of rats fed N-2-fluorenylacetamide was studied and compared with that of normal liver cells. Measurements of unscheduled DNA synthesis were carried out by the use of a primary liver cell culture system. Livers were perfused with collagenase, the cells from individual hyperplastic nodules, and/or from the whole liver aspirated and plated onto plastic Petri dishes. Simultaneous histochemical measurements of beta-glucuronidase were carried out in the cultured cells as an aid in distinguishing functional cell types. The cells from hyperplastic nodules obtained from the liver during carcinogen feeding survived much longer than normal liver cells in culture. The level of unscheduled DNA synthesis was determined radioautographically after exposing cells to ultraviolet light and incubating with [3H]thymidine. [3H]Thymidine labeling was variable among individual nodules or animals and fluctuated as a function of the number of days in culture. In general, however, the level of unscheduled DNA synthesis in the cells from hyperplastic nodules was always higher than or similar to that of normal liver cells. Thus, the cells of hyperplastic nodules are not more readily transformed into the malignant state than normal cells as a result of their lowered DNA repair mechanisms.


Synthetic and naturally occurring hydrazines as possible cancer causative agents. The various synthetic substituted hydrazines, which cause tumors in animals, are briefly enumerated. To date, 19 of them have proved to be tumorigenic in animals. A number of these chemicals are found today in the environment, in industry, in agriculture, and in medicine, and the human population is exposed to a certain degree to some of them. Hydrazine also occurs in nature in tobacco and tobacco smoke. The three other naturally occurring hydrazine compounds are N-methyl-N-formylhydrazine, which occurs in the wild edible mushroom, Gyromitra esculenta, and beta-N-[gamma-L(+)-glutamyl]-4-hydroxymethylphenylhydrazine and 4-hydroxymethylphenylhydrazine, which are found in the commonly eaten cultivated mushroom, Agaricus bisporus. Tumorigenesis studies with the naturally occurring hydrazines are in progress.


Effects of 4-nitrobenzofurazans and their N-oxides on synthesis of protein and nucleic acid by murine leukemia cells. A series of benzo-2,1,3-oxadiazoles (benzofurazans) and their N-oxides (benzofuroxans) inhibit incorporation of precursors into nucleic acids and protein by murine leukemia cells. At slighly higher levels, substantial single- and double-strand DNA breakage was observed. At still higher concentrations, inhibition of phosphorylation of uridine and thymidine was found. Structure-activity relationships show that only compounds bearing appropriate substitutions at positions 4 and 7 were effective inhibitors of biosynthetic pathways. Such compounds appear to interact with a wide variety of biological systems and may be useful in elucidating modes of macromolecule synthesis.


The inhibitory activities of 2-acetamido-2,3-dideoxy-D-hex-2-enonolactones on 2-acetamido-2-deoxy-beta-D-glucosidase. Treatment of 2-acetamido-2-deoxy-D-mannono-1,4-lactone with dicyclohexylamine in ethanolic solution afforded an unsaturated 1,4-lactone, 2-acetamido-2,3-dideoxy-D-erythro-hex-2-enono-1,4-lactone (1), in good yield. 2-Acetamido-2,3-dideoxy-D-threo-hex-2-enono-1,4-lactone (2) was similarly prepared from 2-acetamido-2-deoxy-D-galactono-1,4-lactone. An unsaturated 1,5-lactone, 2-acetamido-2,3-dideoxy-D-threo-hex-2-enono-1,5-lactone (4), was obtained through the oxidation of 2-acetamido-2-doexy-4,6-0-isopropylidene-D-galactopyranose with silver carbonate on Celite, followed by mild hydrolysis. The inhibitory activity of four isomeric 2-acetamido-2,3-dideoxy-D-hex-2-enonolactones [1, 2, 4, and 2-acetamido-2,3-dideoxy-D-erythro-hex-2-enono-1,5-lactone (3)] was assayed against 2-acetamido-2-deoxy-beta-D-glucosidase from bull epididymis. Only the erythro lactones 1 and 3 are weak competitive inhibitors, whereas the threo lactones 2 and 4 are practically inactive. The 1,4-lactone 1 inhibited 2-acetamido-2-deoxy-beta-D-glucosidase more strongly than the 1,5-lactone 3. The lactones 1-4 were found to be quite stable in aqueous solution or under inhibitory-assay conditions. In addition, two 2-acetamido-2-deoxy-D-glycals, 2-acetamido-1,5-anhydrohex-1-enitol (7) were tested; both are 10 times as active as 1.


