Research and demonstration issues in self-care: measuring the decline of medicocentrism. Emergence of consumer health self-care is a reflection of the increased commitment of health professionals to patient education, growing consumer awareness that they are capable of sophisticated self-help, and a variety of social, economic and technological currents. These currents are reviewed and a survey of existing medical self-care programs is summarized. The attempts and potentials to evaluate these programs are critically examined. A number of important research and demonstration issues are raised including the determination of behavioral outcomes, technical limits, and manpower implications. A federal program of replicative studies on such issues would provide substantive knowledge in the self-care field, generalizable to the larger field of health education, but the hazards of undermining the voluntaristic and non-establishment character of the programs must be considered in designing evaluative studies.


The pattern of loci on Drosophila chromosomes. In this study, we have attempted to detect and describe patterns in the arrangement of gene loci within chromosomes of a metazoon. Known loci (842 of them) on the first three chromosomes of Drosophila melanogaster have been characterised according to a list of 45 properties and the distributions of these properties have been examined systematically. For all these properties, there is little, if any, evidence of clustering between known loci that are not close enough to belong to the same 10-locus group. Our analysis has, however, revealed evidence confirming that some properties show a tendency to cluster within a 10-locus group. Even this tendency is not strong, except for some of the morphological properties. However, it is apparently at variance with the findings of Elston and Glassman (1967), based on fewer data.


Selection acting upon slow-migrating ADH alleles differing in enzyme activity. Relative egg to adult viability was compared in three stocks of Drosophila melanogaster differing in alcohol dehydrogenase (ADH) activity. Two of these, Grell's standard ADHS and a mutant ADHS strain producing half the normal number of ADH molecules, had the same electrophoretic mobility. The experiment demonstrated a correlation between survival in alcohol-supplemented media and enzyme activity. This supports the hypothesis that enzyme activity per se, rather than some other attribute of allelic differences, may be responsible for the previously observed selective advantage of high activity alleles on alcohol media.


Polymorphism at two loci through selection for linear metric deviation. A model of phenotypic stabilising selection in which the fitness of an individual depends solely on its phenotype, and not directly on its genetic constitution, is explored algebraically for a system of two linked loci of unequal effect. It is found that selection for metric deviation gives rise to polymorphic gametefrequency equilibria for a variety of  fitness regimes. Stability of non-trivial equilibria occurs for a wide range of parameter sets. Stability is facilitated by close linkage and inequality between gene effects. It is suggested that, in general genetic variation may be maintained under stabilising selection when the fitness of double heterozygotes exceeds that of the phenotypically intermediate homozygotes.


Genetical composition, temperature, density and selection in an enzyme polymorphism. Selection in egg-to-adult viability was investigated at the Esterase-6 locus of Drosophila melanogaster. A factorial experiment was carried out with three variables, temperature, density and genetical composition; there were four temperatures (15 degrees, 20 degrees, 25 degrees and 30 degrees), three densities (115 ml, 10 ml and 2-5 ml of food). Strong frequency dependent selection was observed in many environments. The strength of the selection was very much modified by an interaction between temperature and density. The results are discussed in relation to evidence for a chemical mechanism which mediates frequency dependent selection, by conditioning, in the larval environment.


Esophageal patch grafting as a treatment for esophageal stricture in a horse. Esophageal patch grafting, using the sternocephalicus muscle, was used for treatment of cervical esophageal stricture in a mare. The mare was maintained during the initial healing phase by total parenteral nutrition.


Effects of serum lipid content on the binding of minocycline. Minocycline was added to normal and hyperlipemic serum samples in concentrations of 1 approximately 10 mcg/ml. These specimens had similar protein contents. Chemically extractable minocycline was quantitated fluorometrically. Hyperlipemic serum (cholesterol 480 mg/100 ml; triglycerides 321 mg/100 ml) yielded an average of 50% less minocycline than did normal serum (cholesterol 170 mg/100 ml; triglycerides 114 mg/100 ml). When ultrafiltrates of serum containing 6, 12 and 20 mcg/ml minocycline were assayed microbiologically, it was evident that variations in serum triglyceride and cholesterol levels did not alter the ratio of bound to free drug. Minocycline appears to be reversibly associated with, and/or soluble in, triglyceride-cholesterol components of serum.


The treatment of enteric fever with mecillinam. In vitro studies carried out in this Department confirmed the high activity of mecillinam against Salmonella spp. As a result we set up a study to investigate the efficacy of mecillinam in patients suffering from enteric fever. To date 13 out of 15 patients with typhoid or paratyphoid fever have been successfully treated with this new amidinopenicillanic acid antibiotic.


In vitro studies with mecillinam on Escherichia coli and Pseudomonas aeruginosa. The ability of a beta-lactam antibiotic to inhibit the growth of Gram-negative bacteria depends on three main properties: ting sites in the bacterial cell; the ability to penetrate through the outer layers of the bacterial envelope to these sites; and the ability to resist destruction by beta-lactamases that may be encountered on the way to the target. This article describes the properties of mecillinam with respect to the last two of these properties. Although able to hydrolyse mecillinam under some conditions, the beta-lactamases present in many Gram-negative species are unlikely to be very effective at protecting the bacteria in vivo because of their relatively low affinity for this penicillin and the good penetrative properties of the antibiotic.


Inhibition and facilitation of transfer among Pseudomonas aeruginos R plasmids. Esamining 12 plasmids in Pseudomonas aeruginosa, we found two types of interaction in their transfer (inhibition and facilitation), using donor cells carrying two compatible plasmids. (i) Ten plasmids representing incompatibility groups P-1, P-2, P-5, P-6, and P-7 were all transmissible at a high frequency, 10-2 to 10-1, except for one with a lower frequency of about 10-3. The transfer of P-5 plasmids was inhibited by P-2 plasmids reciprocally or unilaterally, and the unilateral transfer inhibition was observed in other combinations between plasmids belonging to groups P-1, P-2, P-6, and P-7. It was characteristic of Pseudomonas plasmids that most plasmids with high transferability inhibited the transfer of other coexisting plasmids without distinct inhibition of their own transfer. (ii) Two plasmids, Rms149 of P-8 group and Rlb679, which was not classified, were transmissible at an exceptionally low frequency of 10-7 to 10-6, but their transfer was facilitated by plasmids with high transferability.


Myxospore coat synthesis in Myxococcus xanthus: in vivo incorporation of acetate and glycine. Myxospore coat synthesis in Myxococcus xanthus was studied by incorporation of [(14)C]acetate into intermediates in the biosynthesis of coat polysaccharide and into acid-insoluble material during vegetative growth and after glycerol induction of myxospores. During short labeling periods at 27 degrees C, the radioactivity was shown to be located primarily in N-acetyl groups rather than sugar moieties. Two hours after glycerol induction, the pools of N-acetylglucosamine 6-phosphate and uridine 5'-diphosphate-N-acetylgalactosamine (UDPGalNAc) plus uridine 5'-diphosphate-N-glucosamine increased about twofold and were labeled at twice the rate measured for vegetative cells. The increased rate of synthesis of UDPGalNAc and its precursors could be correlated with increased enzyme activities measured in vitro. Controlled acid hydrolysis revealed that the galactosamine portion of the myxospore coat was N-acetylated. After glycerol induction, the incorporation of acetate into acid-insoluble material increased threefold. This enhanced incorporation was sensitive to neither penicillin nor d-cycloserine. In contrast, bacitracin inhibited the incorporation of [(14)C]acetate into acid-insoluble material more effectively 2 h after myxospore induction than during vegetative growth. Chloramphenicol added to cells 90 min after induction blocked further increase in the rate of [(14)C]acetate incorporation. Since the myxospore coat contains glycine, polymer synthesis was also measured by chloramphenicol-insensitive [(14)C]glycine incorporation into acid-insoluble material. Although protein synthesis decreased after glycerol induction, glycine incorporation increased. Two hours after induction, glycine incorporation was only 75% inhibited by chloramphenicol and rifampin. The chloramphenicol-insensitive rate of incorporation of [(14)C]glycine increased during the first hour after myxospore induction and reached a peak rate after 2 to 3 h. The chloramphenicol-resistant incorporation of [(14)C]glycine was resistant to penicillin but sensitive to bacitracin.


Modulation of deoxyribonucleic acid polymerase III level during the life cycle of Bacillus subtilis. Deoxyribonucleic acid (DNA) polymerase III is not detectable in Bacillus subtilis spores; the enzyme activity appears 20 to 30 min after spore activation and rapidly increases just before the onset of the first round of DNA replication (30 min later); the level of polymerase III further increases and reaches its maximum (on a per-genome basis) when the cells enter the vegetative phase of growth; this level is six- to eightfold higher than the one observed during germination. In the stationary phase, the polymerase III drops to levels comparable to those found in germinating spores at the first round of replication. On the contrary, DNA polymerase I is present at appreciable levels in the dormant spore; it increases during vegetative growth by a factor of three and, during the stationary phase, reaches its maximum level which is sixfold higher than that observed in the spores. The block of protein synthesis during vegetative growth does not cause an appreciable reduction of the two enzymes (in absolute terms), showing that the regulation of their levels is probably not due to a balance between synthesis and breakdown. These results indicate that polymerase III is probably one of the factors controlling the initiation of DNA synthesis during spore germination.


Specific alteration of the 30S ribosomal subunits of Bacillus subtilis during sporulation. Active 30S ribosomal subunits were isolated from vegetative and sporulating cells of Bacillus subtilis. Both subunits were able to function in polyuridylic acid of phage phie messenger ribonucleic acid-dependent protein synthesis in vitro. The sporulation 30S subunits were highly active in polyuridylic acid-dependent polyphenylalanine synthesis but showed a reduced activity in the presence of natural messenger ribonucleic acid as compared with their vegetative counter-parts. The reduced activity was independent of the source of 50S particles and initiation factors (vegetative or sporulation). The alteration of the 30S sporulation subunits appears to be related to the sporulation process, since the same subunits isolated from stationary-phase cells of an asporogenic mutant did not show any impairment in protein synthesis in vitro.


Hydrolysis of phenyl beta-maltoside catalyzed by saccharifying alpha-amylase from Bacillus subtilis. 1. The hydrolytic reaction of phenyl beta-maltoside catalyzed by saccharifying alpha-amylase [EC 3.2.1.1] of Bacillus subtilis was studied at 25 degrees C and pH 5.4, by measuring the total reducing power and the amount of phenol liberated, and by thin layer chromatography. 2. The enzyme hydrolyzed phenyl beta-maltoside at the glucosidic linkage between the two glucose residues to form D-glucose and phenyl beta-D-glucoside. Besides these products, maltose, maltotriose, and phenyl beta-maltotrioside were also observed as reaction products. The identification of phenyl beta-maltotrioside is described in detail. The formation of these products was attributed to the transglycosylation reaction of the enzyme. The time course of reaction as followed by reducing power measurement showed an induction period of several minutes.


