Study on the fixation method of microcirculation system for electron microscopy.

1) Although glutaraldehyde solution was injected into the superior mesenteric artery, vessels and blood cells were not fixed immediately. 2) Glutaraldehyde was considered not to be suitable for the fixation of vessels and blood cells. 3) To observe the physiological and topographical correlation of vessels and blood contents, drop osmium fixation method was better to fix the microcirculatory units immediately. 4) The perfusion of glutaraldehyde was supposed to be necessary for the fixation of the solid tissue in which microcirculatory systems exist in deeper places like liver. However, the findings observed by the perfusion method might reflex microcirculatory disturbances occured by the perfusion of glutaraldehyde itself.


Changes in the acyl lipid composition of photosynthetic bacteria grown under photosynthetic and non-photosynthetic conditions.

The acyl lipids and their constituent fatty acids were studied in the photosynthetic bacteria Rhodospirillum rubrum, Rhodopseudomonas capsulata and Rhodopseudomonas sphaeroides, which were grown under photosynthetic and non-photosynthetic conditions. The major lipids were found to be phosphatidylethanolamine, phosphatidylglycerol and cardiolipin in each bacterium. The two Rhodopseudomonas species also contained significant quantities of phosphatidylcholine. Other acyl lipids accounted for less than 10% of the total. On changing growth conditions from non-photosynthetic to photosynthetic a large increase in the relative proportion of phosphatidylglycerol was seen at the expense of phosphatidyl-ethanolamine. In Rhodospirillum rubrum the fatty acids of the major phospholipids showed an increase in the proportion of palmitate and stearate and a decrease in palmitoleate and vaccenate on changing growth conditions to photosynthetic. In contrast, the exceptionally high levels (>80%) of vaccenate in individual phospholipids of Rhodopseudomonas capsulata and Rhodopseudomonas sphaeroides were unaffected by changing growth conditions to photosynthetic. Analysis of the lipids of chromatophores, isolated from the three bacteria, showed that these preparations were enriched in phosphatidylglycerol. The large increase in this phospholipid, seen during growth under photosynthetic conditions, appeared, therefore, to be due to a proliferation of chromatophore membranes. Possible roles for acyl lipids in the formation and function of the photosynthetic apparatus of bacteria are discussed.


Studies on the mechanism of hepatic microsomal N-oxide formation. N-oxidation of NN-dimethylaniline by a reconstituted rabbit liver microsomal cytochrome P-448 enzyme system.

The N-oxidation of NN-dimethylaniline was studied by using a reconstituted rabbit liver microsomal enzyme system consisting of highly purified cytochrome P-448, NADPH-cytochrome c reductase and lipid factor. Both cytochrome P-448 and NADPH-cytochrome c reductase were required for optimum N-oxygenating activity; the catalytic capacity of the reductase fraction for supporting N-oxide formation varied with the isolation procedure applied. Addition of microsomal lipids to the assay media stimulated N-oxidation of the arylamine. N-Oxide formation appeared to be not generally controlled by electron transfer from cytochrome b5 to cytochrome P-448. The present work confirms that cytochrome P-448 can mediate about 44% of the rabbit liver microsomal N-oxidation of NN-dimethylaniline, thus reinforcing the existence of at least two distinct tertiary amine N-oxidases, i.e. haemoprotein and flavoprotein oxidase, in liver microsomal fractions.


[Studies on cellular protein synthesis in synovial fluid following intraarticular gold administration (author's transl)].

Na-aurothiomalate was applied intraarticularly in the conentration range between 2 and 40 mg to joints of patients with rheumatoid arthritis. Between 2 and 20 mg a constant inhibition of 3H-aminoacids incorporation in proteins was observed. At a dosage of 40 mg of this gold salt a strong inhibition could be detected. In the low concentration range an additional degradation of structurally changed proteins could be the cause of the lower incorporation rate of 3H-aminoacids. From 40 mg on a real inhibition of protein synthesis seems to occur.


Reinforcing properties of nicocodine in the rhesus monkey.

The present study demonstrates that nicocodine is self-administered by the rhesus monkey in cross self-administration experiments. The minimum reinforcing dose is 10 times higher than that of codeine and 100 times higher than that of heroin. At reinforcing doses the rate of self-administered infusions of nicocodine is comparable with those of codeine. Further experiments are necessary to rank order nicocodine as to its positive reinforcing properties among the opiate-like compounds. On the other hand the present experiments demonstrate that nicocodine is an opium-like reinforcing compound.


Influence of anticoagulant substances on phosphoserine and taurine of human blood.

The influence of heparine (Liquemin) and sodium-EDTA on phosphoserine and taurine of human blood was investigated. Phosphoserine which was not or nearly not present in serum increased after addition of both the anticoagulant substances. The heparine used contained phosphoserine as the only ninhydrinic substance in low concentrations. After addition of heparine phosphoserine in human plasma does not increase proportionally to that change. A mechanism is assumed which leads to the release from cells or which stimulates the synthesis of the substance. A similar effect was found with sodium-EDTA but not with sodium citrate. An opposite effect was detected with taurine in human blood. In serum the highest concentration was measured which decreased slowly under heparine as well as under sodium-EDTA addition.


Antimicrobial agents in rats. II. Serum levels of oral cephalosporins.

Cephalexin, cephradine and 7 beta-(D-2-amino-2-[(1,4-cyclohexadienyl)-acetamido])-3-methoxy-3-cephem-4-carboxylic acid (CGP 9000) were tested for their pharmacokinetic behaviour in rats. The cephalosporin serum concentrations were determined at certain times after oral administration of 150 mg/kg by the agar-well-diffusion method. The experiments revealed that the serum levels of cephalexin and cephradine did not differe essentially from one another. They maintained maximum serum concentrations of 30 microgram/ml to 40 microgram/ml during the first hour and than declined with a half-life of 2.5 h. CGP 9000 reached peak concentrations 60 to 90 min postdose and was eliminated with a half-life of 3.5 h. The area under the curve was double as large as those of cephalexin and cephradine. This may be a reason of favourable results in experimental chemotherapy with CGP 9000.


[Combined pituitary stimulation test using LHRH, TRH and insulin in subjects with short stature: evaluation of the response of pituitary tropins at 8 a.m. and at 8 p.m].

The effect of a combined test (insulin, TRH, LHRH) on plasma levels of GH, LH, FSH, PRL and Cortisol was studied in 5 subjects with short stature. Two test were performed at 8 a.m. and at 8 p.m. In all subjects the GH, FSH, LH, TSH, PRL and Cortisol levels showed no relevant response during the two tests.


[Plasma LH, FSH, TSH, prolactin and cortisol during a combined test of pituitary reserve in prepubertal subjects with short stature].

The effect of a combined test (insulin, TRH, LHRH) on plasma concentrations of LH, FSH, PRL, Cortisol was studied in 18 normal subjects in prepubescent state and in 65 patients in prepubescent state with short stature. In 6 subjects with short stature there was no change of LH, and in 7 subjects with short stature there was no change of FSH. In conclusion the combined test for pituitary stimulation provides an useful method for localizing the lesion in disorders of hypothalamo-pituitary axis.


Differential effects of metal ions on Rhodospirillum rubrum ribulosebisphosphate carboxylase/oxygenase and stoichiometric incorporation of HCO3- into a cobalt(III)--enzyme complex.

Mg2+ or Mn2+ ions supported both the carboxylase and oxygenase activities of the Rhodospirillum rubrum ribulosebisphosphate carboxylase/oxygenase. For the carboxylase reaction, Mn2+ supported 25% of the maximum activity obtained with Mg2+; oxygenase activity, however, was twice as great with Mn2+ as compared to that with Mg2+. A further differential effect was obtained with Co2+. Co2+ did not support carboxylase activity and, in fact, was a strong inhibitor of Mg2+-dependent carboxylase activity, with a Ki of 10 microM. Co2+ did, however, support oxygenase activity, eliciting about 40% of the Mg2+-dependent oxygenase activity. No other divalent cations supported either activity. With high concentrations of Mg2+ or Mn2+, maximum carboxylase activity was seen after a 5-min activation period; activity decreased to about half of maximum after 30-min activation. A similar time dependence of activation was observed with Mn2+-dependent oxygenase activity but was not seen for Mg2+- or Co2+-dependent activity. Both carboxylase and oxygenase activities were inactivated by the oxidation of Co2+ to Co(III) with the resultant formation of a stable Co(III)--enzyme complex. In the presence of HCO3- (CO2), Co(III) modification was stoichiometric, with two cobalt atoms bound per enzyme dimer. Carbon dioxide was also incorporated into this Co(III)--enzyme complex, but only one molecule per enzyme dimer was bound, indicative of half-the-sites activity. These results thus indicate that there are substantial differences in the metal ion sites of the carboxylase and oxygenase activities of R, rubrum ribulosebisphosphate carboxylase/oxygenase.


Subunit association and side-chain reactivities of bovine erythrocyte superoxide dismutase in denaturing solvents.

The copper- and zinc-containing superoxide dismutase of bovine erythrocytes retains its native molecular weight of 32 000 in 8.0 M urea for at least 72 h at 25 degrees C, as evidenced by sedimentation equilibrium analysis. Subsequent to prolonged exposure to urea, the dimeric enzyme could be dissociated by sodium dodecyl sulfate in the absence of reductants, indicating the absence of unnatural disulfide cross-links. The sulfhydryl group of cysteine-6 was unreactive toward 5,5'-dithiobis(2-nitrobenzoic acid) or bromoacetic acid in both neutral buffer and 8.0 M urea. The histidine residues of the enzyme were resistant to carboxymethylation in neutral buffer and 8.0 M urea. However, when the enzyme was exposed to bromoacetic acid in the presence of 6.0 M guanidinium chloride and 1 mM (ethylenedinitriol)tetraacetic acid (EDTA), both sulfhydryl and histidine alkylation were observed. Guanidinium chloride (6.0 M) increased the reactivity of the sulfhydryl group of cysteine-6 and allowed the oxidative formation of disulfide-bridged dimers. This was prevented by 1 mM EDTA. It follows that 8.0 M urea neither dissociates the native enzyme into subunits nor produces a conformation detectably different than that possessed under native conditions.


Energy transfer among the chromophores in phycocyanins measured by picosecond kinetics.

Energy-transfer processes in the algal light-harvesting proteins, the phycocyanins, have been studied by means of picosecond absorption spectroscopy. After excitation at 530 nm, the absorption at several wavelengths in the range 480--669 nm decayed with a short time constant (picosecond) and a long time constant (greater than 1 ns). For C-phycocyanin, energy transfer from the beta to the alpha subunits is interpreted as being a likely candidate for the short time constant; the long time constant probably is the excitation lifetime of the chromophore on the alpha subunits. The time constants for energy transfer in monomers, trimers, and hexamers of C-phycocyanin extracted from a blue-green alga, Phormidium luridum, were measured as approximately 85, approximately 56, and approximately 32 ps, respectively. The corresponding time constant in the cryptomonad phycocyanin 645 from Chroomonas species was found to be less than 5 ps.


