Phase III clinical evaluation of gadoteridol injection: experience in pediatric neuro-oncologic MR imaging.

Twenty-two pediatric patients with known CNS neoplasms underwent magnetic resonance (MR) imaging before and after intravenous injection of 0.1 mmol/kg gadoteridol injection as part of a Phase IIIB open label multicenter clinical trial. Intravenous administration of this neutral, nonionic contrast agent was found to be safe in children. No clinically relevant changes in vital signs or laboratory values (including complete blood count, blood chemistry, serum electrolytes, thyroid and metabolic panel and clotting function) were attributed to the administration of gadoteridol injection. There were no systemic complaints. The imaging characteristics of gadoteridol in pediatric CNS disease appeared similar to those of gadopentetate dimeglumine. Contrast enhancement was present in 17 of 22 patients (77%). The administration of gadoteridol injection provided additional clinically relevant information including improved visualization and delineation of the primary lesion, detection of additional lesions, determination of tumor recurrence and narrowing the list of differential considerations in all 17 enhancing studies as well as in 2 of 5 studies without signal intensity enhancement. The very low toxicity, inherent to this nonionic low osmolal paramagnetic contrast formulation may allow administration of increased doses at increased infusion rates for an increased number of indications with improved sensitivity.


Sonographic findings of testicular teratoma with pathologic correlation.

Testicular teratoma is a rare tumor in children. We recently encountered two boys, aged 18 months and 2 years respectively, with this tumor. Sonography revealed an intratesticular mass in both. The first revealed a predominantly cystic lesion with echogenic components along its wall. The second showed a complex mass occupying most of the right testicle, and containing several areas of calcifications and cysts. Excellent pathologic correlation was obtained in both. Preoperative diagnosis of testicular teratoma is possible if the characteristic features are recognized. The prognosis is good following orchiectomy in the pediatric patients.


Contributions to the history of psychology: LXXXV. Jean Piaget, student of Pierre Janet (Paris 1919-1921).

This article provides information about an important period of Piaget's formative years, those he spent in Paris from 1919 to 1921, during which he came into contact with eminent members of the French scientific and philosophical community of the time. Among these was the psychiatrist and psychologist Pierre Janet, whose genetic approach to the psychology of behaviour and ideas about the hierarchical organization of psychological functions converged with Piaget's early scientific interests and provided Piaget a conceptual framework within which to work and develop his own project of studying the genesis of knowledge.


Locus of control and perceptions of human aggression.

116 college students were given a locus of control scale and a measure of misconceptions about human aggression. A negative correlation of -.52 indicated that as feelings of mastery and control increased, belief in a human propensity for aggression decreased.


Conservation of distance in African children from rural and urban schools.

This paper contains a review of the small literature on the association of rural/urban residence and attainment of distance conservation among African children and an account of a study of the age at which children in Sierra Leone, West Africa, attain conservation. 150 children from the ages of 8 to 18 years, from rural and urban schools, were assigned Piagetian tasks to judge their conservation of distance. Analysis showed that at 11 + years, conservation was observed and that there was no significant difference between the urban sample and the rural one. Several confounds in the data limit conclusions but use of the children's own languages was possible in some experiments.


Toe graphaesthesia as a discriminator of brain impairment: the outstanding feet for neuropsychology.

Because of the vulnerability of the medial surfaces of the cerebral hemispheres to the consequences of shear forces, we hypothesized that quantitative sensori-motor deficits for the feet would be strong indicators of general brain dysfunction. On the basis of the Halstead-Reitan Impairment Index, 28 adults who had received closed head injuries were assigned to perfectly normal, normal, or (mildly to severely) impaired groups. Foot tap and finger tap as well as agnosia and graphaesthesia for the fingers and toes were measured. Deficits in toe graphaesthesia were the most powerful group discriminator. Potential usefulness of haptic/motor, finger/toe comparisons for recording the progress of degenerative diseases, such as AIDS, is also suggested.


Symmetry in building block design for learning disabled and nonlearning disabled boys.

This study examined perceptual differences in symmetry with and without a model between 21 learning disabled boys with reading deficits and 28 learning disabled boys with deficits in mathematics. 55 nonlearning disabled boys served as controls. All boys were in Grades 3, 4, or 5. Without a model, all learning disabled boys built significantly more asymmetrical building block designs than nonlearning disabled boys. However, with a model, all learning disabled boys could imitate the model, but it took them longer to complete the task successfully. All boys appeared to have difficulty in modeling asymmetrical tasks, taking more time for task completion after seeing an asymmetrical model. This study suggests modeling perceptual tasks might be an effective teaching strategy for such children.


Improving African children's cognitive development through training.

According to Piaget, cognitive development can be realized with maturation and environmental experience, but he attached some significance to training as a means of facilitating cognitive development. This paper gives brief summaries of eight studies carried out in Africa clearly showing that cognitive development can be facilitated by training.


Hemisphere differences in event-related potentials (ERPS) to monaural presentations of simple speech sounds.

