Effect of mutations in the qa gene cluster of Neurospora crassa on the enzyme catabolic dehydroquinase. Catabolic dehydroquinase, which functions in the inducible quinic acid catabolic pathway of Neurospora crassa, has been purified from wild type (74-A) and three mutants in the qa gene cluster. The mutant strains were: 105c, a temperature-sensitive constitutive mutant in the qa-1 regulatory locus; M-16, a qa-3 mutant deficient in quinate dehydrogenase activity; and 237, a leaky qa-2 mutant which possess very low levels of catabolic dehydroquinase activity. The enzymes purified from strains 74-A, 105c, and M-16 are identical with respect to behavior during purification, specific activity, electrophoretic behavior, stability, molecular weight, subunit structure, immunological cross-reactivity, and amino acid content. The mutant enzyme from strain 237 is 1,500-fold less active and appears to have a slightly different amino acid content. It is identical by a number of the other criteria listed above and is presumed to be a mutant at or near the enzyme active site. These data demonstrate that the qa-1 gene product is not involved in the posttranslational expression of enzyme activity. The biochemical identity of catabolic dehydroquinase isolated from strains 105c and M-16 with that from wild type also demonstrates that neither the inducer, quinic acid, nor other enzymes encoded in the qa gene cluster are necessary for the expression of activity. Therefore the combined genetic and biochemical data on the qa system continue to support the hypothesis that the qa-1 regulatory protein acts as a positive initiator of qa enzyme synthesis.


Characteristics of a glycerol utilization mutant of Neurospora crassa. A mutant of Neurospora crassa able to grow on liquid minimal glycerol medium without evidence of conidiation and with high cell yields has been isolated and shown to be allelic to ff-1. The glycerol-specific induction of glycerokinase and glycerol-3-phosphate dehydrogenase was similar in both wild-type and mutant cells, although higher specific activities as well as higher glycerokinase cross-reacting material levels were found in fully induced mutant cells. After growth in minimal glycerol medium there is a significant reduction in wild-type cells of the activities of both pyruvate dehydrogenase and dihydrolipoyl transacetylase. This evidence indicates a relationship between the conditional acetate requirement by wild-type cells grown on glycerol medium and the levels of the pyruvate dehydrogenase complex.


Purification and properties of the fatty acids synthetase complex from Neurospora crassa, and the nature of the fas-mutation. A procedure is described for the purification of the fatty acid synthetase complex (FAS) from Neurospora crassa. The enzyme complex has a molecular weight of 2.3 times 10(6), contains 6 mol of 4'-phosphopantetheine per mol, and on polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate gives a single band, or a closely spaced doublet, which comigrates with standard myosin (molecular weight, 2 times 10(5)). Since the slightly retarded component in the doublet accounts for all protein-bound 4'-phosphopantetheine, the complex appears to be made up of 11 to 12 equally sized subunits, 6 of which carry the acyl carrier protein function. In this unusual arrangement, notably the lack of the low-molecular-weight acyl carrier protein component seen in other FAS systems, as well as in its enzymatic properties, the Neurospora FAS complex is quite similar to the yeast enzyme. The FAS complex of a saturated fatty acid-requiring mutant, previously disignated cel-, contains less than 2% of the 4'-phosphopantetheine prosthetic groups found in the wild-type complex. The leaky phenotype of this mutant, here designated fas-, is accounted for by a residual fatty acid synthesizing activity in its FAS complex, which is several-fold higher than expected from its residual content of 4'-phosphopanthetheine.


Effect of histidine on purine nucleotide synthesis and utilization in Neurospora crassa. Histidine affects de novo purine nucleotide synthesis and purine nucleotide pool utilization in Neurospora crassa. The former effect was assessed qualitatively by the presence or absence of purple pigment production in ad3B and ad3A mutants. Tryptophan also affected the de novo purine nucleotide synthesis. The effect of histidine on purine nucleotide pool utilization resulted in stimulated germination of ad8 and ad4 mutant conidia in adenine-deficient medium. Increased germination was correlated with increased net levels of nucleic acids in these strains. Possible mechanisms for the dual action of histidine are discussed.


Glycerophospholipid variation in choline and inositol auxotrophs of Neurospora crassa. Internal compensation among zwitterionic and anionic species. The glycerophospholipids of cultures of Neurospora crassa were extracted, deacylated, and analyzed. In addition to a wild-type strain, several auxotrophic mutant strains were examined: chol-1 (defective S-adenosylmethionine: phosphatidylethanolamine methyltransferase), chol-2 (defective S-adenosyl methionine:phosphatidylmonomethylethanolamine (dimethylethanolamine) methyltransferase), and inos (defective myoinositol-1-phosphate phosphatase). In addition, a double mutant strain, chol-1;chol-2, was constructed. Cultures of the mutant strains grown with concentrations of supplement(s) just adequate to support growth had bizarre phospholipid compositions. By appropriate choice of mutant and supplement(s), it was possible to vary the relative level of every phospholipid of the organism, with the exception of cardiolipin. The maximum ranges encountered for the zwitterionic species, expressed as per cent of total phospholipid phosphorus, were lecithin (0.9 to 53.1%), phosphatidyldimethylethanolamine (0.0 to 55.5%), phosphatidylmonomethylethanolamine (0.0 to 53.9%), and phosphatidylethanolamine (9.8 to 43.3%). For the anionic species, the ranges were phosphatidylserine (1.7 to 10.4%) and phosphatidylinositol (3.6 to 25.1%). Despite the wide variation of the relative proportions of the individual phospholipid species, five quantities remained constant: the cardiolipin content, the total phospholipid content, the total content of the zwitterionic species, the total content of the anionic species, and the ratio of the zwitterionic to anionic totals. The data suggest the existence of an internal compensation mechanism, the net effect of which is maintenance of a fairly constant contribution by the phospholipid components to the over-all membrane charge.


A re-examination of the cleavage of fibrinogen and fibrin by plasmin. Three Fragment D species (D1, D2, D3) were isolated with time from a plasmin digest of fibrinogen and had molecular weights of 92,999, 86,000 and 82,000 by summation of subunit molecular weights from sodium dodecyl sulfate polyacrylamide gel electrophoresis. Their molecular weights by sedimentation equilibrium ultracentrifugation were 94,000 t87,000, 88,000 to 82, 000, and 76,000 to 70,000 depending on the values calculated for the partial specific volumes. Each of the Fragment D species contained three disulfide-linked subunits derived from the Aalpha, Bbeta, and gamma chains of fibrinogen and differed only in the extent of COOH-terminal degradation of their gamma chain derivatives. Plasmin cleaved Fragment D1 to release the cross-link sites from its gamma' subunit of 38,000 molecular weight; however, the beta'' subunit of 42,000 molecular weight and the alpha'' subunit of 12,000 molecular weight were resistant to further digestion by plasmin. Fragment D isolated from highly cross-linked fibrin had a dimeric structure due to cross-link formation between the gamma' subunits of two fibrinogen Fragment D species. The molecular weight of fibrin Fragment D was 184,000 by summation of subunit molecular weights and 190,000 to 175,000 by sedimentation equilibrium. Cross-linking the gamma chain, as well as incorporating the site-specific fluorescent label monodansyl cadaverine into the gamma chain cross-link acceptor site, prevented its COOH-terminal degradation by plasmin. Therefore, only one species of fibrin Fragment D, as well as only one species of monodansyl cadaverine-labeled fibrin Fragment D monomer, was generated during plasmin digestion. These results show unequivocally that each fibrinogen Fragment D contains only three subunit chains and therefore the digestion of fibrinogen by plasmin must result in the production of two Fragment D molecules from each fibrinogen molecule. The recently proposed model of fibrinogen cleavage that postulates the generation of a single Fragment D with three pairs of subunit chains from each fibrinogen molecule is incorrect. Incorporation of monodansyl cadaverine into the cross-link acceptor sites of the alpha chain did not alter its cleavage by plasmin detectably. A series of monodansyl cadaverine-labeled peptides, which ranged in molecular weight from 40,000 to 23,000, were cleaved from the alpha chain of monodansyl cadaverine-labeled fibrin monomer during the early stages of plasmin digestion. These peptides were degraded progressively to a brightly fluorescent plasmin-resistant peptide of 21,000 molecular weight and a weakly fluorescent peptide of 2,500 molecular weight. Thus both alpha chain cross-link acceptor sites are contained within a peptide segment of 23,000 molecular weight.


Transport and inhibitory Ca2+ binding sites on the ATPase enzyme isolated from the sarcoplasmic reticulum. Ca2+ binding sites located on the Ca2+-dependent ATPase purified from the fragmented sarcoplasmic reticulum (Ikemoto, N (1974) J. Biol. Chem. 249, 649) have been further studied. At 0 degrees there are three classes of binding sites denoted as alpha (K congruent to 3 times 10(61 M-1), beta(K congruent to 5 times 10(4) M-1), and gamma (K congruent to 1 times 10(3) M-1) sites. At 22 degrees there is no beta site but there are about two alpha sites per 10(5) daltons, while at 0 degrees there is one alpha and one beta site. The change is reversible. The parallelism between the temperature-induced changes in the alpha site and the reported (Sumida, M., and Tonomura, Y. (1974) J. Biochem. 75, 283) temperature dependence of the ratio of Ca2+ transport and ATP cleavage (deltaCa2+/deltaATP is 2 at 22 degrees and 1 at 0 degrees) suggests the involvement of the alpha site in transport. Studies at a low ATP to enzyme ratio (0.5 to 2.5 mol of ATP/10(5) g of ATPase unit) permitting the separate investigation of the phosphorylation and dephosphorylation process show that concomitantly with the formation of the phosphorylated enzyme (E approximately P) bound calcium is released from, and concomitantly with the dephosphorylation it is rebound to, the alpha site. Binding of Ca2+ to the E approximately P moiety inhibits the liberation of Pi. Analysis by use of a Hill plot of the Ca2+ dependence of the inhibition suggests the involvement of two sites with an average affinity of approximately 10(3) M-1. These have tentatively been identified as alpha (low affinity form) and gamma sites.


Variable nature of cartilage proteoglycans. Cartilage proteoglycan aggregates are separated from collagen and other non-proteoglycan protein by preparative rate zonal sedimentation under associative conditions. Dissociative rate zonal sedimentation produces sedimented proteoglycan of lower protein content with a corresponding increase in the amount of less sedimentable protein-rich proteoglycan. An extensive number of sequential rate zonal sedimentations discloses that the proceess of disaggregation involves the separation of proteoglycans varying continuously in composition with no apparent discontinuities in distribution to indicate the presence of distinctively different macromolecules. The variations encompass proteoglycans of low protein content containing less than 2% keratan sulfate and proteoglycans with keratan sulfate as the predominant polysaccharide (present in concentrations greater than 2-fold that of the chondroitin sulfate) and more than a 10-fold increase in protein content.