Adsorption of monovalent cations to bilayer membranes containing negative phospholipids.

The electrophoretic mobilities of multilamellar phosphatidylserine vesicles were measured in solutions containing monovalent cations, and the xi potentials, the electrostatic potentials at the hydrodynamic plane of shear, were calculated from the Helmholtz--Smoluchowski equation. In the presence of 0.1 M lithium, sodium, ammonium, potassium, rubidium, cesium, tetraethylammonium, and tetramethylammonium chloride, the xi potentials were -60, -62, -72, -73, -77, -80, -82, and -91 mV, respectively. Similar results were obtained with phosphatidylglycerol vesicles; different results were obtained with cardiolipin, phosphatidylinositol, and phosphatidic acid vesicles. The phosphatidylserine results are interpreted in terms of the Stern equation, a combination of the Gouy equation from the theory of the diffuse double layer, the Boltzmann relation, and the Langmuir adsorption isotherm. Evidence is presented that suggests the hydrodynamic plane of shear is 2 A from the surface of the membrane in solutions containing the alkali metal cations. With this assumption, the intrinsic association constants of the above monovalent cations with phosphatidylserine are 0.8, 0.6, 0.17, 0.15, 0.08, 0.05, 0.03, and 0 M-1, respectively. The validity of this approach was tested in two ways. First, the xi potentials of vesicles formed from mixtures of phosphatidylserine and a zwitterionic lipid, phosphatidylcholine, were measured in solutions containing different concentrations of sodium. All the data could be described by the Stern equation if the "relaxation" of the ionic atmosphere, which is predicted by classic electrostatic and hydrodynamic theory to occur at low salt concentrations and high potentials, was circumvented by using only large (diameter greater than 13 micrometers) vesicles for these measurements. Second, the fluorescent probe 2-(p-toluidinyl)naphthalene-6-sulfonate was used to estimate the potential at the surface of phosphatidylserine and phosphatidylglycerol vesicles sonicated in 0.1 M NaCl. Reasonable agreement with the predicted values of the surface potential was obtained.


Asymmetry of the site of choline incorporation into phosphatidylcholine of rat liver microsomes.

[14C]Choline was incorporated into microsomal membranes in vivo, and from CDP-[14C]choline in vitro, and the site of incorporation determined by hydrolysis of the outer leaflet of the membrane bilayer using phospholipase C from Clostridium welchii. Labelled phosphatidylcholine was found to be concentrated in the outer leaflet of the membrane bilayer with a specific activity approximately three times that of the inner leaflet. During incorporation of CDP-choline and treatment with phospholipase C the vesicles retained labelled-protein contents indicating that they remained intact. When the microsomes were opened with taurocholate after incorporation of [14C]choline in vivo, the labelled phosphatidylcholine behaved as a single pool. Selective hydrolysis of labelled phosphatidylcholine in intact vesicles is not, therefore, a consequence of specificity of phospholipase C. These results indicate that the phosphatidylcholine of the outer leaflet of the microsomal membrane bilayer is preferentially labelled by the choline-phosphotransferase pathway and that this pool of phospholipid does not equilibrate with that of the inner leaflet.


Selective inhibition of precursor incorporation into ribosomal RNA in gamma-irradiated Tetrahymena pyriformis.

Sublethal doses of gamma radiation are known to inhibit total RNA synthesis in the ciliate protozoan Tetrahymena. To determine if the synthesis of a particular class of RNA is preferentially inhibited, pulse-labeled RNA was isolated from normal exponentially growing cells, irradiated cells, and cells in which total RNA synthesis had recovered to the pre-irradiation level. The RNAs were analyzed by SDS-polyacrylamide gel electrophoresis and oligo(dT)-cellulose column chromatography. Inhibition of RNA synthesis primarily involves ribosomal RNA. However, radiation does not cause a delay in the processing of precursor rRNA or a preferential loss of either of the mature rRNAs. Following irradiation, poly(A)-containing RNA [poly(A+)RNA] is synthesized at a rate up to three times greater than the control rate. The elevated poly(A+)RNA synthesis occurs during the period of depressed rRNA synthesis and even after rRNA synthesis has recovered to its pre-irradiation rate. While the sizes of the total cellular ribonucleoside triphosphate pools are depressed in the irradiated cells, these pools probably do not represent the actual compartments containing the precursors for RNA synthesis, and the observed changes cannot explain the modifications in macromolecular synthesis in irradiated Tetrahymena.


Distribution pattern and enzymic hypermethylation of inverted repetitive DNA sequences in P815 mastocytoma cells.

A specific class of DNA sequences, the inverted repetitive sequences, forms a double-stranded structure within a single linear polynucleotide chain in denatured DNA. The reassociation process is unimolecular and occurs very fast. Quantitative analyses have shown that in mouse P815 cells these sequences comprise about 4% of the nuclear DNA and are interspersed within sequences of other degrees of repetitiveness. After labeling the cells with L-[Me-3H]methionine and [14C]deoxycytidine, relative rates of enzymic DNA methylation were computed on the basis of radioactivities found in pyrimidine residues of the nuclear DNA. The results indicate that in P815 cells, DNA of inverted repetitive sequences is methylated to a level about 50% higher than the normal repetitive DNA sequences and to about 300% higher than the unique and intermediary intermediatry sequences. The biological function of the inverted repetitive sequences, as well as of the role of enzymic methylation of DNA remains unknown.


Characterization of UDP-galactosyl:asialo-mucin transferase activity in the Golgi system of rat liver.

UDPgalactosyltransferase activity (UDPgalactose:mucopolysaccharide galactosyltransferase, EC 2.4.1.74) was measured in a well-characterized fraction of Golgi membranes in the presence of UDPgalactose and exogenous acceptor sites. Substrate saturation for 0.05 mg Golgi protein was achieved at a concentration of 4.6 mM UDPgalactose. Desialylated mucin proved to be the most suitable acceptor protein. Access to galactose acceptor sites was not rate limiting for the reaction when 20 mg of asialo-mucin/ml of incubation mixture was used. With these concentrations of substrates the use of nucleotides to inhibit pyrophosphatases and of detergents to perturb the membrane structure was not necessary and proved, in fact, to be inhibitory to galactose transfer. UDPgalactosyl:asialo-mucin transferase activity in Golgi membranes was 230 nmol galactose transferred/mg Golgi protein per 30 min.


Pyrimidine-degrading enzymes. Purification and properties of beta-ureidopropionase of Euglena gracilis.

In photoorganotrophically grown, mid-log phase cells of Euglena gracilis, enzymes of pyrimidine degradation including uracil reductase, dihydrouracil dehydrogenase, dihydropyrimidinase, and beta-ureidopropionase, were detected in a crude extract. beta-Ureidopropionase (N-carbamoyl-beta-alanine amidohydrolase, EC 3.5.1.6) was purified 100-fold by heat treatment, ammonium sulphate fractionation and chromatography using Sepharose 6B and DEAE-Sephadex A-25. The enzyme follows Michaelis-Menten kinetics (Km of beta-ureidopropionase for beta-ureidopropionate 3.8 . 10(-5) M, Hill coefficient n = 1). Other enzyme properties are: pH optimum 6.25, temperature optimum 60 degrees C, stimulation by Mg2+, inhibition by Cu2+, Mr approximately 1.5--2 . 10(6). beta-Ureidoisobutyrate, the intermediate of thymine degradation, and beta-ureidopropionate are competing substrates of beta-ureidopropionase (Ki = Km of beta-ureidopropionase for beta-ureidoisobutyrate 1.8 . 10(-5) M). Structural analogues of beta-ureidopropionate, isobutyrate and propionate are competitive inhibitors (Ki of beta-ureidopropionase 0.3 and 0.16 mM, respectively). There were no indications of regulatory function of beta-ureidopropionase in pyrimidine degradation.


Photosensory transduction in the flagellated alga, Euglena gracilis I. Action of divalent cations, Ca2+ antagonists and Ca2+ ionophore on motility and photobehavior.

1. The flagellated alga, Euglena gracilis, swims forward essentially in a straight path under constant light intensity. Strong motility of the cells can be supported by Mg2+ alone but optimum motility is found in the presence of Mg2+, Ca2+ and K+. 2. Ca2+, Co2+, Mn2+ and Ba2+ induce a concentration-dependent increase in the rate at which the cells change the direction of their swimming path (a klinokinesis). Ni2+ immobilizes the flagellum. 3. On perception of a reduction ('step-down stimulus') in blue light intensity in their environment, Euglena rotate in place (tumble) for a finite period (the step-down photophobic response). 4. The duration of the tumbling is enhanced in the presence of divalent cations following the series Ca2+ greater than Ba2+ greater than Mn2+ greater than Co2+ greater than Mg2+ = Ni2+ = 0. 5. Neither the tumbling response in the presence of low concentrations of Ca2+ or the Ca2+-stimulated response is altered by verapamil (a Ca2+ conductance antagonist). The Ca2+ conductance/active transport antagonist, ruthenium red, is also inactive. 6. The Ca2+ ionophore, A23187, has little effect on flagellar activity in the absence of extracellular Ca2+. However, in the presence of A23187, Ca2+ induces a specific light-independent, concentration-dependent discontinuous tumbling response of the cells. 7. The data support a role for Ca2+ and Mg2+ in control of flagellar activity. However, blue light-induced tumbling behavior would not appear to be the direct result of a light-mediated alteration in the Ca2+ conductance of the flagellar membrane to affect flagellar reorientation. The results are discussed in connection with previous theories on control of flagella activity in green alga.


Chemical basis of the electrophoretic variation observed at the alcohol dehydrogenase locus of Drosophila melanogaster.

The amino acid substitution responsible for the different electrophoretic mobility of the ADHs alleloenzyme and the ADHf alleloenzyme of the alcohol dehydrogenase from a Nigerian population of Drosophila melanogaster has been established as lysine (ADHs) for threonine (ADHf). This result is discussed with reference to the charge state model of electrophoretic variation, in conjunction with other know substitutions at this locus. It is concluded that electrophoretic methods should be capable of distinguishing many alleloenzymes which have identical isoelectric points without recourse to explanations involving conformational variability.


Application of linear system analysis to the horizontal vestibulo-ocular reflex of the alert rhesus monkey using pseudorandom binary sequence and single frequency sinusoidal stimulation.

Horizontal eye movements of the alert rhesus monkey resulting from both pseudorandom binary sequence (PRBS) and single frequency sinusoidal rotational stimulation were analyzed using a PDP 11/40 computer in order to generate gain, phase, and coherence estimates at discrete frequencies between 0.008 and 1.28 Hz. A computer simulation of vestibular induced eye movements was used to validate our analysis procedures and to determine the effects of digital noise. Frequency domain transfer functions derived from gain and phase estimates revealed that the responses to PRBS stimulation and to single frequency sinusoids were not appreciably different. PRBS testing was accomplished in approximately one third the time required for sinusoidal testing and yielded highly reproducible data. We conclude that PRBS stimulation is a reliable and efficient method for assessing linear system parameters of the horizontal vestibulo-ocular reflex. PRBS testing may be particularly advantageous in studies of vestibulo-oculomotor plasticity in which rapid assessment of alterations in system dynamics is essential.