Coordination chemical studies on metalloenzymes. II. Kinetic behavior of various types of chelating agents towards bovine carbonic anhydrase. In order to investigate the kinetics and mechanism of the removal of zinc ions from bovine carbonic anhydrase [EC 4.2.1.1] (BCA), several chelating agents with various stability constants were used to remove zinc from BCA. The second-order rate constants (kaap) of zinc removal from BCA were found to be in the following order; 2,6-pyridinedicarboxylic acid greater than 2-pyridinecarboxylic acid greater than 2,4-pyridinedicarboxylic acid greater than 2,3-pyridinedicarboxylic acid greater than or approximately 1,10-phenanthroline greater than or approximately 5-methyl-1,10-phenanthroline greater than 2,2'-bipyridine. With similar chelating agents the greater the stability constant, the faster was the rate of removal of zinc ions from BCA. With EDTA, trans-1,2-cyclohexanediaminetetraacetic acid, and nitrilotriacetic acid, the rate of zinc ion removal from the native enzyme was governed by the rate of spontaneous dissociation of zinc enzyme. The rate constants for the removal of zinc ions from BCA were governed by the affinity of the chelating agents for the metal ion and the conformation of the chelating agents. Based on these findings, reaction pathways for various chelating agents are proposed.


Purification and properties of two acid phosphatases from midgut glands of abalone Haliotis discus. Midgut glands of abalone Haliotis discus contained two acid phosphatases [orthophosphoric-monoester phosphohydrolase (acid optimum), EC 3.1.3.2] separable by phosphocellulose column chromatography. They were designated as acid phosphatases I and II in order of elution and were purified 99- and 290-fold, respectively. Purified acid phosphatase II was nearly homogeneous as judged by polyacrylamide gel electrophoresis. The substrate specificity of acid phosphatase I was narrow, whereas that of acid phosphatase II was broad. Good substrates for acid phosphatase I included p-nitrophenyl phosphate, phosphoenolpyruvate, inorganic pyrophosphate, and nucleoside di- and triphosphates. The acid phosphatases did not require any metal ion for  maximum activity and were inhibited by Zn2+, Cu2+ and Hg2+. Fluoride and arsenate were potent inhibitors of both enzymes. The pH optima of acid phosphatases I and II were 5.9 and 5.5, respectively. The molecular weights of acid phosphatases I and II were estimated to be 28,000 and 100,000, respectively, by gel filtration on Sephadex G-100. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis suggested that acid phosphatase II consists of two identical subunits.


On the heterogeneity and organ specificity of chicken tropomyosins. 1. Tropomyosins from chicken cardiac, skeletal, and gizzard muscles were each resolved into two subunits by polyacrylamide gel electrophoresis in a system containing sodium dodecylsulfate (SDS), urea and sodium borate, and were designated C1 C2, S1 S2, and G1 G2, respectively, in descending order of mobility on electrophoresis. S1, S2, G1, and G2 were prepared as pure samples by electrophoresis. 2. The apparent molecular weights of C (C1 + C2), S1, S2, G1, and G2 were calculated to be 36,000, 36,000, 37,500, 36,000, and 40,000, respectively, based on SDS gel electrophoretic mobility according to the method of Weber and Osborn. C and S1 showed nearly the same mobility in all electrophoretic systems tried. S1 and G1, which comigrated in an SDS-sodium borate system, showed different mobilities upon addition of 5 M urea to the system. 3. Immunological evidence presented indicates that each subunit has a specific antigenic site(s) in addition to an identical one(s) in common with the others. 4. As each tropomyosin subunit formed two precipitin lines with the homologous antiserum, as many as ten kinds of subunits may exist in chicken muscles.


Alteration in two enzymatically active forms of valyl-tRNA synthetase during the sporulation of Bacillus subtilis. Two enzymatically active forms of valyl-tRNA synthetase [EC 6.1.1.9] were found in the cells of Bacillus subtilis. The aminoacylation activities of the two forms were altered during the sporulation of B. subtilis. The tRNA'S acylated by these enzymes were analyzed by methylated albumin-Kieselguhr column chromatography.


Comparative characterization of four purified lysine-specific transfer ribonucleic acids from chicken embryos. Four purified tRNALys species from 13-day-old chick embryo muscle have been characterized with respect to the following properties: qualitative oligoribonucleotide composition (polyacrylamide gel electrophoresis after RNase T1 digestion), anticodon response towards AAG and AAA (equilibrium dialysis and polylysine synthesis), strength of the aminoacyl bond (de-esterification kinetics), sedimentation coefficient, and temperature-dependent double helix-to-coil transition. The results confirm the existence of four molecularly independent lysine-specific tRNA's in this eukaryotic system.


Isolation of testosterone-binding globulin from bovine serum by affinity chromatography and its molecular characterization. The testosterone-binding globulin (TeBG) from bovine serum was purified by affinity chromatography and hydroxylapatite chromatography. The affinity column used was prepared by coupling 17 alpha-carboxyethynyl-17-hydroxy-4-androsten-3-one to aminoethyl-Sepharose. The compound was replaceable by 17alpha-carboxyethynyl-17-hydroxy-5alpha-androstan-3-one, but not by testosterone 17-hemisuccinate, estradiol 17-hemisuccinate, or testosterone 3-(O-carboxymethyl)oxime. The TeBG isolated was homogeneous on analytical polyacrylamide gel electrophoresis and equilibrium centrifugation. The protein was a glycoprotein having a molecular weight of 89,500 and a carbohydrate content of 17%. The association constant (M-1) at 4 degrees C was 1.1 X 10(8) and the number of binding sites per molecule was 0.8. Treatment with guanidine-HCl dissociated the protein into subunits having a molecular weight of 28,400 (about one-third of that of the original molecule). SDS-gel electrophoresis showed that two of the three subunits were slightly larger than the other. The dissociation into subunits could also be accomplished by GEDTA treatment with concomitant loss of testosterone-binding activity. The activity and molecular size were reversibly restored by incubation with excess Ca2+.


Immunochemical studies on non-precipitating guinea pig antibody produced by administration of an excessive dose of Bacillus subtilis alpha-amylase. Administration of an excessive dose of Bacillus subtilis alpha-amylase [EC 3.2.1.1, alpha-1,4-glucan 4-glucanohydrolase] (BalphaA) induced the production of non-precipitating (non-ppt) IgG2 antibody in guinea pigs, whereas immunization with a normal dose produced precipitating (ppt) IgG1 and IgG2 antibodies. The non-ppt IgG2 antibody thus produced could be isolated from the coexisting ppt IgG2 antibody by means of the precipitin reaction at maximum precipitation. The non-ppt antibody was incapable of forming a precipitin arc with BalphaA in a conventional agar plate. In the presence of 4% polyethylene glycol (PEG), however, it formed a single arc which fused completely with those of the ppt IgG1 and IgG2 antibodies. The non-ppt antibody could not fix complement, but inhibited BalphaA activity, though with less efficiency than the ppt antibodies. These properties of the non-ppt IgG2 antibody may be due to a low affinity for BalphaA, since both gel filtration and precipitation of soluble antigen-antibody complexes with 20% PEG showed that the antibody was easily dissociable from BalphaA.


Use of rabbit antiboty IgG bound onto plain and aminoalkylsilyl glass surface for the enzyme-linked sandwich immunoassay. Rabbit antibody IgG was bound onto aminoalkylsilyl or plain glass rods by simple adsorption. For comparison, rabbit antibody IgG was also bound onto glutaraldehyde-activated aminoalkylsilyl glass rods. These antibody-glass rods were tested by the sandwich procedure using Fab' fragments of rabbit antibody conjugated with beta-D-galactosidase from Escherichia coli. The glutaraldehyde-activated aminoalkylsilyl glass showed the largest capacity to bind antigen and the plain glass showed the smallest. However, the antibody-glass rods prepared by simple adsorption were as useful for the sandwich immunoassay of macromolecular antigens as those prepared with glutaraldehyde. With all the antibody-glass rods prepared, 0.1 to 10 fmol of ornithine delta-aminotransferase from rat liver and 2,4-dinitrophenyl human IgG were measurable. More than 10 fmol of the antigens may be measurable with larger amounts of the antibody-beta-D-galactosidase complexes, although the non-specific binding of the complexes to the solid phase increases to limit the sensitivity of the immunoassay.


Activation of a new plasma enzyme by antigen-antibody reaction. The change of enzyme activity in immunized rabbit plasma after addition of the homologous antigen was examined. The activities of N alpha-tosyl-L-arginine methyl ester (TAMe) and N alpha-tosyl-L-lysine methyl ester (TLMe) hydrolysis increased about 15 to 18 days after immunization. This increase was especially marked before the maximal rise of antibody content, and is thought to be related to the IgM antibody not to the IgG antibody. Enzyme activation was strongly inhibited by chelation of Ca2+ with 5 mM disodium ethylenediamine tetraacetate (EDTA), but not by other protease inhibitors, such as epsilon amino-caproic acid (epsilon-ACA), bovine lung kallikrein inhibitor (Trasylol) or soybean trypsin inhibitor (SBTI).


Enzymic reactions of phosphate analogs.

Enzymic reactions of phosphate analogs. A variety of phosphonates (XPO32-; X = H-, CH3-, CL3C-, CH3CH2-, and phenyl-) as well as methylarsonate have been shown to be suitable phosphate analogs for the reactions catalyzed by yeast glyceraldehyde-3-phosphate dehydrogenase and calf spleen purine nucleoside phosphorylase. The reactivity of the phosphate analogs with these two enzymes is independent of the pKa of the analog.


Mutants of Bacillus megaterium resistant to uncouplers of oxidative phosphorylation. Mutants of Bacillus megaterium displaying malate-stimulated ATP synthesis resistant to uncouplers of oxidative phosphorylation were isolated and partially characterized. ATP synthesis in such mutants was resistant to carbonyl cyanide m-chlorophenyl hydrazone as well as to other uncouplers including 2,4-dinitrophenol, pentachlorophenol, and sodium azide. ATP synthesis in the wild type and in resistant mutants was sensitive to N,N'-dicyclohexylcarbodiimide, tributyltin, valinomycin plus potassium, and potassium cyanide. Active transport of glycine and glutamine which are sensitive to uncouplers in the wild type was also uncoupler-sensitive in the mutants.