Antigenic differences in (Na+, K+)-ATPase preparations isolated from various organs and species. Antisera to purified (Na+, K+)-ATPase raised in rabbits and in sheep were purified by an absorption procedure employing purified canine kidney (Na+, K+)-ATPase. The antibodies were fractionated into two components, one which inhibited catalytic activity, and a second which inhibited ouabain binding. Under certain conditions, the fraction that inhibited ouabain binding also inhibited catalytic activity, and the effectiveness of both was dependent to some extent on the ligands present in the incubation medium. Thus, both antibody fractions appeared to detect conformations of the enzyme that depended upon ligand-induced perturbations. When the antibody raised against catalytic activity was incubated with erythrocyte membrane fragments, an inhibition of the (Na+, K+)-ATPase occurred, but only minimal or no effect on potassium influx or on digoxin-induced inhibition of potassium flux in intact erythrocytes was noted. In a similar experiment, however, the antibody against ouabain binding significantly inhibited potassium influx, suggesting specificity in terms of the macromolecular surfaces of the pump which were exposed to the external medium. We concluded that there may be organ and species differences among (Na+, K+)-ATPase preparations. Antibodies prepared in rabbits and sheep were fractionated by absorption to dog brain enzyme. Both the antibody fraction which bound to the brain enzyme and that which did not bind inhibited the dog kidney (Na+, K+)-ATPase, but only the former inhibited dog brain (Na+, K+)-ATPase. When the two fractions were recombined, inhibition was restored to the extent of the unfractionated antibody.


The gamma protein specified by bacteriophage gamma. Structure and inhibitory activity for the recBC enzyme of Escherichia coli. The protein encoded by the gam gene of bacteriophage lambda ("gamma protein") is a specific inhibitor of the recBC enzyme of Escherichia coli. The lambda protein has been purified approximately 2,000-fold, and its structure and inhibitory activity have been characterized. It appears to be composed of two identical subunits of 16,500 daltons, inhibits all of the catalytic activities of the recBC enzyme with apparently equal efficiency, but has no effect upon any other E. coli or lambda-DNase tested. Inhibition does not occur unless recBC enzyme is exposed to gamma protein prior to reaction of the enzyme with DNA. The inhibitory activity is independent of temperature, and no catalytic activity has been detected that might fulfill the inhibitory function. It appears instead that the inhibition involves a stoichiometric, rather than a catalytic interaction between gamma protein and the enzyme. Reaction kinetics for the recBC enzyme inhibited by gamma protein show no anomalous protein--only a depressed rate. Inhibition is not competitive and does not appear to affect the enzyme's affinity for DNA. The enzyme remains inhibited after it is separated from "excess" gamma protein by gel filtration or sedimentation in a glycerol gradient, and inhibited enzyme has a reduced electrophoretic mobility compared to that of uninhibited enzyme. Gamma Protein inhibits recBC enzyme which has been reconstituted from cell-free extracts by complementation in vitro, but at least one of the complementing factors present in extracts from recB- cells does not by itself form a complex with gamma protein. The mechanism of inhibition and the implications of these results from gamma replication and recombination are discussed.


Structural studies of sodium and potassium ion-activated adenosine triphosphatase. The relationship between molecular structure and the mechanism of active transport. Sodium and potassium ion-activated adenosine triphosphatase is the enzyme responsible for the active transport of sodium and potassium across the plasma membrane. Strophanthidin, from the external surface of the membrane, and an antibody, from the cytoplasmic surface, bind simultaneously to the large polypeptide subunit of the enzyme. These results demonstrate that this polypeptide chain must span the plasma membrane, having different surfaces exposed on each side. When (Na+ + K+)-ATPase is incubated in the presence of cupric phenanthroline, a reagent which catalyzes the oxidation of cysteine residues to form intermolecular and intramolecular disulfide bonds, a covalent dimer of the larger chains is formed. Several characteristics of this dimerization reaction are consistent with the proposal that at least a noncovalent dimer of large chains exists in the native enzyme. These conclusions are discussed in the context of a specific description for the molecular mechanism of active transport.


Effect of the purified (Mg2+ + Ca2+)-activated ATPase of sarcoplasmic reticulum upon the passive Ca2+ permeability and ultrastructure of phospholipid vesicles. The passive Ca2+ permeability of fragmented sarcoplasmic reticulum membranes is 10(4) to 10(61 times greater than that of liposomes prepared from natural or synthetic phospholipids. The contribution of membrane proteins to the Ca2+ permeability was studied by incorporating the purified [Ca2+ + Mg2+]-activated ATPase into bilayer membranes prepared from different phospholipids. The incorporation of the Ca2+ transport ATPase into the lipid phase increased its Ca2+ permeability to levels approaching that of sarcoplasmic reticulum membranes. The permeability change may arise from a reordering of the structure of the lipid phase in the environment of the protein or could represent a specific property of the protein itself. The calcium-binding protein of sarcoplasmic reticulum did not produce a similar effect. The increased rate of Ca2+ release from reconstituted ATPase vesicles is not a carrier-mediated process as indicated by the linear dependence of the Ca2+ efflux upon the gradient of Ca2+ concentration and by the absence of competition and countertransport between Ca2+ and other divalent metal ions. The increased Ca2+ permeability upon incorporation of the transport ATPase into the lipid phase is accompanied by similar increase in the permeability of the vesicles for sucrose, Na+, choline, and SO42- indicating that the transport ATPase does not act as a specific Ca2+ channel. Native sarcoplasmic reticulum membranes are asymmetric structures and the 75-A particles seen by freeze-etch electron microscopy are located primarily in the outer fracture face. In reconstituted ATPase vesicles the distribution of the particles between the two fracture faces is even, indicating that complete structural reconstitution was not achieved. The Ca2+ transport activity of reconstituted ATPase vesicles is also much less than that of fragmented sarcoplasmic reticulum. The density of the 40-A surface particles visible after negative staining of native or reconstituted vesicles is greater than that of the intramembranous particles and the relationship between these two structures remains to be established.


A coupling factor from sarcoplasmic reticulum required for the translocation of Ca2+ ions in a reconstituted Ca2+ATPase pump. 1. During purification of the Ca2+ATPase from sarcoplasmic reticulum of rabbit muscle, different fractions with similar Ca2+ATPase activity were found to vary greatly in their ability to catalyze 45Ca2+ translocation in reconstituted liposomal systems. 2. A heat-stable fraction isolated from the fraction most active in Ca2+ translocation enhanced several-fold the Ca2+ translocation rate of the least active fraction. It also increased the ratio of Ca2+ translocation to ATP hydrolysis over 5-fold. The properties of the coupling factor resemble those of the proteolipid previously described by MacLennan et al. (MACLENNAN, D.H., YIP, C. C., ILES, G. H., and SEAMAN, P. (1972) Cold Spring Harbor Symp. Quant. Biol. 37, 469-478). 3. When the heat-stable factor was added to either sarcoplasmic reticulum fragments or to liposomes after, rather than before, reconstitution, it acted as an ionophore abolishing Ca2+ translocation.


Use of diazido ethidium bromide as a specific probe for mitochondrial functions. The diazido derivative of ethidium bromide has been synthesized as a potential photoaffinity label and shown to be at least as effective as a mitochondrial mutagen as the parent compound, with a similar mode of action. Exposure of mitochondria of Saccharomyces cerevisiae to the compound, followed by ultraviolet-irradiation, which converts it to the highly reactive dinitrene, results in its specific binding to a single component which has been tentatively identified as the smallest polypeptide (subunit 9) of the membrane-bound ATPase. An analogus reaction is also obtained with the soluble, oligomycin-sensitive ATPase complex but not with the F1-ATPase itself. The reaction with the ATPase complex can also be monitored by fluorescence enhancement and by this attribute, as well as by other criteria, diazido-ethidium bromide, ethidium bromide itself, euflavine, N,N'-dicyclohexylcarbodiimide, 2,4-dinitrophenol, and 2-azido-4-nitrophenol all appear to compete for the same, lipophilic, binding site. A mitochondrial mutation (73/1) (see Flury, U., Feldman, F., and Mahler, H.R. (1974) J. Biol. Chem. 249, 6630-6637) produces a photoaffinity product with an altered electrophoretic mobility and molecular weight.


Kinetic studies on rat liver and beef heart mitochondrial ATPase. Evidence for nucleotide binding at separate regulatory and catalytic sites. Mitochondrial ATPases from rat liver and beef heart were used to study the effects of guanylylimidodiphosphate (GMP-P(NH)P) and adenylylimidodiphosphate (AMP-P(NH)P) on the kinetics of MgATP, MgITP, and MgGTP hydrolysis. AMP-P(NH)P was a noncompetitive inhibitor of hydrolysis of all substrates with the rat liver enzyme, whether activating anions were present or not. Also with the liver enzyme, AMP-P(NH)P caused only MgATP hydrolysis to appear to have positive cooperativity. With the beef heart enzyme, AMP-P(NH)P was a competitive inhibitor of ATPase activity and caused positive cooperativity; it gave noncompetitive patterns with GTP or ITP as substrates. In both enzyme systems, GMP-P(NH)P gave complex inhibition patterns with MgATP as the substrate, but was a competitive inhibitor of MgITP and MgGTP hydrolysis. These results are interpreted as indicating the existence of two types of nucleotide binding sites, with varying degrees of specificity and interaction on the ATPase molecules from both sources. It is postulated that MgATP and AMP-P(NH)P bind to regulatory site while MgATP, MgGTP, Mgitp, and GMP-P(NH)P bind to the catalytic site.


Effect of F-actin upon the binding of ADP to myosin and its fragments. The effect of F-actin upon the binding of ADP to rabbit skeletal muscle myosin, heavy meromyosin, and subfragment 1 was studied by equilibrium dialysis, ultracentrifuge transport, and light scattering techniques. Both myosin and H-meromyosin (HMM) bind a maximum of approximately 1.6 mol of ADP/mol of protein, while S-1 binds approximately 0.9 mol of ADP/mol of protein. The affinity for ADP of all three preparations was similar at a given ionic strength (approximately 10(6) M-1 at 0.05 M KCl) and decreased with increasing ionic strength. Under conditions similar to those used for the measurement of ADP binding, the binding sites of myosin, HMM, and subfragment 1 (S-1) are saturated with actin at molar ratios of 2, 2, and 1 mol of actin monomer/mol of protein, respectively, as determined by light scattering, ultracentrifuge transport, and in the case of myosin by ATPase measurements. F-actin was found to inhibit ADP binding, but even at an actin concentration at least twice that required for saturation of myosin, HMM, or S-1, significant ADP binding remained. This ADP binding was inhibited by 10(-4) M pyrophosphate. The observations are consistent with the formation of an actomyosin-ADP complex in which actin and ADP are bound to myosin at distinct but interacting sites.