[Isolation and properties of highly purified C1. botulinum toxin type E].

A new method of isolation of highly purified Cl. botulinum toxin of E type from the cultural fluid of strain 188 centrifugates was developed. The method allows to isolate the toxin both in a precursor and in activated forms with a yield of 10--15%. The method includes fractionation by ammonium sulfate, ultrafiltration and subsequent column chromatography on DEAE-cellulose, Sephadex G-200 and DEAE-Sephadex A-50. The preparations were found homogeneous during polyacrylamide gel electrophoresis and immunoprecipitation in agar with antitoxic horse serum. The potential specific toxicity of the preparations is 1--1,2.10(7) DLM/mg of protein. The molecular weight of the toxin is about 160 000; the molar extinction coefficient is equal to 278 nm. The isoelectric point lies around pH 6.0. The highly purified Cl. botulinum toxin of E type was found stable upon storage.


The time course of the behavioral effects of amphetamine and their reversal by haloperidol in a primate species.

A group of six marmosets was administered amphetamine (Phase I), amphetamine plus haloperidol (Phase II), and then amphetamine alone (Phase III) over consecutive periods of 27, 51, and 33 days after which drug treatment was terminated (Phase IV). The animals' behavior was monitored during these periods and during a predrug treatment control period. Five mutually exclusive categories of behavior were assessed during the experiment. Social contact between animals was significantly suppressed and inactivity was increased throughout Phases I--III but both measures returned to normal values during Phase IV. Locomotion was significantly decreased towards the end of Phase I and initially during Phase II. Rapid head movements (termed checking) were significantly increased at the beginning of Phase I and again when the haloperidol was withdrawn at the beginning of Phase III. Towards the end of Phase I the animals developed destructive self-grooming habits. The time course of the effects of amphetamine and haloperidol on the different behavioral categories suggests that different mechanisms may be involved in each case. Viewed as a model of schizophrenia, the time course of haloperidol in reversing amphetamine-induced suppression of locomotion most closely resembles the time course of the antipsychotic effect of neuroleptics in man. Some effects of amphetamine (e.g., suppression of social interaction) are not reversed by haloperidol, and some effects of withdrawal of haloperidol (e.g., precipitation of checking movements not present when haloperidol was commenced) do not have an obvious counterpart in the clinical situation.


Identification of branched chain fatty acids in baboon liver lipids.

A total of 26 monomethyl branched saturated fatty acids was identified in baboon liver lipids; these included a novel anteiso component with an odd number of carbon atoms in the chain (13-methylpentadecanoic acid).


[Formation of epileptic activity complexes under the effect of the determinant focus in the cortex isolé].

In experiments on cats foci of enhanced excitation working in independent regimes were created in the neuronal-isolated cortex using weak strychnine solutions. Creation of a hyperactive focus by means of application of strychnine concentrated solutions or crystals leads to an increase in the amplitude and discharge frequency in other foci, synchronism in discharges of these foci and their unification into a single functional complex, working in the hyperactive focus regime. The latter, in such a way, presented the determinant structure. Under the influence of the determinaant focus there was also a marked generalization of the convulsive activity manifested by an embrace with epileptical discharges of the cortex area intact from strychnine. The results obtained indicate that the established relations between the foci of hyperactivity in the cortx as well as the effects of the determinant structure may be realized by the cortex mechanisms proper.


[Thiophosphamide-induced chromosome aberrations in embryonal liver and bone marrow cells of A/He and C57BL/L mice].

Chromosomal aberrations were studied in cells of embryonic liver and bone marrow of A/He and C57BL/6 mice following mutagenic treatment by the alkylating agent thiophosphamide in g1-S periods of the cell cycle. Higher sensitivity of chromosomes to aberration induction was found in A/He mice.


Hemopoietic stress and fetal hemoglobin synthesis: comparative studies in vivo and in vitro.

Baboons exposed to acute hemolytic stres increase their production of fetal hemoglobin (HbF). Although the maximal in vivo HbF levels attained in 5 treated animals varied from 6.4% to 34.8%, their cultured bone marrow erythroid cells reverted to the fetal pattern of hemoglobin synthesis. These data suggest that HbF synthesis is modulated by the interaction of inhibiting and promoting factors, which is different among animals in vivo but equal in the cultures of their bone marrow erythroid cells.


A comparison of the anti-anaphylactic activities of salbutamol and disodium cromoglycate in the rat, the rat mast cell and in human lung tissue.

1 Salbutamol and disodium cromoglycate were compared for anti-anaphylactic activity against passive anaphylaxis in rat skin and peritoneum in vivo and in rat mast cells and human lung fragments in vitro.2 Salbutamol administered intravenously to rats inhibited cutaneous anaphylaxis, but also inhibited cutaneous responses to histamine and 5-hydroxytryptamine. Salbutamol administered intraperitoneally inhibited the release of slow reacting substance of anaphylaxis (SRS-A) but not the release of histamine in the peritoneum. It was a very weak inhibitor of histamine release from rat mast cells in vitro.3 Disodium cromoglycate administered intravenously to rats inhibited cutaneous anaphylaxis. Disodium cromoglycate administered intraperitoneally to rats inhibited the release of histamine and, to a lesser extent, SRS-A in the peritoneum. It was an effective but short-acting inhibitor of histamine release from rat mast cells in vitro.4 Salbutamol was a potent inhibitor of the anaphylactic release of histamine and SRS-A from fragments of human lung.5 Disodium cromoglycate was a weak inhibitor of the anaphylactic release of histamine and SRS-A from fragments of human lung. The inhibition was variable and not dose-related.6 The concentration of salbutamol required to inhibit anaphylaxis in human lung is of the same order as that required to relax human bronchial muscle. It is suggested that salbutamol may be more effective in allergic asthma if given in a prophylactic regimen.


A comparison of the abilities of acetylsalicylic acid, flurbiprofen and indomethacin to inhibit the release reaction and prostaglandin synthesis in human blood platelets.

1 A quantitative comparison has been made of the abilities of acetylsalicylic acid, flurbiprofen and indomethacin to inhibit the adenosine diphosphate (ADP)-induced platelet release reaction and to inhibit the synthesis of prostaglandins from arachidonic acid. 2 Experiments were carried out on human platelets that had been incubated with the agents in vitro and on platelets obtained from volunteers who had ingested standard doses of the drugs. 3 The results obtained for acetylsalicylic acid show that there is a close relation between the release reaction and the synthesis of prostaglandins in platelets. 4 Flurbiprofen and indomethacin appear to inhibit the release reaction rather more effectively than they inhibit the synthesis of prostaglandins. It is possible that these agents inhibit the release reaction by another mechanism.


Thyrotrophin releasing hormone stimulates release of [3H]-dopamine from slices of rat nucleus accumbens in vitro.

Thyrotrophin releasing hormone (TRH) (25 to 100 microM) was found to stimulate the efflux of [3Hu-dopamine from small slices of rat nucleus accumbens, but not from similar slices of rat caudate nucleus. Uptake inhibition was not responsible for this action, since at 10 and 50 microM TRH had no effect on the ability of small slices of nucleus accumbens to accumulate radioactivity when incubated with 10(-7) M [3H]-dopamine. In addition the hormone had no effect on basal or dopamine-stimulated adenylate cyclase, nor did it displace [3H]-spiperone binding, in membrane preparations from nucleus accumbens.


Treatment settings in psychiatry: long-term family and social findings.

Treatment in a psychiatric unit, as distinct from a psychiatric hospital, held no benefits in the long term (5-8 years after admission) as far as the patient's mental and behavioural status and employment, or the family's burdens, health needs or attitude were concerned. Schizophrenic patients from both settings received more hospital treatment and medication, were more often unemployed, and had more adverse effect on relatives' health than neurotic or depressive patients. Hospital attendence identified relatives who expressed a need for support.


Downward gaze in monkeys: stimulation and lesion studies.

Ten monkeys were stimulated unilaterally and bilaterally through bipolar electrodes placed stereotactically on each side of the midline under light barbiturate anaesthesia. Bilateral simultaneous stimulation elicited straight downward binocular movements from a core of tissue about 40 mm3 on each side which included the fields of Forel, zona incerta, subthalamic nucleus, oral pole of the red nucleus, fasciculus retroflexus and 'area tegmentalis'. Unilateral stimulation of the same points yielded downward eye movements in only 25 per cent of the instances. Upward deviation of the globes could be elicited by bilateral stimulation of tissue located more caudal, ventral and medial than that from which downward movements were obtained. Bilateral electrolytic lesions within the region outlined above caused significant defects in downward gaze both in saccadic and slow pursuit binocular movements. Passive bending of the head backwards, however, resulted in downward deviation of the globes (oculocephalic reflex). Optokinetic nystagmus and after-nystagmus downward were abolished. Oblique (45 degrees) optokinetic stimulation elicited a perverted response in the horizontal plane. Vestibulo-ocular reflexes elicited by bilateral warm irrigation of both ear canals with the monkey in the erect position, or by turning the animal while lying on one side, caused a strong tonic deviation upward with absence of nystagmus downward. Some of these monkeys showed additional alterations in upward gaze but they were less severe in intensity and duration than those of downward gaze. All eye deviations in the horizontal plane were consistently normal. Recovery occurred in all types of vertical binocular movements except in the rapid motions (saccades and quick phases of nystagmus) below the horizontal meridian. A unilateral lesion had no effect. The minimal damage producing downward gaze defects was about 1.7 mm in diameter, cetred in the prerubral fields, rostral and medial to the red nuclei with minimal involvement of the oral pole of these structures. The nuclei of Cajal, Darkschewitsch and interstitialis of the posterior commissure, as well as the fasciculus retroflexus and the posterior commissure, were spared by this lesion. The so-called rostral interstitial nucleus of the medial longitudinal fasciculus and the nucleus campi Foreli appear to be destroyed. These structures are known to receive an input from the paramedian pontine reticular formation and project on to the oculomotor nerve nucleus. These results demonstrate that the prerubral fields contain structures which are critical for rapid eye movements downward, and therefore an isolated downward gaze palsy is a strong indicator of a bilateral lesion of this zone. The findings in the few reported cases with this sign and available pathological analysis suggest that our conclusions from the experimental monkey apply to man as well. The concept of bilateral innervation for vertical eye movements is amply confirmed for the downward vectors...


Sexual behavior in male rhesus monkeys elicited by electrical stimulation of preoptic and hypothalamic areas.