Developmentally regulated lectin from embryonic chick pectoral muscle. Purification by affinity chromatography. A lectin, whose specific activity in soluble extracts of embryonic chick pectoral muscle increases strikingly between 8 and 16 days of development, has been purified by affinity chromatography on derivatized Sepharose 4B coupled to p-aminophenyl-beta-D-lactoside. After affinity chromatography the lectin is pure except for minor contamination with another protein possibly representing a second muscle lectin. The latter can be completely removed by preparative isoelectric focusing. The purified lectin has an apparent molecular weight of 30,000 and an apparent subunit molecular weight of 15,000. Its isoelectric point is 4.0. The most potent saccharide inhibitors tested were thiodigalactoside and lactose. An antibody has been raised to the pure lectin. Studies with this antibody indicate that the lectin is present both on the surface of and within myoblasts.


Hageman factor substrates. Human plasma prekallikrein: mechanism of activation by Hageman factor and participation in hageman factor-dependent fibrinolysis. Two molecular forms of prekallikrein can be isolated from pooled normal human plasma. Their approximate molecular weights by sodium dodecyl sulfate-gel electrophoresis are 88,000 and 85,000. The two bands observed are shown to represent prekallikrein by functional, immunochemical, and structural criteria. Both forms are cleaved by activated Hageman factor, they appear to share antigenic determinants, they are not interconvertible upon incubation with activated Hageman factor or kallikrein, and the ratio of kinin-generating, and plasminogen-activating activities of the preparations are independent of the relative proportion of each band. Activated Factor XII converts prekallikrein to kallikrein by limited proteolysis and two disulfide-linked chains designated kallikrein heavy chain (Mr = 52,000) and kallikrein light chains (Mr = 36,000 or 33,000) are formed. The active site is associated with the light chains as assessed by incorporation of [3H]diisopropyl fluorophosphate. No dissociable fragments were observed in the absence of reducing agents. However, kallikrein could digest prekallikrein to diminish its molecular weight by 10,000. In addition, two factors capable of activating plasminogen to plasmin have been isolated; one is identified as kallikrein. The second principle fractionates with Factor XI and is demonstrable in normal and prekallikrein-deficient plasma.


Induction of fatty acid cyclooxygenase activity in canine kidney cells (MDCK) by benzo(a)pyrene. Canine kidney cells (MDCK) in which [3H]arachidonic acid was esterified in the cellular lipids released increased levels of radioactive prostaglandins and arachidonic acid into the medium when cultured in the presence of benzo(a)pyrene. When MDCK cells were cultured in the presence of benzo(alpha)pyrene and 7,8-benzoflavone, this increased release was not observed. MDCK cells incubated with benzo(a)pyrene also converted exogenous arachidonic acid into prostaglandins more effectively than cells grown in its absence. 7,8-Benzoflavone inhibited this benzo(a)pyrene effect. Microsomes, prepared from benzo(alpha)pyrene-treated MDCK cells synthesized prostaglandin F2alpha from arachidonic acid more effectively than nontreated cells.


Antibacterial activity of antibiotics in acrylic bone cement. The release of various penicillins and other antibiotics from two brands of polymerised bone cement has been studied in vitro and in vivo in mice. Bone cement plugs containing antibiotics demonstrated antibacterial activity as a result of diffusion of antibiotic from the plugs into the surrounding medium. With all antibiotics tested, from 2-5 to 10 per cent of the antibiotic in the cement was released in vitro in active form within twenty-four hours. Most of the activity appeared within three hours of the start of the test, but in some cases low levels of activity were detected after four days. Antibiotic cement plugs implanted in mice and rats produced low concentrations of antibiotic in the blood up to two hours after implantation, but activity was seldom detected subsequently. In general, penicillins and non-penicillin antibiotics showed similar diffusion characteristics, and the pattern of release in vitro and in vivo was consistent with the leaching of antibiotic from, or near, the surface of the bone cement.


Incorporation of purine nucleosides in cultured fibroblasts from a patient with purine nucleoside phosphorylase deficiency and associated T-cell immunodeficiency. Cultured skin fibroblasts from a patient with T-cell immune deficiency and an absence of purine nucleoside phosphorylase activity in red cells were assayed for their capacity to metabolize inosine and guanosine. The cultured fibroblasts were lacking activity of nucleoside phosphorylase and, compared to normal fibroblasts, could incorporate only 2% and 4% of 14C-inosine and 3H-guanosine, respectively, into acid precipitable material. Autoradiography visually confirmed the failure of the NP deficient cell line to incorporate the nucleosides into nuclear material. The physiological mechanism by which the deficiency of purine nucleoside phosphorylase causes T-cell dysfunction remains unclear.


Induced alteration in uptake properties of Tetrahymena and its association with the development of mating competency. The rate of uridine uptake in Tetrahymena declines by an order of magnitude by two hours after shiftdown to a non-nutrient buffer. This alteration in uptake properties cannot be accounted for by an increase in the intracellular pool of uridine, an increase in apparent Km for uptake or a decline in the rate in which uridine is processed intracellularly. It is argued that the decrease in uridine uptake is due to a reduction in numbers of functional transport molecules exposed at the cell surface and is a reflection of a developmentally related cell surface transformation. In addition, the putative decline in functional transport molecules cannot be entirely explained by metabolic turnover of these molecules in the absence of replacement, nor does it require the synthesis of new protein. We discuss the possibility that a shift in equilibrium between accessible and inaccessible transporters is operating. Finally, a close correlation between conditions which elicit the transport alteration and those which allow the development of mating competency suggests that the two phenomena may be coordinately regulated.


Mg2+-dependent electrical control of flagellar activity in Euglena. When a Euglena gracilis in a Ca2+-containing medium is impaled with a microelectrode, the flagellum is instantly ejected. In a Ca2+-free medium to which 1 mM EGTA has been added, the flagellum remains attached to the organism, but it loses activity upon impalement. Externally added ATP at a concentration of 10 mM will sustain normal flagellar activity (at approximately 20 Hz) of an impaled Euglena. If negative direct current of several tenths of a microamp is injected through the impaling microelectrode, the flagellar activity is stopped or much reduced. When the current injection is turned off the flagellum returns to its initial activity. This cycle can be repeated many times on the same animal, independent of whether Mg2+ is present in the external medium or not. If 1 micrometer of gramicidin is added to Ca2+-free medium containing 1 mM EGTA and 10 mM ATP, the flagellar activity becomes dependent on external Mg2+. Without external Mg2+ no flagellar activity is present after one or two current injection cycles as described above. With 1 mM Mg2+ present in the external medium many cycles (up to 10) can be produced. This Mg2+-dependent flagellar activity shows a smooth dependence on the amount of current injected. Observations taken by high speed cinemicrography show that in the third injection cycle the average frequency of the flagellar motion is 16-3 Hz at 0 muA, is 8 Hz at 0-2 muA, and is approximately 0 at 0-6 muA of negative current. The injection of positive current results in an increase in flagellar frequency dependent on the amount of current injected. The data indicate that the control of motility of Euglena flagella is dependent on an electrically activated Mg2+ pump.


Electrophoretic mobility of microsomes from rat liver. The electrophoretic mobilities of rough and smooth microsomes were studied using free electrophoresis in a sucrose gradient. Rough microsomes have a higher net negative surface charge but removal of the ribosomes decreases their mobility to that of smooth microsomes. Treatment with neuraminidase and phospholipases C and D does not affect the mobility of total smooth microsomes, but this mobility is increased by approximately 20% after trypsin and papain treatment and by approximately 12% after phospholipase A treatment. Further treatment of trypsin-digested smooth microsomes with phospholipase C re-establishes the original mobility. This effect is not caused by the removal of lipid phosphate groups, but by the liberation of negatively charged protein species that are normally buried under trypsin-sensitive proteins. Low concentrations of trypsin also solubilize enzyme proteins from smooth liver microsomes of phenobarbital-treated rats, but the electrophoretic mobility is not increased, indicating structural differences between induced and control membranes.


Mitotic spindles of Drosophila melanogaster: a phase-contrast and scanning electron-microscope study. Mitotic spindles have been isolated from the blastema stage of Drosophila melanogaster embryos using modified tubulin-polymerizing medium. 'Clean' spindles, relatively free of contaminating cytoplasmic material, are obtained. Under phase contrast, mitotic stages appear remarkably similar to those seen in situ, as reported in early literature. This preservation of morphological integrity, coupled with relative structural simplicity due to low chromosome number (2n = 8), makes these spindles ideal subjects for study. Use of the scanning electron microscope provides excellent visulization of their general structural organization, changes in whole spindle structure during the course of mitosis, and higher resolution viewing of surface detail than is permitted with light microscopy.


Proportional regulation of body form and cortical organelle pattern in the ciliate Dileptus. Morphometric and ultrastructural studies of shape and pattern regulation were performed on Dileptus anser. In this ciliate 2 body regions can be distinguished: proboscis and trunk. When a large portion of the trunk is excised, shape regulation occurs and the proper proboscis/trunk ratio is restored through elongation of the remnant of the trunk and shortening of the proboscis. When there is nothing but proboscis left, the trunk is formed from the proximal portion of proboscis and again the right proportion is restored. Regulation of the ciliary pattern is based mainly on resorption of some ciliary elements. The resorption of somatic ciliature is especially intense within narrowing regions, where otherwise overcrowding of cilia would occur. The resorption of oral ciliature was found to occur within an area located at the apex of the proboscis. Oral structures when damaged by cutting may be repaired in situ. Changes in the microfibrillar system of the cell were found during regulation of shape of the posterior region of the cell, as well as during regulation of size of oral structures. Regulatory processes in Dileptus are compared to those known for other ciliates, and possible differences between the regulation of shape and the regulation of pattern are discussed.


A quantitative assay to study cell movement in the myxobacteria. A simple quantitative assay has been developed to test the rate of cell movement of myxobacteria. The assay employs an agar surface and at no time are the cells cultured in a liquid environment. Isolation of a rate-increasing substance(s) from fruiting Myxococcus xanthus is reported. The understanding of the aggregative process in these bacteria will be aided by characterization of the chemotactic system.


Analysis of contractile vacuole pore morphogenesis in Tetrahymena pyriformis by 180 degree rotation of ciliary meridians. The contractile vacuole pores (CVPs) in Tetrahymena pyriformis are usually found to the left of ciliary meridians (CVP meridians). When the CVP meridians are experimentally rotated 180 degrees ('inverted'), the CVPs are now found to the right of the rotated CVP meridians. In other words, the CVP is almost invariably found on the same side with respect to the CVP meridian. Thus, the 2 sides of the CVP meridian have different morphogenetic properties and such differences are determinative in the asymmetrical fine-positioning of the CVP. The number of CVPs per animal and their general placement on the animal (i.e. with which ciliary meridians the CVPs are associated) are known to be a function of the total number of ciliary meridians possessed by the animal. Rotation of the CVP meridians affects both the number of CVPs per animal and their general placement. The specificity of such effects appears to depend on whether both, or one, or which one, of the CVP meridians is rotated. Rotation of ciliary meridians may be used as a tool in analysing the mechanism determining the number of CVPs per animal and their general placement.