The aim of the present study was to compare ERPs over the left and right hemispheres to monaural consonant-vowel (CV) syllables. It was predicted that, if the contralateral auditory representation is stronger than the ipsilateral one, then ERPs over the left hemisphere should be larger for right-ear stimulus input. Furthermore, if the hemispheres differ in efficiency in processing of speech sounds, then ERPs recorded at any given site should vary as a function of the ear to which the sound is presented. Twelve right-handed subjects participated. The CV-syllables used were /Ba/ and /Pa/ with 15 presentations of each syllable to each ear in a randomized order. EEG was recorded from F3, Fz, and F4 with linked ears as reference. The results showed no significant asymmetry in the ERP-leads. N1- and N2-amplitudes were, however, larger at Fz than at F3 and F4, and N1-latency was shorter for right-ear presentations, which also interacted with the /Pa/-syllable presentations. P3-latency was longer to the /Pa/-syllable compared to the /Ba/-syllable, while N4-latency was longer to the /Ba/-syllable. N4-amplitude was more negative for the /Pa/-syllable presented to the left ear. The results are discussed in terms of phonemic differences between the unvoiced /Pa/ and voiced /Ba/, and early versus late stages of processing. The results are also seen in relation to ear differences in dichotic listening.


Confirmation of Tabachnick and Klugman's hypothesis that the death instinct is constant.

In reporting the accident death rate and the chronic liver disease death rate for 1980, the Bureau of the Census divided the United States into nine areas. To test Tabachnick and Klugman's hypothesis that the amount of death instinct per capita remains constant across regions, the 1980 death rates for accidents and chronic liver disease were correlated. Contrary to earlier studies, the present study gave support for Tabachnick and Klugman's hypothesis.


Perspectives of research participants, psychologist investigators, and institutional review boards.

This study investigated the views of 100 student research participants, 107 psychologist investigators, and 45 members of Institutional Review Boards (Human Subject Committees). Participants in these three groups responded to questions regarding the ethical parameters of a fictitious psychological research protocol. Psychologist investigators' endorsements are similar to those of Institutional Review Board members when the debriefing procedure of the protocol and its potential benefits to others are questioned. The views of psychologist investigators are similar to those of students when the topic of risks to participant is at issue. Implications of these findings for review of research proposals in psychology by Institutional Review Boards are discussed.


Willingness to take legal action in wrongful dismissal cases: perceptual differences between men and women.

146 business students with full-time work experience participated in a study of dismissal from employment. Based on self-ratings, men were more likely than women to favor court action in the event of dismissal.


Relation of anxiety about social physique to location of participation in physical activity.

The purpose of this study was to examine the relationship of anxiety about social physique to location of participation in physical activity. 37 nursing students completed the Social Physique Anxiety Scale and answered questions relating to the location of the physical activity in which they participated. Women were assigned to either a high- or low-anxiety group based on these scores. An examination of the reported location where participation in physical activity occurred showed that more high than low scorers reported a tendency to exercise privately than publicly; the number was higher than expected. Perhaps high scorers prefer exercise settings that provide less opportunity for their physiques to be evaluated.


Biochemical and galvanic skin responses to music stimuli by college students in biology and music.

The purpose of this study was to examine biochemical and physiological responses to musical stimuli. Specifically, university music and biology students' plasma levels of norepinephrine, endorphin, and cortisol, and their galvanic skin responses were measured before and after listening to two different musical selections in an anechoic chamber and during controlled silence. The results indicated that biochemical variables changed significantly in both groups during listening to music but were not different during the controlled silence. These data suggest that music majors may listen more analytically to music. GSR responses were significantly higher for music majors than biology majors, and plasma cortisol increased in music students but decreased in biology students. Music which elicits specific emotions induces physiological changes which may be beneficial to relaxation and behavioral therapies.


Fundamental frequency variability in elderly women during production of stressed and unstressed words.

The purpose of this investigation was to examine intrasubject and intersubject fundamental frequency (F0) variability in a group of young adult and elderly female speakers during the production of stressed and unstressed words. While both groups exhibited greater intersubject variability during stressed versus unstressed productions, stressed productions were more variable in the elderly women than in the young adults. Also, the elderly women exhibited greater intrasubject variability during stressed and unstressed productions relative to young adults. Implications of these findings were discussed.


Effects of listeners' experience on two measures of intelligibility.

10 speech-language pathologists with extensive experience in judging speakers' intelligibility and 10 control subjects with no such previous experience provided written identification and magnitude-estimation scaling judgments of the intelligibility of nine audiotaped speech samples. Analysis indicated no significant main effect for experience on either the written identification or the magnitude-estimation scaling tasks. Implications for the continued use of magnitude-estimation scaling as a measure of speech intelligibility are discussed.


Art training and the Rey figure.

A 55-yr.-old woman who had had a meningioma removed many years previously performed well on various neuropsychological tests but showed apparent gross loss of gestalt in copying the Rey figure, and poor subsequent recall of the figure. Further investigation suggested that her copying style was at least partly attributable to her training in art and that she was capable of copying in a more usual fashion with subsequent improvement in recall.


Consistency of interform content for the Gordon Test of Visual Imagery Control.