Electrical stimulation of the preoptic area, lateral hypothalamus or dorsomedial nucleus of the hypothalamus (DMH) has produced sexual behavioral responses in 11 male rhesus monkeys. At a short latency following the onset of each train of stimulation, the stimulated males would mount estrogen-treated, receptive females. Quantitative measures of the elicited sexual behavior, in 4 males, were applied to describe differences between the effect of preoptic and DMH stimulation on male sexual performance. As compared to spontaneous sexual activity, stimulation of the DMH produced mounts of longer duration, with more thrusts per mount, higher thrusting rates and a greater number of ejaculations per test session; refractory periods between successive ejaculatory episodes were shortened as a result of stimulation-induced mounting behavior. Preoptic stimulation also induced mounts of longer than normal duration characterized by a slightly increased number of thrusts per mount. In contrast to DMH stimulus effects, however, thrusting rate was decreased significantly by preoptic stimulation and ejaculation did not occur even after multiple stimulus-induced mounts. Penile erections were elicited by stimulation of the anterior hypothalamus, ventromedial nucleus and the posterior hypothalamus in socially isolated or restrained males; however, stimulation of these same sites was ineffective for provoking sexual behavioral responses when tests were performed while the males were paired with receptive females.


Vestibular projections to the monkey thalamus: an autoradiographic study.

Vestibulothalamic projections were studied in the monkey (macaca mulatta) by injecting anerograde trace substances (radioactive leucine and proline) into the vestibular nuclear complex. Terminal labelling was found bilaterally mainly in the nucleus ventroposterior lateralis pars oralis (VPLo) and to a lesser extent in the nucleus ventroposterior inferior (VPI) and nucleus ventralis lateralis pars caudalis (VLc). The labelling was sparse, and scattered over wide areas. The vestibular origin of this projection was confirmed by injecting retrograde tracer substances (horseradish peroxidase and 125I wheat germ agglutinin) into VPLo. In the autoradiographic study no labelling was found in the posterior group.


Disordered mineral metabolism produced by ketogenic diet therapy.

Vitamin D and mineral metabolism status was examined in five children maintained chronically on combined ketogenic diet-anticonvulsant drug therapy (KG), and the results compared to those obtained in 18 patients treated with anticonvulsant drugs alone (AD) and 15 normal controls. KG patients exhibited biochemical findings of vitamin D deficiency osteomalacia: decreased serum 25-hydroxyvitamin D (25OHD) and calcium concentrations, elevated serum alkaline phosphatase and parathyroid hormone concentrations, decreased urinary calcium and increased urinary hydroxyproline excretion, and decreased bone mass. Although the KG and AD groups demonstrated similar reductions in serum 25OHD concentration, the KG patients exhibited a significantly greater reduction in bone mass. In response to vitamin D supplementation (5000 IU/day), mean bone mass in the KG group increased by 8.1 +/- 0.9% (P less than 0.001) over a 12-month period. These results suggest that ketogenic diet and anticonvulsant drug therapy have additive deleterious effects on bone mass and that these effects are partially reversible by vitamin D treatment.


On the solid state of bone.

Photovoltaic effect (infrared and visible light) is observed in bone and its two major components, collagen and apatite, at room temperature. A dimunition in the magnitude of photovoltage is observed after exposure to ultraviolet light in all the cases. The drift mobility of the charge carriers is obtained by measuring I versus V relationships in sandwich samples and relating them to the permittivity of the medium. Lifetime of the injected carriers is measured in the usual way. The data are consistent with the hypothesis that the effects are due to protonic conduction phenomena.


X-ray diffraction of the calcified tissues in Polypterus.

Pyrolyzed scales, fin spines, and bone from the ray-finned bony fish Polypterus (Actinopterygii) showed two mineral phases on X-ray diffraction: hydroxyapatite (HA), Ca5(PO4)3OH, and whitlockite, Ca3(PO4)2. The ratio of HA/whitlockite varied with the structure (scale, spine, bone) within each individual fish. The relative proportions of HA to whitlockite in pyrolyzed samples reflected the Ca/P ratio of the sample. Whitlockite appears after pyrolysis when the Ca/P is lower than 1.67. Among the five fish investigated, for each structure a general trend was noted. The proportion of HA relative to whitlockite increased with size (age) of the fish. Thus the smallest fish, a juvenile, exhibited a low Ca/P mineral in its calcified tissues, whereas the larger fish had progressively more HA and less whitlockite.


Studies on the biology of fish bone. III. Ultrastructure of osteogenesis and resorption in osteocytic (cellular) and anosteocytic (acellular) bones.

The comparative ultrastructure of fish bone osteogenesis and resorption induced by scale removal was described in the osteocytic (cellular-boned) Carassius auratus and the anosteocytic (acellular-boned) Tilapia macrocephala. Osteocytes, present in osteocytic bone, were lacking in anosteocytic bone. In osteocytic bone the osteoblast secreted a collagenous preosseous matrix in which it became enmeshed and then was termed a preosteocyte. When the preosseous matrix mineralized, the preosteocyte was termed an osteocyte and was completely surrounded by bone. In anosteocytic bone the osteoblasts receded from the mineralizing front and never became trapped as osteocytes. During resorption, types A and B resorptive cells, present in both bone types, invaded the matrix and demineralized the osseous zone. These cells were characterized by large amounts of granular endoplasmic reticulum and intracellular inclusions containing crystal-like material. Although functionally similar to mammalian osteoclasts, these cells lacked a characteristic ruffled border and were not multinucleated. The osteocytes of cellular bone did not appear to be involved during demineralization.


Passive accumulation of magnesium, sodium, and potassium by chick calvaria.

Four-day-old chick calvaria were used to determine the passive concentrations of magnesium, sodium, and potassium in metabolically poisoned bone. When incubated in buffers containing the blood levels of sodium and magnesium, these calvaria contained sodium and magnesium at the identical concentrations found in freshly dissected calvaria. Calvarial sodium and magnesium levels could be varied by altering the buffer concentrations of these cations. The potassium content of metabolically poisoned calvaria incubated in buffers containing 4 mM potassium was less than 20% of the content of freshly dissected calvaria. When the buffer concentrations of sodium and potassium were systematically varied, ouabain-poisoned calvaria concentrated these cations in the bone extracellular fluid by a factor of approximately two above buffer cation levels. Presumably, the hydroxyapatite crystal zeta potential is responsible for this concentrative phenomenon. These results are discussed in terms of the control of the ionic content of the bone extracellular fluid by the postulated "bone membrane."


Matrix vesicles in chicken epiphyseal cartilage. Separation from lysosomes and the distribution of inorganic pyrophosphatase activity.

The extracellular matrix vesicles from epiphyseal cartilage of chickens were isolated by differential centrifugation. The matrix vesicles obtained showed considerable activity of lysosomal enzymes. This appears to have been due to lysosomal contamination because when we used a new density gradient medium (Percoll), the lysosomal enzyme activities and the activity of alkaline phosphatase could be totally separated. Electron microscopy of the alkaline phosphatase-rich fraction showed matrix vesicle-like structures. Phosphatase activities of the cells and matrix vesicles were further studied by Sephadex G-200 gel filtration. Specific magnesium-activated inorganic pyrophosphatase, distinct from nonspecific alkaline phosphatase, could be demonstrated in the cellular fraction. No such separate activity could be demonstrated in the matrix vesicle fraction, and it is supposed that the pyrophosphatase activity in the matrix vesicles originates from the alkaline phosphate.


Effect of human salivary proteins on the precipitation kinetics of calcium phosphate.

Inhibition of calcium phosphate precipitation in saliva, and prevention of the formation of mineral accretions on tooth surfaces, has been ascribed to the existence of inhibiting salivary macromolecules. Marked reductions in the crystal growth rate of hydroxyapatite (HA) seeds were measured in supersaturated solutions containing either of two proline-rich proteins, PRP1 or PRP3, or statherin; the three macromolecules were isolated from human parotid saliva. The reductions were also observed when the HA seeds were pretreated with solutions of the macromolecules before adding them to the supersaturated calcium phosphate solution. This effect was very similar in the case of the two PRPs and it was directly related to the extent of adsorption site coverage of these proteins on the HA seeds. The effect of statherin was larger than anticipated from its adsorption behavior. However, comparison on the basis of number of moles adsorbed per unit area of HA shows that the PRP are more effective inhibitors than statherin. The macromolecule concentrations used were considerably lower than those in the salivary secretions, therefore these macromolecules could readily prevent mineral accretion on tooth surfaces through their adsorption onto the enamel surface.


Effect of ethane-1-hydroxy-1,1-diphosphonate and vitamin D on bone mineralization.

Administration of large quantities of ethane-1-hydroxy-1,1-diphosphonate to growing chicks resulted in a decrease in percent bone ash and an increase in percent osteoid. The degree of inhibition of bone mineral accumulation was a function of both duration and quantity of ethane-1-hydroxy-1,1-diphosphonic acid administration. The inhibition of bone mineral accumulation could be partially corrected with administration of 1,25-dihydroxyvitamin D3. Administration of high levels of ethane-1-hydroxy-1,1-diphosphonate also resulted in inhibition of intestinal calcium absorption. This could be reversed or prevented by the administration of 1,25-dihydroxyvitamin D3.


Formation and serum disappearance of fragments of parathyroid hormone in the infused dog.

Radioiodinated parathyroid hormone [125I-PTH(1-84)] was infused into intact dogs. At various times venous samples were chromatographed to determine the levels of intact hormone (1-84) and large fragments in the circulation. Both bioactive (electrolytically iodinated) and inactive (chloramine-T-labeled) preparations were used. In both instances, plateau concentrations of intact hormone and of large metabolite(s) were quickly reached. After cessation of infusion the levels of intact hormone and metabolite(s) quickly declined. Clearly, in the dog, the peripheral formation and disappearance of large fragments of exogenous PTH occur at rates comparable to the clearance of the intact hormone itself.


Identification of O-phosphoserine, O-phosphothreonine and gamma-carboxyglutamic acid in the non-collagenous proteins of bovine cementum; comparison with dentin, enamel and bone.

O-phosphoserine [Ser(P)], O-phosphothreonine [Thr(P)], and gamma-carboxyglutamic acid (Gla) have been identified in native, calcified cementum and in non-collagenous proteins which can be extracted from the tissue in EDTA at neutral pH. The concentrations of Ser(P) and Thr(P) and the amino acid composition of the EDTA extractable proteins are more similar to those found in bone than in dentin or enamel. The concentration of Gla in cementum is lower than it is in bone and higher than it is in enamel, which contains essentially no Gla. Based on the contents of Gla in these mineralized tissues and the distribution of alkaline and acid phosphatases in these tissues, it is speculated that Gla may be part of these or other proenzymes rather than being involved directly and structurally with the deposition of the mineral phase.


Temporal cell-mediated immune responses of cattle following experimental and natural exposure to living Brucella abortus.

A study on cell-mediated immune responses in cattle with different exposure experiences to Brucella abortus was conducted by an in vitro lymphocyte stimulation assay. The purpose of this study was to determine how soon the cell-mediated immune responses would be detected following experimental exposure to B. abortus and to study the cell-mediated immune trend following experimental and natural exposure of cattle to B. abortus. The first positive cell-mediated immune responses occurred one to two weeks after experimental inoculation with living B. abortus strain 2308. The cell-mediated immune responses in these animals appeared at least one week before the appearance of of B. abortus serum agglutinating antibodies. Animals which were naturally infected with B. abortus biotypes 1 and 2 demonstrated positive cell-mediated immune responses throughout the study.