The ultrastructure of the somatic cortex of Pseudomicrothorax dubius: structure and function of the epiplasm in ciliated protozoa. The ultrastructure of the somatic cortex of the ciliate Pseudomicrothorax dubius is studied with emphasis on the epiplasm layer which lies immediately under the inner alveolar membrane and is continuous with the terminal plates of cortical basal bodies. In addition to a clearly demonstrable cytoskeletal role, the epiplasm appears to function as a comenting substance which integrates numerous cortical fibres and membranes. The kinetodesmal, postciliary and transverse fibre systems which originate at the proximal ends of basal bodies extend toward the cell surface and end at or in the epiplasm. Inner alveolar membranes and trichocyst membranes are attached to the epiplasm. Basal bodies are anchored into the epiplasm via their terminal plates. The epiplasm appears to be morphogenetically important as a matrix into which newly formed basal bodies insert. Electron-opaque arms occur at the terminal plate level of new basal bodies, and these arms fuse with the epiplasm when basal body insertion occurs. The position of trichocysts in the cortex is specified by the epiplasm. Evidence from numerous other ciliates tends to confirm both structural and morphogenetic roles of the epiplasm.


Preferential inhibition of rDNA transcription by 5-bromodeoxyuridine. On a chemically defined growth medium the degree of substitution of thymidine with 5-bromodeoxyuridine (BUdR) in DNA of Tetrahymena pyriformis was controlled by the concentration of tetrahydrofiolic acid, BUdR and thymidine in the medium. A correlation between the degree of BUdR substitution in DNA and the reduction in rate of total RNA synthesis has been established. It was found that the reduction of total RNA synthesis results from inhibition of transcription of all RNA species which have been measured. However, independent of the degree of BUdR substitution in DNA, a preferential inhibition of the synthesis of 25s and 17s ribosomal RNA was found. It is concluded that the various genes may respond differently to BUdR substitution with respect to transcription.


Use of ethyl ethers, deuteriomethyl ethers and cyclic n-butylboronates of hydroxychlorobiphenyls in identification of metabolites of polychlorinated biphenyls. The mass spectra and gas chromatographic properties of a number of chlorobiphenylols, chlorobiphenyldiols, chlorophenols and naphthols, as well as their methyl, deuteriomethyl and ethyl  ethers and some cyclic n-butylboronates have been investigated. Metabolism experiments with 4'-chloro-4-biphenylol showed that the selective use of ethylation and methylation is most effective in both detection and structure elucidation of metabolites partly methylated by metabolic processes. The usefulness of deuteriomethylation in such studies seems to be limited. The formation of cyclic n-butylboronates provides specific information on o-dihydroxy derivatives of chlorobiphenyls.


Sensitive enzyme-linked immunosorbent assay for detection of antibodies against typhus rickettsiae, Rickettsia prowazekii and Rickettsia typhi. An enzyme-linked immunosorbent assay (ELISA) has been developed for the titration of rickettsial antibodies in human and animal sera. Two preparations of soluble typhus-group antigens were obtained from Rickettsia typhi and Rickettsia prowazekii by ether extraction: a standard antigen from infected yolk sacs (YS antigen) and one free of yolk sac contaminants from Renografin-purified rickettsiae (PR antigen). Rabbit, mouse, and guinea pig sera were obtained by immunization with viable purified R. typhi or R. prowazekii. Human sera were obtained from individuals who had recovered from laboratory infections with either typhus rickettsia months or years previously. Goat-derived anti-immunoglobulins were conjugated to alkaline phosphatase with glutaraldehyde. Although the PR and YS antigens gave equivalent antibody titers in the complement fixation test, the PR antigen was clearly superior in the ELISA. With this antigen, the titration curves of all antisera were linear over a wider range of serum concentrations and the titers were higher than with the YS antigen. With YS and PR antigens, ELISA titers were higher than those obtained by complement fixation by one and two orders of magnitude, respectively. In human sera, immunoglobulin G and immunoglobulin M antibodies were demonstrated by their respective anti-immunoglobulins and by differential susceptibility to ethanethiol. ELISA titers showed some type specificity, whereas none was observed in complement fixation tests. The ELISA is highly sensitive, reproducible, and easily adaptable to the various requirements of clinical and research laboratories.


Preservation of mycobacteria st -70 degrees C: persistence of key differential features. The advantages of long-term preservation of suspensions of mycobacteria by storage at -70 degrees C established in earlier studies are reinforced by present evidence that freezer storage does not alter key taxonomic features used to identify mycobacteria. Occasional discrepancies in biochemical test characteristics of mycobacteria that have been stored in the freezer and reconstituted at 37 degrees C reflect only sluggish metabolic activity, which is restored to normal on repeat testing.


Modified indirect fluorescent-antibody test for aspergillosis. Diffuse fluorescence in an indirect fluorescent-antibody test for antibody to Aspergillus fumigatus was eliminated by using germlings, rather than hyphae, as antigen. The modified method yielded diagnostic titers larger than or equal to 1:32 in 24 of 25 known cases, was highly specific, and showed good correlation with clinical status and results of immunodiffusion. This technique, which uses the germling antigen, inactivation of test serum at 59 +/- 1 degrees C, and anti-immunoglobulin G conjugate, is a sensitive, specific, and reliable quantitative indicator of aspergillosis and is useful in following the course of the disease.


Application of freeze-dried monkey erythrocytes to measles viral hemagglutination and hemagglutination-inhibition tests. Erthrocytes collected from monkey species including grivet, rhesus, and cynomolgus monkeys were stabilized by fixation with glutaraldehyde of a low concentration and were freeze-dried in vacuo. These freeze-dried erythrocytes were compared with fresh erythrocytes for measles viral hemagglutination and hemagglutination inhibition and could be used in both tests instead of fresh erythrocytes. They maintained their initial appearance and sensitivity to measles viral hemagglutinins after storage at 4 degrees C for 4 months more.


Nontypable group B streptococci isolated from human sources. The present study was done to determine whether so-called nontypable (NT) group B streptococci from human sources possess as yet unrecognized type antigens. Antisera were raised in rabbits against several NT strains and then tested with hydrochloric acid extracts of 53 NT group B streptococci. One serum was strain specific, another was nonspecific in that it contained only R-protein antibodies, and a third (NT1), although apparently type specific, reacted with only five strains. These results do not justify using NT1 serum in the group B typing system.


Thyroid dysfunction in chronic renal failure. A study of the pituitary-thyroid axis and peripheral turnover kinetics of thyroxine and triiodothyronine. Thyroid function was evaluated in 46 patients with end-stage kidney disease and 42 normal subjects. Patients were studied before and after the institution of maintenance hemodialysis (HD) and after renal transplantation (RT). Serum total triiodothyronine concentrations (TT(3), ng/100 ml, mean+/-SD) were 63+/-17 and 83+/-22 in the non-HD and HD groups, respectively. Values from normal subjects were 128+/-25 and from RT patients 134+/-20. The TT(3) was in the hypothyroid range (<78 ng/100 ml; 2 SD below normal mean) in 80% of non-HD and 43% of HD patients. Mean serum total thyroxine concentration (TT(4)), although within the normal range, was lower than the control value. T(4)-binding globulin capacity was also slightly lower but the difference was not statistically significant. Among patients whose TT(4) was 1 SD below the normal mean, the free T(4) index was equally depressed, suggesting that factors other than decreased binding capacity might be responsible for the low TT(4). In addition, there was a 37% incidence of goiter. Mean serum thyroid-stimulating hormone (TSH) was not elevated and the TSH response to thyrotropin-releasing hormone (TRH) was distinctly blunted, suggesting the possibility of pituitary dysfunction as well. In vivo (125)I-l-T(4) and (131)I-l-T(3) kinetics during 0.2 mg/day of l-T(4) replacement showed marked reduction in T(3) turnover rate in the uremic patients, both before and during HD; the values (mug T(3)/day, mean+/-SD) for the different groups were as follows: normal, 33.8+/-6.1; non-HD, 13.5+/-2.6; HD, 12.9+/-3.1; and RT, 30.3+/-7.1. The low T(3) turnover rate was due to impaired extrathyroidal conversion of T(4) to T(3). The mean percent+/-SD of metabolized T(4) converted to T(3) was 37.2+/-5.8 in normal subjects, 15.7+/-3.1 in non-HD, 12.8+/-1.7 in HD, and 34.0+/-14.7 in RT patients. In contrast, thyroidal T(3) secretion rate was not different between the control and the three patient groups. Thus, it appears that uremia affects thyroid function at several levels: (a) subnormal pituitary TSH response to TRH; (b) possible intrathyroidal abnormalities as suggested by slightly decreased TT(4) and high incidence of goiter; and (c) abnormal peripheral generation of T(3) from T(4). Restoration of renal function with RT resulted in normalization of all parameters of thyroid function with the exception of blunted or absent TSH response to TRH. The latter may be a direct consequence of glucocorticoid administration.


Purine nucleoside phosphorylase deficiency. Evidence for molecular heterogeneity in two families with enzyme-deficient members. Purine-nucleoside phosphorylase (NP) deficiency is associated with severely defective thymus-derived (T)-cell and normally functioning bone marrow-derived (B)-cell immunity. In this study, two unrelated families with a total of three NP deficient members were investigated. High pressure liquid chromatography of the plasma of the three patients showed inosine levels greater than 66 muM. This nucleoside was absent from the plasma of their parents and control samples.NP was purified from normal human erythrocytes by affinity chromatography and an antiserum prepared in rabbits was used to study the NP variants in the two families. In family M the patient had no detectable erythrocyte NP activity and no detectable immunological-reacting material (irm) to the NP antibody. The parents, who are second cousins, had less than one-half of normal enzyme activity and approximately 14% irm attributable to a variant protein. Their electrophoretic patterns revealed a series of isozymes with slower than normal migration. In family B the patients had 0.5% residual enzyme activity and about one-half normal irm. Their electrophoretic pattern showed faintly staining bands which migrated faster than normal NP. The mother of the patients had one-half normal enzyme activity, 11% irm attributable to her variant protein, and a normal electrophoretic pattern. The father had less than one-half normal enzyme activity, equal amounts of normal and variant irm, and an electrophoretic pattern that showed increased activity of the more rapidly migrating isozyme bands.The combined use of immunological and electrophoretic techniques has shown the presence of three separate mutations; one in family M and two in family B associated with severely defective T-cell function.