A total of 116 undergraduates (49 men, 67 women) completed the Gordon Test of Visual Imagery Control followed immediately by another form. Scores were higher for women than for men and on the original than on the new version. Alternate-form reliability coefficients were .517 for the complete sample, .712 for men, and .210 for women. It was concluded that consistency across content was lower than desirable.


Comment on Nelson, et al.: 'Comparative time estimation skills of Hispanic children'.

Recently Nelson, Smith, Dodd, and Smith reported a dissipation of a cross-cultural difference in time estimation. A number of difficulties in this study are noted and a recommendation for further research is provided. The difficulties include an inappropriate jump to a conclusion about future perspective and the restriction of the dissipation of the cross-cultural difference to Grade 5 children. Research on children beyond Grade 5 would demonstrate the durability of the dissipation reported.


Comparison of Kaufman's short form of the McCarthy Scales of Children's Abilities and the Stanford-Binet Intelligence Scales--Fourth Edition.

21 kindergarten children completed the Stanford-Binet-IV (Binet-IV) and the McCarthy Scales of Children's Abilities. Kaufman's (1977) estimated General Cognitive Index (GCI) was computed from the entire McCarthy scales. There was only one significant difference among the Binet-IV and estimated GCI scores. Abstract/Visual Reasoning was significantly lower than the Kaufman GCI. Correlations of the five Binet-IV Standard Age Scores and the Kaufman GCI ranged from .29 to .67. The correlation between the GCI and the Kaufman estimated GCI was .91.


Addressing the psychosocial needs of orthopedic pain patients.

Analysis of comprehensive examinations of 258 patients with orthopedic pain yielded 5 types of pain responders. Research is under way to analyze the response to pain and effectiveness of response treatment for each type of pain responder.


Differential diagnosis of disturbed adolescents with the Minnesota Percepto-Diagnostic Test-Revised.

Two groups of psychiatric adolescent patients (19 schizophrenics, 22 nonschizophrenics) on an acute inpatient ward of a general hospital were compared for their performance on the Minnesota Percepto-Diagnostic Test--Revised. The effects of admission on test performance were also investigated. Results supported the efficacy of the test in differentiating between the two groups. The schizophrenic adolescents scored significantly less well (higher rotation scores) than the nonschizophrenics. It was shown that the groups' test performance significantly improved from admission to nine days later.


Hemisphericity style, sex, and performance on a mirror-tracing task.

60 right-handed college students classified as preferring either a style of left or right hemisphericity mirror-traced a star pattern once with each hand. Speed of tracing and number of deviations outside the star pattern were the dependent measures. Analysis showed no statistically significant differences in performance between right- and left-style groups or between men and women. However, subjects classified as showing a right-hemisphericity style mirror-traced the star pattern significantly faster and more accurately with the left hand than with the right. The left-hemisphericity-scoring group displayed a nonsignificant tendency to perform better on the reverse pattern.


Perception of loudness and musical preference: comparison of musicians and nonmusicians.

To estimate the relationship between musical preference and perceived loudness, 25 subjects, musicians and nonmusicians, were asked to match the loudness of a neutral stimulus to a given musical selection. Ten different types of music were used; each was rated for likability by the subject. Over-all analysis of loudness ratings indicated that nonmusicians were more accurate in matching loudness of a neutral stimulus with the musical selections. This finding suggests differences in loudness perception between musicians and nonmusicians. Both groups were most accurate on the selection which could be inferred as most familiar.


Effects of four physical education teaching methods on development of motor skill, self-concept, and social attitudes of fifth-grade children.

The purpose of this study was to examine the effects of four teaching methods on several measures of motor skill, self-concept, and social attitudes of 130 fifth-grade children (67 boys, 63 girls), who were randomly chosen from five elementary schools in one area. Teaching methods were systematically applied for 60 teaching days (20 weeks). Measurements were assessed on three occasions (pre-, mid-, and post-experiment). A 4 x 2 x 3 multivariate analysis of covariance showed the 'combined' method most effective in improving the students' motor ability, while 'indirect' and 'game-oriented' methods contributed to the improvement of self-concept and social attitudes, respectively. Since each of the four methods had different effects on various measures made to evaluate whether aims of physical education had been met, the teacher should be aware of many methods and able to implement them in various combinations, depending upon the special features and purposes of a lesson.


Binocularity and photophobia in intermittent exotropia.

The relationship between photophobia and binocular function in intermittent exotropia was examined in 42 subjects (21 patients and 21 controls). In all subjects fusional amplitude and strength of binocular sensory status were tested before and during exposure to bright light. An explanation of photophobia in intermittent exotropia was suggested as impairment in binocular cooperation, since the indicators of binocular function were significantly altered in the patients, particularly during light exposure.


Empirical evidence for assumptions underlying time orientation in undergraduates.