Long-term culture of canine peripheral blood monocytes in vitro.

Various cultural conditions were assessed for their ability to maintain canine peripheral blood monocytes in vitro. Approximately ten days after incubation of peripheral blood leukocytes in Earle's minimum essential medium supplemented with homologous red cell lysates and normal horse serum, virtually a pure macrophage culture was obtained which could then be maintained for about two months. This culture was judged to be pure by surface marker analysis and their phagocytic activity. The number of monocytes could be increased by injecting the dogs with a chloroform extract from Listeria monocytogenes prior to collection of the blood.


Actions of various gastrointestinal peptides on the isolated amphibian spinal cord.

The effects of a number of peptides which are found in the gastrointestinal tract have been ascertained on the direct current recorded dorsal and ventral root responses of the isolated hemisected toad spinal cord. Motilin, substance P, bombesin, neurotensin, and thyrotropin releasing hormone had potent depolarizing actions on dorsal root terminals and motoneurons. These substances evoked discernable effects at concentrations as low as 10--7 M, or even lower with motilin. The effects of motilin, neurotensin, and thyrotropin-releasing hormone were greatly reduced or abolished by perfusion of the preparation with tetrodotoxin. Adrenocorticotrophic hormone, secretin, and pancreozymin (cholecystokinin) also depolarized dorsal root terminals and motoneurons. The effects of secretin and cholecystokinin were not abolished by tetrodotoxin. Leu- and Met-enkephalin had weak hyperpolarizing actions on the dorsal and ventral root potentials of repetitively stimulated preparations. Gastrin, gastric inhibitory peptide, glucagon, and somatostatin had no apparent effects on the responses of the preparation. Angiotensin and vasopressin both had rather weak depolarizing effects on the dorsal and ventral roots.


Effects of MK-771 on the isolated amphibian spinal cord: comparison with thyrotropin-releasing hormone.

Topically applied MK-771 (pyro-2-aminoadipyl-histidyl-thiazolidine-4-carboxamide), a novel thyrotropin-releasing hormone (TRH) analog, was found to be equipotent with TRH in depolarizing the ventral roots of the isolated, hemisected amphibian (Bufo marinus) spinal cord. The 3-methyl-histidyl analog of TRH was approximately 10 times more potent than MK-771 and TRH. MK-771 is known to be equiactive with TRH in their actions on the pituitary gland. Taken together these findings suggest that the previously observed enhanced potency of systemically administered MK-771 over TRH in in vivo central nervous system (CNS) test paradigms is not likely to be due to a difference in the agonist requirements of CNS as compared with pituitary receptors for TRH.


Randomized sequential trial of parenteral nutrition in healing of colonic anastomoses in man.

Adults who underwent elective colonic resection with anastomosis for both benign and malignant disease were randomly assigned to control or treated groups. Treated patients received daily 50 ml/kg of amino acid solution containing either 10 or 25 g/dl glucose by central or peripheral vein plus 500 ml of commercial soybean emulsion for 24 hours before operation and for 5 consecutive days after. Patients in both groups were examined by roentgenography after a water-soluble contrast enema, between 10 and 14 days after operation, and the integrity of the anastomosis was assessed radiologically without knowledge of the treatment given. Results were assessed by sequential analysis. After 21 pairs of subjects were entered,a boundary was crossed making the area favourable to treatment inaccessible, and the trial was terminated. Forty-seven patients were included in the trial: there were four anastomotic leaks in 23 control subjects and eight leaks in 24 treated subjects. The authors conclude that intensive perioperative parenteral nutrition in patients who undergo routine elective colonic resection does not improve the rate of healing of colonic anastomoses.


Hypospadias: a review.

Hypospadias is a common congenital anomaly with a prevalence estimated to be as high as 1 in 125 live male births. Its etiology is not entirely resolved but is embryologic in origin and has been attributed to fusion defects. Several authors believe it is a manifestation of intersex. Experience to date has produced a general consensus with regard to indications, timing and surgical principles for repair of the hypospadiac meatus. While two-stage procedures have been the mainstay of therapy, successful single-stage procedures have recently been developed. The prevention and management of the complications of persistant chordee, fistula and stricture make hypospadias repair one of the more challenging urologic procedures.


Monitoring the critically ill surgical patient.

Critically ill surgical patients account for approximately half the patients in an active multidisciplinary critical care unit. Hypovolemia and sepsis are common in such patients and affect a number of organ systems. Monitoring these systems provides therapeutically relevant information that may decrease morbidity and improve patient survival. Circulatory hemodynamics may be assessed by direct measurement of the arterial blood pressure, central venous and pulmonary artery pressure monitoring and cardiac output determination; the data thus obtained are valuable in guiding fluid replacement in the hypovolemic individual. The respiratory status may be assessed by bedside spirometry and measurement of arterial blood gas tensions to gauge pulmonary function and the need for assisted ventilation. Renal dysfunction is common in such patients; careful analysis of both urine and blood may identify prerenal as opposed to renal and postrenal factors. Monitoring of the gastrointestinal tract, especially for hemorrhage, is important. Finally, careful attention to nutritional status and provision of adequate protein and energy intake by mouth or by vein is a vital component of the optimal care of these patients.


External irradiation in the management of stage II endometrial carcinoma.

Twenty-nine patients with stage II endometrial carcinoma were reviewed and the possible risk factors involved in state II disease are presented. Twenty-four patients received external irradiation as part of their treatment with or without intracavitary or intravaginal radium and/or TAH BSO. The 5-year actuarial survival in our series was 81.4%. The data showed that preoperative external irradiation can be effectively administered without undue complication. A strong argument against the traditional use of preoperative intracavitary radium is presented. Preoperative external irradiation administered with a 4-field box technique to deliver a minimum dose of 5000 rad in 5--6 weeks to all the structures at risk is the recommended treatment for stage II endometrial carcinoma.


Levels of histaminase and L-DOPA decarboxylase activity in the transition from C-cell hyperplasia to familial medullary thyroid carcinoma.

The distribution of calcitonin, L-dopa decarboxylase, and histaminase is studied in sectioned total thyroid glands from patients with different stages of familial medullary thyroid carcinoma. In 5 glands with gross carcinoma and in 3 with early microscopic carcinoma the distribution of all three parameters positively correlates (p less than .01 for each correlation). In contrast, in 6 glands with C-cell hyperplasia only the distribution of calcitonin and L-dopa decarboxylase correlates (r = 0.64, p less than .01) while those for histaminase vs. calcitonin (r = .17, p = N.S.) and histaminase vs. dopa decarboxylase (r = .03, p = N.S.) do not. In the glands with microscopic carcinoma the peak levels of histaminase occur in the areas of disease as defined by immunohistochemical staining of calcitonin; mean histaminase activity is the only one of the three parameters measured that distinguishes between C-cell hyperplasia and microscopic carcinoma (p less than .005). Immunohistochemical staining of histaminase shows positive cells in glands with gross and microscopic carcinoma, but in none of the glands with hyperplasia alone. Histaminase is thus found in high amounts in some malignant C-cells only and may be a useful marker to distinguish between hyperplasia and malignancy in thyroids with early C-cell proliferative disorders.


Depressed PHA transformation and acute leukemia in macroglobulinemia.

Two out of fifty-three patients with macroglobulinemia developed acute leukemia following chemotherapy. The Phytohemagglutinin (PHA) Transformation Index performed prior to the appearance of acute leukemia was found to be markedly depressed in these two patients in comparison with ten other patients with macroglobulinemia. In addition, a clone with monosomy 7 was detected in one of the currently reported patients when the leukemic process became apparent. The cytogenetic analysis was normal in the same patient 48 months earlier, when macroglobulinemia was diagnosed. The low PHA Transformation Index and the chromosomal hypodiploidy are of interest and their clinical significance merits further investigation.


Isolation and identification of a human serum fibronectin-like protein elevated during malignant disease.

A human DNA-binding protein, designated MAD-2, has recently been found to be elevated in the serum from patients with malignant diseases. MAD-2 has been purified approximately 500-fold from peritoneal and pleural fluids collected from cancer patients. Immunodiffusion studies have indicated that MAD-2 is immunochemically identical to human plasma fibronectin. The purified material has been resolved by sodium dodecyl sulfate gel electrophoresis into two major protein chains with molecular weights in the range of 200,000 to 210,000 in either the presence or absence of disulfide bond-reducing agents. These results suggest that MAD-2 is a fibronectin fragment which has been generated through proteolysis. A quantitative assay system capable of detecting ng quantities of MAD-2 has been developed and used to verify the presence of elevated MAD-2 levels in DNA-binding protein fractions isolated from the serum of individuals with malignant diseases.


Induction of osteogenic sarcomas and tumors of the hepatobiliary system in nonhuman primates with aflatoxin B1.

The carcinogenicity of aflatoxin B1 (AFB1) has been under evaluation in nonhuman primates for the past 13 years. A total of 47 Old World monkeys, chiefly rhesus and cynomolgus, have received AFB1 i.p. (0.125 to 0.25 mg/kg) and/or p.o. (0.1 to 0.8 mg/kg) for 2 months or longer, and 12 are currently alive and without evidence of tumor. Thirteen of the 35 monkeys necropsied to date (37%) developed one or more malignant neoplasms, yielding an overall tumor incidence of 28%. Five of the neoplasms were primary liver tumors (2 hepatocellular carcinomas and 3 hemangioendothelial sarcomas), and 2 cases of osteogenic sarcoma were found. Other tumors diagnosed were 6 carcinomas of the gall bladder or bile duct, 3 tumors of the pancreas or its ducts, and one papillary Grade I carcinoma of the urinary bladder. The tumors developed in animals receiving an average total AFB1 dose of 709 mg (range, 99 to 1354 mg) for an average of 114 months (range, 47 to 147 months). Fifteen of the 22 necropsied monkeys (68%) without tumor showed histological evidence of liver damage, including toxic hepatitis, cirrhosis, and hyperplastic liver nodules. These animals had received an average total AFB1 dose of 363 mg (range, 0.35 to 1368 mg) for an average of 55 months (range, 2 to 141 months). Our results indicate that AFB1 is a potent hepatotoxin and carcinogen in nonhuman primates and further support the hypothesis that humans exposed to this substance may be at risk of developing cancer.


Effects of transplacental exposure to diethylstilbestrol on carcinogenic susceptibility during postnatal life in hamster progeny.