Some effects of calcium on the activation of human factor VIII/Von Willebrand factor protein by thrombin. When Factor VIII/von Willebrand factor (FVIII/vWF) protein is rechromatographed on 4% agarose in 0.25 M CaCl(2), the protein and vWF activity appear in the void volume, but most of the FVIII procoagulant activity elutes later. Recent evidence suggests that the delayed FVIII procoagulant activity is a proteolytically modified form of FVIII/vWF protein that filters anomalously from agarose in 0.25 M CaCl(2). To test whether or not thrombin is the protease involved, the effect of 0.25 M CaCl(2) on FVIII/vWF and its reaction with thrombin was examined. About 30% of the FVIII procoagulant activity was lost immediately when solutions of FVIII/vWF protein were made 0.25 M in CaCl(2). When FVIII in 0.15 M NaCl was activated with 0.04 U thrombin/ml and then made 0.25 M in CaCl(2), the procoagulant activity of a broad range of FVIII/vWF protein concentrations remained activated for at least 6 h. But, in 0.25 M CaCl(2), the increase in FVIII procoagulant activity in response to thrombin was much more gradual and once activated, the procoagulant activity was stabilized by 0.25 M CaCl(2). When thrombin-activated FVIII/vWF protein was filtered on 4% agarose in 0.15 M NaCl, there was considerable inactivation of FVIII procoagulant activity; however, the procoagulant activity that did remain eluted in the void volume. In contrast, when thrombin-activated FVIII/vWF protein was filtered in 0.25 M CaCl(2), the FVIII procoagulant activity eluted well after the void volume and remained activated for 6 h. The procoagulant peak isolated by filtering nonthrombin-activated FVIII/vWF protein on agarose in 0.25 M CaCl(2) was compared to that isolated from thrombin-activated FVIII/vWF protein. Both procoagulant activity peak proteins had about the same specific vWF activity as the corresponding void volume protein. Before reduction, the sodium dodecyl sulfate gel patterns for the two procoagulant activity peak proteins were the same. After reduction, the gel pattern for the nonthrombin-activated procoagulant activity peak protein contained bands of 195,000, 148,000-120,000, 79,000, 61,000, 51,000, and 18,000 daltons whereas the pattern for the reduced thrombin-activated procoagulant activity peak protein always lacked the higher molecular weight bands, but consistently showed the four lower molecular weight bands to be well resolved. Taken together, these results imply that thrombin generates the FVIII procoagulant activity that is stabilized by 0.25 M CaCl(2) and elutes aberrantly from 4% agarose in that solvent.


Cells of origin and terminal distribution of descending projections of the rat somatic sensory cortex. The retrograde, horseradish peroxidase technique has been used to demonstrate the cells of origin of corticofugal fiber systems arising in the rat somatic sensory cortex and projecting to the striatum, diencephalon, brainstem, and spinal cord. Correlative experiments conducted with the anterograde, autoradiographic method have been used to confirm the terminal distribution of many of these fiber systems. Corticofugal pathways directed to subcortical structures arise in the first and second somatic sensory areas exclusively from pyramidal cells of the infragranular layers, V and VI. Fibers which descend to the midbrain, pons, medulla and spinal cord arise exclusively from the largest pyramidal cells, the somata of which are found in the deep part of layer V (layer VB). There is some evidence for a sublaminar organization of the different classes of efferent cells within this layer. Fibers projecting to the diencephalon arise from somata situated throughout layer VI and to a lesser extent in layer V. Corticostriatal fibers arise only from cells with somata in layer V, but the somata are more superficially situated than those of the other classes of corticofugal neurons. The laminar distribution of the somata of corticofugal neurons differs considerably from that of commissural and ipsilateral corticocortical neurons.


Factors influencing sexual performance in male rhesus monkeys. A 5-yr retrospective study of the sexual behavior of eight adult male rhesus monkeys showed that sexual vigor declined over the years but testosterone levels in peripheral vein plasma did not. Two prospective experiments were then carried out on these males during the sixth year. (a) The four poorest performers were injected daily for 28 days with testosterone propionate (1 mg/kg of body weight). There was no significant increase in level of performance, and behavior was not correlated with plasma levels of testosterone either before or 24 hr after the last hormone injection. (b) All eight males were exposed to novel nonspecific sensory stimulation during tests of sexual behavior. Eight different adult male rhesus strangers-present in the room but not in the test cage-were used as stimuli, one for each experimental test. Sexual behavior during experimental and control tests did not differ.


Task dependence of slowing after pyramidal lesions in monkeys. The effect of lesions in a medullary pyramid was studied in two experiments. (a) Monkeys were trained to press a key with a rear-projected circle, presented together with different numbers of keys with ellipses. Two to eight choices were presented in a random sequence. Discrimination was measured with a titration schedule, and short choice reaction times were reinforced selectively. Choice reaction time and the function relating it to the number of choices were unaffected by lesions reducing the area of a transverse section of the pyramid up to 93%. (b) A repetitive response controlled by a fixed-ratio schedule was performed simultaneously with a holding response with the other hand, and the two hands changed function after each food pellet. Lesions reducing the pyramid only 25% increased interresponse times of the phasic response with the contralateral hand. Experiment 1 and previous studies are interpreted to indicate that slowing after pyramidal lesions is dependent on behavioral context. Experiment 2 showed that a simple repetitive movement is affected when the rate is high. The static response was more clearly affected than the phasic response, which indicates a role for the pyramidal tract in posture.


Immune neutrality of calf skin collagen gel used to stimulate revitalization in pulpless open apex teeth of rhesus monkeys. A collagen mineral gel has recently been shown to stimulate revitalization of pulpless open apex teeth when syringed into debrided root canals. Experiments were conducted to examine possible stimulation of the immune system by the experimental procedure. Ring tests (interface precipitation analysis), Ouchterlony plates and hemagglutination tests on plasma samples taken from experimental rhesus monkeys throughout the healing process indicate a complete lack of humoral immune responsiveness to the injected collagen.


Toward organism identification through analysis of bacterial colonies. The clinical problem of bacterial identification has been approached by applying pattern-recognition techniques to multi-wavelength surface-scattering and reflectance data derived from real-time scans of isolated colonies. Preliminary results, obtained from blood-agar plates inoculated with a mixture of staphylococci, streptococci and escherichieae, indicate that these organisms can be differentiated with better than 90% certainty, provided the colonies are physically separated and their growth conditions closely controlled. Data collection and classification characteristics of the experimental system are briefly described; it is felt that the technique, possibly expanded to include boundary characteristics of the colonies, may offer a viable means of identifying clinically important bacteria.


[Qualitative and quantitative investigations of the area striata in an ontogenic series of male Tupaia belangeri]. The cyto-, myelo- and fibrilloarchitectonics of the area striata and its laminae were described in 36 male Tupaia belangeri at an age of 36 to 536 days of ontogenesis. The fresh volumes of the area striata and its laminae were determined and the growth of the fresh volumes described with the 6-parametric growth-function: (formula: see text) Likewise, the matter lying directly under the area striata were determined and these data were analysed with the logistic growth-function: (formula: see text) 1) During ontogenesis, the area striata and its laminae show a growth over-shoot of the fresh volumes with differing procentual reduction, which is greatest (45%) at lamina IIIc and the least (18%) at lamina VI. The points in time for maximal fresh volume and the reduction speeds are different. Initially, the reduction of lamina VI is finished and that of lamina I, lastly. 2) The associated white matter of the area striata shows a nonlinear a monotonous increase in fresh volumes.


Measurements of skinfold thickness and musculature in the assessment of growth and nutritional status of malnourished children. Measurements of the mid-upper arm circumference and the triceps skin-fold thickness were made at six montly intervals in 15 malnourished children. From these measurements, values were obtained for 'muscle bulk' and 'fat bulk' using local standards. Over a 12 month period, both measurements increased among the overtly malnourished children. The fat increased most in the children who were initially underweight (marasmic children). All these indicated an improvement in the nutritional status of these children. The Oje group of children deteriorated in all the parameters without any evidence of acute illness. A possible reason for this situation was a false sense of confidence on the part of the children's mothers and consequently, inadequate provision of food. The use of subcutaneous fat as an index of inadequate intake, and skeletal muscle for evidence of protein depletion, was also highlighted.


Immunologic classification of Neisseria gonorrhoeae with micro-immunofluorescence. A reproducible immunologic classification of Neisseria gonorrhoeae strains has been achieved by the micro-immunofluorescence (Micro-IF)3 method by using formalinized whole organisms as test antigens and mouse antisera prepared by i.v. immunization with the whole organisms as antibody. Immunologic differences among Neisseria species were also distinct in this test system. Immunologic differences among gonococcal strains were not influenced by gonococcal colony type. Classification of gonococci was facilitated by use of antisera absorbed with an antigenically unique gonococcus strain. Of 180 gonococcal strains, 175 could be classified into three immunotypes: A, B, and C. Each type was further divided into subtypes designated A1, A2, A3, B1, B2, B3, C1, and C2. Minor antigenic differences still exist within each subtype. The two gonococcal isolates from each of 17 pairs of sexual contacts fell into the same subtype. Seventy-one of 73 isolates which required arginine, hypoxanthine, and uracil for growth (Arg-Hyx-Ura-) and seven of 107 other auxotypes belonged to subtypes A2 and A3. Marked geographical differences in distribution of gonococcal immunotypes were observed among those available for testing. Subtypes A2 and A3 were predominant in Seattle whereas types B and C were predominant in Southeast Asia. The only Arg-Hyx-Ura- isolates not belonging to subtypes A2 or A3 were the only two that were serum sensitive. This Micro-IF immunotyping appears potentially useful for future immunologic, epidemiologic, and genetic studies of N. gonorrhoeae.


Structural evidence for an unblocked VHI sub-subgroup of human heavy chains. The N-terminal amino acid sequence of an unblocked human heavy chain of an IgGK paraprotein from patient Tho is reported. This unblocked VH sequence belongs to the VHI subgroup and has striking structural homology to an unblocked VHI sequence previously reported for the IgGK paraprotein isolated from the serum of patient Bro. Direct sequence comparison of the Tho and Bro proteins reveals complete structural identity in 25 of the N-terminal 27 residues, with unique amino acid substitutions shared at the N-terminus and positions 16, 18, and 24. This remarkable sequence homology suggests that the two VH sequences represent examples of an unblocked VHI sub-subgroup. The Tho and Bro sequences possess an ala-glu-val basic triplet at positions 9 to 11 in common with the other previously reported VHI proteins which help to further establish this sequence triplet as a recognizable VHI subgroup-specific region. Moreover, in this regard, the unblocked sequences of Tho and Bro emphasize the importance of extending an unblocked VHI sequence beyond positions 9 to 11 before a subgroup assignment can be determined.