Mechanism of action of "ruthenium red" compounds on Ca2+ ionophore from sarcoplasmic reticulum (Ca2+ + Mg2+)- adenosine triphosphatase and lipid bilayer. Sarcoplasmic reticulum (Ca2+ + Mg2+)-ATPase was previously shown to have Ca2+-dependent and -selective ionophoric activity when tested in oxidized cholesterol lipid bilayer membranes (Shamoo, A. E., and MacLennan, D. H. (1974) Proc. Natl. Acad. Sci. U. S. A. 71, 3522). ruthenium red, a known inhibitor of (Ca2+ + Mg2+)-ATPase, is found to inhibit the Ca2+-ionophoric activity associated with (Ca2+ + Mg2+)-ATPase. Furthermore, ruthenium red alone acts as an anion-selective ionophore in lipid bilayers with the the following selectivity sequence for anions: l- greater than Cl-, Br- greater than F- greater than NO3-. The PCl-/PNa+ ratio was approximately 4/l. The presence of ruthenium red in excess of Ca2+ ionophore in lipid bilayer experiments converts the cation selectivity of the bilayer due to Ca2+ ionophore into anion selectivity.


Porous hydrophilic polymer: good and bad news in the orthopedic application of cruciate ligament substitution. In the configuration used, the Hydron sponge did not enhance the ingrowth of bone into the Dacron for prosthesis anchorage. In fact, the presence of the Hydron seemed to retard such ingrowth, even though there was bony incorporation of portions of the Hydron polymer. Fixation was more rigid when Dacron was implanted bare. Hydron sponge does not appear to remain intact within a joint. It would not seem suitable for intra-articular protection of a prosthesis or local delivery of antibiotics here. We did not search further for the polymer in the regional lymph nodes. Hydron sponge is capable of eleciting an unusual phenomenon of woven bone formation. This is "good news" for its potential, but realization of such potential will certainly require additional study. Double and triple interval fluorochrome labelling would be especially helpful in further studying the localization and rate of this bone formation.


Design and fabrication of two types of artificial tendons for evaluation of surgical techniques. Double-loop tendons of fixed and adjustable lengths, and tendons with inner porous tapes both for anastomosis to tendon stumps have been made from readily available materials. By using stainless steel wire and polymer fibers, breakloads have been obtained that range from 20 to 100 lb and elongations have been reduced to less than 2%. Pore sizes up to 50 times 250 mu2 were achieved by using woven, knitted, and braided polyester tapes. For all tendons, Silastic was used to isolate inner fibers and to effect smooth gliding. Preliminary evaluation of these devices, after being implanted in the feet of chickens for 4 weeks, indicates that anastomotic strength of unions formed by use of loops is about 1000 g compared to more than 2000 for unions formed by tissue ingrowth.


[Comparative study of 4 types of polyester arterial prostheses]. The authors report a comparative study of 4 types of arterial prosthesis made of polyester. After implantation in the dog, the superiority of unfolded Rhodergon velvet prostheses was shown. They have excellent qualities for tissue reconstruction owing to presence of histological structures characteristic of large caliber arteries and absence of histological changes and hemorrhagic suffusion after the 12 th month following insertion.


Simple amidase test for identification of mycobacteria. A modified amidase test for differentiation of mycobacteria is described. A total of 224 atypical mycobacteria, 154 Mycobacterium tuberculosis, and 26 M. bovis strains were classified by this procedure. Of the 404 strains of various species studied, 400 exhibited an amidase spectrum identical to the established pattern. The simplicity of this method may promote its application in routine examinations.


Experiences and observations with the typing of Staphylococcus aureus phage 94. During a 28-month period, 5,078 cultures from a variety of anatomical sites were received for staphylococcal phage typing. Of these, 503 (10%) were not suitable for the procedure requested. Of the 4,575 viable cultures, 1,030 (23%) of the microorganisms were nontypable at both the routine test dilution (RTD) and at 100 times RTD. Of the 3,545 typable organisms, 3,061 (86%) were lysed at RTD, whereas 484 (14%) were typed only at 100 times RTD. Observations pertaining to the typing efficacy of staphylococcal phage 94 indicate that 651 (18%) of the typable microorganisms were lysed only by phage 94 at RTD or at 100 times RTD. Without the addition of this new phage to the international basic set, the number of nontypable strains (1,030 or 23%) would have been 1,681 (39%). Data regarding the geographic distribution of Staphylococcus aureus phage 94 point to the occurrence of the host strain in 13 (68%) of 19 states and 18 (62%) of the 29 hospitals submitting specimens to this laboratory. The assumed origin and speculated mode of dissemination of this microorganism are discussed.


Two serotypes of exfoliatin and their distribution in staphylococcal strains isolated from patients with scalded skin syndrome. Two serotypes of exfoliatin, A and B, previously reported by Kondo et al. (1974) were examined for their presence in 43 strains of Staphylococcus aureus, most of which were isolated from patients with Ritter's disease and impetigo. The tested strains consisted of 24 strains of phage group II and 19 strains not of phage group II. Twenty-four strains were found to produce an exfoliatin of either type A or type B, but 16 strains produced both types and 3 strains produced neither. No relationship was found between the serotype of an exfoliatin and the phage type of the exfoliative strain, although the single producers of exfoliatin A were all found to belong to phage group II and those of exfoliatin B to the other phage group.


Alteration of T cell function in healthy persons with a history of thymic x-irradiation. The possible late effects of x-irradiation to the infantile thymus were investigated by studying immune functions in 12 healthy persons with a history of thymic x-irradiation and healthy control subjects. No differences were found in serum immunoglobulin values, humoral antibody levels, lymphocyte counts, and lymphocyte reactivity to phytochemagglutinin, vaccinia virus, purified protein derivative (PPD), and allogeneic cells. The irradiation group exhibited cellular hyperresponsiveness to streptoskinase-streptodornase (SK-SD). In contrast, mean skin and in vitro lymphocyte responses to Candida albicans were depressed in the patients with thymic irradiation. A dissociation of these two Candida responses was found in only 1 of 14 healthy control subjects but in 7 of 12 irradiated individuals. While thymic irradiation did not result in impaired immunologic defenses leading to clinical disease, it caused alterations in T cell responses similar to those reported in patients with chronic mucocutaneous candidiasis.


Enzymes of the cholinergic system in islets of Langerhans. Quantitative radiometric assays were employed to measure activities of choline acetyltransferase and acetylcholinesterase in freeze-dried pieces of islets of Langerhans and exocrine tissue from rat pancreas. The activities of both enzymes were about an order of magnitude higher in islets than in exocrine tissue. This difference in activity was found in rats made diabetic with streptozotocin as well as in the controls. Although the enzyme activities in islets from diabetic rats averaged about 30-40% higher than those in islets from control rats, the differences were statistically only marginally significant. Since the islets of diabetic rats are probably much smaller than those of control rats, it is suggested that cholinergic elements associated with pancreatic islets are lost following induction of streptozotocin diabetes.


In vitro human reactivity to staphylococcal phage lysate. The cell-mediated reactivity of normal individuals to staphylococcal phage lysate (SPL) were tested in vitro in the lymphocyte stimulation (LS) and leukocyte migration inhibition (LMI) assays. There were 95% positive responses in LS (stimulation ratio larger than or equal to 3 with p less than 0.01) and 67% positive responses in LMI (migration index less than or equal to 0.80). Enriched subpopulations of T and B lymphocytes were prepared with rosette formation and density gradient centrifugation. SPL stimulated lymphoproliferative responses in both T and B cell subpopulations whereas phytohemagglutinin (PHA) stimulated only the T cell subpopulation. Cord blood leukocytes were tested in the LS assay and 41% gave positive responses to SPL, 81% to PHA, and 17% with SLO. SPL appears to be a useful reagent for the in vitro study of cell-mediated reactivity, and may provide somewhat different information from that obtained with other mitogens or antigens.


Genetically controlled autologous immune complex glomerulonephritis in rats. A study was conducted on the disease susceptibility of inbred strains of rats to experimental autologous immune complex glomerulonephritis (AIC). Three strains representing two different H-1-haploytpes developed severe glomerulonephritis within 3 months in response to a single injection with equal doses of an autologous primary tubular epithelial fraction and complete Freund's adjuvant (Lewis, AS (H-11)) and Lew.BDV (H-1d)).By contrast, two strains of the H-1 haplotype H-1n (BN and Lew.BN) showed no proteinuria and no immunohistologic findings during that time. Hybrids of Lew.BN and Lewis, subjected to the same immunizing procedure, showed a later onset of the disease as compared to the responder parent. The possible relationship between responder status and the major histocampatibility complex is discussed.


Lymphoid cell subpopulations. II. Characterization of cell populations responsible for syngery in the mixed lymphocyte interaction. Mixtures of isogeneic lymph node cells (LNC) and thymocytes (TC) exhibit far greater responsiveness in the murine MLI, as measured by proliferation and development of cytotoxic effector cells, than either cell type cultured alone. Pretreatment of either lnc or TC with mitomycin-C or ultraviolet irradiation completely abolished their synergistic interaction. Administration of cortisone acetate to cell donors 20 hr before sacrifice reduced the capacity of LNC and enhanced the capacity of TC to synergize. The LNC and TC populations participating in synergy, were found to be thymus dependent. LNC were shown to be responsible for the bulk of proliferative and effector activity observed in synergizing cultures, whereas TC appeared to amplify the activation of LNC. These findings provide the basis for a three cell model of MLI responsiveness.