Prenatal exposure to a single dose of diethylstilbestrol (DES) produced a significant increase in carcinogenic response of hamster progeny that were subsequently subjected to the carcinogenic stimulus of 7,12-dimethylbenz(a)anthracene (DMBA) during postnatal life. The compounds were administered according to the following schedules. The pregnant animals (second group) received a single dose of DES, 10 mg/kg, on Day 14 of gestation. Postnatally, at 6 weeks of age, the progeny were given DMBA, 25 mg/kg p.o., twice weekly for 8 weeks. The first group received DMBA at 6 weeks of age, 30 mg/kg p.o., twice weekly for 18 weeks. The progeny exposed to DES prenatally and DMBA postnatally (DES-DMBA-exposed progeny) developed a greater multiplicity of tumors per tumor-bearing animal (p less than 0.001) and higher rates of neoplasms of the reproductive tract, e.g., ovarian and uterine tumors, mammary gland and forestomach tumors, and dermal melanomas. The prenatally DES-exposed progeny also had significantly higher incidences of malignant tumors, e.g., carcinomas of the mammary gland (p less than 0.001) and carcinomas of the forestomach (p less than 0.001), than did the hamsters given DMBA alone during postnatal life. Endocrine imbalance produced by exposure in utero may heighten the sensitivity of the progeny to development of neoplasms after a challenge with carcinogenic stimuli in adult life. The significance of these experimental data to the human situation is discussed.


Enhancement of endotoxic shock by N-acetylmuramyl-L-alanyl-(L-seryl)-D-isoglutamine (muramyl dipeptide).

We described elsewhere that the synergistic antitumor activity of endotoxic extracts from Re mutants of gram-negative bacteria and trehalose mycolate against guinea pig syngeneic line 10 tumor was abrogated after peptide substances accompanying these extracts had been removed. This activity could be restored by combining peptide-free endotoxin either with cell wall skeleton from Bacillus Calmette-Guérin, a polymeric mycolic acid-arabinogalactan-mucopeptide complex, or with a combination of two separate components, trehalose dimycolate and N-acetylmuramyl-L-alanyl-(L-seryl)-D-isoglutamine (MDP). We report here that when a combination of endotoxin (150 microgram) and a mixture of MDP (150 microgram) and trehalose dimycolate (150 microgram) was inoculated into established dermal tumors, a significant number of the animals died, presumably of endotoxic shock. All surviving animals suffered severe but temporary lethargy. When administered alone intradermally in the dose levels tested, none of the components caused severe lethargy or lethality. The lethal effects of 159 microgram of MDP also occurred in combination with relatively weak endotoxic products, such as Pseudomonas vaccine (Pseudogen), and these effects did not depend upon the presence of malignant tissue. Guinea pigs inoculated i.v. were even more susceptible inasmuch as the addition of as little as 6 microgram of MDP to 150 microgram of Pseudogen, itself not lethal, caused the death of 80% of the animals.


Quantitative assessment of generalized epithelial changes in tracheal mucosa following exposure to 7,12-dimethylbenz(a)anthracene.

Sequential morphological changes occurring after brief carcinogen exposures of heterotopic tracheal transplants in rats were semiquantitatively studied. Tracheas were exposed to 7,12-dimethylbenz(a)anthracene for 1, 2, or 4 weeks, during which time means of 138, 152, and 160 microgram 7,12-dimethylbenz(a)anthracene, respectively, were delivered. The first two types of exposures resulted only in generalized epithelial changes; these included hyperplasia and early metaplasia, both of which regressed rapidly, and persistent atrophic alterations. No focal epithelial lesions or tumors developed. The third type of exposure (160 microgram 7,12-dimethylbenz(a)anthracene delivered in 4 weeks) resulted in the appearance of generalized mucosal changes with long-lasting, severe inhibition of mucus production. In addition, focal metaplastic lesions reappeared at 4 to 8 months after exposure, and invasive carcinomas developed after 1 year with an incidence of 9%. Overall carcinoma incidence, including carcinoma in situ, was 15%. The studies emphasize the importance of the duration of carcinogen exposure, and they demonstrate the emergence of focal lesions when effective carcinogenic exposures are being used. The possible significance of epithelial atrophy in the pathogenesis of cancer in this experimental model is discussed.


Renal tubular transport of methotrexate in the rhesus monkey and dog.

The mechanism and localization of renal transport of methotrexate (MTX) were studied in the rhesus monkey and the dog. It was found that in both animals MTX was bound with plasma protein in a range of 50 to 68% varying with the MTX plasma concentration. Paper chromatographic analysis showed that a negligible amount of MTX was metabolized. The excretion of MTX in rhesus monkey was mainly by tubular secretion which was blocked by probenecid, but in the dog a bidirectional transport mechanism for MTX was indicated. Tubular secretion was localized in the proximal tubules, and a tubular reabsorptive process was in the distal section. Simultaneous administration of folic acid blocked the tubular reabsorption of MTX, resulting in an increase of renal excretion. Maximum tubular excretory capacity determination showed that a maximum tubular excretory capacity value of approximately 5 mumol/100 ml of glomerular filtrate was observed in the rhesus monkey at a plasma concentration of 0.07 mM and a value of 2 mumol/100 ml of glomerular filtrate for the dog. Studies with renal cortical slice technique also indicated that the monkey kidney can accumulate greater amounts of MTX than can the dog kidney.


Effect of catechol estrogens on rat mammary tumors.

The effect of 1,3,5(10)-estratriene-3, 16 alpha, 17 beta-triol (estriol), 1, 3, 5(10)-estratriene-2,3-diol-17-one (2-hydroxyestrone), and 1,3,5(10)-estratriene-2,3,17 beta-triol (2-hydroxyestradiol) on the growth of dimethylbenz(a)anthracene-induced mammary tumor and of R3230AC-transplantable mammary tumor was compared with that produced by estradiol benzoate treatment. Estriol showed minimal inhibition of tumor growth in dimethylbenz(a)anthracene-induced tumor and no effect on R3230AC tumor while 2-hydroxyestrone showed no effect of tumor inhibition. On the other hand, 2-hydroxyestradiol showed appreciable inhibition of tumor growth in both tumors studied. That 2-hydroxyestradiol has been found to bind to estrogen receptors in mammary tumors and is uterotropic suggests that the inhibition of tumor growth by 2-hydroxyestradiol may be similar to the mechanism of inhibition of mammary tumors by high concentrations of estradiol.


Biochemical basis for cytotoxicity of 7,12-dimethylbenz(a)anthracene in rat liver epithelial cells.

When the effects of 7,12-dimethylbenz(a)anthracene (DMBA) on normal and malignant rat liver epithelial cells were compared in a colony inhibition assay, this carcinogen showed a preferential cytotoxic action on the normal cells. In investigations of the biochemical basis of this selective toxicity, it was found that both cell lines were similarly effective in binding DMBA to DNA and that both cell lines had the capacity to metabolize this carcinogen. However, the hepatoma cells were more efficient than were the normal cells in generating very polar metabolites (not organic solvent extractable). These studies suggest that the basis of the resistance of the hepatoma cells to the toxicity induced by DMBA lies in their ability to detoxify biologically active metabolites. Several phenols were examined as possible toxic metabolites of DMBA, but these were not toxic at dose levels at which DMBA kills most of the normal cells.


Disposition and metabolism of pentamethylmelamine and hexamethylmelamine in rabbits and humans.

The disposition and metabolism of pentamethylmelamine (PMM) and hexamethylmelamine (HMM) were studied in the rabbit, and the disposition of PMM was studied in humans. Parent compound and metabolites were identified by thin-layer chromatography, gas chromatography, and gas chromatography/mass spectrometry analyses. Plasma elimination in both species following i.v. administration of each drug was best described by a two-compartment open model. Both compounds were extensively demethylated with less than 1% of the total dose administered recovered in the urine over 24 hr. The areas under the plasma time-concentration curves of PMM and HMM following p.o. administration to rabbits were 5 and 25% of the areas following i.v. administration. Gastrointestinal absorption was rapid and efficient with 75 to 89% of drug equivalents recoverable in the urine after p.o. administration of [ring-14C]PMM or [ring-14C]HMM to rabbits. Reduced bioavailability of PMM and HMM p.o. appears to be a consequence of rapid metabolism presumably in the liver.


An appraisal of the role of radiation therapy in the management of nonseminomatous germ-cell tumors of the testis in the era of effective chemotherapy.

Patients with nonseminomatous germ-cell tumors of the testis can be divided into two broad groups. The first includes patients with negative lymphograms or small-volume metastases confined to the abdominal nodes. The overall cure rate with orchiectomy and nodal irradiation is 80%. A policy of early detection of relapse and treatment with chemotherapy is advocated. Adopting this approach, no deaths have occurred in this group of patients treated in 1976 and 1977, and only one (associated with acute myeloblastic leukemia) occurred in 1975. The second group consists of patients with other stage categories who receive chemotherapy as initial treatment, followed in stage II and II and selected stage IV patients by radiation therapy to sites of initial involvement and surgery. Preliminary experience has shown this to be a practicable and promising approach. The prognosis for stage IV patients depends upon metastatic site and volume; in those patients with limited lung disease 80% are surviving disease-free.


Somatostatinergic nerves in the cervical spinal cord of the monkey.

Somatostatinergic nerves in the spinal cord of the monkey were investigated utilizing immunohistochemistry with various antibodies against synthetic somatostatin. In contrast to earlier investigations, it is shown that somatostatinergic nerve endings occur in most of the areas of the grey matter of the spinal cord. The somatostatinergic axons are, however, characteristically distributed in three main regions: (1) Densely-packed endings are seen in lamina II of the substantia gelatinosa, forming a crescent-shaped pattern in the columna dorsalis. Somatostatin immunoreactivity is also seen in lamina I and in the Lissauer tract. (2) A fine network of fibers is observed around the central canal; the endings are concentrated on special cell bodies. Some single perikarya are also stained in this region. (3) A loose network of single fibers is found ending on perikarya of the columna lateralis or ventralis. The perikarya of the nerve axons, with the exception of those terminating in the columna dorsalis, have as yet not been identified. In order to better understand the somatostatinergic system of the spinal cord, these newly-detected somatostatinergic nerves must be studied and their exact pathways analyzed.


The nephrotoxicity of p-aminophenol. I. The effect on microsomal cytochromes, glutathione and covalent binding in kidney and liver.

p-Aminophenol administration lowered the microsomal cytochrome P-450 and b5 content and decreased the activity of NADPH cytochrome c reductase in kidney, but not in liver. Kidney GSH was depleted to 29% of the control value at 2 h, and only partly restored (50% of control) at 24 h. Liver GSH was transiently decreased, the lowest levels (77% of control) occurring at 30 min. The maximum level of covalently bound radioactivity was at two hours when 16.8% of the total radioactivity in kidney, 1.5% in liver and 3.6% in plasma was protein bound. At this time 81% of the total radioactivity in kidney and 95% of that in the liver was present in the soluble fraction.


Effects of membrane perturbants on UDP-glucuronosyltransferase activity in rat-liver microsomes. Circular dichroism studies.