Cell-mediated immune response to lymphocytic choriomeningitis and vaccinia virus in rats. The parameters of cell-mediated immune responses of rats to infection with lymphocytic choriomeningitis virus or vaccinia virus were assessed by measuring primary footpad swelling, increased weights of the local lymph nodes, increased numbers of lymphocytes per lymph node, and the course of virus-specific cytolytic activity by these lymphocytes. Except for lack of a defined swelling caused by vaccinia virus injected into the hind footpads of rats, the kinetics of all these responses correlated and were in accord with the usual time course of cellular immune responses. Starting 3 days after infection, peaking at 5 to 7 days, and disappearing after 10 to 12 days, the responses by rats to both viruses were comparable to those found in mice. The phagocytes of these infected rats inhibited the growth of Listeria monocytogenes in vivo, indicating activation of the macrophages by virus-specific cellular immunity. The rat cytotoxic lymphocytes were thymus derived as judged by various criteria: inactivation by an absorbed rabbit anti-rat brain antiserum plus C, susceptibility to anti Thy 1.1 plus C, restriction of the lytic activity within inbred strains and probably by the Ag-B locus, and the kinetics of the response. The cytotoxic T lymphocytes were virus specific since they killed only target cells infected with the same virus but not uninfected cells, or targets that were infected with an unrelated virus.


Analysis of the diversity of murine antibodies to dextran B1355. II. Demonstration of multiple idiotypes with variable expression in several strains. We have developed radioimmunoassays that detect idiotypic (variable region) differences among the alpha(1 leads to 3) dextran-binding meyloma proteins U102, J558, and M104 as well as an assay that detects variable region determinants common to all three proteins. Using these assays, we have examined 7S and 19S anti-alpha(1 leads to 3) dextran antibodies induced in five murine strains of the a1 IgCH linkage group and the recombinant strain BAB/14. All idiotypes were expressed in both 19S and 7S antibodies from all strains, but with considerable strain-specific variability in penetrance. In two strains, one additional type of antibody, which lacked all four idiotypic determinants, generally constituted the bulk of total anti-alpha(1 leads to 3) dextran antibodies.


Immunoepidemiology of meningococcal disease in military recruits. I. A model for serogroup independency of epidemic potential as determined by serotyping. One hundred twenty strains of Neisseria meningitidis were serotyped with use of cross-absorbed rabbit antisera in a bactericidal test. Fifty-eight epidemic strains of serogroup B, C, and Y that occurred simultaneously among military recruits at two basic training centers during a period of epidemic meningococcal disease were compared with 62 strains of serogroups A, B, C, and Y isolated worldwide. Antisera to the six original antigenic factors of the Gold serotyping schema were adequate for typing 94% of strains, including all of the epidemic strains. The array of serotyping factors in the epidemic strains differed from those in the nonepidemic strains. Epidemic strains were almost exclusively of two serotypes, with type CII predominant among strains of groups B and C. Concurrent strains of groups B and C were invariably of the same serotype. A model for the epidemic potential of meningococcal strains, which is based on their serotype and serogroup antigens, and a modification of the original Gold typing schema are presented.


Ultrastructural study of cervical gonorrhea. An ultrastructural examination of the squamocolumnar junction of the cervix of patients infected with Neisseria gonorrhoeae is described. Gonococci were found to become firmly attached to stratified squamous epithelium, a process that appeared to be initiated by activity of the cytoplasmic membrane of superficial squames. By contrast, gonococci were not found attached to, or even closely associated with, mucus-secreting columnar epithelium. Gonococcal growth, as evidenced by numbers of organisms and surface vesicle formation, appeared most active in cervical secretions, chiefly on exfoliated squames, but also lying free. Survival after phagocytosis by polymorphonuclear leukocytes was found not to be unique to gonococci, and it is suggested that continual phagocytic recycling minimizes the significance of this occurrence. It seems probable that persistence of gonorrhea in the female depends upon the adherence of gonococci to stratified epithelium, where they are protected from phagocytosis, and the infectivity of gonorrhea arises from the ability of gonococci to divide rapidly on the surface of exfoliated squames, from where they are released into secretions.


Antifungal activity of four tetracycline analogues against Candida albicans in vitro: potentiation by amphotericin B. The antifungal activities of four tetracycline analogues in combination with amphotericin B (AmB) were determined against 20 strains of Candida albicans. When a microtiter checkerboard technique was used, minocycline (less than or equal to 10 microgram/ml) acted synergistically with AmB against all strains, whereas doxycycline had a reduced effect, and demeclocycline and tetracycline had no potentiating effect at this concentration. Killing-curve experiments with two strains of C. albicans demonstrated that the combination of minocycline and AmB produced a decrease in number of colony-forming units (cfu) of greater than 2 logs in 4 hr and a 4-log decrease in cfu in 24 hr at concentrations (minocycline, 0.64 microgram/ml; AmB, 0.1 microgram/ml) that were subinhibitory when each agent was used alone and that are readily achieved in human serum and body fluids with conventional doses. The killing-curve technique indicated that doxycycline had an intermediate degree of synergistic activity, whereas tetracycline had no synergistic activity at clinically relevant concentrations.


Successful prophylaxis against rabies in mice and Rhesus monkeys: the interferon system and vaccine. Addition of interferon to ineffective rabies virus vaccines by the local injection of either exogenous interferon or a potent interferon inducer (a complex of polyriboinosinic-polyribocytidylic acid containing poly-L-lysine and carboxymethylcellulose) into the footpads of mice previously challenged with rabies virus dramatically reduced the mortality rate. A significant reduction in mortality rate was also noted when the interferon system was administered to rhesus monkeys, but only when treatment was given 6 hr after challenge with rabies virus. Since the monkeys were given an overwhelming challenge of virus, the treatment had to be given quickly to obtain results comparable to those in mice.


Protection against infection with Neisseria gonorrhoeae by immunization with outer membrane protein complex and purified pili. Some of the antigens that are capable of producing strain-related immunity to gonococcal infection in the guinea pig are located on the outer membrane of Neisseria gonorrhoeae. This finding has been demonstrated by immunization of guinea pigs with isolated outer membranes from two different strains of N. gonorrhoeae prior to challenged. Isolated principal outer membrane protein complex proved a better protective immunogen than purified pili from the same strain of N. gonorrhoeae. Principal outer membrane protein appears to react in antibody-complement-mediated killing of gonococci, whereas antibodies to pili are only weakly bactericidal. Pili-mediated attachment of N. gonorrhoeae to human cells is inhibited by antibodies to pili, and maximal inhibition occurs when antibodies are directed at pili antigenically identical to the pili mediating the attachment.


Surface components affecting interactions between Neisserai gonorrhoeae and eucaryotic cells. Interactions of Neisseria gonorrhoeae with nonleukocytic eucaryotic cells including tissue culture cells (either primary or continuous lines), human sperm, and buccal mucosal cells appear to be influenced primarily by the presence or absence of pili on the bacteria. In this context, piliation enhances attachment to or association of microorganisms with the eucaryotic cells. Pili, on the other hand, appear to reduce interaction of N. gonorrhoeae with mouse peritoneal macrophages. Other groups have demonstrated that pili reduce phagocytosis of gonococci by neutrophils, but in out studies this affect is secondary to that dependent on "leukocyte-association" factor. The presence of such a factor is correlated with a particular protein found by polyacrylamide electrophoresis. Adherence among N. gonorrhoeae varies and results in colony forms of different coloration or aggregation characteristics. These different coloration forms correlate with the protein patterns of gonococci as found in polyacrylamide electrophoresis and also with the susceptibility of gonococci to killing by trypsin.


Formation of antibody in the newborn mouse: study of T-cell-independent antibody response. The ontogeny of immune responsiveness, as assayed by antibody formation in vitro, of mouse spleen lymphocytes to thymus-independent antigens is reviewed. Responsiveness to trinitrophenyl (TNP)-lipopolysaccharide and TNP-Brucella abortus appear soon after birth and one to two weeks before TNP-Ficoll or capsular polysaccharide of Streptococcus pneumoniae (SSS-III) elicits significant antibody formation. This hierarchy of responsiveness to antigens is also apparent in the CBA/N mutant mouse strain, which has a bone marrow-derived (B-) cell maturation arrest and fails to respond to either TNP-ficoll or SSS-III. These findings are interpreted to suggest sequential maturation of different populations or lines of B-lymphocytes, each of which can respond to a defined class of thymus-independent antigens. The implication for vaccine use in humans is that a late-appearing subclass of B-cells may be required for adequate immune responses to polyaccharide antigens.


Interim report of a controlled field trial of immunization with capsular polysaccharides of Haemophilus influenzae type b and group C Neisseria meningitidis in Mecklenburg county, North Carolina (March 1974-March 1976). Approximately 16,000 children, from two months to five years of age, were vaccinated with the capsular polysaccharide of either Haemophilus influenzae type b or group C Neisseria meningitidis. Immunizations were carried out in a double-masked, randomized manner; the doses of immunogens used were 10 microgram of H. influenzae type b polysaccharide and 25 micron g of the group CN. meningitidis polysaccharide. Immunogenicity of the two vaccines was measured in single, random specimens of blood taken from vaccinees of all ages at various intervals after immunization. A positive effect on formation of serum antibody was observed in children of all ages vaccinated with N. meningitidis polysaccharide, but increased levels of serum antibody to H. influenzae type b were observed only in recipients of that vaccine who were three years of age or older. No untoward reactions to either vaccine were noted, and both vaccines retained their original molecular size after storage for three years. Too few cases of disease have been studied for a definitive assessment of vaccine efficacy; however, a slightly protective effect against meningitis was observed for the H. influenzae type b vaccine in infants up to one year of age.


Sialic acid-containing polysaccharides of Neisseria meningitidis and Escherichia coli strain Bos-12: structure and immunology. A polysaccharide antigenically related to that of group C Neisseria meningitidis was isolated from Escherichia coli strain Bos-12 (O48: K91:NM). Like the polysaccharides of groups B and CN. meningitidis, the Bos-12 antigen was shown to be a pure polymer of sialic acid. The 13C-nuclear magnetic resonance studies of the groups B and C polysaccharides indicated that the former consists of units of sialic acid joined in alpha-2 leads to 8 linkages, whereas the latter contains sialic acid residues linked by alpha-2 leads to 9 glycosidic bonds. Chemical and nuclear magnetic resonance studies of the polysaccharide of E. coli strain Bos-12 established that it is a heteropolymer containing both alpha-2 leads to 8 and alpha-2 leads to 9 linkages. Physical parameters including partial specific volume, reduced viscosity, diffusion and sedimentation coefficients, shape, weight-average molecular weight, and the Stokes radius of the polysaccharides of groups A, B, and C N. meningitidis have been determined. The results indicate that the polysaccharides are highly asymmetric and exist in solution as rigid rods; aggregates are formed by the association of these rods. The polysaccharides appear to have "restricted" length.