282 undergraduate students between 17 and 68 years of age were asked to list the 5 most significant experiences of their lives, to assign the time zones to these experiences, and to provide estimates of emotional valence corresponding to each significant life experience they listed. Subjects also provided judgements of time perspective on a Life Line. The sample showed a near-past orientation and a positive emotional valence across the experiences reported. However, the first experiences reported were distant past experiences significantly more frequently than expected by chance, while the last experiences in the Experiential Inventory were significantly more often located in the distant future. While this result validates the prevailing assumption of a unidirectional flow of past to future, empirical evidence was also found for the larger magnitude and variability of future as compared with past perspective. This suggests a bidirectional model of time should be invoked to explain the differing character of the opponent temporal processes of recall and anticipation in human experience.


Prison norms for Raven's Standard Progressive Matrices.

Prison norms for the Raven's Standard Progressive Matrices were developed using 1126 male inmates in a prison in Nevada. 556 of the men were white, 480 black, 55 Mexican, 19 Cuban, 9 Asian, and 7 Native-American. Norms were provided for three age categories--under 35 years, age 36 to 54, and all ages combined. Normative information was presented for white inmates, black inmates, and all ethnicities combined. There was substantial overlap in distribution of scores by black and white inmates.


Knowledge and motor performance.

Traditionally, motor skill acquisition has implied that the performance of a given individual on a particular skill is dependent on the amount of prior practice of that skill. However, concepts such as schema theory, or kinetic formulae, or the strategic allocation of resources imply that, even when practising specific skills, performers gain knowledge about their own motor performance which can be used or applied to related or novel situations. An attempt was made to relate the performance of a complex psychomotor task to differing levels of motor skill expertise or knowledge (athlete and nonathlete). 20 subjects performed (1600 responses) on a novel pursuit or tracking task. Analysis indicated that the athletes performed significantly better. Their main advantage appeared to be more in their ability to control and produce fast, accurate movements than in their decision-making. Accepting Henry and Rogers' 1960 proposition that there is no such thing as a general motor ability or coordination factor does not imply that the only alternative is for all motor skills to be specific. It is argued that the differences in the present study arose from the athletes' greater knowledge (schema, kinetic formulae) related to their understanding of their own motor capabilities.


Context effects in recall and recognition at various retention intervals.

This study investigated the differential effects of cuing in a number of contexts on recall and recognition at various retention intervals. Subjects were shown cue-target word pairs and were asked to recall the target words under three sets of cues, varying in relation to the original cue. All 80 subjects completed a recognition task following the final recall task. Results suggest type of cuing does not have differential effects on recall and recognition; both recall and recognition are hindered by cuing in a number of different contexts, and the ability to recall improves over time, cuing, and exposure to the list.


Use of subliminal stimulation to enhance learning mathematics.

24 students, who participated in a summer session mathematics enrichment program, viewed subliminally presented messages. Prior to each class, the experimental group of 16 viewed a subliminally presented psychoanalytic merging message and the control group of 8 viewed a neutral message. After 20 sessions students were retested. Experimental students had higher mathematics scores than controls. These data are compared with results obtained by other investigators.


Comparison of parents' leadership styles: perceptions of parents and student leaders.

The Parent Leadership Style instrument was completed by 99 students enrolled in a leadership program and also by 129 of their parents. Data were analyzed and reported according to predominant leadership style (Telling, Selling, Participating, and/or Delegating) and effectiveness of leadership. A comparison was made between the parents' self-perceptions and how their children as student leaders perceived their parents. Some suggestions for parents are made relative to the development of leadership skills in students.


Differentiation between valproate-induced anticonvulsant effect, teratogenicity and hepatotoxicity. Aspects of species variation, pharmacokinetics, metabolism and implications of structural specificity for the development of alternative antiepileptic agents such as delta 2-valproate.

Valproate is metabolized into a large number of compounds via various metabolic routes. Metabolic profiles depend on species and age. Hepatotoxicity may be correlated with abnormal metabolism, especially in young age. Teratogenicity is associated with specific structural requirements: a free carboxyl atom connected to a carbon atom which also carries a hydrogen, and two carbon chains. This provides a clue for the development of alternative antiepileptic agents.


Influence of co-medication on the metabolism of valproate.

Valproate is extensively metabolized in the liver and at least six main pathways which produce about 50 metabolites have been identified in man. The enzyme-inducing antiepileptic drugs phenobarbital, primidone, phenytoin and carbamazepine increase total valproate clearance by 30-85%, whereas cimetidine and the new anticonvulsant compound striripentol display a small inhibitory effect (10-20%). Both carbamazepine and phenytoin induce a two-fold increase in the formation of delta 4-valproate and stimulate omega-oxidation and omega-1-oxidation. Acetylsalicylic acid causes a fall of 60-70% in the content in the urine of the metabolites of the beta-oxidative pathway, i.e. delta 2-valproate, 3-OH-valproate and 3-oxo-valproate, and an increase of glucuronidation (approximately 30%) and delta-dehydrogenation (approximately 20%). Stiripentol inhibits the formation clearance of delta 4-valproate by 30%. In the light of the possible therapeutic and toxic effects of some valproate metabolites, drug interactions with valproate at metabolic level may have important clinical implications.


Pharmacokinetics of valproate in pregnancy: mother-foetus-newborn.