Rat-liver microsomes were treated with two non-ionic detergents, Triton X-100 and Lubrol WX, with phospholipase A2, or with aqueous acetone solution. The activity of the membrane-bound UDP-glucoronosyltransferase (UDPGT, EC 2.4.1.17) was measured after the treatment with these perturbants. At the same time, modifications of the secondary structure of the microsomal proteins were followed and studied by circular dichroism (CD) spectroscopy. The detergents greatly activated UDPGT, maximally at a 1 mM concentration of either detergent. The maximally activating Triton X-100 treatment did not greatly change the ellipticity of the microsomes at 222 nm ((theta)222), whereas that with Lubrol WX affected the secondary structure of the membrane proteins more strongly. UDPGT activation also occurred in phospholipase A2-treated microsomes. Maximal activation was obtained after 1--5 min of incubation and was stable throughout the experiment. Phospholipase A2 at the ratio of microsomal protein to phospholipase 250 : 1 (w/w) slightly increased (theta)222 after 10 min of incubation and did not change it further even after 30 min of incubation. Treatment of liver microsomes with a 10 : 90 (v/v) aqueous acetone solution removed 90% of the total membrane phospholipids, particularly phosphatidylcholine and phosphatidylethanolamine. The UDPGT activity was decreased in lipid-depleted microsomes, and the enzyme was not reactivated when phosphatidylcholine-lysophosphatidylcholine liposomes were added at a low temperature. An even greater decrease was obtained when the lipid binding was carried out at 37 degree C. Lipid-depleted microsomes had a high (theta)222 associated with a red-shift of 2 nm, indicating partial aggregation of membrane proteins and an increase in the alpha-helical content of the protein after acetone extraction. However, this particular protein structure was partially reversible, since a binding of phospholipids to lipid-depleted microsomes gave a (theta)222 close to that found in control microsomes. The UDPGT activity was not dependent on the secondary structure of the membrane proteins.


Nucleic acid base and carcinogen metabolite specificities during intercalative interactions between DNA and 4-nitroquinoline 1-oxide.

Intercalation of the carcinogen 4-nitroquinoline 1-oxide (4-NQO) or its metabolic intermediate forms, probably precedes the covalent bond formation of the ultimate carcinogenic form with DNA. A 'complete' empirical-potential energy description of the base-sequence and metabolite specificities of the 4-NQO intercalation process is presented in this work. The important force and structural interaction components are depicted via decomposed energy functions. Energy-minimized intercalated complexes are presented and indicate several interesting characteristics. It is clear that the various intercalated quinoline-metabolites do not generally enter into 'strictly' parallel-planar stacked orientations (unlike the structurally rigid ethidium-intercalated complexes). Intercalation is energetically permitted for six of seven quinoline-metabolites (QMS) studied, although, intercalation into to Pyr(3'-5')Pur sequences is preferred over Pur(3'-5'1Pyr sequences. The three quinoline-metabolites that are more energetically favoured to undergo intercalation than the parent form are also known to enter into the greatest amount of covalent interactions with DNA and its constituents. Thus the present work further suggests the existence of a two-step binding mechanism: intercalation followed by covalent reaction.


Characteristics of aryl hydrocarbon hydroxylase activity in rat mammary epithlial cells grown in primary culture.

Rat mammary epithelial cells grown in primary culture contain the microsomal enzyme, aryl hydrocarbon (benzo[a]pyrene) hydroxylase (AHH), which catalyses the oxidative conversion of polycyclic aromatic hydrocarbons (PAH) to more polar derivatives. Constitutive AHH activity, measured with an established fluorometric method, was 46 pmol/mg protein/h in homogenates of rat mammary epithelial cells after 5 days in culture. The addition of dimethylbenz[a]anthracene (DMBA), benz[a]anthracene (BA), or 3-methylcholanthrene (3-MC) to the cell culture medium increased AHH activity 5.3-, 4.7- and 2.4-fold, respectively. Kinetic studies revealed that maximal hydroxylase induction occurred 16 h after 1 micro M DMBA was added to the culture medium. The decay of the DMBA-induced hydroxylase was biphasic: one component had a t1/2 of 15--30 min and another a t1/2 of 4 h. Norepinephrine, 17 beta-estradiol and 5,6-benzoflavone also increased AHH activity in mammary epithelial cells in vitro, however, sodium phenobarbital had no effect. Fetal bovine serum (FBS), previously shown to be a potent in vitro inducer of AHH activity, had no effect on either constitutive or DMBA-induced mammary epithelial hydroxylase activities following treatment with 1% activated charcoal. Metyrapone and 7,8-benzoflavone, inhibitors of microsomal mixed function oxidase activity, reduced both constitutive and DMBA-induced AHH activities when added to homogenates of untreated and DMBA-treated mammary epithelial cells. The addition of 7,8-benzoflavone reduced both constitutive and DMBA-induced hydroxylase activities by approx. 80%, whereas metyrapone addition inhibited these activities by 20%. The study demonstrates several in vitro factors which alter AHH activity in primary cultures of rat mammary epithelial cells.


The formation of dihydrodiols in the chemical or enzymic oxidation of dibenz[a,c]anthracene, dibenz[a,h]-anthracene and chrysene.

The formation of trans-dihydrodiols from dibenz[a,c]anthracene, dibenz[a,h]anthracene and chrysene by chemical oxidation in an ascorbic acid-ferrous sulphate-EDTA system and by rat-liver microsomal fractions has been studied using a combination of thin-layer (TLC) and high pressure liquid chromatography (HPLC) to separate the mixtures of isomeric dihydrodiols. The 1,2- and 3,4-dihydrodiols of dibenz[a,c]anthracene, the 1,2-,3,4- and 5,6-dihydrodiols of dibenz[a,h]anthracene and the 1,2-, 3,4- and 5,6-dihydrodiols of chrysene were formed in chemical oxidations. These dihydrodiols were also formed when the three parent hydrocarbons were metabolized by rat-liver microsomal fractions and, in addition, dibenz[a,c]anthracene yielded the 10,11-dihydrodiol. The 1,2- and 3,4-dihydrodiols of dibenz[a,c]anthracene have not been reported previously either as metabolites of the hydrocarbon or as products of chemical syntheses and the 5,6-dihydrodiol of chrysene was not detected in earlier metabolic studies.


Regulation of the binding of 7,12-dimethylbenz[a]-anthracene to DNA of cultured murine epidermal cells at metabolic level: competition of the binding by polycyclic aromatic hydrocarbons and by other precarcinogens.

The binding of labeled carcinogen [3H]DMBA to murine epidermal cells (MEC) DNA in culture has been studied. The influence of unlabeled noncarcinogenic and carcinogenic polycyclic aromatic hydrocarbons (PAH), several PAH metablites, and various directly and indirectly acting non-PAH carcinogens on the binding of [3H]DMBA to MEC DNA has been examined. All the carcinogenic PAH and some of non-carcinogenic PAH effectively inhibit the binding of [3H]DMBA to MEC DNA. The non-PAH chemical carcinogens requiring metabolic activation also reduce the binding of labeled DMBA to MEC DNA; however, a higher concentration of these compounds is required for 50% inhibition of binding than the concentrations of PAH for the same degree of inhibition of binding of [3H]DMBA to MEC DNA. The directly acting carcinogens do not significantly inhibit the binding of [3H]DMBA to DNA. The relationship between structures of PAH and their ability to inhibit the binding of [3H]DMBA to MEC DNA is also discussed. Thus, it appears that the binding of DMBA to cellular DNA is primarily controlled at a level of metabolism and to some extent at the level of binding of reactive metabolites to DNA.


[Role of vitamin B 12 in the incorporation of ribonucleosides into the DNA of various Periplaneta americana cell lines].

Hemocytes and non hemocytes in vitro cell lines from Periplaneta americana, have been tested for their ability to incorporate, in the presence of vitamin B 12, various nucleic acid precursors into their DNA. Evidence was shown for enhanced transformation of all ribonucleotides into deoxyribonucleotides in the presence of vitamin B 12. Moreover, in hemocyte cell lines, it was demonstrated that vitamin B 12 produced an activation of thymidine kinase activity.


[Plasma estrogens in Pleurodeles waltlii. Comparison between diploid and triploid females].

Estrogens have been examined in the plasma of diploid and triploid newts Pleurodeles waltlii. Estradiol-17 beta (E2) and estrone (E1) were determined by radioimmunoassay, before and after enzymatic hydrolysis of the conjugates. Total (t) and unconjugated (u) E2 levels were positively correlated (E2u = 0.478 35 E2t + 0.579 98; r = 0.883), but no correlation was detected between E1 levels. No statistical difference was found for the estrogen levels between the different experimental lots of diploid newts (E2t = 7.5 +/- 0.37 ng/ml, E2u = 4.3 +/- 0.20 ng/ml, E1t = 2.19 +/- 0.08 ng/ml, E1t = 0.41 +/- 0.02 ng/ml) but every estrogen level was lower in the triploid group (E2t = 1.8 +/- 0.60, E2u = 1.0 +/- 0.18, E1t = 1.4 +/- 0.13, E1u = 0.3 +/- 0.04 ng/ml). This difference is discussed in relation to lower fertility of the triploid females.


[2 new esterase loci Es-14 and Es-15 in mice (genus Muc L.) : characterization by different substrates and inhibitors].

Allelic differences at an esterase locus designated Es-14 exist between Mus musculus (both inbred strains and wild Mice) and Mus spretus. This locus is expressed in a variety of tissues. Mus musculus has been assigned the allele Es-14r while Mus spretus carries the alternate alleles Es-14l. Each phenotype manifests itself as a single band of enzyme activity and F1 hybrids show two bands corresponding to the parental forms. Allozyme variation was also detected at another esterase locus, temporarily disignated as Es-15 until confirmed by current linkage analysis. A slow variant Es-15 was recorded in a Mus musculus population from Greece while inbred strains carry the Es-15m allele. Study of a Mus spicilegus population from Greece revealed the presence of a third allele, Es-15r. Substrate and inhibition characteristics are provided for both loci.


[Inhibition of 9-amino-2-methoxy-6-chloroacridine fluorescence by human serum albumin].

Study of the fluorescence quenching of 9-amino 2-methoxy 6-chloro acridine upon serum albumine additions reveals a single protein fixation site. Characteristic values of thermodynamic functions are obtained from experiments at different temperatures.


[Factor VIII-related antigen (F VIII R : Ag). Purification by immuno-affinity chromatography].

Immuno-affinity chromatography on immunoglobulins from a rabbit immunised with the plasma of a patient devoid of factor VII R : Ag allows the elimination of certain contaminants from preparations already enriched in factor VIII. The biological activity and criteria of purity of the factor VIII R : Ag preparation obtained are presented.


[Dissolution of the pulmonary collagen of the normal rat by repeated pepsin action].

A non-denaturating method of extracting collagen from Rat lungs was developed. It consists in dissolving the collagen by repeated action of pepsin. Depending on the age of the animal, efficiency values between 50 and 90% are obtained.


[Vitamin A consumption in pregnant and non pregnant rats, and in castrated rats receiving or not receiving estradiol or pregnenolone; relation to tocopherol and cytochrome P-450].

