Counterimmunoelectrophoresis in determination of prostatic acid phosphatase in human serum. We evaluated counterimmunoelectrophoresis for use in measuring prostatic acid phosphatase in detection of prostatic cancer. After staining for acid phosphatase, we could detect as little as 0.3 ng of purified enzyme standard complexed with antibody by this technique. However, when serum samples were used as antigen, the method was less sensitive (1.5-2.0 ng) because some of the serum proteins migrate with the phosphatase and decrease the intensity of the stain for acid phosphatase. For this reason we could not detect the phosphatase in serum samples of normal persons; only patients with moderately (or greater) increased activity in their serum showed positive results. In contrast, by radioimmunoassay as little as 1.0 ng of the phosphatase can be detected in serum.


Evaluation of the Harleco Micro CO2 system for measurement of total CO2 in serum or plasma. In the Harleco Micro CO2 System a volumetric procedure is used to measure total CO2 content in a small volume (0.10 ml) of serum or plasma. CO2 gas is released by mixing specimen and acid, either manually or with a "Micro CO2 Shaker." Within-batch CV's by manual and machine shakings were, respectively, 7.7 and 5.6% for a total CO2 value of 8 mmol/liter, decreasing to 2.0 and 1.4% for a value of 45 mmol/liter. Between-batch CV's were 8.0 and 7.1% at low total CO2, decreasing to 2.4 and 2.0% at high total CO2. Measurements were linear for values from 5 to 60 mmol/liter. Analytical recoveries at low and high concentrations were 97.7 to 98.9%. Correlation coefficients for results obtained with the Technicon "SMA Plus" autoAnalyzer and the Radiometer ABL 1 blood gas analyzer were 0.992 and 0.996, respectively. The Harleco Micro CO2 System appears to be a quick, reliable, and inexpensive method for measuring serum or plasma total CO2.


The peak monitor of the Technicon SMAC system. The peak-monitor software of the Technicon SMAC system has been investigated by experiments designed to test its ability to detect abnormally shaped peaks, deliberately produced by presenting serum samples of inadequate volume for analysis. The software provided by the manufacturer is shown to detect faulty peaks inefficiently, and a set of modified parameters has been identified which greatly improves the performance of the peak-monitor software.


Simple determination of high urinary excretion of 5-hydroxyindole-3-acetic acid with ferric chloride. We describe a method for determination of urinary 5-hydroxyindole-3-acetic acid, a metabolite of serotonin and tryptophan and a useful indicator of metastasizing carcinoid tumor. The analysis is based on the color development with ferric chloride added to the residue obtained on evaporating a diethyl ether extract of the sample that has been acidified with sulfuric acid and saturated with sodium chloride. The color is measured by 510 nm. The reaction was made highly specific by the acidification so that nonspecific reactions with catecholamines and with other phenolic compounds such as salicylic acid could be eliminated.


Plasma glucose measurement with the Yellow Springs Glucose Analyzer. The Yellow Springs Glucose Analyzer, a device for the quantitative measurement of glucose concentrations, involves the use of immobilized glucose oxidase, incorporated on a membrane covering a hydrogen peroxide sensor. Operation of the instrument is simple. After an initial calibration, 25 microliter of plasma is injected into a reaction chamber. At 45 s a digital result for glucose is displayed. The within-batch coefficient of variation for the method is 1.2% or less for glucose concentrations of 0.94 to 3.98 g/liter. The between-batch coefficient of variation is 5.8% or less for glucose concentrations of 0.29 to 2.91 g/liter. Concentration and readout are linearly related to at least 4.6 g/liter. Analytical recoveries ranged from 100 to 102%. Carry-over was negligible. Values for glucose concentration obtained with the instrument compared well (r = 0.997) with those obtained with the Beckman Glucose Analyzer.


Variability among commercially available digoxin radioimmunoassay kits in cross reactivity to dihydrodigoxin. We evaluated four commercially available 125I-digoxin radioimmunoassay kits with regard to their ability to cross react with the digoxin metabolite dehydrodigoxin. We prepared dihydrodigoxin serum samples in digoxin-free serum over the concentration range 0.4 to 5.0 microgram/liter and assayed them with each kit according to the manufacturer's instructions. The metabolite was able to displace the 125I-labeled digoxin derivative from the antibody supplied with all four kits. However, the extent of the cross reactivity depended on the kit, ranging from essentially zero to a high degree of interference. Dihydrodigoxin is the only metabolite of digoxin to have been quantitiated in human serum, and may comprise up to 30% of total glycosides. Over the clinical and therapeutic range of serum digoxin concentrations, enough dihydrodigoxin can be produced to interfere in the determination of serum digoxin concentrations by this method. We suggest that laboratories evaluate their specific kit with regard to cross reactivity to this metabolite.


Urinary free methyldopa determined by reversed-phase high-performance liquid chromatography. We describe a chromatographic method, in which 3,4-dihydroxybenzylamine is used as the internal standard, for determining free methyldopa in human urine. The drug was adsorbed onto alumina, eluted, and the eluate directly injected onto a reversed-phase column (octadecyl-bonded silica stationary phase), with dilute acetate buffer (pH 2.7) as the mobile phase and ultraviolet detection at 280 nm facilitated. Methyldopa is well separated from other urinary biogenic amines present in the alumina extract, and other commonly used antihypertensives and diuretics do not interfere with the analysis. The sensitivity of the method is adequate to quantify 8.0 mg of methyldopa per liter in 30 ml of sample; the lower limit of detection is 25 ng. Analytical recovery for methyldopa varied from 95 to 102% with within-run and day-to-day coefficients of variation of 2.7 (n = 10) and 3.8% (n = 5), respectively. This procedure is readily adaptable for use in studies of the pharmacokinetics of methyldopa and to routine clinical laboratory use.


Automated procedure for kinetic measurement of total triglycerides (as glycerol) in serum with the Gilford System 3500. An automated procedure is used on the Gilford 3500 Computer-Directed Analyzer to measure serum triglycerides indirectly by using aqueous glycerol standards. Most enzymatic methods require long hydrolysis or awkward saponification. The method of Bucolo and David [clin. Chem. 19, 476 (1973)], in which lipase and glycerol kinase are used, is modified. The kinetic procedure described eliminates the need for a serum blank. It uses "Eskalab" bulk reagents and reduces both time and cost per test by measuring the decreasing NADH concentration from the glycerol kinase reaction at 340 nm after enzymatic hydrolysis at room temperature. The change in absorbance of the standard during a 14-s measuring time is used in the ratiometric calculation of the unknowns. The use of a stable aqueous standard that can be reliably and accurately prepared makes this method ideal for the traceability desired by many organizations. Reagent blank drift did not affect the results. A correlation coefficient of 0.991 for comparison with the manual endpoint method and a typical CV of 2.25% show this method to be accurate and reliable.


Improved radiochemical assay for carnitine and its derivatives in plasma and tissue extracts. We describe the measurement of carnitine and short-chain and long-chain acylcarnitines in perchloric acid extracts of liver and muscle. [1-14C]Acetyl-CoA is used, and labeled acetylcarnitine is measured with a sensitivity of less than 1 to 50 nmol of L-carnitine per 50-microliter sample. Samples frozen at -20 degrees C lost no carnitine during several months. Analytical recovery of carnitine (5 nmol) added to 50-microliter perchloric acid extracts of liver was 103 +/- 2%. This method may be suitable for clinical applications such as screening patients with muscle and metabolic disorders.


Sensitive gas-chromatographic determination of fluphenazine in human plasma. We describe a method for quantitative determination of fluphenazine in human plasma, based on gas chromatography of its trimethylsilyl derivative and the use of a structurally similar internal standard. We determined its concentrations in plasma of four patients who were receiving the drug intramuscularly as its decanoate, in doses ranging from 12.5 to 250 mg; the concentrations in the plasma of these patients ranged from 0.25 to 11.8 microgram/liter. The precision (CV) of the method over the range 2 to 17 microgram/liter was 6.8% when fluphenazine was extracted from 5 ml of plasma. Of 18 other drugs tested, only thioridazine and the cis and trans forms of flupenthixol interfere with the determination.


Analysis for 1,3-bis(2-chloroethyl)-1-nitrosourea by chemical ionization mass spectrometry. We used chemical ionization mass spectrometry to quantify the cancer chemotherapeutic agent 1,3-bis(2-chloroethyl)-1-nitrosourea in biological fluids. A stable isotope-labeled internal standard was added to the fluid being analyzed. Protonated molecular ion intensity ratios were determined by selected ion monitoring of sample extracts inserted directly into the chemical ionization source. Peak ratios were calculated for chlorine isotope ratios in their natural abundance, to verify the integrity of the data acquired, and to compute the drug concentration. The assay was applied to measure the compound in rat blood and plasma and in human plasma, and to monitor in vitro metabolism in rat-liver microsomal preparations. For the analyses we used a quadrupole mass spectrometer interfaced to a commercial computer system. Data-system characteristics are described.


Gas-liquid chromatographic determination of nicotine and cotinine in plasma. We report a procedure for determining nicotine and cotinine in plasma. Nicotine is extracted from 1 ml of plasma with diethyl ether, back extracted, and analyzed by gas-liquid chromatography with a nitrogen/phosphorus detector. Nicotine and its internal standard, modaline, had retention times of 1.9 and 2.9 min, respectively. Cotinine is then extracted from the same plasma with dichloromethane and similarly analyzed. Cotinine and its internal standard, lidocaine, had retention times of 3.8 and 4.9 min. Day-to-day reproducibilities (CV) within 14% for nicotine and within 6% for cotinine are attainable for the respective concentration ranges 1-100 microgram/liter and 1-200 microgram/liter. Nornicotine and related alkaloids do not interfere. The sensitivity was such that less than 0.1 microgram (0.62 nmol) of nicotine and 0.1 microgram (0.62 nmol) of nicotine and 0.1 microgram (0.57 nmol) of cotinine could be detected per liter.


New automated dye-binding method for serum albumin determination with bromcresol purple. We describe a new automated dye-binding method for serum albumin determination with bromcresol purple (BCP) that has several advantages over an existing bromcresol green (BCG) method. The continuous-flow method is sensitive, linear, and precise, with negligible sample interaction at an analytical rate of 60 samples per hour. Unlike BCG, BCP did not react with an albumin-free serum globulin preparation or pure human transferrin solutions. Reaction with serum was instantaneous; in contrast, BCG exhibits a slow nonspecific reaction with some specimens. The specificity of BCP was demonstrated by good agreement with results of "rocket" immunoelectrophoresis (EIA) where y(BCP) = 0.95X(EIA) + 1.72. The BCG method overestimated serum albumin concentration where y(BCG) = 1.01X(EIA) + 6.77. Precipitation, which affects the BCG method, was not observed with BCP. Blank corrections were negligible, salicylate did not interfere, and bilirubin affected the method only if present in very high concentration. The method offers a solution to the poor accuracy of existing BCG methods while retaining many of their desirable features.