Strucutres of the capsular polysaccharides of Neisseria meningitidis as determined by 13C-nuclear magnetic resonance spectroscopy. The application of 13C-nuclear magnetic resonance spectroscopy to the structural determination of the capsular polysaccharide antigens of Neisseria meningitidis is described. Complete assignments of the spectra of the polysaccharides of serogroups A, B, C, W-135, X and Y were made and were based mainly on previous assignments made for the monomer units of the respective polysaccharides. This technique provides information on all structural aspects of the polysaccharides including composition, mode of linkage, location of O-acetyl substituents, sequence, and conformation. In addition, nuclear magnetic resonance is nondestructive, extremely rapid, and has enormous potential in bacteriological research.


Role of protein serotype antigens in protection against disease due to Neisseria meningitidis. Evidence is provided for the protective effects of antibody to noncapsular antigens in immunity to meningococcal disease. Serotype 2 strains were associated with 50% of disease due to group B Neisseria meningitidis in Europe. The amount of serotype antibody in normal individuals is related to age, and adult levels are reached by about 10 years of age. Children under two years of age who have recovered from disease due to groups B and CN. meningitidis of serotype 2 make antibody to serotype 2; this fact indicates the potential immunogenicity of the serotype antigen in young children. The isolated serotype 2 antigen injected into rabbits elicited bactericidal antibody that was only weakly opsonic. Guinea pigs that were infected via spring implants with serotype 2 N. meningitidis developed antibody to serotype 2. After clearance of infection these animals became resistant to challenge with serotype 2 strains of groups B and C N. meningitidis. This serotypes-specific immunity to reinfection persisted for at least four months.


Quantitative measurement of the bactericidal capability of neutrophils from patients and carriers of chronic granulomatous disease. Previous studies examined the ability of polymorphonuclear leukocytes (PMN) from patients and carriers of chronic granulomatous disease (CGD) to kill small numbers of bacteria in vitro. The purpose of the present study was to accurately measure the number of microorganisms killed by CGD PMN when the initial number of bacteria per neutrophil was increased. Bactericidal capacities of PMN from seven patients and five proven carriers of CGD were examined with five test ratios ranging from 1.25 to 100 bacteria/neutrophil. CGD PMN killed a limited number of catalase-positive S. aureus but the number of bacteria killed increased progressively as the initial test ratio was advanced. At the highest ratio (100:1) patient PMN killed a mean of nine S. aureus per cell (normal 48 +/- 13.2 SD). PMN from patients with CGD killed increasing numbers of catalase-negative S. faecalis with normal efficiency, reaching a mean of 14 S. faecalis killed per cell at 100:1. Individual variation in the ability of CGD PMN to kill S. Aureus was apparent but the degree of PMN bactericidal effectiveness did not correlate with the patient's clinical course. PMN from asymptomatic carriers of proven sex-linked CGD also had a wide spectrum of staphylocidal activity which ranged from normal to levels comparable to CGD patients.


Audiometric findings in 34 patients with Waardenburg's syndrome. The dominant hereditary form of deafness as part of Waardenburg's syndrome was found in 12 (35%) of 34 patients never previously described with Waardenburg's syndrome among 5 families in the Netherlands. Extensive audiometric examination could be performed in 11 patients, 5 patients with deafness on both sides and 6 patients with hearing deficiency on one side. The hearing loss could be divided into 4 types: (type I) (sub) total deafness on both sides; (type II) serious lack of hearing on both sides; (type III) (sub) total deafness on one side; (type IV) moderate hearing deficiency on one side, particularly in the low frequencies. An extensive review was made of the more than 1,000 patients with Waardenburg's syndrome described in the literature. This revealed that deafness in both ears (the most serious expression of the syndrome) occurred in about 25% of the patients with Waardenburg's syndrome type I (including dystopia canthorum) and in 50% of the patients with Waardenburg's syndrome type II (without dystopia canthorum). This striking difference is connected with the close relationship between deafness and pigmentary disorders.


Deafness, ichthyosiform erythroderma, corneal involvement, photophobia and dental dysplasia. Early childhood deafness, congenital non-bullous ichthyosiform erythroderma, corneal involvement, photophobia, chronic blepharoconjunctivitis, hypotrichosis, anhidrosis, hyperkeratosis of the nails and dental dysplasia were the symptoms observed in the 19-year-old girl described in this paper. The literature comprises reports on 8 patients with this syndrome, which McKusick (1975) listed as no. 24215.


Methods for the study of antidiarrheal agents. Study of commonly used protective and adsorbent agents. Several agents were employed to induce diarrhea in squirrel monkeys: (1) diarrhaogenic diets, (2) various doses of cholera toxin, (3) prostaglandin derivatives, (4) bile, (5) lactulose, (6) phenolphthalein, (7) castor oil. Kaolin, pectin, Kaopectate and placebo were used as antidiarrheal treatment. Evaluation was based on (a) frequency, (b) consistency, (c) total and dry weight of the stools, and (d) electrolyte loss. The spectrum of procedures employed appeared to be suitable for the evaluation of antidiarrheal agents of the protective and adsorbent class.


Isolated hapten-binding receptors of sensitized lymphocytes. II. Receptors from nylon wool-enriched rabbit T lymphocytes lack serological determinants of immunoglobulin constant domains but carry the A locus allotypic markers. Hapten-binding receptor material was isolated from sensitized rabbit lymphocytes by a method described previously for murine receptor material. The material was separated into a fraction expressing immunoglobulin determinants (anti-Ig+ fraction) and a fraction lacking known class and type-specific determinants of Ig constant domains (anti-Ig- fraction). We present evidence in support of the notion that--in analogy to the mouse system--the anti-Ig+ fraction is B-cell-derived, whereas the anti-Ig- fraction originates from T lymphocytes. Receptors of the anti-Ig- phenotype are found to express a locus allotypic determinants and, thus, appear to carry variable portions of immunoglobulin heavy chains.


[The use of primate tissue homogenates for quality control in the creatine kinase-MB determination with inhibiting antibodies (author's transl)]. The creatine kinase isoenzymes CK-MM, CK-MB and CK-BB from Rhesus monkey heart and brain tissue homogenates were separated chromatographically. Thereafter it could be demonstrated that the activity of the simian subunit CK-M is completely inhibited by anti-inhibitory-CK-M serum. Thus control sera from simian tissue are in principle suited for quality control in an immunological determination of creatine kinase-MB. The intra-assay variance and interassay variance were n = 56, -/x = 29.2 U/1, SD = 3.2 U/1, CV = 11.1% and n= 12, -/x = 166.7 U/1, SD = 5.0 U/1, CV = 3.0% respectively. It is desirable to develop control sera with catalytic concentrations of creatine kinase-MB in a lower range.


[Occurrence of purine deoxyribosides in urine (author's transl)]. It was shown that purine deoxyribosides are normal constituents of human urine. Healthy probands excrete about 25 microgram (41% of the total urinary deoxyribosides) per 24 h. Purine deoxyribosides were determined as the acid-labile fraction of the total deoxyribosides by the microbiological assay with Lactobacillus acidophilus R 26.


Diffusion of carbon dioxide through lipid bilayer membranes: effects of carbonic anhydrase, bicarbonate, and unstirred layers. Diffusion of (14)C-labeled CO(2) was measured through lipid bilayer membranes composed of egg lecithin and cholesterol (1:1 mol ratio) dissolved in n-decane. The results indicate that CO(2), but not HCO(3-), crosses the membrane and that different steps in the transport process are rate limiting under different conditions. In one series of experiments we studied one-way fluxes between identical solutions at constant pCO(2) but differing [HCO(3-)] and pH. In the absence of carbonic anhydrase (CA) the diffusion of CO(2) through the aqueous unstirred layers is rate limiting because the uncatalyzed hydration-dehydration of CO(2) is too slow to permit the high [HCO(3-)] to facilitate tracer diffusion through the unstirred layers. Addition of CA (ca. 1 mg/ml) to both bathing solutions causes a 10-100-fold stimulation of the CO(2) flux, which is proportional to [HCO(3-)] over the pH range 7-8. In the presence of CA the hydration- dehydration reaction is so fast that CO(2) transport across the entire system is rate limited by diffusion of HCO(3-) through unstirred layers. However, in the presence of CA when the ratio [HCO(3-) + CO(3=)]:[CO(2)] more than 1,000 (pH 9-10) the CO(2) flux reaches a maximum value. Under these conditions the diffusion of CO(2) through the membrane becomes rate limiting, which allows us to estimate a permeability coefficient of the membrane to CO(2) of 0.35 cm s(-1). In a second series of experiments we studied the effects of CA and buffer concentration on the net flux of CO(2). CA stimulates the net CO(2) flux in well buffered, but no in unbuffered, solutions. The buffer provides a proton source on the upstream side of the membrane and proton sink on the downstream side, thus allowing HCO(3-) to facilitate the net transport of CO(2) through the unstirred layers.


Juvenile Gaucher's disease with horizontal gaze palsy in three siblings. Three children in a West African family had Gaucher's disease of juvenile onset (Type 3), and all showed an identical neurological disorder. The diagnosis was substantiated by histochemical demonstration of Gaucher cells in bone marrow, liver, and spleen, the finding of an excess of glucosyl ceramides in a liver extract, and a deficient activity of the enzyme beta-glucosidase in cultured skin fibroblasts. The neurological picture was characterised by myoclonic epilepsy, muscle wasting, hypotonia, pyramidal signs, some intellectual deterioration, and a striking disturbance of eye movements. The latter appears to result from specific involvement of the supranuclear pathways subserving lateral gaze. The distinctive features of this clinical syndrome are emphasised.


Proprioceptive pathways of the spinal cord. In the Macaque, surgical lesions were made in the dorsal funiculus, in the dorsolateral funiculus, and through half of the spinal cord. The somatosensory and motor capacity of the animal were examined neurologically and electrophysiologically. The exact lesion was then confirmed pathologically in detail. The results of these experiments indicate that limb position information from the distal limb and proximal limb are relayed to the brain in two different fashions. Distal limb position information, especially the cortical representation of the limbs' volar surface as it moves in space, is drastically impaired by dorsal funiculus or posterior white column lesions. Proximal limb position may or may not be impaired by similar lesions, for this information while initially in the dorsal or posterior white columns is sorted out (as it ascends in the spinal cord) to the dorsolateral funiculus or white columns. For example, in the lower thoracic spinal cord, both distal and proximal hind limb sensation are impaired by posterior white column damage; in the cervical cord, only distal sensation is impaired by the same lesion, and proximal information is spared. We refer to this neuroanatomic rearranging as "fibre sorting", and we believe that it is clinically significant in spinal cord disease.