Spermatogenetic clones developing from repopulating stem cells surviving a high dose of an alkylating agent. We investigated stem cell renewal and differentiation in 10- and 15-days-old spermatogonial clones developing in mouse seminiferous epithelium after an extremely large cell loss, inflicted by high doses of the alkylating agent Myleran. The spermatogonial clones arise from cells that resemble the Ais spermatogonia but have a larger nuclear diameter. In spite of their mitotic activity these 'repopulating stem cells' lie mainly isolated or in pairs. This explained by migration and differentiation. Migration appeared to occur at random in all directions along the basement membrane of the seminiferous tubule. After one or more divisions of the stem cells, a second type of cell appears, which is called the 'differentiating spermatogomium'. The time elapsing before this type of cell appears, depends on the dose of Myleran: the larger the dose the later differentiation starts. A relation could be demonstrated between the stage of the cycle of the seminiferous epithelium and the start of differentiation. Differentiating cells were found isolated or in groups of two, four, eight or sixteen cells. Hence we concluded that at least up to their fourth division differentiating cells divide synchronously without degenerations. Three types of division of repopulating stem cells were distinguished, producing (1) two repopulating stem cells, (2) one repopulating stem cell and one cell starting spermatogonial differentiation, or (3) two differentiating cells. Type 1 divisions were found most frequently.


The fine structure of the epidermis of two species of salmonid fish, the Atlantic salmon (Salmo salar L.). and the brown trout (Salmo trutta L.). The fine structure of epidermal mucous cells of two species of salmonid fish has been described. Mucous cells are, next to filament-containing cells, the most commonly encountered cells in fish epidermis. The development of the cells as they progress to the periphery has been characterised. They are initially difficult to distinguish from filament-containing cells: later, they can be recognised by the presence of much smooth-surfaced E.R. The mucigenesis and the subsequent secretion of mucus has been observed and it is essentially comparable to that which occurs in the mucous cells of the mammalian intestine. The mucous layer of the epidermal surface seems to mainly comprise of the products of these mucous cells and the "cuticle" seen in other species has not yet been observed in the salmonid species investigated here.


Chromaffin cells in the glomerular zone of adult rat adrenal cortex. The occasional presence of islets of chromaffin cells in the glomerular zone of the adrenal cortex of adult rats, is reported in this light and electron microscope study. A possible error in organogenesis of the gland and the possible persistence of some foetal characteristics in these ectopic cells are discussed.


Interaction of ethidium bromide with DNA as studied by kinetic spectrophotometry. The interaction of ethidium bromide (EB) with DNA has been investigated using the pulse radiolysis technique. In particular, the absolute rate constant for the reaction of hydrated electrons, generated by single pulses of high-energy electrons, with EB is shown to drop dramatically in the presence of DNA. This drop in diffusion-limited reactivity results from the interaction of EB with DNA, effectively immobilising it, thus lowering the reaction cross-section or probability. Analysis of the resulting kinetic spectrophotometric data shows that they are consistent with a reversible interaction of EB with DNA as described by the law of mass action. The Scatchard-type plots obtained are linear, and give quantitative information on the extent and degree of association, comparable with that obtained by more conventional methods. The potential of the pulse radiolysis technique for studying different types of interactions between small molecules and various biopolymers has been demonstrated.


Investigations into the mechanism of action of anti-tumour platinum compounds: time- and dose-dependent changes in the alkaline sucrose gradient sedimentation profiles of DNA from hamster cells treated with cis-platinum (II) diamminedichloride. Under the conditions of low speed centrifugation used in this study, the proportion of radioactively labelled DNA from Chinese hamster V79-379A cells sedimenting to the 700S region of an alkaline sucrose gradient was increased in a dose-dependent manner by prior treatment of the cells for 2 h with cis-platinum (II) diamminedichloride [cis-Pt(II)]. This increase was at the expense of material sedimenting in the 400-650S region. This profile was not modified by a 2-h post-treatment incubation prior to centrifugation. 6 h after treatment, the DNA from treated cells sedimented in a narrow band at a position corresponding to 350S. 21 h after exposure to the drug, a dose-dependent restitution of the DNA species sedimenting in the 450-650S range was observed. These results combined with other data relating to platinum binding allow the following conclusions to be reached: (1) cis Pt(II) treatment does not lead to the rapid formation of single-stranded breaks or alkali-labile sites in cellular DNA. (2) The time-dependent changes in sedimentation rate of DNA from treated cells may reflect the transient appearance of gaps following endonuclease attack at platinum-bound sites in DNA. (3) The likely ratio of inter to intra-strand DNA-platinum interactions suggests that such endonuclease attack is primarily at platinum induced inter-strand DNA cross-links.