Liquid-chromatographic procedure for tricyclic drugs and their metabolites in plasma. We describe a procedure for determining amitriptyline and imipramine and their active metabolites nortriptyline and desipramine, respectively, at therapeutic concentrations in human plasma by use of liquid chromatography. The drugs are extracted at pH 10.5 into hexane/isoamyl alcohol, which is evaporated and the residue chromatographed. Protriptyline is used as the internal standard. As little as 10 microgram of each drug per liter could be detected in plasma, the limit being established by variability in drug-free plasmas. The day-to-day coefficient of variation for each drug at a concentration of about 100 microgram/liter was about 7%. Doxepin and diphenhydramine interfere with the analysis of amitriptyline. Total analysis time for a single sample is 20 min.


Applications of a new instrument, the potentiophotometer. The Potentiophotometer is a new instrument for measuring the absorption characteristics of solutions. A brief description of its construction and theory of operation is presented, with illustrations of its use in the colorimetric quantitative analysis for inorganic phosphorus and total protein in serum. The potentiophotometric results correlate well (r = 0.97 to 0.98) with measurements made of the same solutions with either of two conventional spectrophotometers, the Gilford 300N and the Coleman Jr. II.


Polyamine compartmentalization in various human disease states. Whole blood was separated into preparations of erythrocytes, mononuclear leukocytes, polymorphonuclear leukocytes, platelets and plasma. Each preparation was analyzed for the concentration of the polyamines putrescine, spermidine and spermine. This was done in 17 controls, 14 patients with psoriasis, four patients with hereditary elliptocytosis, two patients with chronic lymphocytic leukemia and one patient each with lung cancer, non-Hodgkin's lymphoma, sickle cell anemia with mild psoriasis, and progeria. In patients with elevated blood polyamine levels, absorption onto erythrocytes was relatively common, and the spermine/spermidine ratio was useful in localizing abnormalities and characterizing the nature of the polyamine alteration. Proliferative states were associated with elevated spermine/spermidine ratios relative to controls while this relationship was reversed in erythropathies such as hereditary elliptocytosis and sickle cell anemia.


A practical routine gas chromatographic determination of phenobarbital, primidone and diphenylhydantoin. A gas chromatographic method for the routine determination of phenobarbital, primidone and diphenylhydantoin using alkaline extraction and on-column methylation has been developed. The procedure has been optimized for recovery of all three anticonvulsant drugs with particular attention to primidone. Correlation with photometric methods for phenobarbital and diphenylhydantoin show the technique offers significant advantages in both accuracy and precision over other methods.


Doubling time alpha-aminoisobutyrate transport and calcium exchange in cultured fibroblasts from cystic fibrosis and control subjects. Population doubling time, kinetics of transport of alpha-aminoisobutyrate (AIB) and calcium (Ca) exchange were studied in skin fibroblast monolayers obtained from 5 subjects with cystic fibrosis (CF) and 5 age- and sex-matched controls. Population doubling time as estimated from cell count, protein and DNA was no different in the two groups. KM, Vmax, maximal uptake and time of half maximal uptake of AIB were no different in the two groups. Intracellular Ca pool size based on exchange of 45Ca with unlabelled Ca was significantly greater in monolayers from CF subjects.


A simplified method for the determination of steroids in diets and faeces. A simplified method for the quantitative isolation and determination of sterols and bile acids for application in sterol balance studies in humans consuming mixed general food is described. The recovery of steroids from diets and from faeces was found to be quantitative. By application of temperature programmed gas-liquid chromatography (GLC), thin-layer chromatographic (TLC) pre-separation could be omitted from the multistep analytical procedure. Furthermore, it was possible to use simple internal standards, 5alpha-cholestane and/or 5beta-cholanic acid, instead of labeled compounds for the calculation of overall recoveries.


Human serum alpha-L-fucosidase.

Human serum alpha-L-fucosidase. Human serum alpha-L-fucosidase has been purified 241 200-fold with 35% yield by an affinity chromatographic procedure utilizing agarose-epsilon-aminocaproyl-fucosamine. Isoelectric focusing of the purified enzyme indicated the presence of several forms, with the form at pI 5.0 comprising the majority of the activity. Assay of the purified alpha-L-fucosidase showed only trace amounts of contaminating glycosidases present, with beta-galactosidase being the largest contamnant (0.5% by activity). Sodium dodecyl sulfate polyacrylamide gel electrophoresis indicated the presence of two subunits with very similar molecular weights (56 500 and 54 000). Using the p-nitrophenyl substrate, the purified serum alpha-L-fucosidase has an apparent Michaelis constant of 0.52 mM and a broad pH optimum centered around pH 4.8 with a second, minor optimum at pH 6.1. Gel filtration on Sepharose 6-B indicated an apparent molecular weight of 296 000 +/- 30 000. Preincubation with antibodies made previously against purified liver alpha-L-fucosidase led to quantitative immunoprecipitation of the purified serum alpha-L-fucosidase. Assay of the purified serum alpha-L-fucosidase for sialic acid indicated the presence of 1.7 microgram sialic acid per 100 microgram enzyme, about twice that previously found for the purified liver enzyme.


A selective method for determination of methylguanidine in biological fluids. Its application in normal subjects and uremic patients. A selective analytical method for the determination of methylguanidine in plasma in biological fluids has been developed. Methylguanidine is extracted in a column to dichloromethane as an ion pair with hexanitrodiphenylamine (dipicrylamine). It is isolated from coextracted compounds by partition chromatography as the picrate ion-pair. The methylguanidine fraction is collected and after reextraction to a buffer solution the methylguanidine content is quantitatively determined photometrically as picrate. An absolute recovery of 95 +/- 5% was obtained in the concentration range 1.5-10 microgram/ml plasma. The concentration of methylguanidine in plasma was higher in uremic patients, (44.4 +/- 5.71 mumol/l in conservatively-treated and 42.4 +/- 7.87 mumol/l in dialysis-treated patients) than in normal subjects, (4.0 mumol/l), but still lower than reported by other investigators using non-specific methods and also lower than the concentrations found to be toxic in experimental animals. There was a significant correlation between methylguanidine and creatinine concentration but no correlation between methylguanidine and urea concentration in plasma. No obvious relation was found between plasma methylguanidine concentration and various uremic symptoms, mode of treatment or protein intake.


A simplified radioimmunoassay procedure for serum progesterone. A simple, rapid radioimmunoassay for progesterone in unextracted serum is described. Cortisol is used to inhibit progesterone binding to corticosteroid binding globulin. The method has been optimised for the determination of progesterone in pregnancy sera or in the luteal phase of the menstrual cycle.


Prevalence of unsuspected urinary bacterial contamination: effects of screening tests for detection of inborn errors of metabolism. Hospitals and doctors submitting urine samples to a central reference laboratory screening for inborn errors of metabolism had been given precise instructions on sample handling designed to prevent growth of contaminating bacteria. 140 consecutive samples were examined bacteriologically and 22% had a heavy growth of bacteria (greater than 10(5) organisms/ml). Contamination was commonest in samples from children under 2 years. Normal human urine samples spiked with various amino acids were inoculated with pure cultures of different urinary pathogens and then incubated, with control samples, at 37 degrees C. Urinary amino acids were measured either semi-quantitatively by high-voltage electrophoresis or quantitatively by ion-exchange chromatography. Very wide-ranging changes in amino acid levels were noted in some inoculated samples after incubation. In two urine samples spiked with 8 mg/ml arginine, this had disappeared after 24-48 h. Unsuspected urinary bacterial contamination may be a more important source of false positive and false negative results in the investigation of disorders of amino-acid metabolism than previously supposed.


Dipeptidyl peptidases in human muscle disease. With the use of selective inhibitors of arylamidase, four dipeptyl peptidases (I, II, III, and IV) capable of hydrolyzing the beta-naphthylamides of Gly-Arg, Lys-Ala, Arg-Arg and Gly-Pro, respectively, were distinguished in homogenates of human muscle. Dipeptidyl peptidase I showed maximum activity at pH 5.0-6.0. Dipeptidyl peptidase II was maximally active at pH 5.0 and inhibited by cations. Dipeptidyl peptidases III and IV were most active at pH 8.5 and 7.5, respectively. When compared to controls, significant increases in muscle dipeptidyl peptidases I and II were observed in patients with muscular dystrophies and polymyositis. Dipeptidyl peptidase III was not altered in the neuromuscular disease examined. Dipeptidyl peptidase IV showed marked increase in a variety of muscle wasting conditions. The increase in dipeptidyl peptidases I and II may be attributed to lysosomal activation that is known to occur in conditions of muscle degeneration. However, the striking increase in a variety of muscle diseases of dipeptidyl peptidase IV, an enzyme shown to be associated with microsomal membranes in other tissues, suggest that in addition to lysosomes other sources also contribute to the total hydrolytic potential of diseased muscles. Dipeptidyl peptidases II and IV were found to be present in human serum. Their levels were not altered in serum of patients with Duchenne dystrophy.


Quantitative evaluation of insulin antibodies by radiochemical analysis. A method is described for determining insulin antibodies, and some applications are reported, concerning 45 patients, 30 with diabetic pathology, 15 of them receiving insulin. The diagnostic tool proved useful.


Heme synthetase activity in normal human and rat erythroid cells and in sideroblastic anemia. The enzyme heme synthetase, involved in the final step of the biosynthesis of heme, has been assessed in rat and human bone marrow and peripheral blood. The pH optimum of the enzyme in bone marrow appeared to be pH 7.6, whereas the Michaelis constant in human and rat bone marrow was found to be 1.6 micrometer and 0.6 micrometer, respectively. Rat reticulocytes showed approximately 100-times higher heme synthetase activities than did rat erythrocytes. By contrast, human reticulocytes did not show significantly higher activities than human erythrocytes. This difference between rat and human reticulocytes could be confirmed by in vitro experiments with intact cells in which iron uptake and heme synthesis of human and rat cells were compared. Finally, heme synthetase activity was assessed in bone marrow cells of two patients with sideroblastic anemia. In both cases the enzyme activities were found to be comparable to those in control bone marrow.