An increased risk of seizures during and immediately after labour has been observed in epileptic women, and it is recognized that serum levels of antiepileptic drugs may decrease in pregnancy. Several studies have suggested that total valproate levels fall, but that free fractions increase during pregnancy. Recent findings suggest that the actual metabolism of valproate is not altered by pregnancy and that the changes of the plasma clearance are due primarily to decreased protein binding. The levels of free drug will not change significantly as pregnancy advances. However, dose reduction after delivery may be necessary to avoid toxicity. Valproate and its metabolites undergo placental transfer. In the foetus the plasma level of valproate and the protein binding are higher than in maternal plasma, and the half-life of valproate following placental transfer is considerably longer than in adults. Only small amounts of valproate appear in breast milk and those are not likely to cause any problems. During pregnancy and the first month after delivery preferably both total and free valproate serum levels should be closely monitored to determine the lowest effective dose.


Valproate hepatotoxicity syndrome: hypotheses of pathogenesis.

Therapeutic use of the anticonvulsant valproate (VPA) has been associated with a rare, but severe and often fatal hepatotoxicity. Cases usually present with lethargy, anorexia, and vomiting with rapid progression to coma. Liver histopathology is characterized by steatosis with and without necrosis. In some instances only necrosis was present. Several hypotheses of pathogenesis have been postulated. These deal mainly with biochemical systems that are known to be affected by VPA, or with the possible idiosyncratic production of toxic VPA metabolites, especially delta 4-VPA. At present, no hypothesis entirely explains the diverse characteristics of the disorder.


Metabolism of valproate to hepatotoxic intermediates.

A number of lines of evidence indicate that metabolites of valproate rather than the parent drug, mediate the microvesicular steatosis which characterizes valproate-associated liver injury. In this article, two mechanisms are discussed whereby valproate may cause hepatic steatosis through interference with the process of fatty acid beta-oxidation. In the first, valproate itself enters the mitochondrion where it completes for the enzymes and/or co-factors involved in the beta-oxidation of endogenous substrates, while in the second, valproate is metabolized via the hepatotoxic terminal olefin, delta 4-valproate, to a variety of chemically reactive intermediates which inhibit key enzymes in the beta-oxidation cycle.


Effects of valproate on xenobiotic biotransformation in rat liver. In vivo and in vitro experiments.

Male Wistar rats were in vivo exposed for 2 weeks to 100 micrograms/ml sodium valproate by subcutaneous implantation of osmotic pumps and hepatocytes were isolated. As an in vitro model co-cultures of rat hepatocytes with epithelial cells were daily treated with valproate (25, 50, 100, 200 micrograms/ml) for 2 weeks. In both models the cytochrome P-450 content and the enzymatic activities of 7-ethoxycoumarin O-deethylase, aldrin epoxidase and glutathione S-transferase were determined in valproate-treated hepatocytes, in controls and in phenobarbital-induced cells. It appeared that in both systems the cytochrome P-450 content and the 7-ethoxycoumarin O-deethylase activity increased significantly after valproate treatment. On the other hand, the activities of aldrin epoxidase and glutathione S-transferase decreased. A cDNA probe, encoding rat P450IIB2 was used to determine whether mRNAs encoding the P450IIB subfamily were induced by valproate. It became clear that the inducing effect of valproate was even more pronounced in vitro than in vivo.


Strategies for identifying and developing new anticonvulsant drugs.

The identification of new anticonvulsant drugs depends on the use of different animal models of epilepsy. The models should be mechanism-independent, able to screen a large number of compounds, at limited cost and technical expertise. Primary screening models include genetic or reflex models of epilepsy and electrically and chemically induced seizures. Once active compounds have been identified, more advanced mechanistic and seizure-specific models are needed to refine the choice of a lead compound. These can be either in vivo or in vitro models. Models known to interact with specific receptors or the production of the putative neurotransmitters of neural excitability or inhibition are valuable in assessing possible mechanisms of action. In vitro models have evolved as important tools in correlating changes in electrical phenomena and therapeutic spectrum. The use of the hippocampal slice and the cultured neuron permits classification of anticonvulsant activity based on cellular actions of the drug. Interactions by the experimental drugs with specific subcellular fractions of the central nervous system augment information on possible mechanisms of action. The final choice of compounds for development requires synthesizing and comparing all of the pharmacodynamic information with the pharmacokinetic and toxicologic data. In the final analysis, no single animal model of epilepsy known today can assure the development of better drugs for all treatment of the epilepsies.


Pharmacological, toxicological and neurochemical effects of delta 2(E)-valproate in animals.