Measurement by radioimmunoassay of unconjugated estriol in pregnancy serum. Antisera to estriol 6--onjugates were tested for suitability in the direct radioimmunoassay of unconjugated estriol in extracts of pregnancy serum. Assessment of specificity through titration with competing steroids allowed a selection to be made within the group of antisera. The measurement was then standardized by checking the absence of analytical blank (solvent and sample blank) and the extent of binding variability associated with the use of charcoal-dextran as a separating agent. Validation was made by means of the usual recovery and dilution tests, and by a cross-comparison of analytical data obtained with different antisera; reproducibility of calibration curve and within-and between-assay variability was evaluated under routine conditions. The validity of the clinical information was assessed by assaying 202 samples randomly collected throughout normal pregnancy from the 16th week to term: both trend and levels of unconjugated estriol concentration were found to be in good agreement with the literature data.


An enzymic method for the determination of the glycine/taurine ratio of conjugated bile acids in bile. A method is described in which the ratio of the glycine- to taurine-conjugated bile acids (G/T ratio) in bile is determined. After pretreatment of the bile for removal of the lipids, the bile acids are deconjugated enzymically with choloylglycine hydrolase (EC 3.5.1.24). The amino acids taurine and glycine are liberated in this way, after appropriate isolation, taurine and/or glycine are then determined with ninhydrin, enabling the establishment of the G/T ratio. A nearly complete hydrolysis was obtained for 6 conjugated bile acids, while the recovery of these acids when added to hog or ox bile was quantitative. The mean G/T ratio for hog bile, ox bile and human B-bile was 6.3, 2.5 and 2.0, respectively. The amount of total, free and conjugated bile acids can be determined by this method, combined with the 3alpha-hydroxysteroid dehydrogenase technique for bile acid determination described by Iwata and Yamasaki [1]. A high G/T ratio was observed in 3 cases of Crohn's disease in the small bowel, but the extent of deconjugation in B-bile was lower than in duodenal fluid. The determination of the G/T ratio can be complementary to our knowledge of the metabolism of bile salts in certain gastro-intestinal disorders.


Late streptokinase therapy in thrombotic microangiopathy: a case study. A 42 year woman presented with malignant hypertension, anuria and hemolytic anemia with schistocytosis. The diagnosis of thrombotic microangiopathy was confirmed by early renal biopsy. Purely symptomatic treatment (peritoneal dialysis and hypotensive drugs) was supplemented by administration of heparin and Dipyridamole. Gastro-intestinal bleeding prevented early thrombolytic therapy. Microangiopathic anemia rapidly disappeared but anuria persisted. Three months later a second renal biopsy showed persistence of active lesions and absence of irreversible parenchymal damage. Streptokinase treatment was then instituted and followed by a rapid return of urinary output. Hemodialysis was stopped and renal function continued to improve over the following months. Two years later the patient remains well despite persistence of hypertension difficult to control. Creatinine clearance is stable at 20 ml/min. This observation suggests that late thrombolytic therapy may be effective in patients with thrombotic microangiopathy when histological findings do not indicate extensive irreversible lesions.


Psychological aspects of treatment and rehabilitation of the spinal cord injured person. The various phases of the usual or normal reaction to the experienced loss which follows a spinal cord injury include: denial, withdrawal, internalized hostility, externalized hostility, and reaction against dependence. There are also various implications of the major categories or pre-existing psychopathology as they interact with a spinal cord injury in the rehabilitation processes. The major forms of pre-existing psychopathology are: depression, schizophrenia, personality disorders, and neuroses. The psychological management of pain problems is facilitated by a phenomenological classification of pain sensations from the paralyzed portion of the body. Sexual functioning is an early concern of the patient and the rehabilitation program should consider his individual requirements. The family is an important part of the total rehabilitation program.