The effect of glucose oral administration on hyperammonemia in cirrhotics. In order to investigate the controverted effect of glucose on hyperammonemia the diet of eight advanced cirrhotics was supplemented hourly, between 9 a.m. and 5 p.m., with 20 g of glucose orally. Plasma insulin and arterial and median cubital venous ammonia levels were measured hourly and the results were compared to those of a control test performed in the same patients without glucose supplementation. In the control test the lunch (protein meal) induced an identical rise in arterial and venous ammonia levels (+40 +/- 3 and +36 +/- 5 microgram/100 ml, respectively). With glucose supplementation plasma insulin rose significantly and the arterial ammonia increase produced by lunch (+42 +/- 3 microgram/100 ml) did not differ from that observed in the control test; but the rise in venous ammonemia was lower (+12 +/- 4 microgram/100 ml; p less than 0.01) with a significant increase in arterio-venous ammonia difference. These results suggest that oral glucose administration increases the peripheral muscular ammonia uptake through a mechanism which remains to be elucidated but which is inefficient for arterial hyperammonemia.


Nepsilon-(beta-Aspartyl)lysinuria in children with various pathological conditions. The isolation, identification, and quantitative determination of an unusual urinary dipeptide, Nepsilon-(beta-aspartyl)lysine, is described, as well as its synthesis. This compound was observed in children in various disease states, but without any correlation with a particular symptom. Its origin is discussed.


A new method for determination of oxalate in blood serum by gas chromatography. A method for the determination of blood oxalate by gas chromatography is described. After clarification of the serum with acetone, the oxalate was extracted, ethylated and analysed on a gas chromatograph. The concentration of oxalate in the sample was determined by comparison with the elution pattern of diethylmalonate used as an internal standard. Using this technique, the recovery of added oxalate was more than 80%. Normal subjects were found to have blood oxalate levels of 27 mumol/l or less.


The variability of metabolite excretion in propionicacidaemia. Random urine samples from eight patients with propionicacidaemia were analyzed by gas chromatography and mass spectrometry in order to see if a consistent metabolite pattern with a high diagnostic value could be found. However, wide variations were observed. The presence of 3-hydroxypropionate and/or methylcitrate were considered to be diagnostic of propionyl-CoA carboxylase deficiency. In addition, samples from ketotic periods frequently contained 3-hydroxy-n-valerate and 3-oxo-n-valerate.


The design of a training program for adolescent medicine. Thoughts of fellows in training. Program design for training in Adolescent Medicine is still in an evolutionary state. Postgraduate fellows and program directors have different perceptions with regard to the importance of various components of the ideal training program. Fellows indicate a higher priority for problems associated with the female reproductive system than do their mentors. Inpatient experience received lesser emphasis by the Fellows. Adolescent Medicine as a subspecialty discipline is discouraged.


Diabetic day. Setting goals for a child-directed ambulatory program. The techniques of management by objective have been used to coordinate and evaluate the activities of a non-traditional program for the medical care and education of children with juvenile diabetes mellitus. "Diabetic Day" is an ambulatory child-directed clinic program in which health professionals work together to improve the child's ability to cope with diabetes.


Caring for parents of sick newborns.

Caring for parents of sick newborns. This paper is concerned with the care needed by parents who give birth to a "high risk" infant. Interviews with eight families demonstrate the diversity of responses to this anxiety-producing situation.


Who should accept primary responsibility for the encopretic child? A successful pediatric program based on dietary control, bowel training, and family counseling. Such a program was used to treat 18 encopretic children; 17 improved and 14 became completely free of soiling. Most encopretic children can be managed by pediatricians and only those children with severe emotional disorders require psychiatric referral.


The utility of clonazepam in epilepsy of various types. Observations with 22 childhood cases. Clonazepam, a new anticonvulsant, appears to be useful for childhood minor motor seizures and for petit mal refractory to Ethosuximide and Trimethadione. It appears less effective in infantile spasms though may be beneficial when there is no response to steroids. It is variably effective in partial complex and focal epilepsy and may exacerbate tonic seizures. A transient disadvantage is the high incidence of side effects, especially lethargy and ataxia, though these may be transitory. Aggressivity and hyperkinesis may necessitate medication withdrawal. Some children who initially respond to therapy and then relapse may respond again to a higher dosage.


Trichorhinophalangeal dysplasia (Giedion syndrome). A case report. This rare syndrome is characterized by short stature, sparse, fine scalp hair, a pear-shaped nose, long philtrum, normal intelligence, and cone-shaped epiphyses of the phalanges with deformities of the fingers.


A psychological perspective for double-blind trials. Blind breaking is likely to occur in a long-term clinical trial involving subjects at risk. Double-blind conditions require subjects to live with ambiguity regarding their health status. It is unrealistic to expect them to tolerate this ambiguity over an extended period of time. A logical step for a subject is to break the blind and surreptitiously seek the best medical care available. Moreover, the staff assigned to monitor protocol adherence may further contribute to blind breaking because of ambiguity regarding their proper clinical role. Several steps are recommended to manage blind breaking within a trial, including improving the quality of medical care, more rigorous screening procedures, and maintaining a clinical environment that promotes accurate disclosure of blind breaking.


Hemodynamic effects of trimazosin in patients with left ventricular failure. Trimazosin is a new quinazoline derivative vasodilator previously thought to act primarily on resistance vessels. Vasodilators characteristically improve hemodynamics in patients with left ventricular failure. Trimazosin in single oral doses of 100 to 300 mg was given to 6 patients with class III-IV congestive heart failure due to ischemic or primary cardiomyopathy and hemodynamics were monitored for 4 hr. Onset of action was apparent at 1 hr, with peak effects occurring at 1 to 3 hr and significant effects persisting at 4 hr. At peak, mean systemic arterial pressure fell from 91.2 +/- 4.8 (SEM) to 82.5 +/- 2.5 mm Hg (p less than 0.05), heart rate was unchanged, and cardiac index (by dye dilution) was up to 2.12 +/- 0.27 from 1.76 +/- 0.33 L/min/m2 (NS). Right heart pressures (by Swan-Ganz catheterization) were reduced, right atrial, 11.3 +/- 2.1 to 8.7 +/- 1.6 mm Hg (p less than 0.02), mean pulmonary arterial pressure, 40.7 +/- 4.6 to 33.8 +/- 4.8 mm Hg (p less than 0.01), and pulmonary arterial wedge, 30.7 +/- 4.5 to 23.8 +/- 5.0 mm Hg (p less than 0.01). Forearm venous capacitance, by occlusion plethysmography, increased from 0.83 +/- 0.18 ml/100 gm to 1.12 +/- 0.22 ml/100 gm after trimazosin (p less than 0.01). Trimazosin is an orally effective vasodilator that acts on both resistance and capacitance vessels in patients with congestive heart failure.


Serum digoxin and empiric methods in identification of digitoxicity. A prospective study of consecutive patients admitted to a medical service was undertaken to evaluate the prevalence of cardiac digitalis toxicity and the relative diagnostic values of serum digoxin versus an empiric method, based on calculations of digoxin dose in relation to kidney function and body weight. Of 711 patients admitted, 109 were treated with digitalis on admission. Sixteen of the patients developed cardiac arrhythmias consistent with digitalis intoxication. Five of these, none with serum digoxin above 1.6 ng/ml, were not toxic. The remaining eleven patients, all with serum digoxin levels above 1.6 ng/ml, were either definitely or possibly toxic. A similar borderline between intoxicated and nonintoxicated patients could not be established on the basis of calculations based on body weight and renal function. In all cases in which suspicion of digitalis intoxication was raised, serum digoxin measurements could discriminate between the toxic and the nontoxic patients.


Determination of procainamide acetylator status. Variation in renal function can obscure the measurement of acetylator status for compounds such as procainamide in which appreciable active drug or acetyl metabolite is excreted in urine. Computer simulations and patient studies were used to compare two common metabolite/drug ratio methods and a proposed clearance technique for phenotyping acetylation rate. The calculation of apparent acetylation clearance from steady-state serum concentrations of procainamide and urinary excretion rates of N-acetyl-procainamide provides the most definitive discrimination between fast and slow acetylators.


Binding of quinidine to plasma proteins in normal subjects and in patients with hyperlipoproteinemias. Quinidine in the plasma binds to various lipoproteins as well as to albumin. With the use of computer simulations the effect of varying the serum level of each of the lipoproteins as well as albumin was examined. The results suggested that the overall binding of quinidine is relatively insensitive to changes in any one serum protein provided the others are present in normal concentrations. Evaluation of the degree of protein binding of quinidine in plasma from normal subjects and patients with types IV and IIa hyperlipoproteinemias supported the computer predictions. The degree of protein binding in plasma from normal subjects and patients with types IV and IIA hyperlipoproteinemias were 71.2% +/- 11.4%, 75.9% +/- 8.7%, and 69.9% +/- 15.0%, respectively. The quinidine binding in plasma of patients with hyperlipoproteinemias did not differ statistically from that in the normal subjects.


Effect of pleural effusion on high-dose methotrexate kinetics. The kinetics of methotrexate were followed in a patient given two 6-hr infusions of 400 mg/kg in the presence and absence of a pleural effusion. Although the decline in serum concentrations during the first 30 hr after the infusion was similar for the two treatment courses, the half-life beginning 30 hr after the infusion was 6.7 hr without the pleural effusion and 14.4 hr with the pleural effusion. Comparison of the intercompartment distribution rate constants indicated slower movement of the drug back into the central compartment from the peripheral compartment when a pleural effusion was present. Pleural fluid methotrexate concentrations were consistently higher than serum concentrations. These data indicate that the increased risk of toxicity following high-dose methotrexate in patients with pleural effusions is due to changes in methotrexate kinetics resulting in delayed excretion.


A clinical safety trial of stroma-free hemoglobin. A stroma-free hemoglobin (SFH) solution was prepared which was sterile, pyrogen free, and contained only 1.2% of the stromal lipid present in unpurified hemolysate, 250 ml of which was administered slowly intravenously to 8 healthy men. Two control subjects received 250 ml of serum albumin. The SFH infusions were generally well tolerated by 7 of the 8 men. One subject developed abdominal pain and costovertebral angle tenderness after infusion, which disappeared within 48 hr. Bradycardia and a mild increase in blood pressure was present during ths SFH infusions and for 4 to 5 hr thereafter. A decrease in urine output and endogenous creatinine clearance appeared during the SFH infusions and for 2 to 4 hr after infusion. A mild prolongation of the activated partial thromboplastin time developed immediately after infusion. Gross hemoglobinuria appeared as expected during the SFH infusions and completely disappeared by 6 to 10 hr after infusion. All the cardiovascular, renal, and clotting changes were present for only a few hours after the SFH infusion, during the hemoglobinemia (free Hb in plasma). At 24 hr and 7 days after infusion all measurements were normal, and 6 mo follow-up showed no abnormalities or hepatitis.