The E isomer of 2-ene-valproic acid (delta 2(E)-VPA) is the major active metabolite of the antiepileptic drug valproate (VPA) in various species, including humans. Experimental studies on delta 2(E)-VPA and VPA indicate that delta 2(E)-VPA may be a useful antiepileptic drug itself. delta 2(E)-VPA has the same wide spectrum of anticonvulsant activity as VPA with a somewhat higher anticonvulsant potency in rodent and dog models of different seizure types. As VPA, delta 2(E)-VPA increases presynaptic gamma-aminobutyric acid (GABA) levels in the brain, presumably by an effect on GABA synthesis and/or GABA degradation. delta 2(E)-VPA is a much more potent inhibitor of the human brain GABA-degrading enzyme than VPA. In high doses delta 2(E)-VPA is more sedative in rodents than is VPA; LD50 values are about the same. In mouse and rat models for teratogenicity, delta 2(E)-VPA does not induce teratogenic effects, whereas VPA is teratogenic in these models. Pilot rat studies on liver toxicity of VPA and VPA metabolites suggest that delta 2(E)-VPA is not hepatotoxic. In view of the rare but serious hepatotoxicity and teratogenicity of VPA in humans, delta 2(E)-VPA obviously merits interest as a valuable alternative drug in antiepileptic therapy.


Delta 2-valproate biotransformation using human liver microsomal fractions.

The metabolism of 2-n-propyl-2-pentenoate (delta 2-VPA) was evaluated in human hepatic microsomal fractions. Two biotransformation pathways have been particularly investigated. In the presence of the cytochrome P-450 co-factor, NADPH, the main metabolites recovered were delta 3-VPA, delta 2,4-VPA and VPA. The glucuronidation of delta 2-VPA was also studied on various hepatic microsomal fractions using Brij 35 as activator and UDP-glucuronic acid as co-factor. A large interindividual variability occurred in this metabolic pathway. Km and Vmax were 0.85 mmol/l and 1.75 nmol.min-1.mg-1, respectively, for delta 2-VPA and 1.11 mmol/l and 5.71 nmol.min-1.mg-1 for VPA, respectively. The good correlation (r = 0.82; p less than 0.001) observed between the glucuronidation of VPA and delta 2-VPA as well as the mutual inhibition of each other's glucuronidation strongly suggests that (a) common single UDP-glucuronosyltransferase isoenzyme(s) was (were) involved in this glucuronidation step. The glucuronidation of specific substrates for various UDP-glucuronosyltransferase isoenzymes showed a good relationship between the glucuronidations of delta 2-VPA and morphine, a substrate for UDP-glucuronosyltransferase-2B. Moreover, morphine competitively inhibits delta 2-VPA glucuronidation. It seems the same isoenzyme or, at least, (a) very closely related isoenzyme(s) belonging to UDP-glucuronosyltransferase-2 isoenzyme, is involved in delta 2-VPA glucuronidation.


Single-dose tolerance and pharmacokinetics of 2-n-propyl-2(E)-pentenoate (delta 2(E)-valproate) in healthy male volunteers.

2-n-Propyl-2(E)-pentenoic acid (delta 2(E)-valproate) was administered to healthy volunteers in oral doses of 50-800 mg. The drug was tolerated well and no significant adverse effects were observed. Pharmacokinetic parameters were determined. Valproate and 3-keto-valproate were detected as metabolites.


A double-blind, placebo-controlled multicentre trial of ticlopidine in patients with peripheral arterial disease in Argentina. Design, organization and general characteristics of patients at entry. The EMATAP Group.

Patients with atherosclerotic disease of the lower limbs are at high risk of thrombotic events not only in the peripheral circulation, but also in the coronary and cerebral arteries. Ticlopidine, a potent inhibitor of platelet aggregation, can reduce the incidence of arterial thrombosis in patients suffering from intermittent claudication, as shown by a meta-analysis performed in 1987. In order to confirm these findings, we undertook a randomized, stratified, placebo-controlled, double-blind multicentre trial in Argentina in two parallel groups of patients suffering from intermittent claudication. Twenty-one clinical centres participated in the trial and 615 patients were enrolled. Patients were eligible if they had presented obstructive arterial disease of the upper part of the lower limb (popliteal or above) for at least 12 months, confirmed by either angiography or Doppler studies, and intermittent claudication (stage II) assessed by treadmill testing. Non insulin treated diabetic patients were eligible if they fulfilled the other inclusion criteria and two strata, diabetic and non diabetic, were established. Patients enrolled were treated with either ticlopidine (500 mg/day) or placebo for 24 weeks. In this paper, we report the design and organization of the study and the baseline characteristics of the patient population at inclusion.(ABSTRACT TRUNCATED AT 250 WORDS)


Decreased G3PDH binding to erythrocyte membranes in sickle cell disease.

Several membrane abnormalities have been described in red cells from patients with sickle cell disease, responsible for chronic hemolytic anemia. We describe here a 35-50% inhibition of the binding of glyceraldehyde-3-phosphate dehydrogenase (G3PDH) to the membrane of sickle red cells. Varying the phosphorylation state of the membrane proteins did not change their affinity for the enzyme. Protein band 3 and the cytoplasmic domain of this protein isolated from sickle red cells showed normal interaction with the enzyme. The inhibition observed with intact membranes is not due to short term oxidation of membrane proteins, as various procedures inducing acute oxidative stress in normal membranes did not reproduce the inhibition of G3PDH binding. We conclude that the alteration of the binding of G3PDH to the membrane of sickle erythrocytes is probably related to long term processes involving cycles of HbS polymer formation.


Age and sex matched analysis of Hb Lepore trait in a new population in Spain.