Human chorionic gonadotropin: effects of crude and purified preparations on lymphocyte responses to phytohemagglutinin and allogenenic stimulation. The factors that prevent maternal immunologic rejection of the histoincompatible fetus are not understood. High levels of human chorionic gonadotropin (HCG) are present in the placenta, and several reports have noted suppresion of mitogen-induced lymphocyte transformation when cultures were supplemented with crude preparations of HCG. Purified HCG and multiple lots of crude HCG obtained from different suppliers were examined for their ability to suppress lymphocyte transformation produced by phytohemallutinin (PHA) or allogeneic stimulation. Crude preparations of HCG produced suppression of the lymphocyte stimulation induced by low doses of PHA, but the suppression could be overcome completely by increasing the PHA dose. The purified preparations of HCG produced no suppression of lymphocyte  responses, even at the lower PHA dose. Purified HCG did not give a dose-related suppression of allogeneic lymphocyte responses, and crude lots of HCG gave highly variable results. One lot of crude HCG produced spontaneous stimulation of lymphocytes. Isoelectric focusing of HCG preparations demonstrated multiple bands, and lymphocyte suppression may be secondary to these additional unidentified proteins. The failure of pruified HCG to suppress lymphocyte responses makes it unlikely that the absence of maternal rejection of the fetus is due to high placental levels of HCG.


Functional activities of rosette separated human peripheral blood leukocytes. Subpopulations of human peripheral blood leukocytes were isolated by rosette formation and tested for functional activity. E -rosette-forming cells (E-RFC) and EAC-RFC were separated from non-resetting cells by sedimentation on Ficoll-Hypaque gradients. The efficiency of the method and the purity of the resulting subpopulations were high. E-RFC responded to PHA Con A, allogeneic leukocytes, and PPD, with higher levels of proliferative reactivity than the unseparated lymphocytes while E-RFC depleted, EAC-RFC, and null cells showed only low levels of reactivity. Reactivity to PWM and tetanus toxoid was also restricted to the E-RFC subpopulation, but was lower than that of unseparated cells. A staphylococcal antigen preparation triggered lymphoproliferative reactivity in the E-RFC, E-RFC depleted, EAC-RFC, and the null cell subpopulations. 51Cr release lymphocyte cytotoxicity against a human lymphoblast target cell line was found in the E-RFC and null cell fractions but was not observed with the EAC-RFC subpopulation.


Propionibacterium levels in patients with and without acne vulgaris. Propionibacterium species were quantified on the foreheads and cheeks of persons with and without acne in three age groups: 11 to 15, 16 to 20, and 21 to 25. Propionibacteria were virtually absent in the pubertal non-acne group compared to a geometric mean density of 114,800 per sq cm in the acne group. A similar sharp difference existed between the acne subjects and normals in the age range of 16 to 20 years: 85,800 organisms per sq cm compared to 588 per sq cm. Patients with acne and normal subjects over age 21 showed no difference in Propionibacterium levels. In acne patients, while there was a trend for lower levels, no significant difference was seen as the severity of inflammation increased.


Persistent atrial standstill, report of three cases. Three cases, of which two are brothers, of persistent atrial standstill are reported. The diagnosis was made by the lack of P wave in routine 12 leads and right atrial cavity lead, no response of atrium to electrical stimulation and absence of "a" wave in right atrial pressure curve.


Synthesis of dehydroepiandrosterone and dehydroepiandrosterone sulphate by the human adrenal. The role of pregnenolone sulphate in adrenal steroid biosynthesis and the ability of the human adrenal gland to synthesize and secrete dehydroepiandrosterone (DNA) and dehydroepiandrosterone sulphate (DHA sulphate) was investigated. The presence of pregnenolone sulphate and DHA sulphate was demonstrated by measuring their concentrations in human adrenal tissue. Pregnenolone sulphate was metabolized in vitro mainly to free steroids, including DHA and cortisol, as well as directly to DHA sulphate in some cases. Similar results were obtained upon perfusion of the adrenal gland in situ with [14C]pregnenolone and [13H]prenenolone sulphate as the substrates and isolating the metabolites from the adrenal venous blood. Dehydroepiandrosterone sulphate was derived mainly from the sulphation of free DHA. The hydrolysis of DHA sulphate did not appear to make a significant contribution to the amounts of DHA synthesized under these conditions. The adrenal secretion of DHA and DHA sulphate by eight patients undergoing adrenal-ectomy was determined by measuring the concentrations of these compounds in samples of adrenal and peripheral venous blood taken simultaneously. In one patient secretion of DHA and DHA sulphate was equivalent whilst in the remainder there was much greater secretion of DHA.


Generation of cytotoxic T lymphocytes in vitro. IV. Functional activation of memory cells in the absence of DNA synthesis. Re-exposure of day 14 mixed leukocyte culture (MLC) cells to the original stimulating alloantigens (secondary response) has previously been shown to result in significant proliferation and in rapid reappearance of high levels of cytolytic T-lymphocyte (CTL) activity within the next 4 days. Moreover, evidence has been presented that CTL precursor cells in day 14 MLC populations, while they derived from cells were large at peak of the primary response (day 4) were themselves small lymphocytes which developed into large CTL after restimulation. In this study, inhibition of DNA synthesis by cytosine arabinoside (ARA-C) was used to investigate whether CTL formation could be dissociated from proliferation during the secondary response. It was found that within the first 24 h after restimulation (a) CTL activity increased 6-to-20-fold, (b) 60-70% of the small T lymphocytes became medium- to large-sized cells, and (c) both events were independent of DNA synthesis. By using two successive cell separations by velocity sedimentation at unit gravity, before and after stimulation of day 14 MLC cells for 24 h in the presence or absence of ARA-C, direct evidence was obtained that small CTL precursor cells developed into large CTL, irrespective of DNA synthesis. The presence of ARA-C for periods longer than 24 h inhibited any further increase in CTL activity, in contrast to a parallel increase in lytic activity and cell number from day 1 to day 4 in control restimulated cultures. Taken together with the finding that 90% of the medium- and large-sized lymphoid cells in control restimulated cultures underwent DNA synthesis within 24 h, these results thus suggest that during a secondary MLC response there is initially a differentiation step leading to the formation of CTL which, although it can be clearly dissociated from DNA synthesis, is under normal conditions followed by proliferation of these effector cells.


Measles infection of human mononuclear cells. I. Acute infection of peripheral blood lymphocytes and monocytes. A study of the susceptibility of human peripheral blood mononuclear cells to measles virus infection and replication is reported. Resting lymphocytes obtained from adults showed very low levels of infection and virus replication while lymphocytes activated by plant mitogens or allogenic lymphocytes supported mononuclear cells obtained from the umbilical cord of healthy neonates were more susceptible to measles virus infection than those of adults; however, activated cord lymphocytes supported viral replication in the range observed with adult activated lymphocytes. Monocytes obtained from adults were relatively resistant to measles virus infection and replication while neonatal cord blood monocytes supported viral replication to the degree observed with activated lymphocytes. It is hypothesized that infection of acitivated lymphocytes may explain the depression of cell-mediated immunity seen during acute measles virus infection. The significance of the finding that neonatal monocytes are more susceptible to viral infection and replication than adult monocytes is discussed.


Heterogeneity in the development of cytotoxic T lymphocytes in vitro revealed by sensitivity to hydrocortisone. In the present study we used hydrocortisone (HC) treatment in vivo as a probe to analyze two different in vitro systems for the regeneration of cytotoxic T lymphocyte (CTL), namely the antifibroblast reaction (AFR) and the mixed lymphocyte culture (MLC) system. We found that cells remaining in the thymus after HC treatment had increased reactivity in these two systems. However, the same treatment in the spleen severely depressed the MLC reactivity in both the proliferative and the cytolytic phases, while markedly increasing the AFR reactivity. These findings demonstrate heterogeneity of CTL precursors and/or their pathways of differentiation into effector cells. In addition, MLC-reactive cells in the thymus appear to be distinct from such cells in the spleen, as judged from their differential sensitivity to HC.


Relation of Na-K-ATPase to acute changes in renal tubular sodium and potassium transport. Renal Na-K-ATPase activity changes adaptively in response to chronic alterations in sodium reabsorption or potassium secretion, but the role of this enzyme in rapid adjustments of renal tubular Na and K transport is not known. To evaluate this question, microsomal Na-K-ATPase specific activity and kinetics were determined in the rat and guinea pig kidney after massive but short-term (3 h) sodium or potassium loading. In other experiments renal sodium handling was evaluated in hydropenic and saline-loaded rats in which enzyme synthesis was prevented by the concurrent administration of actinomycin D or cycloheximide. Saline loading increased net sodium reabsorption in both rats and guinea pigs, but microsomal Na-K-ATPase from the outer medulla (where the reabsorptive increment is greatest) did not change significantly in either species. In vitro [3H]ouabain bidint to guinea pig microsomes and apparent Km for sodium of rat microsomal Na-K-ATPase, both from outer medulla, were also unaltered. Actinomycin D and cycloheximide failed to increase sodium excretion and microsomal Na-K-ATPase remained unchanged. KCL loading resulted in a 10-fold increase in K excretion but again Na-K-ATPase specific activity (in cortex, outer medulla, and papilla), and its apparent Km for potassium were not affected. Taken together these results suggest that rapid adjustments in remal tubular Na or K transport are mediated by mechanisms that do not involve the Na-K-ATPase enzyme system.


Patterns of neuropsychological test performance in Huntington's disease. Neuropsychological test performance of Huntington's disease (HD) and a mixed brain-damaged group were compared. Wechsler patterns in the present samples were compared to those of HD patients in the study of Boll et. al (1), and some correspondence specific to HD is suggested. Minnesota Multiphasic Personality Inventories (MMPIs) were found not to differ between the HD and mixed brain-damaged groups.


Histochemical and histopathological changes in skeletal muscle in late-onset hereditary distal myopathy (Welander). Histochemical and histopathological staining methods were applied to muscle biopsy material from 13 patients with distal myopathy of late onset. Six cases showed slight to moderate histopathological changes and the normal distinction between Type I and Type II muscle fibres, based on their staining characteristics for myofibrillar ATPase, was well preserved. A selective Type I atrophy and an irregular distribution of oxidative enzyme and fat staining in Type I fibres were evident. In the other 7 cases, with moderate to advanced histopathological changes, there was a marked blurring of the normal difference observed in ATPase activity between Type I and TYpe II fibres. Thus, both types of fibre exhibited a high intensity of staining for myofibrillar ATPase at pH 9.4 without inhibition by acid preincubation (pH 4.3). These changes in phosphatase activity were found not only in atrophic fibres but also in normal-sized fibres without other signs of degeneration. Nuclear proliferation in chains and "ring fibres" were found. The early histopathological and histochemical changes in distal myopathy are strikingly similar to those of myotonic dystrophy.