Renal function studies in normal and toxemic pregnant sheep. Renal blood and plasma flow, glomerular filtration rate (GFR) and maximal tubular transport of PAH (TmPAH) were measured in nonpregnant and twin-pregnant sheep. Twin-pregnant animals were studied during normal pregnancy as well as during ovine pregnancy toxemia artificially produced by starvation. All animals were surgically prepared with aortic, post caval and renal vein cannulas at least one week prior to experimentation. Total renal blood and plasma flow was found to be elevated during pregnancy, but if expressed on the basis of body weight no changes were noted. Starvation and the resultant development of hypoglycemia and hyperketonemia caused a 25-30% decline in renal blood and plasma flow. GFR in pregnant fed sheep (193 ml/min or 2.7 ml/kg.min) was significantly higher (P less than .001) than that of nonpregnant ewes (118 or 2.3 ml/kg min). During ovine pregnancy toxemia the GFR was significantly (P less than .001) diminished (142 ml/min or 2.0 ml/kg min). TmPAH also was significantly higher (179 mg/min or 2.5 mg/kg min) in pregnant animals when compared to nonpregnant ewes (98 mg/min or 1.9 mg/kg min.), but starvation had no effect on Tm PAH in pregnant sheep. It thus appears that a functional renal hypertrophy occurs during pregnancy which is similar to that which follows unilateral nephrectomy or renal disease. During ovine pregnancy toxemia the diminution of renal function probably results from the metabolic derangements and is thus not comparable to human preeclampsia.


Relationships between teat-end bacteria and intramammary infections. Teat-end populations of staphylococci and streptococci were studied on teat apices of an experimental herd. Individual cows were found to have higher numbers of some of these microorganisms than other cows and certain teats had consistently higher populations than other teats in the same individual. A positive relationship was found between the numbers of staphylococci on teat-ends and presence of intramammary infections of the quarters. High teat-end bacterial numbers were present prior to the laboratory recognition of intramammary infections.


Anencephaly in calves.

Anencephaly in calves. Anencephaly occurred in four calves and was characterized by cranioschisis, absence of the diencephalon with the cerebral hemispheres and rostral midbrain, various forms of eye defects, and relatively normal development of caudal brain stem, cerebellum and spinal cord. Amorphous dysplastic vestiges of the cerebral tissue protruded into the cranial defects. Morphologic features varied but were essentially similar. The cerebellum was absent in one case. Anencephaly in calves at least those we observed and defined in this study is localized defect confined to the brain, eye and skull. No spinal defect was observed in the calves.


An outbreak of tularemia in mink.

An outbreak of tularemia in mink. An outbreak of tularemia in farm raised mink is reported. Twenty-six of approximately 5000 mink succumbed within a 10 day period. Prodromal signs were minimal. Necropsy revealed necrotic nodules scattered in the parenchyma of the lungs, liver, spleen, and mesenteric lymph nodes. Francisella tularensis was isolated from spleens, livers and lungs.


Serological types of Pasteurella haemolytica in Kenya. The 12 known serotypes of P. haemolytica and two biotypes A and T were found to occur in Kenya. Biotype T was more common in disease conditions than biotype A, which was more common in the nasal passages of healthy animals. Only biotypes A strains were recovered from cattle and the majority were serotypes 1 and 2, but serotypes 4 and 11 were also isolated. All serotypes were found to occur in sheep and goats, but serotypes 3, 4, 6, 8 and 10 were more commonly associated with pneumonia. It was observed that chickens could harbour both biotypes A and T in pathological conditions. Biotype A serotype 2 was isolated from an adult wildebeest, but the prevalence of P. haemolytica in wild animals needs furter investigation.


A Robertsonian translocations in the female cells of a bull, co-twin to a freemartin. A case of 1/29 chromosome translocation in a male calf of the Brown Swiss breed is reported. Except for the aggressive behavious, the subject showed clinically nothing peculiar. A post-mortem examination did not reveal any internal macroscopic malformation. Since many reports indicate that such a type of translocation, found in a wide range of breeds of Bos taurus of several countries, seems to be linked with a lowering of fertility, the authors feel that chromosome screening might play an important role in breeding programs in the immediate future.


Agenesis of corpus callosum in calves.

Agenesis of corpus callosum in calves. Agenesis of the corpus callosum is a rare brain defect in cattle, and was described in two calves. It was characterized by the total absence of the corpus callosum, septum pellucidum and hippocampal commissure. Fornices were present but separated. Both calves also had moderate internal hydrocephalus, micrencephaly and a CSF-filled cyst in the longitudinal fissure dorsal to the thalamus. Clinical signs were not specific. Etiology and pathogenesis are unknown. No evidence of inflammation or inheritance was found as cause of the defect.


Tri-minocycline: a minocycline complex for parenteral use. I. Antibacterial activity and toxicity data. Aqueous solutions of tri-minocycline (equivalent to 20--50 mg/ml minocycline HCl) and preconstituted ready-for-use injections were prepared at the pH of blood; these exert high antibiotic activities equal to those of the parent substance. The chloroform/water distribution coefficients of tri-minocycline in the different types of injections were somewhat lower, but in the same order of magnitude as that of the parent substance. The MIC values of tri-minocycline proved to be very low and the LD50 values very high, their separation covering more than 3 orders of magnitude.


Inhibition of growth of mouse ehrlich ascites by normal tissue extracts. A dialyzable component from the aqueous extracts of mouse skeletal muscle and liver inhibited in vitro growth of a mouse Ehrlich ascites. A similar component was not detected in extracts of spleen, kidney, lung, skin, serum or small intestine. The muscle component appeared to be different from that of the liver in its resistance to heat and its stability in the culture medium. Both components however were stable on storage at 4, 23 and 37 degrees C for 72 h. Intraperitoneal injection of the muscle and liver component into mice previously innoculated with Ehrlich ascites significantly decreased tumor incidence in these animals as compared with the control.


Antibiotic X-63 in experimental candidiasis of mice. X-63, a polyene antibiotic of heptaene series showed activity against experimental infection with Candida albicans in mice, by several routes. The oral, intraperitoneal and intravenous activities ranged between dosages of 3.1--50, 0.06--0.5 and 0.25--0.5 mg/kg, respectively. Seven days treatment by intraperitoneal route caused significant progressive reduction of colony-forming units in kidney homogenates of infected mice, which was comparable to amphotericin B. The regression of infection was further demonstrated histopathologically. In vitro, antibiotic X-63 was similar to nystatin against nine strains of Candida used. It also proved active against amphotericin B-resistant strains of C. albicans, C. tropicalis, C. pseudotropicalis, C. krusei, and C. parakrusei. The use of combined drug therapy in candidiasis has been discussed.


Failure of morphine to increase striatal 3,4-dihydroxyphenylacetic acid in fasted rats. Morphine given to rats fed ad libitum increased the striatal levels of DA and DOPAC while naloxone had no effect. Conversely, after prolonged fasting, morphine failed to increase both DA and DOPAC, while naloxone markedly decreased striatal DOPAC concentration. Intermediate sensitivity to the DOPAC increasing effect of morphine and to the DOPAC decreasing effect of naloxone was present in rats trained to eat their daily meal within 2 h, at different times after feeding. It is concluded that striatal DA synthesis in fasted rats is sustained by an increased endorphin-like activity.


Imipramine antagonism of apomorphine-induced hypothermia: a non-dopaminergic interaction. Imipramine antagonized high dose apomorphine-induced hypothermia, and did not modify small dose apomorphine-induced hypothermia. It is suggested that apomorphine-induced hypothermia is the result of two effects. The first, induced by small doses of apomorphine, and antagonized by pimozide and sulpiride, is probably related to dopaminergic receptor stimulation. The second, induced by high doses of apomorphine, and antagonized by imipramine, is probably not related to dopaminergic receptor stimulation.


Naloxone as a GABA antagonist: evidence from iontophoretic, receptor binding and convulsant studies. From the following three lines of evidence, it is proposed that at least part of the convulsant activity of naloxone is a result of GABA receptor blockade. Firstly, iontophoretic naloxone reversibly antagonized GABA-evoked depression of firing rate in 21 of 27 neurons tested in the rat olfactory tubercle-nucleus accumbens region, without blocking inhibition evoked in the same cells by glycine (15 cells) or morphine (6 cells). Secondly, i.p. naloxone in high doses caused convulsions in mice, and potentiated the convulsant activity of bicuculline, but not that of strychnine. Diazepam, which protected mice against convulsions elicited by bicuculline, but not by strychnine, also protected mice against naloxone. Thirdly, naloxone, morphine, levorphanol and its non-analgesic enantiomer dextrorphan displaced 3H-GABA from GABA receptor sites in homogenates of human cerebellum, all with comparable low potencies (IC50 = 250--400 micron). There was no correlation with affinities at the stereospecific receptor sites that mediate opiate-induced analgesia, since the potent opiates etorphine and diprenorphine were relatively inactive (IC50 greater than 3 mM). In addition naloxone displaced 3H-GABA from receptor sites in rate forebrain and cerebellum, with similar low potency.


Dantrolene sodium: effects on smooth muscle. The effects of dantrolene sodium on smooth muscle were evaluated in vitro using the guinea pig ileum and the guinea pig vas deferens preparations. Dantrolene irreversibly decreased the strength of the responses of the ileum to acetylcholine, histamine, potassium chloride and electrical stimulation. Doubling the concentration of calcium in the bathing media reduced the effect of dantrolene on all agonists. Dantrolene irreversibly decreased the response of the vas deferens to epinephrine, norepinephrine, acetylcholine and electrical stimulation. Doubling the concentration of calcium in the bathing media reduced the effect of dantrolene on all agonists. Dantrolene seems to depress smooth muscle as it does skeletal muscle. The mechanism of action probably involves the reduction of calcium flux into the muscles.


Antiarrhythmic potency of procainamide and N-acetylprocainamide in rabbits. The antiarrhythmic potency of procainamide (PA) and N-acetylprocainamide (NAPA) has been investigated in rabbits using isolated atrial preparations and ouabain-induced ventricular fibrillation in vivo. At concentrations in the range 3 x 10(-5) to 1 x 10(-3) M, both PA and NAPA decreased the maximum following frequency (MFF) of isolated atria. The dose--response curves were not parallel but at a concentration of 10(-3) M, NAPA had only one tenth of the potency of PA. Threshold level voltage of the atria were increased by PA but NAPA had no significant effect on this parameter. When atria were preincubated with NAPA (1.6 x 10(-4) or 8.0 x 10(-4) M), the dose--response curve for PA on MFF was displaced to the right. Pretreatment of anaesthetised rabbits with either PA (25 mg/kg i.v.) or NAPA (75 mg/kg i.v.) prolonged the time to onset of ouabain-induced ventricular fibrillation. NAPA (25 mg/kg) did not affect the response to PA (25 mg/kg). The data support the view that NAPA is less potent than PA and suggest that, under certain circumstances, NAPA may antagonise the actions of PA.