A group of subjects with Hb Lepore trait has been found in the region between Extremadura and Toledo in Spain. Clinical, radiological and hematological studies were carried out on 81 cases from 23 families. Asthenia was the sole complaint in seven of forty cases. Abdominal echography showed no cholelithiasis in 16 children under 16 years. Hb Lepore mean was 10.81 +/- 1.97%, range 6.5 to 16.1%, Hb A2 levels were normal and Hb F values were high. Globin chain synthesis in reticulocytes showed a total alpha/beta ratio of 1.89 +/- 0.3. Hematological values from Hb Lepore trait subjects were analyzed according to age and sex and the data compared to beta thalassemia and delta-beta thalassemia cases of matched age and sex. Hb Lepore trait patients had a milder form of thalassemia minor than beta thalassemia patients, with higher levels of hemoglobin, MCV and MCH for all three groups: children under 13 years, males over 14 years and females over 14 years. Children and females with Hb Lepore had higher hemoglobin levels than those with delta-beta thalassemia minor, while no significant difference was found in males.


Chromosome studies in stimulated lymphocytes of 50 patients with B-cell chronic lymphocytic leukemia.

The chromosome constitutions of stimulated lymphocytes from 50 B-cell chronic lymphocytic leukemia patients were studied using different stimulation systems, i.e., TPA alone or associated with different cytokines. Adequate metaphases were obtained in 44 subjects (88%). Among 20 patients with abnormal karyotypes (45.5%), 7 had trisomy 12. The most frequent structural abnormality was a 14q+ resulting from translocations including one t(11;14) and two t(14;17), while deletions on the long arms of chromosomes 6 and 13 constituted a second common alteration. The most important finding in this series was the recurrence of a t(18;22) observed in two cases.


Inhibition of human endothelial cell growth by human monocytes in coculture.

Previous studies conducted in different experimental models have shown that human monocytes (Mo) possess both stimulating and inhibiting factors for endothelial cell growth. Since Mo are frequently found in the vicinity of endothelial cells, we examined the direct effect of human peripheral blood Mo on the proliferation of human umbilical vein endothelial cells (HEC) in in vitro culture systems. Mo obtained either by counter centrifugation elutriation or by selective adhesion on gelatin-autologous plasma or purified fibronectin coated plastic surfaces inhibited HEC growth as determined by HEC count or 3H-methyl thymidine incorporation. This growth inhibitory effect was dependent on the number of monocytes: 50% inhibition of 3H-thymidine uptake by HEC was achieved for a Mo-EC ratio of 1:4. Mo isolation by selective adhesion on gelatin-plasma resulted in a significantly higher inhibitory effect (p less than 0.02) than that observed with Mo isolated on fibronectin coated surfaces or by elutriation techniques, suggesting a possible stimulation of Mo by contact with gelatin-plasma. Endothelial cell growth factor plus heparin did not reverse this inhibition of HEC growth. Results obtained in diffusion chamber and coculture systems showed that soluble products of Mo origin can inhibit HEC proliferation but the inhibition was greater when Mo and HEC were in contact. The effect was not due to cytotoxicity as evaluated by cell counting and 51Chromium release from HEC. These in vitro results suggest that normal monocytes could exert a regulatory role on the proliferation of endothelial cells.


Streptococcal bacteremia in neutropenic adult patients.

In order to define the circumstances of occurrence, clinical presentation and prognostic factors of streptococcal bacteremia in neutropenic patients, we retrospectively reviewed 60 cases of streptococcal bacteremia following intensive chemotherapy for acute myeloblastic leukemia (AML), acute lymphoblastic leukemia (ALL) or allogeneic bone marrow transplantation. Causative streptococcal species included streptococcus viridans (42 cases), streptococcus faecalis (12), streptococcus pneumoniae (4) and other streptococci (2). All patients were febrile and 32% presented diffuse pneumopathy with frequent isolation of streptococci from bronchoalveolar washing fluids. There were no statistical differences in presentation between the bacteremia caused by different streptococcal species. Death occurred in 22% of the patients and factors favoring poor prognosis included pneumopathy (p less than 0.001), more than 2 positive blood cultures (p less than 0.01) and initial chemotherapy for AML (p less than 0.01). Pneumopathy occurred more frequently after chemotherapy for AML. It is concluded that streptococcal bacteremia is to be prevented in patients undergoing intensive chemotherapy, particularly if cytarabine is used.


A national Italian survey in hemocytometry: first year report.

The paper presents the results of the first year of an Italian nation wide quality control programme for hemocytometric tests, involving public laboratories and concerning the following parameters: hemoglobin, erythrocytes, leukocytes (nucleated cells), mean corpuscular volume, platelets and hematocrit. Results are compared principally with those obtained from the French national programme owing to the use of identical blood standards.


Plasma prekallikrein, coagulation and fibrinolytic parameters in myeloma patients.

Plasmatic prekallikrein, factor XII, factor XI, C1-inactivator and alpha 2-macroglobulin were evaluated in myeloma patients. Results suggest no modification in the contact phase since there was no significant difference between patients and controls. Other parameters of blood coagulation and fibrinolysis (APTT, TT, fibrinogen, factor V, ELT and LAFP) gave findings consistent with those described in the literature.