Growth of diseased human muscle in combined cultures with normal mouse embryonic spinal cord. Normal and diseased human muscle cells have been grown in combined cultures with 12-14 day embryonic mouse spinal cord explants. Nerve endings on myotubes were found by light and scanning electron microscopy, and motor end-plates were identified by a histochemical reaction for acetylcholinesterase. Contracting myotubes, never observed in cultures of human muscle alone, were found in a culture from normal muscle. Histochemical studies demonstrated the presence of strongly and weakly reacting myotubes for both ATPase pH 9.4 and NADH-TR, but could not be related to the development of fibre types. No differences in morphology or histochemical reactions were found between normal and diseased muscle cells in these combined cultures.


Patch testing of coolant fractions.

Patch testing of coolant fractions. Dermatitis among machine operators, and others who are consistently exposed to cutting and grinding liquids, has presented the industrial physician with the problems of how to protect the machinist from the cause of his dermatitis and how to detect irritants that may be responsible for the skin problem. In order to examine the various components of machine coolant that might be responsible for causing a dermatitis reaction, 13 suspensions were prepared. They consisted of various combinations of New Codol¿ (coolant base), biocides, masking agent, tramp oil, used coolant and metal fragments. Patch test techniques were used to examine these suspensions for their potential to cause a localized skin irritation on plant employees. Two hundred and six New Process Gear employees were tested over a ten-month period. The participants were management, general salaried employees and hourly labor employees. There was not a single positive reaction in any one of the 206 individuals who completed their skin testing. Of the 206 tested, 29 persons were tested and retested a total of 62 times over a period of one to 25 weeks after the initial testing. In neither the testing nor the retesting was a positive patch test reaction -- irritant or allergic -- obtained.


Ultrastructure of the oral epithelium in leukoplakia associated with tylosis and esophageal carcinoma. Ultrastructural studies have been carried out on epithelium taken from the oral lesions of tylosis-related leukoplakia and preleukoplakia in a group of patients known to be at high risk for esophageal carcinoma. Several structural abnormalities are described; in particular, the presence of intranuclear electron dense particles 90 nm in diameter. Similar particles have been found in the nuclei of epithelial cells from an esophageal carcinoma developing in one of the patients under study.


Prognostic implications of left ventricular function in endomyocardial disease in infants and children. Hemodynamic factors measured at cardiac catheterization in 40 infants and children with chronic endomyocardial disease were analyzed in regard to subsequent clinical condition. The patients were followed for periods ranging from 2 months to 11 years (average 4.3 years) after initial cardiac catheterization. There were no statistically significant differences in left ventricular end-diastolic volume among survivors with no symptoms, survivors with persistent congestive heart failure, and nonsurvivors. Ejection fractions were depressed in about four fifths of patients with chronic endomyocardial disease and were significantly lower in the group of patients who died subsequently. LVEDP in patients who did not survive was significantly higher than in asymptomatic survivors, but there was too much overlapping of individual values to be of prognostic value. Left ventricular pressure-volume loops appeared to offer an improved, although more tedious, method of assessing LV function. In addition to offering information on LVEDV, LVEDP, and EjF, LV stroke work may easily be estimated from pressure-volume loops. There was generally good correlation between hemodynamic status assessed from pressure-volume loops and subsequent clinical outcome.


The response to stretch of human intercostal muscle spindles studied in vitro. 1. The discharge properties of human muscle spindles have been studied in vitro in a preparation based on the biopsied external intercostal muscle. 2. The static and dynamic responsiveness of thirty-six endings in twenty visualized and histologically identified spindles have been investigated using amplitudes and velocities of stretch likely to encompass those occurring in vivo. 3. The dynamic index, measured at a stretch velocity of 3 mm/sec, ranged from 3 to 40 impulses/sec and was distributed bimodally, consitent with the presence of primary and secondary endings. 4. The relationship between the dynamic index and the velocity of stretch was approximately linear both for primary and secondary endings up to the maximum velocity tested (10 mm/sec). 5. The frequency/extension relationship was approximately linear for both primary and secondary endings. The mean values of the slope for primary and secondary endings were 16-1 +/- 8-3 S.D. of the observation and 12-1 +/- 6-5 impulses/sec per five per cent extension. 6. The slopes of the frequency/extension relationship for endings lying in the same spindle were positively correlated, significant at the 10% level. 7. It was estimated from the results in vitro that the position sensitivity of human intercostal spindles in vivo ranges from 2 to 21 impulses/sec per millimetre.


General practitioners and social help for the handicapped. A study was undertaken by the Greater London Association for the Disabled in consultation with the Royal College of General Practitioners, to explore the depth of knowledge of the Chronically Sick and Disabled Persons Act and statutory and voluntary social provision, of 22 general practitioners in 16 practices served by one area social services office in a London borough.The doctors were mainly middle-aged, of British or Irish birth and training and had no language barrier. The majority lived in or near their practices. Half the practices were groups or partnerships, half were singlehanded. Only in three groups was there any attached district nursing staff and in only one was there an attached health visitor. More than half the general practitioners had reception staff only during surgery hours. Four practices had no reception staff during National Health Service surgery hours, two of which had no reception staff at all. In no practice was there any privately employed nursing staff. All the practices had private patients.Nine of the 22 doctors in the study had never heard of the Chronically Sick and Disabled Persons Act, and a further five had not mentioned the Act to their patients. Fifty per cent had no knowledge of the extent of functional disability in their practice. More than half the doctors knew no more of the social services than that home helps and meals-on-wheels were available, while six doctors knew of no provision at all. Knowledge and use of the voluntary services was almost non-existent. No meetings with team members were held, other than in the group practices with attached staff, and the team members were largely unknown to most of the doctors.Attempts were made through various channels to extend the knowledge of the general practitioners of the services provided by both statutory and voluntary agencies, and to introduce them and their receptionists to their team, but little use was made of the opportunity.


Synthesis of a bifunctional coordination complex of osmium with curariform activity. Based on the known curariform action of tris(bipyridyl)iron(II) sulfate and other complex ions, two series of bifunctional ligands designed to hold transition metal ions at approximately the same distance apart as the interquaternary ammonium distance in the potent neuromuscular block agents were synthesized. In the first series two 1,10-phenanthrolines (R1) were joined at the 2 position to form four compounds: R1CO-c-N(CH2CH2)2N-COR1, R1CONH-1,2-C6H10-NHCOR1, R1CONH-1,2-C6H4-NHCOR1, and R1CON(CH3)(CH2)2N(CH3)COR1. In the second series two terpyridines (R1) were joined by different chains to give R2(CH2)2CH=CH(CH2)2R2, R2CH2C(CH3)(OH)(CH2)2C(CH23)(OH)CH2R2, R2CH2C(CH3)(OH)C(CH3)(OH)CH2R2, and R2CH2(OH)-1,4-C6H10-(OH)CH2R2. Three other ligands in which the terpyridines were joined by 5-, 60, and 7-methylene groups were also made. The ligands were converted to nickel(II) complexes and the coordination of each nickel ion was completed by adding terpyridine. These were assayed by the intravenous mouse LD50 method. The most potent ligand, the di-hydroxy compound R2CH2(OH)-1,4-C6H10-(OH)CH2R2 was then converted to the bis(pyridinebipyridine)diosmium-(II) coordinated complex and assayed by the iv mouse LD50 method and by the ED50 isolated guinea-pig diaphragm method. By the iv mouse LD50 method, it was about twice as potent as d-tubocurarine and by the isolated diaphragm method, it was 16 times more potent. The compound has been called dihydroxyosmarine tetrachloride or DHO for short. The term "transarine" ions is proposed for transition metal coordination complexes having curariform action. The position of the transarine ions is discussed in the classification of cholinergic ligands, in structure-action relationships, and in relation to some current ideas on receptor mechanisms.


Attitudes of patients and their relatives to Huntington's disease. Reaction to medical, social, and genetic implications of Huntington's disease was evaluated by means of a questionnaire mailed to members of a lay organization concerned with this disease in the United States. One thousand and sixty-five of the approximately 2600 members chose to respond. Patients and those at high risk found physical disabilities most disturbing while mental deterioration and personality change were the most disturbing to spouses of patients. The best source of information regarding the disease for 46% was the lay organization itself. Medical specialists or genetic counsellors were cited as the best source of information by 18%. The transmission risk in Huntington's disease was correctly stated to be one-half by 92% of all respondents including 94% of those who indicated the lay organization as best source and 91% of those who indicated medical specialists and genetic ounsellors as best source. If at risk for Huntington's disease, 86% of respondents would modify their family size; desire for limitation was greatest among those affected but lowest among young adults at high risk. If a screening test were available, 23% at high risk might refuse it.


Oscillations of an electrogenic pump in the plasma membrane of Neurospora. The presence of the poky mutation in Neurospora crassa produces mitochondria which are defective in cytochromes b and aa3 but which compensate by means of an alternate, cyanide-insensitive oxidase. As previously reported (Slayman, Rees, Orchard & Slayman, J. Biol. Chem., 250:396, 1975) cyanide blockade of the poky strain carrying the partial suppressor f results in a metabolic downshift of only 56%, compared with a downshift of 98% in wild-type Neurospora; the downshift is accompanied by exponential decay of ATP in the wild type, but by an undershoot and monotonic recovery of ATP in poky f. Whereas the membrane potential declines with ATP in wild-type Neurospora, it oscillates near the resting level (ca. -- 185 mV) in poky f. Oscillations begin with a depolarizing swing of 30--100 mV, followed by slight hyperpolarization, then by 2--4 damped cycles having a frequency near 1/min. Similar oscillations arise with antimycin, salicyl hydroxamic acid, and several uncoupling agents, and depend on partial maintenance of respiration through either the defective cytochrome chain or the alternate oxidase. Small oscillations (maximally +/- 30% of the control value) in membrane conductance also occur, roughly in phase with the oscillations of membrane potential. The amplitude of these, in comparison with the nonlinearity of the normal current-voltage relationship for the membrane, strongly suggests that they arise as a secondary consequence of the voltage changes. Therefore, since it has previously been argued (Slayman, Long & Lu, J. Membrane Biol. 14:305, 1973) that most of the resting membrane potential in the organism arises from active extrusion of H+ ions, the simolest interpretation of the cyanide-induced voltage oscillations is that current through the H+ pump is modulated cyclically. The ultimate mechanism for this modulation is unresolved, but could plausible involve a metabolic feedback system, oscillations of intracellular pH, or both. In many respects the observed voltage oscillations resemble the well-known oscillations of mitochondrial H+ flux which are produced by sudden metabolic shifts.