Development of insulin sensitivity in rat aorta after chronic propranolol treatment. Spontaneously hypertensive rats and normotensive Kyoto Wistar controls were divided into 3 groups of 10 animals each and treated with phenoxybenzamine (5 mg/kg once daily), propranolol (25 mg/kg twice daily) or saline (once daily). After 5 weeks the in vitro incorporation of D-[U-14C]-glucose into aortic lipids and glycogen was measured in the presence and absence of insulin (1 mU/ml). In both normotensive and hypertensive rats treated with propranolol 14C-incorporation into triglycerides was reduced. Furthermore, insulin significantly stimulated 14C-incorporation into triglycerides, phospholipids and glycogen in propranolol-treated hypertensive rats. This effect was not statistically significant (0.05 less than p less than 0.1) in propranolol-treated normotensives. Phenoxybenzamine treatment did not significantly modify aortic lipogenesis or glycogen synthesis from glucose. Chronic propranolol treatment of spontaneously hypertensive rats resulted in aortic tissue becoming sensitized to insulin. Possible mechanisms and explanations for this are discussed.


Effects of clonidine on vulnerability to fibrillation in the normal and ischemic canine ventricle. Clonidine infusion (10 microgram/kg, i.v.) elicited a 30% increase in repetitive extrasystole (RE) threshold in 6 chloralose-anesthetized dogs. A reduction in heart rate and arterial blood pressure accompanied the increased threshold. Intracisternal injection of clonidine (2 microgram/kg) in 6 dogs caused similar alterations in these parameters. Bilateral vagotomy, performed in 6 dogs prior to intravenous clonidine, prevented the increase in RE threshold but did not prevent the drug-induced bradycardia. Atropine (0.2 and 0.6 mg/kg), however, did not attenuate the effect of clonidine on RE threshold. Clonidine administration did not prevent the reduction in ventricular fibrillation threshold associated with a 10 min occlusion of the left anterior descending coronary artery or following reperfusion. We conclude that: (1) clonidine reduces ventricular vulnerability in the normal but not the ischemic heart, and (2) its protective effect is mediated by enhanced afferent vagal input to midbrain cardiovascular regulatory centers. This central nervous system action causes a reduction in sympathetic tone to the heart.


Effect of ouabain on myocardial metabolic and contractile responses to coronary ligation. The effect of pretreatment with ouabain (40 microgram/kg, i.v.) on myocardial metabolic and contractile responses to regional ischemia induced by coronary artery ligation was studied in the canine left ventricle. In control dogs, ischemia increased activity of phosphorylase a and the levels of glucose-6-phosphate and lactate, and decreased the levels of glycogen and phosphocreatine, without affecting the levels of adenosine triphosphate, adenosine diphosphate, and adenosine monophosphate (AMP). Ouabain increased the activity of phosphorylase a. In ouabain-treated dogs, ischemia did not further increase the phosphorylase a activity but it increased the epicardial AMP level. Other metabolic responses to ischemia in ouabain-treated dogs were similar to those in control dogs. In control dogs, myocardial contractile force decreased by about 10% after ischemia, but blood pressure and heart rate remained unchanged. Ouabain increased contractile force by about 32%. In ouabain-treated dogs, ischemia decreased contractile force by about 54% without affecting blood pressure and heart rate. It is concluded that ouabain increases the activity of the myocardial phosphorylase a and that the inotropic action of ouabain can be nullified by coronary artery ligation.


Effects of cholinesterase inhibitors on the spasmogenic action of acetate esters on rat uterus. Acetate esters, such as phenyl acetate and aspirin, induced atropine-sensitive contractions of isolated uterus only when choline was present. These contractions were selectively and reversibly inhibited by carbamate-type cholinesterase inhibitors, such as neostigmine and eserine, and quaternary ammonium compounds, such as tetraethylammonium and decamethonium. After treatment with organophosphorus cholinesterase inhibitors, such as di-isopropyl fluorophosphate and tetraethyl pyrophosphate, the uterus failed to respond to the acetate esters, even when high concentrations of choline were present. The inhibition of the response of the uterus by organophosphates was effectively removed by pyridine-2-aldoxime methiodide. Pretreatment of the uterus with neostigmine or simultaneous addition of high concentrations of quaternary ammonium compounds prevented the inhibition by organophosphates. The inhibition produced by neostigmine was also reduced by simultaneous addition of quaternary ammonium compounds. These findings suggest that some esterase having an anionic site and an esteratic site, probably cholinesterase, may mediate in the uterine contractions induced by acetate esters in the presence of choline, and that inhibition by organophosphates, carbamates and quaternary ammonium compounds of cholinesterase activity in the preparation may impede the initiation of contractions by the acetate esters in the presence of choline.


Monitoring trends in food utilization: application of an archaeologcial method. Archaeological methods were utilized to monitor trends in food purchase and utilization behavior in Tucson, Arizona households through the analysis of household refuse. As a supplement to traditional methods of collecting data on food consumption, the study of household refuse can provide information on short-term trends in behavior and on food discard. Between 1973 and 1975, households studied decreased the amount of food brought into the home and decreased their purchase of selected animal protein foods. Lower-income tracts, in addition, increased their purchase of grain products, substituted cheaper for more expensive food intems, and decreased food discard. While these data do not reflect nutritional status, they provide a unique view of the socioeconomic and behavioral patterns that in large part determine nutritional status.


Towards representative energy data: the Machiguenga study. Representative energy data for a human population can be produced by combining randomly sampled time allocation observations with activity-specific energy expenditure measurements. Research to produce representative energy data for adults of a population of Machiguenga Indians has recently been conducted in lowland, southeastern Peru. Marked contrast was found between the sexes for average married adults in energy expended on an average day. Men spent about 3,200 kcals and women, about 1,925; ratio: 1.66 to 1. In general, men tended to work at somewhat more energetic activities and for longer periods than did women. In addition to sex-role-related task differences were contrasts in uses of technological items and in respective work settings. These representative behavior data permit direct estimates of population-level energy requirements for average days, seasons, or for 1 year.


Seasonal effects of dietary intake on Kalahari San. San living in the northwestern Kalahari Desert may be grouped according to different kinds and proportions of dietary habits. At one end of a continuous scale are those whose food is obtained almost entirely from wild bush sources; at the other extreme are those who obtain large amounts of food from domestic sources, principally milk and maize meal. The majority fall between these poles. The region is also home to a substantial number of pastoralists and horticulturalists. Data have been collected on the productivity of both the natural and domesticated environments, on resource procurement strategies, and on food consumption during a period of more than 1 year. Seasonal weight fluctuations associated with food production cycles are recorded. Mean weight and serum lipid values are higher in those San who consume substantial amounts of domestic foods than they are in bush food eaters. Seasonal variation in fertility is reported to be coordinated with variations in nutritional status. Some implications for demographic and social processes are mentioned.


Epidermal antihyperplastic effects of contact antiphlogistics. The inhibitory effects of six contact antiphlogistics and two dermatocorticoids on hexadecane-induced hyperplasia of the guinea-pig skin were tested and compared. On the basis of the results obtained, the substances can be ranked in the following order of potency (percent inhibition of surface epithelial proliferation): acetylsalicylic acid (7.9%), phenylbutazone (12.5%), bufexamac (16.4%), bendazac (16.6%), oxyphenbutazone (19.5%), indomethacin (20.0%), hydrocortisone (20.6%) and fluocinolone acetonide (32.0%).


Dyskeratosis congenita with epidermodysplasia verruciformis of Lewandowsky and Lutz. The case is reported of a 30-year-old man suffering from dyskeratosis congenita with severe pancytopenia, in association with generalized verrucosis. Viral particles were detected electron microscopically. The importance of an impaired immunological state, demonstrated in this case by lymphocyte stimulation and rosette tests, is discussed.


Treatment of basal cell carcinoma of the skin with ultrasoft X-rays. The results of a follow-up study carried out in 1975-76 are described. The study involved 454 patients who had received ultrasoft X-ray treatment for basal cell carcinomas during the period 1965-1969. In females, the percentage of recurrence was 4.9, in males, 11.9. The figures indicating the frequency of recurrence are minimum ones; the patients were selected to a certain extent, the large tumors being treated surgically. The most important result of the study was the observation that approximately 30% of the X-ray scars examined were so disfiguring from a cosmetic point of view that they must be characterized as unacceptable. The indications for X-ray treatment of basal cell carcinomas should, consequently, be limited to deep-growing tumors or tumors which are not well demarcated, and to tumors in elderly patients, who should be spared surgery.


Blood flow, collagen components of oral tissue and salivary kallikrein in young to senescent, germfree and conventional rats. A study on the etiologic factors of periodontal disease. In oral tissues of rats, progress of age resulted in a gradual reduction of regional blood flow and in decreased levels of eluable (labile) hydroxyproline from the collagen component. These were paralleled by an increment of salivary kallikrein. The presence of the microbial flora in the oral cavity, portrayed by the comparison of conventional and germfree rats, caused an elevation of blood flow in all age groups, without exerting a marked effect on eluable hydroxyproline. In addition to aging and flora effects, the action of chronic hypoxia and of mechanical stress has been observed on the mentioned parameters. The presently used animal models and experimental design permit a critical study of etiologic agents and characteristic lesions of peridontal disease.


In vivo stimulation of nerve cells by phytohemagglutinin. I. Alterations of some cytological aging parameters in rat brain cells. Phytohemagglutinin P (PHA-P) was administered in form of intralumbar injection to old (24-27 months) female Wistar rats. PHA-P has an inductive effect also on the nerve cells. The thermal denaturation characteristics of the DNA in situ as well as the numerical density of perichromatin granules being connected to the extranucleolar RNA synthesis returned practically to the young levels in the large brain cortical cells and in the cerebellar granular cells 20 h after the injection. The reversal of these aging phenomena was accompanied by a significant decrease of intranuclear and intracytoplasmic K+ content measured by X-ray microanalysis in the dry mass, whereas Na+ and Cl- remained unchanged. If PHA-P causes also some (1-2%) rehydration of the old neurons, the total ionic strength in the cell due to the monovalents may even reach the young level. The possible regulatory role of the ionic strength and the Na+/K+ ratio in the chromatin function is discussed on the basis of experimental data. It is the first time that the reversibility of some cytological aging parameters has been demonstrated in vivo.