Blood flow and oxygen consumption of the focally traumatized monkey brain. The exposed left superior frontal gyrus of the anesthetized macaque brain was focally traumatized by a jet of compressed air. Focal blood flow in tissue around the lesion and total cerebral blood flow was determined before and during the 4 hours after trauma by the hydrogen clearance technique. Blood flow fell in tissue adjacent to the injured brain but the reduction was not statistically significant. Total cerebral blood flow, blood flow in the right superior-frontal gyrus, and oxygen consumption of the brain was unaffected by the trauma. The authors conclude that neither spreading ischemia within uninjured tissue surrounding focally traumatized brain nor posttraumatic diaschisis is readily provoked in the anesthetized brain of the monkey.


Electrophysiological studies of long-term electrical stimulation of the cerebellum in monkeys. Six rhesus monkeys were stimulated on the paravermal cortex for 205 hours (18 days) with different charge densities in order to determine the electrode performance and neural damage that may result from long-term cerebellar stimulation comparable to that being used in man. The electrode-tissue interface was relatively stable and no neural damage was found when the charge/phase (0.5 muC/ph) or charge density (7.4 muC/sq cm/ph) was very low. At all higher charge levels tested (2.4, 4.8, 10, and 22 muC/ph), changes in the electrode-tissue interface, meningeal encapsulation, and neural damage were directly related to the charge density delivered. Unstimulated electrodes on the opposite paravermal cortex exhibited mild tissue reactivity and cell damage, probably due to mechanical compression of the molecular layer and pial vessels. Motor cortex field potentials could be evoked by charges as low as 0.1 muC/ph delivered to paravermal cortex; for a given charge/phase longer pulses were more effective than short pulses. After neural damage resulting from 205 hours of 4.8 muC pulses at 10 per second (total charge 14.76 C), the threshold for the motor cortex evoked potential increased by a factor of four or more. With the charge held constant to different-sized electrodes placed bilaterally in the same monkey, damage was greater under the smaller electtrode. This finding suggests that the charge density to cerebellar cortex must be controlled to avoid neural damage.


Tissue reactions to long-term electrical stimulation of the cerebellum in monkeys. Light and electron microscopic analyses were carried out on the stimulated and unstimulated paravermal cortices of six rhesus monkeys that had electrodes implanted on their cerebella for 2 months. The electrodes and the stimulation regime (10 p.p.s.: 8 min on, 8 min off) were similar to those used to stimulate the human cerebellum for treatment of certain neurological disorders. Mere presence of the electrode array in the posterior fossa for 2 months resulted in some meningeal thickening, attenuation of the molecular layer, and loss of Purkinje cells immediately beneath the electrode array. There was no evidence of scarring. After 205 hours of stimulation (7.35 X 10(6) pulses) over 18 days, a charge of 0.5 muC/ph or estimated charge density of 7.4 muC/sq cm/ph resulted in no damage to the cerebellum attributable to electrical stimulation per se. Such a charge/phase is about five times the threshold for evocation of cerebellar efferent activity, and might be considered "safe" for stimulation of human cerebellum. Charge density/phase and charge/phase were directly related to increased cerebellar injury in the six other cerebellar cortices stimulated. Leptomeningeal thickening increased with increased charge density. Injury included severe molecular layer attenuation, ongoing destruction of Purkinje cells, gliosis, ongoing degeneration of myelinated axons, collagen intrusion, and increased levels of local polysaccharides. In all cases, even with damage that destroyed all conducting elements beneath the electrodes, there was no damage further than 1 to 2 mm from the edges of the electrode arrays.


Effect of ethane-1-hydroxy-1,1-diphosphonate (EHDP) and dietary fluoride on biomechanical and morphological changes in chick bones. There are only a few papers in the literature in which parameters related to bone strength are examined over wide range of fluoride concentrations. In this report, chicks were used and fed diets containing fluoride added at 0, 50, 200, or 800 ppm in preparation for measurements on bone strength. The diets were fed for 20 days. After 10 days, the birds were also injected daily with solutions containing either isotonic saline or 5 mg of ethane-1-hydroxy-1,1-diphosphonate (EHDP) as phosphorus/kg body weight or 10 mg of EHDP as phosphorus/kg body weight. The effects of added fluoride and EHDP on bone morphology and the biomechanical properties of bone were then examined. The rate of bone formation was assessed using histological and tetracycline labeling techniques. Administration of high fluoride and/or EHDP resulted in significant changes in bone morphology. The changes effected by the administration of EHDP appeared independent of fluoride induced changes. Biomechanical studies also indicated that both high levels of fluoride and EHDP caused a significant reduction in bone torsional breaking strength. The reductions in breaking strength were independent of changes in body size.


Kerosene absorption in primates.

Kerosene absorption in primates. Kerosene, labeled with 3H-toluene or 14C-hexadecane, was given to baboons by nasogastric tube after a tracheostomy had been performed. Six hours later the animals were killed and samples of tissues taken for analysis. The radioactive label was recovered from all the tissues analyzed. 3H-toluene appeared to be absorbed and taken up by most tissues to a greater extent than was 14C-hexadecane. No abnormal neurologic signs or behavior was noted in the baboons which were conscious during the study period. It appears that primates absorb kerosene from the gastrointestinal tract, but the volumes are very small and do not cause gross neurologic signs.


Plexiform neurofibromatosis of the ileum in an infant. An infant is reported in whom severe malabsorptive symptoms developed shortly after birth associated with intermittent episodes of intestinal obstruction. Plexiform neurofibromatosis involving the terminal ileum was found at laparotomy at the age of 6 mo. The infant died from gastrointestinal functional impairment at the age of 1 yr.


Application of proton and 13C-NMR spectroscopy to estimation of diastereoisomer ratio in phenethicillin. A procedure for the determination of the diastereoisomer ratio in phenethicillin potassium and its formulations is described. The method utilizes analyses of PMR and 13C-NMR data. Assignments of 13C-chemical shifts in D- and L-phenethicillin potassium also are included.


Factors influencing decomposition rate of amitriptyline hydrochloride in aqueous solution. The degradation rate of amitriptyline hydrochloride in buffered aqueous solution containing various additives was determined. The oxidation was a free radical-mediated process, and the rate was accelerated by the presence of metal-ion contaminants. Glass ampuls, particularly amber ones, in which the solutions were stored were the major source of these contaminants. Edetate disodium stabilized the solution, but the primary antioxidants propyl gallate and hydroquinone were less effective. Sodium metabisulfite accelerated the decomposition, and it is postulated that there was direct attack by metabisulfite at the olefinic double bond in the drug molecule.


Dissolution rates of model gallstones in human and animal biles and importance of interfacial resistance. Cholesterol monohydrate dissolution kinetics in human gallbladder bile were studied to determine the magnitudes of the in vitro dissolution rates, the rate resistances in human gallbladder bile, and the extent that the interfacial resistance is the rate-determining factor. Dissolution rate studies also were conducted using human duodenal bile and animal bile for comparison. The dissolution rate resistance, R, ranged from 10(4) sec/cm for chicken bile to 10(4)-10(6) sec/cm for human bile. Interfacial resistance was the rate-determining factor for essentially all results. Where chemical composition data were obtained, the R values for the human bile samples were consistent with predictions made from the simulated bile studies. In two human gallbladder specimens having low bile acid-lecithin molar ratios (i.e., 2.9 and 2.3), very high R values of 1.9 X 10(5) and 4.1 X 10(5) sec/cm were found. These values were in good agreement with the findings in the simulated bile studies and suggest that stone dissolution in patients with low bile acid-lecithin ratios may proceed very slowly, even when the bile is highly undersaturated with respect to cholesterol.


Time-dependent kinetics III: diurnal oscillations in steady-state plasma valproic acid levels in rhesus monkeys. Valproic acid was administered by constant rate intravenous infusion to catheterized chained rhesus monkeys for 8-10 weeks under controlled environmental conditions. Steady-state plasma levels were monitored at 2-hr intervals for 26 hr (10 am-12 noon on the following day), 1 day/week for 6 weeks. Individual steady-state plasma concentration-time plots exhibited the following characteristics. During Period A (10 am-6 pm), plasma levels remained stable or decreased. During Period B (6 pm-6 am), plasma levels increased, reached a maximum, and remained markedly higher than during Period A. The maximum concentrations were 40-140% higher than the observed minimum concentrations. During Period C (6 am-noon), plasma levels tended to decline from the maximum concentrations achieved in Period B. In most cases, plasma concentrations at 10 am and 12 noon of the 2nd experimental day fell within 10% of their respective values on the previous day. The mean (+/- SD) periods obtained by cross-correlation analysis of individual plasma concentration-time plots were 30.7 (+/- 3.7) and 22.8 (+/- 3.6) hr for Animals 903 and 923, respectively. The corresponding mean (+/- SD) amplitudes were 27.3 (+/- 12.6) and 17.4 (+/- 2.3)%. A circadian rhythm in total body clearance was hypothesized, and its pharmacokinetic implications are discussed.


Quantification of lidocaine and several metabolites utilizing chemical-ionization mass spectrometry and stable isotope labeling. Quantification of the suspected metabolites of lidocaine in humans was carried out using the direct insertion probe and chemicalionization mass spectrometry. Deuterated analogs of the metabolites of lidocaine were added to serial human plasma and urine samples and were used as internal standards following oral administration of 250 mg of lidocaine hydrochloride monohydrate to two male subjects and 202 mg of lidocaine free base to one male subject. The average results after analysis of the 0-24 hr urine samples, before beta-glucuronidase-sulfatase treatment, indicated the presence of seven of the possible metabolites in the following amounts (percent of administered dose based on the free base): lidocaine, 1.95; omega-ethylamino-2,6-dimethylacetanilide, 4.90; omega-amino-2,6-dimethylacetanilide, 0.88; m- and/or p-hydroxylidocaine, 0.73; m- and/or p-hydroxy-omega-ethylamino-2,6-dimethylacetanilide, 0.56; 2,6-dimethylaniline, 0.97; and 4-hydroxy-2,6-dimethylaniline, 63.5. Both N-ethyl- and N,N-diethylglycine were detected in human and Rhesus monkey urine, although quantification was not achieved.