Iatrogenic muscle fibrosis. Arm levitation as an initial sign. Three patients had iatrogenic muscle fibrosis, without weakness or sensory loss. Deltoid muscle fibrosis produced the unique clinical sign of gradual, involuntary, and irreducible arm levitation. The third patient had both levitated arms and levitated legs, a result of injections in the rectus femoris muscles. Repeated intramuscular injections apparently resulted in muscle fibrosis and intramuscular nerve-twig damage. Pentazocine (Talwin) is a particularly offending agent. One patient showed an unusual tissue reaction to foreign material, which may have played a role in her muscle fibrosis. Nevertheless, dramatic recovery of muscle function followed surgical sectioning of the fibrous deltoid bands.


Failure of heparin to affect two types of experimental glomerulonephritis in rabbits. Anticoagulation has been reported to ameliorate antiglomerular basement membrane glomerulonephritis (anti-GBM-GN) while its effect on chronic immune complex glomerulonephritis (IC-GN) as studied in the NZB mouse is unclear. Chronic serum sickness IC-GN was induced in rabbits by injecting bovine serum albumin (BSA) daily. Anti-GBM-GN was induced by i.v. injection of a known amount of heterologous anti-GBM antibody. Heparin was administered beginning at two to six weeks after the first BSA injections or before the administration of anti-GBM antibody, on various schedules from 5000 U every 12 hr to 8000 U every 8 hr. With this dosage the partial thromboplastin time remained greater than 1-1/2 to 2-1/2 times the control at the time of the subsequent heparin injection. Heparinized and nonheparinized groups were matched according to duration of disease, maximum anti-BSA concentrations or anti-GBM antibody dosage--and no significant differences were found in proteinuria; severity of the glomerular histologic lesions; or immunofluorescence patterns of immunoglobulin G (IgG), third component of complement (C3), BSA or fibrinogen-related antigen(s) (FRA). Crescent formation was not prevented. This study shows that heparin in the maximum permissible dosage is ineffective in preventing glomerular FRA deposition or altering the progression of experimental IC-GN or anti-GBM-GN in rabbits.


[The chronically ill and disabled child: a psychologic task of paediatrics. Part I: General problems (author's transl)]. This first out of three articles reviews general psychological aspects and problems of the child with chronic illness and handicap and of his parents and healthy siblings.


[The chronically ill and disabled child: a psychologic task of paediatrics. Part II: Special problems of various diseases (author's transl)]. This second out of three articles deals with some specific psychological problems of chronic illness and handicap in childhood, i.e. epilepsy, physical handicap, diabetes mellitus, congenital heartdisease, hemophilia, dwarfism, hemodialysis and renal transplantation, and malignant disease.


[The chronically ill and disabled child: a psychologic task of paediatrics. Part III: Psychologic treatment (author's transl)]. This third and last paper on psychological aspects of chronic illness is childhood discusses tasks of the physician and the parents as well as aspects of social medicine.


[Syncopal attacks and sudden death due to ventricular fibrillation -- a variant of the Jervell or Lange-Nielsen syndrome (author's transl)]. Syncopal attacks due to ventricular fibrillation or -- flutter with elongation of QT in the ECG and deafness are signs of a congenital condition described by Jervell and Lange-Nielsen 18 years ago. Since then we have learned that the range of variation of symptoms is very wide. There are differences genetically and in age-groups. A boy aged 15 years with supposedly cerebral fits is reported on. Since the right diagnosis was missed he had been treated with anticonvulsants for years. But these were syncopal attacks, Effort produced a tendency to bigeminus. -- A knee-bending exercise repeated 13 times lead to his death from ventricular fibrillation. Post-mortem examination showed no pathologic findings except mild left hypertrophy of the ventricle. His sister also had similar attacks without spasms after excitement or mild physical effort. She too had no extension of the QT interval. Their mother and grandmother also had syncopal attacks and died at the ages of 31 and 24 years. A girl-cousin died suddenly at the age of 15. In addition we have seen a neonate with a mild cardiac murmur on whom an ECG was done on the 16th day. The QT interval was significantly prolonged. There were numerous extrasystoles. He died suddenly 2 days later. Post mortem showed no cardiac pathology. One should now be able to explain some cases of unexpected natural death. But ignorance of the pathogenesis does not yet permit definition of the clinical pictures.


Homologous dura mater cardiac valves. Structural aspects of eight implanted valves. We studied eight homologous dura mater valves removed 1 to 35 months after implantation. Five dura mater valves were placed in the mitral position and three in the aortic position. Two of the valves were obtained after deaths not related to the valve itself and six after reoperations due to immediate or late incompetence. In 2 cases the dura mater valves were entirely normal, both macroscopically and microscopically. In 2 other cases the leaflets were caught by stitches related to the struts, a technical error that caused laceration of the leaflets. Not withstanding this fact, the dura mater was normal on histologic examination. In 3 cases, and probably in 1 other, one or two leaflets were detached partially from the suture line, probably because of a technical error in the manufacture of the valve. These studies demonstrated satisfactory evolution of the homologous dura mater valves until 35 months after implantation. No signs of rejection or degeneration of the dura mater were observed in the present series.


Prosthetic replacement of the esophagus. A tubular prosthesis was initially made of silicone rubber, spot-bonded on the inside and outside to a layer of knitted Dacron. This was covered on the outside by a loose layer of knitted Dacron, attached to the rubber only at the two ends, to allow incorporation by fibrous tissue and fixation of the prosthesis. Anastomoses were made with one layer of continuous sutures. There was initial incorporation of the graft. After 29 to 44 days the tube became loose and migrated, leaving a stricture. The prosthesis was redesigned with two loose layers of wider mesh. Incorporation occurred early with abundant fibrous reaction. Separation of the tube started at 40 days and was complete within 72 to 152 days. When the sutures became disengaged, bacteria entered the space between rubber and incorporated fabric, resulting in collagen lysis and separation of the tube. There were no anastomotic leaks in the entire series.


[Characteristics of filamentous phototrophic bacteria from freshwater lakes]. Filamentous phototrophic bacteria were isolated in pure cultures from fresh-water stratified lakes, and identified as chlorobacteria. The photosynthesizing apparatus of these bacteria is chlorobium-vesicules, the main pigment is bacteriochlorophyll c. The bacteria do not require reduced compounds of sulphur; they are capable of anaerobic photosynthesis on organic compounds or grow under aerobic conditions in the light or in the darkness. They are mesophilic (optimum temperature is 25 degrees C); the optimum of pH is 7.0. A possibility to class these microorganisms as a new species of the genus Chloroflexus is discussed.


Indirect induction of a Staphylococcus aureus prophage by P11de a plasmid phage hybrid. The ability to mediate indirect induction of staphylococcal prophages was found to be a property of the cryptic high frequency transducing phage P11de but not of three other phages tested. P11de is the product of a recombination between a P11 phage and a gamma plasmid. Irradiated P11de preparations could not induce prophage development in strains which contained either a P11 prophage or a gamma plasmid. The establishment of P11de in a strain was not, however, inhibited by the presence of a P11 prophage. It is inferred that the inhibition of indirect induction exerted by the resident P11 prophage occurs at a stage other than the establishment of the P11de replicon.


Genetic and metabolic control of the purine catabolic enzymes of Neurospora crasse. Neurospora crassa can utilize various purine bases such as xanthine or uric acid and their catabolic products as a nitrogen source. Four classes of mutants which affect the purine degradative pathway were isolated and studied. Mutants of the aln-1 class specifically lack allantoinase, while alc-1 mutants lack allantoicase. Mutants designated as xdh-1 cannot utilize hypoxanthine as a nitrogen source and are presumed to be deficient in xanthine dehydrogenase activity. A regulatory mutant, amr, was found to have only very low, uninduced levels of uricase, allantoinase, and allantoicase. None of these genes are closely linked to each other. The three initial enzymes involved in the catabolism of uric acid are controlled in a complex manner by both induction and repression. Several lines of evidence indicate that the true inducer of uricase and allantoicase is uric acid. The use of the newly isolated mutant strains made it possible to demonstrate that neither allantoin nor allantoic acid could act as inducers. Furthermore, hypoxanthine itself was shown to be ineffective as an inducer although it can be metabolized to form an inducer. A non-metabolizable analogue of uric acid, 8-azaxanthine, is a gratuitous inducer of these enzymes. Uricase and allantoicase were found to be synthesized coordinately, but they were not coordinately regulated with allantoinase. Both uricase and allantoicase are stable enzymes and do not undergo turnover; nor are they subject to feedback inhibition by ammonia. Allantoinase, however, is quite labile both in vivo and in vitro. This enzyme was found to turnover in vivo in the presence of cycloheximide with a half-life of approximately 20 minutes. The amr (for ammonia regulation) mutant cannot utilize a wide range of compounds, including purines, nitrate, and many amino acids as a nitrogen source and also displays a multiple enzyme loss. The amr gene appears to play a major role in the control of nitrogen metabolism. It is postulated that the amr locus encodes a regulatory protein which is required to activate transcription of the structural genes for a group of related enzymes involved in nitrogen metabolism.


[ECG evaluation by the computer using the USPHS program (author's transl)]. The efficiency of ECG evaluation by the computer has definitely increased owing to the current improvement of the program. Applying the USPHS program in our material, a wrong diagnosis was made in 14% of the pathological tracings and in 25% of the normal tracings under investigation. These results are certainly encouraging, considering that dissimilar diagnoses are established by different cardiologists in about 20% of the cases. However, we deem the number of wrong diagnoses both in pathological and in normal cases still too high and, therefore, further improvement should be made before applying ECG diagnosis by the computer in practice.


[Pharmacotherapy of hypertension and its significance for the function of the salivary glands (author's transl)]. The secretion mechanism of the large salivary glands at the level of the acinus cells is presented to explain the side effects "dryness of the mouth and initial parotid pain on chewing". The importance of adrenergic alpha- and beta-receptors and a cholinergic receptor in the acinus cell membrane is discussed in connection with the action of antihypertensives on the autonomic nervous system. Experiments on rat salivary glands after treatment with clonidine, guanoxan and propranolol serve as a basis for this.


[Colon diverticulosis and complicating diverticulitis (author's transl)]. Dietary habits in the civilised countries of the West with foods low in bulk and cellulose are blamed as causes of the increase in colon diverticulosis and the complicating diverticulitis in recent times. Of all sections of the colon, the sigmoid is most frequently involved, older patients above the 5th decade being principally affected. While uncomplicated diverticulosis of the colon usually remains symptomless, complicated diverticulitis often shows very uncharacteristic symptoms. For prophylaxis and the treatment of slight complaints, conservative treatment with administration of bulky foods (bran etc.) is indicated. Threatening hemorrhages and perforative peritonitis demand emergency operations. In closed abscess formation, internal or external fistulae and incipient scarring stenosis with imminent ileus, resection of the affected part of the intestine is indicated.