Quantitative studies in the cerebral cortex of aging humans. The capillary network of the human cerebral cortex was morphometrically investigated in a young, aged and aged hypertonic group. In the aging cortex, augmented values of capillary diameter and volume are accompanied by smaller distances between capillaries and an extended length per unit cortex volume. An increased capillary diameter of the aged hypertonic group is probably due to a high blood pressure. The regulating influence of the surrounding astroglia and the extracellular fluid on capillaries seems to lack. In comparison to normotonic brains, higher distances between capillaries and a decreased capillary length were measured.


Total free amino acid levels in adult and senescent rat liver. The total free amino acid content per gram of liver wet weight does not differ significantly between adult and senescent female rats, being 28.8 +/- 3.3 mumol in adult (13-17 months) and 26.2 +/- 2.2 mumol in senescent (23-29 months) animals. This is the first study in which these two age-groups of rats have been so compared. These results combined with a survey of the literature show that amino acid levels in rat liver do not change much, if at all, throughout the lifespan of the animal.


Ageing changes in the human testis: a light-microscopic study. A histologic study of ageing testes, removed surgically from 'healthy' men aged 52-100 years, is presented. The age-related changes noted were: tubular sclerosis, Leydig cell hyperplasia, focal mononuclear orchitis, capsular smooth muscle hyperplasia, and dilatation of the rete testis. Great individual variations were seen but the testes, removed from patients with prostatic carcinoma, showed less severe ageing changes than those removed from patients during herniorrhaphy. The aetiology and pathogenesis of these changes are briefly discussed. A plea is made for more systematic studies of the ageing human testis, combining light microscopy with more sophisticated techniques, such as electron microscopy, histochemistry and cytochemistry.


Age, inactivity and some physiological responses to exercise. The quality of life in old age is crucially dependent upon the ability to pursue a variety of physical activities. Factors which affect the capacity for physical activity in the elderly should therefore be of interest both to individuals and to institutions concerned for the elderly. This is a review of literature pertinent to the problem of maintaining an adequate capacity for physical exercise into old age. It includes information about the interrelations between age, physical condition and physical working capacity; and the effects of training and inactivity upon all these parameters. Reduction in levels of habitual activity with age seems likely to cause or exacerbate deterioration in both exercise capacity and physical condition, setting up a vicious circle which eventually jeopardises the capacity for independent living. If so, efforts should be directed towards maintaining effective levels of habitual activity.


Neutrophilic turnover rate in human age groups evaluated by serum lysozyme activity. Serum lysozyme activity was determined in 135 healthy people classified into five different age groups ranging from 20 to 90 years. A turbidometric method with egg-white lysozyme as standard enzyme using the Fragiligraphy for automatic recording of the serum lysozyme was used. The results show a progressive increase in serum lysozyme activity with age. In the oldest age group of 60-90 years, the increase in serum lysozyme was more than that expected for the diminished glomerular filtration rate in old age. Since the serum level of lysozyme can reflect the rate of neutrophilic turnover, it can be assumed that this rate increases above the age of 60.


25-hydroxy-vitamin D, diet and sunlight exposure in patients admitted to a geriatric unit. Dietary intake, sunlight exposure and 25-hydroxy-vitamin D levels were estimated in 62 patients admitted to a geriatric assessment unit. There was a significant correlation between vitamin D intake and 25-hydroxy-vitamin D levels but none between sunlight exposure and these. This suggests that in old age dietary deficiency may be a more important cause of metabolic bone disease than limited sunlight exposure.


Metabolism of thyroid hormones during aging. The paper presents experimental data on extrathyroidal metabolism of thyroid hormones in rats of various ages (1.5--32 months):: T4-131I distribution in an organism, indices of T4 and T3 kinetics, tissue deiodinative capacity, formation of T4 tissue metabolism products and their clearance from cells and organism, T4 binding with tissue proteins. Essential changes in T4 peripheral tissue metabolism have been found with aging: T4 binding by tissues is decreased; T4 distribution space and metabolic clearance are increased; 125I clearance with urine is decreased; T4 half-time in blood remains unchanged; tissue deiodinative capacity is enhanced. Indices of T3 peripheral metabolism in old rats remain unchanged. All these findings result in shifts of the T4/T3 ratio in favour of the latter and in invariability of its content in tissues, alongside with a decrease of functional activity of the thyroid gland with aging.


Restitution of ATP and creatine phosphate after experimental depletion in young, adult, and old rats. The rates of restitution of skeletal muscle, heart, and brain creatine phosphate and brain ATP, following experimental depletion, were compared in young (3-5 month), adult (12 month) and old (24 month) rats. In skeletal muscle, restitution of creatine phosphate after 3 min of recovery was greatest in the young rats, minimal in the adult animals, and absent in the old animals. In heart muscle, on the other hand, restitution was rapid in all three age groups, and was essentially complete after 60 sec recovery. In brain, restitution of creatine phosphate was rapid and was complete after 10 min recovery in the young and adult rats; restitution was slower and less complete in the old rats. In the old rats there was no restitution of brain ATP during the first 2 min of recovery, after which the rate of restitution paralleled that of the young and adult rats.


Is there a biological basis for geriatric pharmacotherapy? The problem of developing a geriatric pharmacotherapy is briefly discussed. It is stressed that combined scientific knowledge from biological, clinical and pharmacological research should be used as basis. However, in applied (clinical and pharmacological) gerontology, direct relations to basic experimental gerontology are often avoided, because of its seemingly theoretical character. With some examples, it is pointed out that during the recent past, biological-aging research has gained new knowledge, particularly on skeletal muscle and CNS aging, that can be used in the concept of a specific geriatric pharmacotherapy.


Interaction of ergot alkaloids with dopaminergic receptors in the rat striatum and nucleus accumbens. Various ergot alkaloids and derivatives were investigated for their interaction with dopaminergic receptors at the level of the rat corpus striatum and nucleus accumbens. Dihydro(DH)-ergotoxine, DH-ergocornine, DH-ergocryptine, DH-ergocristine, 2-Br-alpha-ergocryptine, ergotamine and DH-ergotamine were shown to inhibit, at micromolar concentrations, the dopamine-stimulated adenylate cyclase activity of rat striatal and nucleus accumbens homogenates. Interestingly, the inhibitory effect of the ergot drugs was higher in the nucleus accumbens than in the striatum. Moreover, the ergot drugs were more active in displacing 3H-haloperidol than 3H-dopamine from striatal membranes. The results, which are in apparent contradiction with previously obtained behavioral, pharmacological and clinical data, are discussed in the light of the possible presence in the central nervous system of distinct dopaminergic receptors with different conformations.


Comparative effects of dihydroergotoxine (DHET) on CBF and metabolism changes produced by experimental cerebral edema, hypoxia and hypertension. Dihydroergotoxine (DHET) perfused in the dog (100 microgram/kg) presenting a cerebral per-hypocapno-anemic syndrome reduces cerebral hyperemia, increases cerebral venous PO2, despite the rise in CMRO2 and favors glucose oxidation by the brain. DHET (20 mg/kg p.o.) is able to drop mean, diastolic and systolic arterial blood pressures in renal-hypertensive rats having a cerebral edema induced by triethyltin intoxication without affecting cerebral water and sodium levels which are increased in the controls. DHET (50 microgram/kg i.v.) can also improve EEG changes produced by a traumatic edema but does not exert (200 microgram/kg i.v.) any effect on EEG changes produced in the rabbit by lithium chloride intoxication.


Action of norepinephrine on microcirculation and PO2 distribution in the isolated perfused rat liver. The addition of a potent vasoconstrictor, such as norepinephrine, induces an influx of sodium and an efflux of calcium and potassium in the parenchymal cells of the perfused liver. This reaction can be reversed by the addition of dihydroergotoxine mesylate (active substance of Hydergine). The example shows clearly that very distinct biological signals are generated under such conditions at the membrane level of the hepatocytes and presumably of other cells of liver tissue. At present, our investigations cannot clearly answer the question of whether or not swelling the shrinkage of parenchymal cells and of endothelial cells can serve as an additional mechanism for regulating microcirculation.


Dihydroergotoxine-induced modulation of spontaneous activity of cultured rat Purkinje cells. We use nerve tissue cultures as a model system for studying the mode of action of centrally acting drugs. Neurons in cerebellar cultures retain a number of organ-specific properties including the formation of complex synaptic networks and characteristic bioelectric activity. Electrical activity may be recorded from Purkinje cells which can be identified by direct visualization. We observed that dihydroergotoxine mesylate (DHET; active substance of Hydergine) modulated the spontaneous activity of cultured neurons. Addition of 10(-8)g/ml DHET to rat cerebellar explants led to a significant regularization of the spontaneous firing of Purkinje cells. Within the 10(-10) - 10(-5)g/ml concentration range used in these experiments, DHET had no detectalbe effect on the average firing rate.


Failure of medullary carcinoma of the thyroid to respond to doxorubicin therapy. We describe 3 patients with metastatic medullary carcinoma of the thyroid who were treated with doxorubicin hydrochloride (Adriamycin). Serum calcitonin was measured before and after doxorubicin therapy. Doxorubicin failed to arrest the progression of the disease in any of the patients. Although serum calcitonin levels dropped in 1 patient during therapy, they remained markedly elevated in all 3 patients. From the present series it appears that medullary thyroid carcinoma often does not have a response to doxorubicin.


On the role of the pineal in thermoregulation in the pigeon. Diurnal rhythm in body temperature of pigeons subjected to different experimental conditions (pinealectomized, sham-operated, melatonin-implanted, cold-exposed) was studied under a 12 h light and 12 h dark regimen. The body temperature of pigeons during photophase was higher than that during scotophase in the normal as well as every treatment group studied. Pinealectomized pigeons showed higher body temperature in the photophase as well as scotophase, than that of the normal and sham-operated birds when examined 2 or 3 weeks following the post-surgical acclimatization to 25 or 3 degrees C. However, subcutaneous implantation of melatonin pellets into pinealectomized pigeons nullified or even reversed the hyperthermic effect of pinealectomy. Exposure of pigeons to--18 degrees C for 280 min during photophase as well as scotophase, produced a marked drop in body temperature in pinealectomized, sham-operated and normal pigeons. The pinealectomized pigeons exhibited a higher body temperature than that of the sham-operated and normal ones when exposed to--18 degrees C during the photophase, but not during the scotophase. It was concluded that while the pineal is not necessary for maintaining the daily thermal rhythm in the avian body, it has a thermoregulatory role, in that it prevents rise in body temperature in warm (25 degrees C) acclimatized and chronic cold (3 degrees C) exposed birds. In acute short-term cold (--18 degrees C) exposure however, the temperature regulatory role of the pineal was not effective during the scotophase.