Effect of phenylalanine and its metabolites on the metabolism of leucocytes and lymphocytes. 1. The pathogenesis of the mental retardation in phenylketonuria remains obscure. Leucocytes have proved of value in the study of other inborn errors of metabolism. The lymphocyte is a suitable model cell for the study of mammalian metabolism, because of its ability to divide in vitro in response to various stimuli. 2. We have examined the effects of phenylalanine, phenylpyruvate, phenyl-lactate and phenylacetate on the human leucocyte and the resting and phytohaemagglutinin-stimulated rabbit lymphocyte. 3. Phenylpyruvate and phenyl-lactate reduced acetate incorporation into leucocyte lipid by 38% and 48% respectively. Only phenyl-lactate reduced acetate incorporation into the resting and stimulated lymphocyte, by 20% and 34% respectively. 4. Glucose incorporation into leucocyte lipid was unaffected by phenylalanine, phenylpyruvate and phenyl-lactate. Only phenyl-lactate inhibited (46%) the production of CO2 from glucose. 5. Phenylalanine and leucine incorporation into trichloroacetic acid-insoluble material of resting and stimulated lymphocytes was inhibited by phenyl-lactate (10-42%), phenylpyruvate (27-57%) and phenylacetate (19-39%). 6. Uridine incorporation into resting and stimulated cells was inhibited by phenyl-lactate (22-26%), phenylpyruvate (42-52%) and phenylacetate (20%). 7. Thymidine incorporation into resting lymphocytes was reduced by phenyl-lactate, phenylpyruvate, phenylacetate and phenylalanine by 12-26%. Incorporation into the stimulated cell was inhibited by phenylpyruvate and phenyl-lactate (90%) and phenylacetate (66%). 8. Phenylalanine inhibited lymphocyte pyruvate kinase and phenylpyruvate inhibited citrate synthetase. 9. These results are compared with published data relating to experimental hyperphenylalaninaemia and the effects of these metabolites on nervous tissue in vitro.


Salt (sodium) deficiency in dairy cattle: polyuria and polydipsia as prominent clinical features. Polyuria in a herd of forty-five dairy cattle was the chief complaint in the first case. Polydipsia was the principle sign in a second case involving one dairy cow. Less conspicuous clinical findings included salt hunger, pica, weight loss and decreased milk production. Clinicopathologic investigation included monitoring the concentrations of sodium, potassium and chloiride in urine, parotid saliva and plasma. These analyses indicated a primary sodium deficiency which responded to sodium chloride ad libitum.


Hematology of the neonatal calf. III. Frequency of congenital iron deficiency anemia. The hematocrits of 258 Holstein-Friesian and Jersey calves were determined during the first 3 days following birth. Hematocrit values of 25% or less were observed in 15.8% of the calves and hematocrits of 20% or less were observed in 6.9% of calves from apparently normal dams. The serum iron concentration of neonatal calves with hematocrits of 25% or less was significantly lower than the value recorded in calves with hematocrits above 25% and there was a direct correlation between PCV and serum iron concentration. The data presented suggested that prenatal development of iron deficiency is an important cause of anemia in neonatal dairy calves. Factors which may be important in pathogenesis are discussed.


Regional mapping of the human No. 1 and X chromosome in interspecific cell hybrids using an X/1 translocation. Fibroblasts from a carrier of an X/1 translocation, 46,XY,t(X;1)(q28;q31), were fused with Chinese hamster cells. The resulting hybrids were analyzed for human No. 1 and X-chromosome markers. The data indicate that the loci for PGM1, PGD, PPH, and GuK1 are situated either in the long arm proximal to a break point in band 1q31 or in the short arm. The loci for Pep-C, FH, and GuK1 are located distal to the break point. HPRT and G6PD are probably situated distal to a break point in band q28 of the X chromosome; alpha-Gal A is situated proximal to the break point, either on the long or short arm of the chromosome.


Treadmill arrhythmias in patients with idiopathic hypertrophic subaortic stenosis. Treadmill tests were performed in 19 patients with previously documented idiopathic hypertrophic subaortic stenosis (IHSS). Arrhythmias occurred in 79 percent (15) of the patients, and new arrhythmias not previously documented occurred in over 50 percent (10) of the patients. Paroxysmal supraventricular tachycardia (PSVT), ventricular premature beats (VPBs) (two or more per minute) or atrial premature beats (APBs) (three or more per minute) occurred in 10 of 19 patients. There was no association between treadmill arrhythmias and clinical symptoms, hemodynamic data, or electrocardiographic features. Propranolol administration resulted in failure of exercise to induce PSVT in one patient and had no effect on PSVT in two others, nor any effect on maximum frequency of APBs or VPBs. Treadmill testing is more productive than retrospective analysis of ECGs for characterizing arrhythmias in IHSS.


Tactile stimulation and behavioral development among low-birthweight infants. The Neonatal Behavioral Assessment Scale was administered to a group of premature infants both prior to and following a nursery program of enriched tactile stimulation. The same instrument was administered twice to a control group of prematures who received no intervening "extra" stimulation. Whereas control infants showed changes of 2 or more points on only 2 of the instrument's 26 scales, the experimental babies showed positive changes of 2 or more points on 11 scales. There were no difference in weight gain between the two groups.