[Repair of prenatally ruptured omphalocele and the paraumbilical abdominal wall defect with the infant's own fetal membranes (author's transl)]. Since 1971, 35 children with prenatally ruptured omphalocele and paraumbilical abdominal wall defects were operated on in the surgical clinic of the Cologne Children's Hospital. 45.7% of them survived compared with 29.2% in earlier years. This improved chance of survival is attributed the use of the child's own fetal membranes.


Striking prevalence of ankylosing spondylitis in "healthy" w27 positive males and females. Ankylosing spondylitis is diagnosed once or twice in each 1000 males and one tenth as frequently in females, but the true prevalence is unknown. Indentification of genetic marker, HL-A W27, for susceptible persons has provided a tool facilitating epidemiologic studies and allowing identification of "control" populations without the marker. Evaluation by postal questionnaires, and pelvic radiography of 78 HL-A 27W-positive blood donors selected from a group of apparently healthy subjects revealed 14 who satisfied the criteria for definite ankylosing spondylitis. The prevalence was similar in both sexes. One hundred and twenty-six W27-negative controls matched for race, sex, and age failed to yield a single case. For a person of either sex with HL-A W27, there appears to be about a 20 per cent chance that ankylosing spondylitis will develop, suggesting a prevalence of 10 to 15 per thousand. Hitherto accepted figures may underestimate the frequency by a factor of 10 to 20.


An improved method for examination of cerebrospinal fluid cells. A new collection technique allows cytologic examination up to 2 weeks after lumbar puncture without loss of cells or morphologic detail. Cerebrospinal fluid (CSF) was allowed to flow directly from a lumbar puncture needle into a solution of Carbowax in ethanol and was then processed with a cytocentrifuge. Two groups of patients were studied -- those undergoing spinal anesthesia and those having myelography for low back pain. Lymphocytes, polymorphonuclear neutrophils, and ependymal cells were regularly found in the CSF; a definite difference was found in the quantity and cell types in the two patient populations. The proposed method may be sensitive enough to aid in the identification of other neurologic diseases with low cell counts.


Extraperitoneal para-aortic node dissection through an upper abdominal incision. A surgical technique for para-aortic lymph node evaluation for metastatic tumor by the extraperitoneal approach is described. The procedure is devised to avoid gastrointestinal complications frequently found after transperitoneal evaluation of the para-aortic area followed by extended field irradiation. The surgical procedure was performed in 20 patients. Nine patients treated with 6000 rads extended field irradiation remained without gastrointestinal complications after 3 to 16 months of follow-up.


The applications of steroid hormone radioimmunoassays to clinical obstetrics. This is a selective survey of recent publications dealing with theoretical and established applications of steroid hormone radioimmunoassay procedures applied to clinical obstetric investigation. This subject is reviewed in three sections: first, basic principles of steroid hormone radioimmunoassay methodology; second, steroidogenesis in normal pregnancy and its relation to patterns of steroid hormones measured in maternal and fetal circulations; third, applied steroid radioimmunoassay technology as it is being used in established and potential clinical applications. It is concluded that steroid hormone radioimmunoassay procedures have been and will continue to be a highly productive technology applied to clinical obstetric investigation.


[Dyskinetic lesions of the left ventricle wall. Electrocardiographic data]. The study of the electrocardiogram in 87 cases of dyskinetic lesions of the left ventricule, including ventricular aneurysms and plaques of post-infarction dyskinesia, all of which underwent surgery, showed a correlation between the size of the dyskinetic lesion and the number of ST elevations in the electrocardiogram which is closer than the correlation between this size and the height of ST elevation.


[Transient malabsorption of Vitamin B12 bound to intrinsic factor in pernicious anemia (author's transl)]. In six patients with pernicious anemia, absorption of vitamin B12 bound to intrinsic factor was low as measured by the Schilling's test. Collection of urine was complete and the activity of intrinsic factor used was checked in other pernicious anemia patients. Nutritional deficiency, drug induced malabsorption, ileal or pancreatic diseases were excluded by clinical, radiological and biological investigations. This vitamin B12 malabsorption was selective and reversible within a few months after treatment with parenteral vitamin B12. Mechanisms of this ileal dysfunction are discussed.


[Isolation of erythroblastic islands. Study by optical and scanning electron microscopy (author's transl)]. 1. A simple technique is described for isolation and estimation of number of erythroblastic islands (EBI) in rat bone marrow. The number of EBI decreases during hypertransfusion polycythemia, and increases during experimentally-induced hemolytic anemia. It has also been possible to maintain EBI in vitro, under culture conditions, for up to 36 hours. 2. Each EBI contains a central cell having the character of a macrophage-histiocyte which adheres to glass and spreads, while still retaining its connections with the erythroblasts. 3. In an EBI the erythroblasts are arranged in the form of concentric rings, with the outer cells being more mature than the inner. The outermost layer is frequently made up of reticulocytes even in the isolated island. The erythroblasts of a single ring appear to be at the same stage of maturation which suggests that they are derived from the same stem cell. 4. Using a coordinate reference system, studies of nuclear expulsion were made, by both optical microscopy and scanning electron microscopy.


Origin of stress macroreticulocytes from macronormoblasts. Stress erythropoiesis was induced in rats and guinea-pigs by graded amounts of bleeding and phenyl-hydrazine administration. Hemoglobin and DNA content was measured by microspectrophometry of single erythroblasts in the process of nuclear extrusion. DNA content was always 2 C. Hemoglobin content was in all cases superior to the normal value, occasionally reaching a value of almost twice normal.


[The in vitro effect of oxidative and analgesic agents on the erythrocyte membrane protein electropheretic pattern (author's transl)]. The in vitro effect of oxidative agents, b-naphthol, p-aminophenol, phenylhydrazine, hydroxylamine, methylene blue, hydroquinone and 1,4-naphthoquinone and of analgesics, acetylsalicylic acid, sodium salicylate and dipyrone, on the electrophoretic pattern of the red cell membrane proteins, is described. The electrophoresis was effected in acid urea-starch gel vertical system. All the agents studied altered the red cell membrane protein electrophoretic pattern, causing a total loss of certain protein bands or a loss of some bands and the diminution of others simultaneously.


[Treatment of hemophilia by self-infusion: clinical and psychological approach (author's transl)]. Twenty-seven hemophiliacs between the ages of 12 and 17 volunteered in a self-infusion training program. An attempt was made to define a teaching method. It consisted of teaching them about the disease so as they would be able to recognise the site of their bleeding and the limitations of self-treatment. They also acquired dexterity in infusing themselves with various antihemophilic factor concentrates. Systematic clinical and psychological studies were undertaken and showed that this method of treatment gave a considerable improvement of the overall disability. At the same time the number of transfusions almost doubled. Self-treatment has been found successful among children over 13 years of age who are strongly motivated, intelligent, sensitive, and able to adapt themselves to concrete situations. This training has failed in 30% of the children in whom psychological disturbances have been found. In spite of this failure, a psychological improvement was noted.


[In vitro incorporation of tritiated thymidine in acute leukemia and response to chemotheraphy (author's transl)]. The incorporation of tritiated thymidine (3H-TdR) has been assessed in 52 cases of adult acute leukemia, during the first phase of the disease, before and after 48 hours of chemotherapy. Two methods were utilized simultaneously: the measurement of the incorporation of 3H-TdR by liquid scintillation counting and the labeling index by autoradiography. These two data allowed to appreciate the mean incorporation per labeled blast cell. A significant correlation has been found between liquid scintillation counting and labeling index on the one hand, between mean incorporation per labeled blast cell and mean grain count on the other hand. The prognostic value of these different data was significant, at least if one consider the chance to get a complemente remission with the various drugs which were utilized. The responders are characterized, on the average, by a high rate of incorporation of 3H-TdR before therapy, with a higher labeling index than the non-responders. The measurement of the variations of incorporation of 3H-TdR, 48 hours after starting chemotherapy and before any morphological change, allows an early evaluation of the therapeutic effect: in the responders there is generally a decrease of the 3H-TdR incorporation and of the labeling index. On the apposite in the non-responders there is an increase of the mean incorporation per labeled blast cell, even if the labeling index decreases. The measurement of the incorporation of 3H-TdR by liquid scintillation counting which gives an earlier response than autoradiography, may be considered as a useful prognostic tool in acute leukemia, mainly in the non-lymphoblastic types and during the first phase of the disease.


[Granulopoiesis: comparison of several methods for studying production and bone marrow cellularity (author's transl)]. Several methods measuring the granulocyte production have been compared in normal subjects, hypoplastic granulopenia, chronic leucopenia and acute leukemia: the granulocyte turnover-rate after injection of in vitro 51Cr labelled autologous granulocytes, the efficiency of 75Se-selenothionine in vivo labelling, the flow-out of the myelocyte compartment after in vitro flash-labelling with 3H-thymidine. The two first methods are correlated and measure the global effective production of granulocytes. The third method is a fractional measure of the bone marrow production and is not correlated with the former ones. The ratio between the granulocyte turnover-rate and the flow-out of the bone marrow proliferative pool is a measure of the bone marrow cellularity. Discrepancies between this measure and the granulocyte turnover-rate has been found only in leukaemic patients.


[Editorial: Discrepancies in granulocyte survival time (author's transl)]. Important discrepancies between the renewal rate of mature granulocytes measured by in vitro labelling with DF 32P and with sodium chromate 51Cr are observed. The possible explanations are reviewed: errors made in the drawing of the experimental curve of the radioactivity disappearance; labelling of long-living lymphocytes with chromium; elution of 32P; toxicity of DFP for the tagged cells. If the renewal-rate measured by using 51Cr is the real one, as it seems to be, currently accepted models of granulopoiesis will need reappraisal.


[Myeloid leukemia with myelofibrosis (author's transl)]. Myelofibrosis is observed in 1/5 of the cases of C.M.L. It consists of reticulin fibers with few collagen and no osteosclerosis. It involves signs which usually indicate the extension of the myelosis to other organs and other types of cells: hepatosplenomegaly, erythroblastosis, thrombocytemia. Its prognosis is always bad. In one third of the cases, myelofibrosis develops early, and in two third it is late. Chimiotherapy is not responsible for it. These forms of C.M.L. with myelofibrosis appear as a special type of myeloproliferative disorder apart from the true C.M.L. and the true osteomyelofibrosis.