Dermatoglyphic studies of myocardial infarction patients. Dermatoglyphic traits were studied in a sample of 834 subjects selected from a cohort of some 8,000 living Japanese men, under a long-term study of heart disease in Hawaii. All of them were born between 1900 and 1919. Among them, 100 subjects had had positive diagnosis of myocardial infarction (MI). The present study included comparisons between the MI patients and the remaining group of all digital dermal pattern types and ridge counts by digit, by hand, and by individual. The MI patients had significantly higher frequency of true whorls, double loops and less ulnar loops and tented arches. Total and absolute ridge counts were significantly higher (less than 0.05) in all digits in favor of the MI patients. Similar trends were observed in analyses by digit and by hand. These observations suggest an antenatal origin of certain types of coronary disease.


Placental enzyme polymorphisms in Canadian populations. III. Alkaline phosphatase. Placental alkaline phosphatase phenotypes have been determined for a large Canadian population. The 'common' alleles, PL1, PL2, and PL3 have frequencies similar to those of European populations. Five new phenotypes are described and the incidence of rare phenotypes is compared for several populations.


Gene differentiation among the Dhangar caste-cluster of Maharashtra, India. Genetic differentiation among the 22 Dhangar castes of Maharashtra, India, is studied using data on several polymorphic serological and biochemical loci employing Nei's distance measures. The intercaste genetic distances and the coefficient of gene diversity among these caste groups are found to be rather small. The relationship between gene identity and geographic distance is also studied empirically from the gene frequency data. All these analyses indicate that genetic differentiation among the Dhangar castes is at its very early stage only. It is also suggested that these caste groups probably originated from a common stock and are in the process of differentiation by fission with very little intercaste migration in the recent past.


Serum protein and red cell enzyme polymorphisms in affective disorders. Frequencies of serum groups (Hp and Gc) and red cell enzyme types (PGM1, 6-PGD and ES D) were studied in 195 patients with affective disorders. The patients were classified into four groups: (1) bipolar (manic-depressive) psychosis; (2) unipolar, recurrent, depressive psychosis; (3) non-psychotic reactive depression, and (4) unclassifiable. The Hp2 gene was increased in reactive and unclassifiable patients, the PGM1 1 gene was increased in bipolar patients and the ES D1 gene in reactive patients. No associations were found between affective disorders and the Gc and 6-PGD systems.


Blood groups and affective disorders.

Blood groups and affective disorders. Frequencies of ABO, Rh, MNSs, P, Kell, Lewis and Duffy blood groups were studied in a total of 219 patients with affective disorders. The patients were classified into four groups: (1) bipolar (manic-depressive) psychosis; (2) unipolar recurrent depressive psychosis; (3) nonpsychotic 'reactive' depression, and (4) 'unclassifiable'. The following statistically significant results were found: an increased frequency of the blood group factor B among psychotic (bipolar and unipolar) patients compared to nonpsychotic patients, a decreased frequency of the SS phenotype in the unclassifiable group and an increased frequency of the K(+) phenotype among the nonpsychotic patients. Previous results concerning differences between bipolar and unipolar patients with respect to the A and O blood types were not confirmed in this investigation.


Hemoglobin S and some other hemoglobinopathies in Eti-Turks. Frequencies of various hemoglobinopathies were examined in a total of 1,922 individuals of Eti-Turk origin by electrophoretical techniques. Hemoglobin A2 (Hb A2) and hemoglobin F (Hb F) determinations were also performed in 651 and 1,642 cases, respectively. Mean hemoglobin S (Hb S) frequency was found to be 15.3%. Variations among the different age groups were insignificant. Hemoglobin E (Hb E) and beta-thalassemia frequencies wer 0.47 and 1.23%, respectively. Hemoglobin Hacettepe and hemoglobin D were found once. Red cell glucose-6-phosphate dehydrogenase (G6PD) deficiency was found in 6.5% of males.


Adenosine deaminase polymorphism among the Semai, Temuan, Semelai, and Jakun groups of West Malaysian Orang Asli. 812 West Malaysian Orang Asli belonging to four ethnic groups were surveyed for adenosine deaminase (ADA; EC 3.5.4.4) using starch gel electrophoresis. Only the common ADA1 and ADA2 alleles were found, with the frequencies of the latter being 0.025, 0.103, 0.115 and 0.028 in the Semai, Semelai, Temuan, and Jakun groups, respectively. A new 'breeding genetic distance' was applied to these gene frequencies and the Semelai and Temuan were found to be more closely related to each other, and to have considerably more evolutionary flexibility on this scale of 'micro-evolution' than the other two groups. The Semai and Jakun were more similar to each other on the basis of these ADA gene frequencies.


The Nubians of Kom Ombo: serum and red cell protein types. Phenotype and gene frequencies are presented for eight polymorphic systems among the Nubians of South Egypt, namely, acid phosphatase, glucose-6-phosphate dehydrogenase, adenylate kinase, 6-phosphogluconate dehydrogenase, esterase D, phosphoglucomutase I, peptidase A, and haptoglobin. Eleven systems, namely, albumin, ceruloplasmin, hemoglobin, lactate dehydrogenase, isocitrate dehydrogenase, phosphohexose isomerase, malate dehydrogenase, peptidase B and C, phosphoglucomutase II, and transferrin were found to be monomorphic. A single electrophoretic variant of phosphohexose isomerase were observed.


Population structure in Kanoya population, Japan. The mean inbreeding coefficients found for Minami-cho (366 couples) and Shinsei-cho (511 couples) were 0.00307 and 0.00191, respectively. The mean inbreeding coefficient decreased and the mean marital distance increased as the year of marriage becomes more recent. The mean distances and their standard deviations between birthplaces of mates, father-offspring, mother-offspring, and sibs are 69.05 +/- 229.64, 73.09 +/- 246.66, 49.81 +/- 158.43, and 39.53 +/- 159.51 km, respectively, at Minami-cho. These values are 188.45 +/- 387.05, 187.79 +/- 562.59, 148.26 +/- 326.35, and 73.93 +/- 225.92 km, respectively, at Shinsei-cho. The dimensionality of migration is closest to one dimension.


Dermatoglyphic studies in rheumatic heart disease. 62 patients suffering from rheumatic heart disease were taken from Institute of Child Health, Niloufer Hospital, Hyderabad, for the present dermatoglyphic investigation. The finger, palm and toe prints were analysed to see if there was an association between rheumatic heart disease and any of the dermatoglyphic traits. The parameters which were significantly different from the controls are: (1) reduced frequency of arches on the finger tips in males and increased frequency of whorls in females; (2) increased frequency of patterns in the III interdigital area in males; (3) decreased d-t ridge count in females, and (4) higher incidence of multiple axial triradii in females.


On the nature of the presumed receptor for IgE on mast cells. V. Enhanced binding of 125I-labeled IgE to cell-free particulate fractions in the presence of protease inhibitors. Cell-free particulate preparations derived from sonicated, purified rat peritoneal mast cells can be radiolabeled with IgE. The IgE concentration-dependence and the kinetics of the binding reaction at pH 6.6 were very similar to those observed with intact mast cells. The number of IgE molecules bound per mast cell equivalent at saturation varied somewhat from one particulate preparation to the next and, on average, was about 50% of the binding capacity of the intact cells. Incubation of intact mast cells or of particulate fractions with low concentrations of IgE in serum-free media resulted in a decrease in the amount of bindable IgE which remained in the supernatants in excess over the amount of IgE which was bound to the cells. Specific binding of IgE to both particles and to intact cells at limiting IgE concentrations was stimulated up to approximately twofold by a variety of antibiotic, synthetic or high molecular weight (protein) protease inhibitors of which soybean trypsin inhibitor and p-nitrophenyl-p'-guanidinobenzoate were the most active. These substances also markedly protected the IgE, leaving more of it bindable to rat basophil leukemic cells in a second incubation. Binding to particulate fractions also occurred at pH 4.8 when bovine serum albumin was added to the incubations. The results are consistent with the view that normal peritoneal mast cells have a membrane-bound protease which has high selectivity for IgE.


Alterations in immune responsiveness in acute measles and chronic post-measles chest disease. Immune responses in 24 children with acute measles (AM) were compared with those in 20 children who had chronic pulmonary complications (CPMC) following measles. The immuno-suppressive effects of acute measles were extensive: total white cells were reduced and this reduction was accounted for entirely by lymphopenia which was equally expressed among the major lymphocyte sub-populations studied; the function of 'T' cells, assessed by radio-isotope incorporation into phytohaemagglutin (PHA) transformed lymphocytes and delayed skin hypersensitivity (DHR) to dinitrochlorobenzene (DNCB), was depressed. Serum IgA was reduced in AM patients. In contrast there was a relative sparing of the measured indices of immunity in patients with chronic post-measles chest disease, with the major defect being an impaired DHR to DNCB. There were minor alterations of complement components in both groups of patients.


Empiric analysis of the interactions involved in the formation of Fc receptor rosettes. Erythrocytes coated with IgG were used in forming rosettes with lymphocytes and polymorphonuclear leukocytes to investigate Fc receptors. The density of the IgG-Fc coating on erythrocytes predictable influenced the number of rosettes formed with leukocytes. This relationship could be empirically described by application of the Hill equation derived from studies of molecular interactions in other biologic systems. The approximate Hill coefficient for Fc receptors was 2.4. These findings suggest at least two possible models for membrane Fc receptor function.


Studies on contact sensitivity to DNCB in guinea pigs by the macrophage migration test. Guinea pigs contact-sensitized to 2,4-dinitrochlorobenzene (DNCB) by foot-pad injections or by topical applications of the hapten were studied by the direct macrophage migration test using DNP-guinea pig albumin (DNP-Alb) and DNP-guinea pig skin extract (DNP-SE) as antigens. Specific inhibition of macrophage migration in the presence of either antigen was found in a significant number of animals exhibiting contact sensitivity to DNCB. However, a wide variation in migration indices was observed in both groups of sensitized animals, with some animals showing a significant enhancement of macrophage migration.


Ehrlich ascites carcinoma: synchronised ultradian rhythms in certain cell parameters. In populations of Ehrlich Ascites Carcinoma (EAC) cells rhythms of near-hourly periods have been demonstrated for several cell parameters - nucleus and cell size, permeability and strength of plasma membranes. These oscillations are characterized by a high degree of synchrony and autonomy.