Consumption of vitamin A and cytochrome P 450 by pregnant Rat is more important than in non pregnant and ovariectomised Rat. Estradiol implant in ovariectomised female has some, but slighter, influence only on retinol. These physiological situations have no action on the hepatic levels of tocopherols (total and alpha). These results are discussed.


[In vivo and in vitro study of the effect of pregnant female serum graft rejection in Salamandra salamandra Lin].

Serum from pregnant female Salamandra salamandra inhibits the cytotoxic reaction from the mother towards its larvae. Such a serum accelerates the allograft rejection reaction. In vitro studies show that a serum from pregnant female inhibits the cytotoxic reaction of host spleen cells towards epithelial cells of the donor of the graft.


Exercise cross-sectional echocardiography in ischemic heart disease.

We performed cross-sectional echocardiograms at rest, during supine bicycle exercise, and after sublingual nitroglycerin administration in 28 patients suspected of having ischemic heart disease. Technically adequate exercise cross-sectional echocardiograms were obtained in 20 patients (71%). Ten patients had new areas of reversible segmental dysynergy, and all 10 had significant stenoses of coronary arteries supplying areas of the heart corresponding to the location of reversible dysynergy. Six of these 10 patients also underwent exercise thallium-201 perfusion scanning, and all six had reversible perfusion defects in the area that demonstrated reversible dysynergy on exercise cross-sectional echocardiography. At least two of the remaining 10 patients who did not have reversible segmental dysynergy on exercise cross-sectional echocardiography probably experienced myocardial ischemia that we did not detect. We conclude that exercise cross-sectional echocardiography is technically difficult but feasible. The mechanical consequences of exercise-induced regional myocardial ischemia can be detected noninvasively by real-time, two-dimensional, cross-sectional echocardiography.


Long-term effects of physical training on coronary patients with impaired ventricular function.

Eighteen patients with coronary heart disease and an ejection fraction of 0.40 or less were entered into an individualized exercise training program. Maximal symptom-limited exercise stress test and cardiac catheterization studies were performed initially and 12--42 months (average 18.5 months) after exercise training. At the time of the follow-up study, the mean functional aerobic impairment (FAI) improved from 32.1 to 23.4% (p less than or equal to 0.01); resting and submaximal heart rates were significantly lower (p less than 0.01 and 0.05, respectively). There was no significant change in the pulmonary artery or left ventricular end-diastolic pressure, cardiac index, stroke index, left ventricular end-diastolic volume or ejection fraction. Exercise training, therefore, can be beneficial even for patients with impaired ventricular function. Increase in physical work capacity was not correlated with improvement of ventricular function; on the other hand, exercise training did not cause deterioration of ventricular function.


Epithelial or dandruff allergen for the diagnosis of dog allergy by RAST?

When used for screening by the RAST technique, dog epithelium provided too few positive results (29.2%) compared with dog dandruff (58.8%), cat epithelium (53.6%) and horse dandruff (52.2%). However, dog dandruff can provide an unexpectedly high incidence of positive RAST results (45.2%) in sera from patients with an allergic history to cat or horse, but not to dog. Dog dandruff allergens were found to cross-react with some cat or horse serum proteins in about 25% of such patients. We conclude that dog dandruff is preferable to epithelium for screening by the RAST technique, but if emotional issues involving a family pet arise then confirmation by a trial separation is likely to be more convincing than a provocation test with a possibly non-specific allergenic extract.


Blood histamine levels after exercise testing.

Histamine has been measured in whole blood and in the plasma of twenty-three subjects following exercise testing of seventeen asthmatics and six normal controls. Whole blood histamine rose in both asthmatics and controls and was associated with a concomitant rise in the basophil count. Plasma histamine in the peripheral venous blood only rose in one patient who developed urticaria after exercise.


IgE antibodies against penicillin as determined by Phadebas RAST.

Serum samples from eighty-one patients with suspected penicillin allergy were investigated with Phadebas RAST using the penicillin derivatives Benzylpenicilloyl-human serum albumin (PBO-HSA) and Phenoxymethylpenicilloyl-human serum albumin (PMPO-HSA) and the results were compared with skin test results and clinical data. Of the sixty-one patients who had anaphylactic shock and/or urticaria as a possible consequence of penicillin administration, reagins against PBO-HAS and PMPO-HSA could be detected in thirty-four cases (56%). Five per cent of these patients, with positive RAST results, showed negative skin tests; in the other 95% both RAST and skin tests were positive. All, except eight, of the RAST-negative patients had had their adverse reactions at least 2 years prior to the blood sampling and in some of these cases skin tests were also negative. RAST and provocation test results agreed in 80% of the cases where exposition was performed. It is concluded that the RAST technique is a valuable diagnostic tool for the detection of immediate type hypersensitivity to penicillin.


Serum creatine kinase BB linked to immunoglobulin G.

We describe an atypical form of creatine kinase in the serum of a woman after myocardial infarction. Electrophoresis on agarose gel showed a single fraction between CK-MM and CK-MB isoenzymes. After ion-exchange chromatography on DEAE-Sephadex A-50 we isolated the atypical CK and CK-BB as detected by electrophoresis of the eluted fractions. Results of our immunological investigations and exclusion chromatography clearly demonstrate that the atypical CK consists of complexes with high molecular masses formed by CK-BB isoenzyme and immunoglobulin G. The clinical significance, if any, of this Macro CK has yet to be determined.


A numerical comparison of the use of T3-uptake values and of TBG levels for the estimation of free thyroxine in serum.

The correlation of the diagnostic indices FTI and T4/TBG with actual free T4 levels (FT4) was investigated by numerical analysis. It was shown that, FT4 being kept constant, both indices vary with altered TBG concentrations. Thus neither index is truly proportional to FT4. In this respect both indices appear doubtful tools for diagnosing patients with abnormal TBG levels. A "map" constructed by plotting T4 versus T3U appeared to offer a higher discriminatory potential with respect to pathologic FT4 values. Such an idealized map was shown to consist of hypo-, eu- and hyperthyroid domains separated by lines governed by the limit values of the FT4 normal range. In practice, of course, these domains are separated by border regions rather than border lines and they should be derived from clinical data.


pH-independent inhibition of plasma angiotensin I degradation: implications for renin assay.

Valid enzymatic assay of plasma renin (EC 3.4.99.19) requires complete inhibition of enzymes that destroy angiotensin I. Historically, the degree of such inhibition has varied with assay pH and with the combination of protease inactivators employed. To circumvent this problem, we studied the effects of several protease inactivators on both the formation and degradation of angiotensin I by human plasma at pH 5.5 and at pH 7.4. While several reagent combinations effectively suppressed the rate of angiotensin I disappearance at one of the two pH values, three combinations of phenylmethylsulfonyl fluoride with chelating agents accomplished this at both pH values. Of these, however, only the combination of 5 mmol/l ethylenediamine tetraacetate, 1.5 mmol/l 8-hydroxyquinoline, and 7.5 mmol/l phenylmethylsulfonyl fluoride also maximized the rate of angiotensin I formation at both pH 5.5 and 7.4. Thus, the use of the latter reagent combination would facilitate comparison of renin assays performed at different pH values and would permit standardization of the protease inactivators employed in such assays.


Longitudinal study of circulating immune complexes in a patient with Staphylococcus albus-induced shunt nephritis.

The direct measurement and partial characterization of circulating immune complexes has been performed in a longitudinal study of a patient with Staphylococcus albus-induced shunt nephritis. The high levels of immune complexes were associated with cryoglobulinaemia and hypocomplementaemia. The activation of complement was found to be via the classical pathway, but the functioning of the alternative pathway may have been impaired in vivo due to very low levels of C3. The host response to the infection was also characterized by the production of a marked macroglobulinaemia, high titres of rheumatoid factor and a typical acute phase increase in the C-reactive protein level. Immune complex levels were persistently elevated many months after the removal of the focus of the infection. A possible explanation for this surprising finding may lie in the nature of the antigens in the immune complexes. It was found that the immune complexes contained both antibodies to and antigens from Staphlococcus albus. In particular, glycerol teichoic acid and staphylococcal nuclease were identified as components of the immune complexes present during the acute phase. Glycerol teichoic acid was also identified in the immune complexes found later although other Staphylococcus albus antigens as yet unidentified were also present and persisted in the circulation for several months.


An IgM monoclonal protein with multiple serological specificities.

An IgM lambda M-component with a false Wasserman reaction and 'false' rheumatoid factor activity from a patient suffering from a well-differentiated lymphocytic lymphoma is presented. The monoclonal protein showed antibody activity against cardiolipin and cross-reacted with acryl particles, not with human IgG. Both these activities were found in the Fab fragment. The reduction of the 19S IgM to 7S IgM subunits was responsible for a strong decrease in activity. The importance of the IgM quaternary structure in determining antibody affinity is emphasized.


Celluar immunoglobulins in human gamma- and alpha-heavy chain diseases.

Proliferating cells from twenty-four patients with alpha- or gamma-heavy chain disease (HCD) were studied by direct immunofluorescence and in several cases by biosynthesis experiments with 14C-amino acid incorporation. In twenty-two patients, the cells contained the HCD proteins only and no light chain synthesis could be detected. Conversely, apparently non-secreted monotypic light chains were found in one case of gamma-HCD and one case of alpha-HCD. The proportion of proliferating cells containing cytoplasmic heavy chains, their appearance and the presence or not of surface heavy chains showed great variation from patient to patient. In some cases, the proliferation predominantly affected either plasma cells or lymphocytes whereas in others the disease seemed to correspond to a proliferation of HCD protein-bearing lymphocytes with persistent maturation into plasma cells. Large cell lymphomas supervening on alpha-HCD belonged to the same proliferating clone as the clone secreting the HCD protein, as shown by surface markers and biosynthesis experiments which demonstrated synthesis but no secretion of HCD proteins. In one patient with gamma-HCD, the cells carried surface gamma and delta chains.


Metabolism of carbaryl by kidney, liver, and lung from human postembryonic fetal autopsy tissue.

Metabolic profiles of carbaryl in human postembryonic fetal autopsy tissue were determined using an in vitro tissue-maintenance technique. 1-Naphthyl-14C or N-methyl-14C-carbaryl was applied to growth medium containing explants of the tissue. Each mixture was incubated for 18 hr and the medium analyzed by DEAE-cellulose column chromatography. Fetal liver performed the metabolic processes of demethylation, hydrolysis, hydroxylation, and oxidation, followed by conjugation, as was found with adult liver. However, the anionics from fetal liver amounts to 20% of those found with adult liver. The kidney made naphthyl glucuronide and naphthyl sulfate, whereas the lung produced naphthyl sulfate from carbaryl. The metabolic activities of the fetal kidney and lung were close to the corresponding human adult tissues based upon the anionic metabolites found and the amount of unmetabolized carbaryl in the medium after 18 hr of incubation. Silica gel chromatography of ether-extractable neutral fractions from DEAE-cellulose revealed 3,4, and 9 ether-extractable metabolites from lung, kidney, and liver, respectively. The present study shows that the in vitro technique is capable of semiquantitatively demonstrating the metabolic activities of specific organs from the human fetus.