[Study of chronic myeloid leukemia by marrow culture in vitro (author's transl)]. The technique of bone marrow cultures in vitro on semi solid agar medium has been applied to the study of 24 cases of chronic myeloid leukemia in chronic phase and/or in acute blastic crisis. At the chronic phase of the disease the colony forming capacity is normal or above the normal range. In blastic crisis the number of colonies is very low. In the same time clusters are prominent.


[Androgens and prolonged complete remissions in acute non lymphoblastic leukemias. Results of a systematic treatment with stanozolol associated with chemotherapy (author's transl)]. An androgen (stanozolol: 0,15 mg/kg/d) was systematically associated to the treatment of acute non lymphoblastic leukemias, since the beginning of induction therapy (vincristin, daunorubicin, prednisone) and throughout the maintenance period (6-mercaptopurine and methotrexate). Thirty-six patients less than 60 years old (median age: 44 years) presenting with acute non-lymphoblastic leukemia were entered to the study. Sixteen achieved complete remission (C.R.), i.e. 44% of the whole and 53% of treated patients. Out of 16 patients with complete remission, 4 relapsed during the observation period which lasted 4-1/2 years. The stability of the hematologic equilibrium in patients in C.R. is the main finding of the present study. The actuarial curve of the duration of the first complete remission reaches a "plateau"; after the 8th month only one relapse was observed in 9 patients. The rate of C.R. at 2 years is 76 +/- 23%. As compared to the results from other schedules of treatment, this rate appears significantly better, specially in the case of immunotherapy (p less than 0,001). A prospective randomized study is now suggested as to confirm this result; its therapeutic and theoretical basis and perspectives are discussed.


[Hereditary hypogranular thrombocytopathic thrombocytopenia. Ultrastructural study of a megakaryocytopathy (author's transl)]. The ultrastructure of megakaryocytes and blood platelets has been studied in 3 members of a Swiss family, in which a haemorrhagic diathesis of varying degree occurred in at least three generations. Blood platelets show an extremely low number of dense granules, slightly increased mean diameter and irregular distribution of glycogen in unusually large clusters. The number of megakaryocytes in bone marrow is normal. Electron microscopy shows asynchronous development of their structural components: a severe defect in maturation of specific granules is followed by retardation and irregularity in the demarcation of the "platelet prospective fields", Golgi membranes have few vesicles, glycogen occurs partly in huge clusters and megakaryocytes with mature platelet fields are very rare. The ultrastructural picture of platelet formation in megakaryocytes indicates that this haemorrhagic syndrome may be understood as a megakaryocytopathy. The results of functional and biochemical analyses performed on blood platelets show a defect in both phases of aggregation and in PF 3 availability, normal survival, decreased total sialic acid content, and absence of the heaviest platelet population ("D") isolated by discontinuous sucrose gradient. The autosomal dominant transmission of this together with its concomitance with blood group O suggest its relationship with the familial thrombopathic thrombocytopenia described in 1968 by Kurstjens and al. in a Dutch family [13].


Necrotaxis: a scanning and transmission electron microscopy study. This paper describes an easy method to determine a "necrotactic rosette forming index (NRI) of leucocytes" after destruction of a red cell by a laser beam. Among the drugs tested, only cytochalasin B (0.5 mug/ml) is found to significantly lower NRI. The lectins PHA and CON-A did not have any effect on the NRI. Patients with infections are found to have increased NRI. Transmission and scanning electron microscopy have permitted the study of fine structurale details of the entire phenomenon.


[Effects of various adsorbants on coagulation factors (author's transl)]. Adsorption of clotting factors by various adsorbants is studied (tricalcium phosphate, baryum sulfate or carbonate or citrate, calcium oxalate, aluminium hydroxyde and several silicate such as: kaolin, celite, bentonite, attapulgite, beidellite, asbestos). The main properties of each adsorbant are listed as well as several applications such as: selective adsorption of fibrinogen, separation between fibrinogen and factor VIII, separation of factor II from the other components of the prothrombin complex. Activation of factors XII and XI by the various silicates, as well as the activation of factor V by attapulgite are studied. Finally, the action of such adsorbants on the fibrinolytic system is summarized.


Oxygen exchange in the gamma-phosphoryl group of protein-bound ATP during Mg2+-dependent adenosine triphosphatase activity of myosin. When ATP binds to myosin in the presence of Mg2+ there follows a rapid cleavage reaction to yield a myosin-product complex whose breakdown is rate-limiting in the overall adenosine triphosphatase reaction at 21 degrees and pH 8.0. Recent kinetic studies on this system have led to the proposal that the cleavage of ATP bound to myosin is reversible. This conclusion is based in part on the observation that when ATP is mixed with an excess of myosin active sites a small amount of tightly bound ATP exists whose life-time coincides with that of the myosin-product complex and implies these two species are in equilibrium during their decay. Previous oxygen exchange studies have shown that phosphate released as free product contains more than one oxygen atom from water. A rapid equilibration between myosin-bound ATP and a myosin-products complex can account for the extra water oxygen incorporation of the product phosphate. Such a model requires that the gamma-phosphoryl group of the bound ATP also exchanges its oxygen atoms with water. Results presented in this paper show that protein-bound ATP labeled in the three terminal oxygen atoms of the gamma-phosphoryl group with 18O exchanges about 75% of its label within 2 s of binding to the active site of myosin. This result provides chemical evidence for a model in which bound ATP undergoes a reversible reaction with water. Incomplete exchange may arise from kinetic and/or structural restraints on the mechanism and plausible models are discussed.


The effect of apomorphine on serotonin neurons in dorsal raphe nucleus and catecholamine nerve terminals in paraventricular hypothalamic nucleus. Histofluorescence studies. Histofluorescent method was employed to study the intensity of serotonin (5-HT) fluorescence in nucleus raphe dorsalis (DR) and catecholamine (CA) fluorescence in nucleus paraventricularis hypothalami (PVH). Apomorphine (APO, 20 mg/kg) induced, after 30 min, the increase in intensity of 5-HT fluorescence in DR and CA fluorescence in PVH. This increase was prevented by the previous dopamine receptor blockade by spiroperidol (2 mg/kg), administered 1 hr prior to APO. After alpha-methyl-tyrosine (alpha-MT) (2 X 400 mg/kg)-induced decrease in CA level, simultaneous intensification in 5-HT fluorescence was observed in DR. APO administered to those rats decreased the intensity of 5-HT fluorescence in DR, down to the level observed in the controls. This decrease in fluorescence was also prevented by pretreatment with spiroperidol. p-Chlorophenylalanine (PCPA)-induced inhibition in 5-HT synthesis decreased the intensity of 5-HT fluorescence and this decrease was considerably intensified by APO. The obtained results support the following concepts: 1) the effect of apomorphine on 5-HT in DR and on CA in PVH seems, to be indirect, namely through dopamine receptor stimulation, 2) the changes in 5-HT fluorescence in DR are probably connected with the acceleration in this amine turnover.


Ethidium bromide inhibits mitochondrial phosphorylating oxidation. Ethidium bromide, in addition to combination with mitochondrial nucleic acids, is a phosphorylation inhibitor during glutamate and succinate respiration by mitochondria. Exhaustive washing of ethidium bromide-treated mitochondria did not relieve the inhibition nor significantly decrease the amount of bound dye. Dialysis against a cation exchange resin at 3 degrees for 17 hr removed about 97% of bound dye. This restored phosphorylating capacity to that of untreated mitochondria which had also been dialyzed against the resin. Since state 3 respiration was diminished and state 4 was unaffected by the presence of the acridine dye, and since neither swelling of mitochondria nor release of latent ATPase was observed, then ethidium bromide was not an electron transport inhibitor nor an uncoupler of oxidative phosphorylation. Inhibition of metabolic processes by ethidium bromide may be due in part to depressed generation of mitochondrial ATP.


[Staphylococcus aureus phage induction by and sensitivity to ultraviolet rays]. The main characteristics of the phiD convertant phage of Staphylococcus aureus have been studied, including the quantitative study of the induction of BSS (phiD) strain by U.V., of the multiplication of the inducted phage, and of its sensitivity to the U.V. irradiation.


Long-term maintenance in vitro of human T cells by repeated exposure to the same stimulator cells. Differences when using repeated stimulation in allogeneic mixed leukocyte culture and when using stimulation with autologous lymphoblastoid cells. Cells from one-way human mixed leukocyte cultures (MLC) which had reverted to small lymphocytes after 2 weeks' incubation responded with accelerated kinetics and higher thymidine incorporation on restimulation with lymphocytes or lymphoblastoid cell line (LCL) cells having relevant antigens. In contrast to fresh lymphocytes, they did not respond to autologous LCL cells. Cultures could be restimulated every second week with relevant allogeneic lymphocytes and could thus be maintained for periods of up to 4 months. Almost all these cultured cells had T-cell characteristics, during stimulation as well as in their reverted phase. The response to phytohemagglutinin (PHA) successively disappeared with repeated allogeneic restimulation, whereas the response to the relevant lymphocytes and cells of related donors was maintained. When lymphocytes had been stimulated with autologous LCL cells, the restimulation response was accelerated, although lower than after the primary stimulation. Restimulated cultures could not be maintained by further restimulation. Allogeneic and autologous LCL were equally efficient restumulators. A low level of stimulation was also achieved with allogeneic lymphocytes. The PHA response was usually reduced.


Role of B cells in the expression of the PPD response of human lymphocytes in vitro. Human blood lymphocytes were tested for their proliferative response to purified protein derivative (PPD) in vitro. Purified T cells showed a lower response than the orginial purified lymphocyte suspension. Purified B cells were unresponsive to PPD. Mixtures of preparations of T and B cells were stimulated to a greater extent by this antigen than anticipated by adding the responses of the two cell types tested separately. This cell synergy could largely be explained by soluble factors released by PPD-stimulated T cells, which were found to be mitogenic for B cells.


Back pain and isometric back muscle strength of workers in a Danish factory. 105 factory workers (38 females and 67 males) have been questioned about their frequency of back pain. 60% of the females and 61% of the males have previously experienced episodes of back pain. 21% of the females and 37% of the males have been absent from work due to back pain. The incidence of back pain is not related to age, height, sort of work, or isometric muscle strength of the back (IS). For the males the incidence rises with increasing weight, i.e. combination of height and obesity, but is not related to any two single factors. For the females there is no correlation between the incidence of pain and weight. IS is correlated to height and age in the males but not in the females. Standards for IS are presented and suggested as a guide to evaluation of the working capabilities of individual subjects with back pain.