Time-course of cowpea chlorotic mottle virus RNA replication. In a synchronous infection of cowpea chlorotic mottle virus in cowpea leaves, the three largest viral RNA components were synthesized throughout the infection at relatively constant ratios of two molecules of component 3 to one each of components 1 and 2. Component 4 (the coat protein message) was produced minimally during the initial stages of the infection, and as the infection progressed the proportion of component 4 continuously increased. The replicative forms of components 1, 2, and 3 were produced with kinetics similar to those of the corresponding single-stranded molecules.


Togaviridae.

Togaviridae. The family Togaviridae is described; it contains four genera--Alphavirus, Flavivirus, Pestivirus and Rubivirus--and additional members. The main characteristics of the family are as follows: single-stranded, linear RNA, molecular weight about 4 X 10(6). Virions have isometric nucleocapsids surrounded by a lipoprotein envelope containing host cell lipid and virus-specified polypeptides including one or more glycopeptides. Virions yield infectious RNA. There are at least 80 members; all the alphaviruses and most flaviviruses are arboviruses in the biological sense.


Stability of mutations conferring temperature sensitivity on tobacco mosaic virus. Among several temperature-sensitive mutants of tobacco mosaic virus, reversion to wild-type phenotype is an event sufficiently rare that essentially pure stocks of mutant virus can easily be maintained. The temperature-sensitive phenotype of many of these mutants is stringently expressed. Our data show that most of our mutants are suitable for use in biochemical experiments.


The effect of protoplast concentration during inoculation on the infection of tobacco protoplasts. When tobacco protoplasts were inoculated with tobacco rattle virus, the proportion infected was inversely related to the protoplast concentration between 0.5 X 10(5) and 4 X 10(5) per milliliter when the inocula contained phosphate or Tris-HCL buffers, but was independent of protoplast concentration when using citrate buffer. The decrease in infection obtained at high protoplast concentrations in phosphate was diminished by increasing the poly-L-ornithine (PLO) concentration. At usual PLO concentrations, inocula in phosphate produced more infection that those in citrate, but at some suboptimal PLO concentrations the position was reversed. These effects may result from differences in the reaction between PLO and the different buffer anions.


Pea enation mosaic virus genoma RNA contains no polyadenylate sequences and cannot be aminoacylated. An active synthetase enzyme preparation from peas (Pisum sativum L.) did not catalyze the aminoacylation of pea enation mosaic virus RNA. The viral RNA was shown not to contain polyadenylic acid sequences.


Characterization of germination and activation of Bdellovibrio bdellocysts. A simple method of assaying germination of bdellocysts in liquid medium has been devised. Bdellocysts can be induced to germinate by any of eight L-amino acids or the monovalent cations K+ and NH4+. L-Glutamine was the best individual inducer of germination, although the resulting rate of germination was much slower than in a complex medium. The use of a defined germination medium containing L-glutamine, KCl, and NH4Cl produced a faster rate of germination than did complex media. Bdellocysts germinated most rapidly at pH 8.0 and at 35 degrees C and required aerobic conditions. Respiration of bdellocysts began to increase at 3 min after the addition of germinants. Germination was inhibited by respiratory-chain inhibitors and by inhibitors of macromolecular synthesis. When bdellocysts were heat shocked at sublethal temperatures for short periods, there was no effect on the rate of germination in the defined germination medium or in the complex medium. However, heat-shocked bdellocysts germinated at a much faster rate in the presence of single inducers of germination when compared to nonshocked bdellocytes.


Biosynthesis of oligosaccharide-lipid in Streptococcus sanguis. An oligosaccharide-lipid containing N-acetyl d-glucosamine (GlcNAc), l-rhamnose, and d-glucose was synthesized when the particulate enzyme from Streptococcus sanguis was incubated with UDP-GlcNAc, TDP-rhamnose, and UDP-glucose. The incorporation of d-glucose into the lipid was dependent on the preincorporation of l-rhamnose, which in turn was dependent on that of GlcNAc. This indicates that the order of sugar incorporation is GlcNAc, l-rhamnose, and d-glucose. The synthesis of GlcNAc-lipid was stimulated twofold by ATP and was inhibited strongly by UDP and slightly by UMP, CDP, and TDP, but not by all other nucleoside diphosphates and nucleoside monophosphates tested. A [gamma-(32)P]ATP labeling experiment indicated that some acceptor lipid was present in nonphosphorylated form. The acid and alkaline stabilities of the GlcNAc-lipid were similar to those of glycosyl undecaprenylphosphate, and the thin-layer chromatographic mobility of the lipid was slightly faster than that of the mannosylphosphorylundecaprenol. The molar ratio of phosphate to GlcNAc in purified GlcNAc-lipid was found to be 0.96:1. These results suggested that the GlcNAc was attached to the lipid moiety, presumably undecaprenol, by phosphodiester bonds. The incorporation of l-rhamnose into the lipid was inhibited by UDP and UMP, respectively, in a manner similar to the incorporation of GlcNAc. This suggested that the oligosaccharide was also linked to the lipid moiety by phosphodiester bonds.


Evolution of Pseudomonas R-plasmids: consequences of Tn1 insertion and resultant partial diploidy to chromosome and Tra- R-plasmid mobilization. Tn1 transposes from pRO161, a Tra- derivative of RP1, to Pseudomonas aeruginosa sex factor FP2. The acquisition of Tn1 by FP2 results in its ability to mobilize pRO161 to other bacteria. Genetic evidence presented here suggests two sequential mechanisms. Initially, transposition of Tn1 results in trans-diploidy for the Tra+ and Tra- plasmids. This subsequently allows mobilization of the Tra- R-plasmid dependent on a host recombination mechanism. Transconjugants from this mating contain either stable cointegrate R-plasmids or aggregates resulting from dissociation of the cointegrates into a Tra+ and Tra- plasmid. These aggregates have lost at least part of Tn1 from their parent FP2:Tn1 component, but now they mobilize the tra- R-plasmid from a recombination-deficient (Rec-) genetic background as well as from Rec+ donor strains. Transconjugants from these retransfer matings are aggregates. These results suggest a contribution of transposons to R-plasmid evolution and dissemination beyond the mere acquisition of resistance to a given antibiotic.


Changes in cytochrome content and electron transport patterns in Pseudomonas putida as a function of growth phase. Optical absorbance difference spectra of membrane vesicles prepared from aerobically grown Pseudomonas putida indicated that, when harvested in logarithmic phase, the cells contained one c-type cytochrome and two or three b-type cytochromes, one of which was cytochrome o. As the cells grew into stationary phase and the oxygen concentration of the medium dropped to essentially zero, an additional component believed to be cytochrome d was produced. Both the o- and d-type cytochromes might function as terminal oxidases. No a-type cytochromes could be detected at any stage of growth. Polarographic measurement of oxygen utilization revealed that cyanide and azide are effective inhibitors of the oxidation of ascorbate coupled with 2,6-dichlorophenolindophenol or N,N,N',N'-tetramethyl-p-phenylenediamine in respiratory particles from either log-phase or stationary-phase cells. Reduced nicotinamide adenine dinucleotide- or succinate-dependent oxygen utilization, however, was sensitive to these inhibitors only in log-phase particles. These results indicate that an alternate terminal oxidase may be synthesized by this organism in response to restricted oxygen availability and that branching of the respiratory system may result.


Tween 80 effect on glucosyltransferase synthesis by Streptococcus salivarius. Streptococcus salivarius (ATCC 25975) produced very low or nondetectable amounts of the extracellular enzyme glucosyltransferase (GTase) when grown in a chemically defined medium. The addition of Tween 80 to this medium resulted in the production of markedly enhanced levels of the enzyme. Oleic acid, the methyl ester of oleic acid, and sucrose each could not substitute for Tween 80 in this regard. The surfactant had no direct activating effect on performed enzyme activity. Tween 80 also stimulated the production of GTase by concentrated cells suspended in defined medium during a time when no measurable growth occurred. Under these conditions, the stimulatory effect of Tween 80 was blocked by chloramphenicol. It was further found that the surfactant dramatically stimulated the differential rate of GTase synthesis. These and other data strongly suggest that Tween 80 stimulates the production of extracellular GTase by acting either directly or indirectly at the level of enzyme synthesis.


Composition and Characterization of tRNA from Methanococcus vannielii. Purified bulk tRNA from Methanococcus vanielii (carbon source, formate) showed variation in the modified nucleoside pattern reported for Escherichia coli as analyzed by both ion-exchange and thin-layer chromatography. Ribothymidine and 7-methylguanosine were absent; 1-methyladenosine, 1-methylguanosine, N2-methylguanosine, N2,N2-dimethylguanosine, thiolated nucleosides, pseudouridine, dihydrouridine, and O2'-methylcytidine were quantitated. In vitro methylation by M. Vannielii extracts with S-adenosylmethionine and undermethylated E. coli tRNA revealed active tRNA methyltransferases for formation of methylated residues found in native M. vannielii tRNA, but none for the formation of 7-methylguanosine or ribothymidine. The native M. vannielii tRNA became methylated in the 7-methylguanosine position by E. Coli extracts, but ribothymidine was not formed. Both M. vannielii and E. coli tRNA methyltransferases produced unidentified methylated residues in tRNA's lacking or deficient in ribothymidine.


Fermentation of fumarate and L-malate by Clostridium formicoaceticum. The fermentation of fumarate and L-malate by Clostridium formicoaceticum was investigated. Growing and nongrowing cells degraded fumarate by dismutation to succinate, acetate, and CO2; on the other hand, only small amounts of succinate were detected when the organism was grown on L-malate. This dicarboxylic acid was mainly converted to acetate and CO2. The fermentation balances were modified if bicarbonate or formate were present in the medium. When C. formicoaceticum was grown in the presence of both dicarboxylic acids, fumarate was consumed before L-malate. The latter was mainly converted to acetate, whereas fumarate was fermented to acetate and succinate. Molar growth yields were determined to be 6 g of dry weight per mol of fumarate and 8 g of dry weight per mol of L-malate fermented.


Early stages of arthrospore maturation in Streptomyces. In the sporogenesis of Streptomyces, two basic stages can be considered: sporulation septum synthesis and arthrospore maturation. Most of the information about the ultrastructural changes accompanying the sporogenesis refer to the first stage of the process, but nothing has been published about the evolution of the sporulation septum during maturation. In a previous paper, proposed three basic types of sporulation septum formation in Streptomyces. Our ultrastructural study on the evolution of the sporulation septum during the early stages of arthrospore maturation in seven species of Streptomyces indicates correlation between the sporulation septum type and its evolution during the arthrospore maturation. In types I and II the material of the annuli was incorporated into the lateral walls of the arthrospore, whereas in types II and III the deposits were lysed during the maturation. Only in type III was the arthrospore wall synthesized de novo. In type I there was total integration and in type II there was partial integration of the septum into the arthrospore wall.