Evaluation of the erythrocyte and platelet morphological analysis given by the hematology analyzer Coulter STKS.

The morphological profiles of red blood cells (RBC) and platelets (Plt) derived from the distribution analysis given by the Coulter STKS were evaluated in two groups of patients suffering from non hematological (n = 293) and hematological (n = 257) conditions. The RBC and Plt flags were studied in terms of sensitivity of the morphological analysis and specificity and significance of each flag. When all RBC and Plt flags were considered, the percentages of false negatives and false positives in our subjects were found to be 4.7% and 13.4% respectively, with a global efficiency of 81.8%. The sensitivity of the alarm system was higher than 90% for all types of abnormality, except microcytosis (81%) and Howell-Jolly bodies (57% over a limited number of 7 cases). The specificity of the STKS response was found to be low except for anisocytosis (88.5%) and macrocytosis (86.1%). It was shown that the flags microcytosis and/or hypochromia and macrocytosis were poorly significant when they appeared in isolation (false positive rates of 86.6% and 84.2% respectively). Thus, these alarms could be eliminated from the review criteria. When considering only the flags anisocytosis, NRBCs, micro RBCs/RBC fragments, dimorphic RBC pop, PLT clumps and giant Plt, the percentage of false positives was 8.1%. However, it must be kept in mind that suspect leucocyte flags remained review criteria, resulting in a final false negative rate of 2.5% for RBC and Plt morphological abnormalities.


Malignant melanoma and hairy cell leukemia. Two cases.

The authors report two cases of malignant melanoma associated with hairy cell leukemia. Skin neoplasia preceded hematological malignancy in the first observation. Among reports concerning the association of malignant melanoma with hematological diseases, chronic lymphocytic leukemia, Hodgkin's lymphoma and non Hodgkin's lymphoma are preponderant. Epidemiological studies would be of value to predict the expected risk of malignant melanoma in hairy cell leukemia.


Juvenile chronic myelocytic leukemia with unusual cytogenetic clonal evolution.

Cytogenetic studies are reported in a case of juvenile chronic myelocytic leukemia with dysmyelopoiesis and skin involvement. The clonal evolution of a 6q-anomaly is described. Hematological and cytogenetic findings suggest a role of hematopoietic stem cell in this patient for whom the outcome was fatal.


Malignant thymic lymphoblastic lymphoma and myasthenia gravis: an exceptional association.

Thymic lymphoblastic lymphoma and myasthenia gravis rarely coexist. Only two cases have been reported and we describe here a third case. A 60 year old man presented a typical history of myasthenia gravis, confirmed by neurological investigations including electromyography. Chest X-ray revealed an anterior mediastinal tumor. At thoracotomy, a 60 mm mass adherent to the pericardium was excised and a lymphoblastic lymphoma was diagnosed. The lymphogram showed enlarged pelvic and abdominal lymph nodes consistent with lymphoma. A m-BACOD chemotherapy regimen gave rapid and complete remission of both lymphoma and myasthenia gravis and the patient is now alive 25 months after the start of chemotherapy with no evidence of disease.


Assessing health needs and measuring patient satisfaction.

This paper describes two aspects of the NHS reforms--an emphasis on needs assessment and patient satisfaction. The latter is illustrated with reference to day surgery.


The effects of various dental procedures and patient behaviors upon nitrous oxide scavenger effectiveness.

This prospective study of 36 children, ages 44-93 months, receiving nitrous oxide-oxygen under standardized conditions during routine dental procedures was conducted to determine what influence eight selected dental procedures and three patient behaviors had on ambient nitrous oxide (N2O) levels in the dentist's breathing zone. Half the children received nitrous oxide-oxygen via a scavenging nasal mask. An infrared spectrophotometer analyzed the ambient N2O level continuously throughout the procedure and the time-weighted average (TWA) for consecutive 15-sec intervals was recorded by a microprocessor. A video camera was used to synchronize the coded dental procedures and patient behaviors to the TWA ambient N2O levels recorded. The results demonstrated that scavenging significantly reduced the dentist's exposure to ambient N2O (P less than 0.05, nonscavenged mean = 284.7 ppm; scavenged mean = 36.6 ppm), but the mean TWA N2O concentration remained significantly higher, P less than 0.05, than the 25-ppm level recommended by the National Institute for Occupational Safety and Health (NIOSH). Supplemental oral evacuation was the only dental procedure capable of reducing ambient N2O to below NIOSH's recommendation when scavenging was employed. The administration of local anesthesia consistently created a significant increase in ambient N2O levels. Patient behaviors of talking, crying, and movement also resulted in significant increases from baseline ambient N2O levels.


Frequency of alveolar bone loss adjacent to proximal caries in the primary molars and healing due to restoration of the teeth.

The frequency of alveolar bone loss adjacent to extensive proximal caries, and the effect of dental restorations on alveolar bone loss and healthy alveolar bone were examined in human primary molars. Proximal caries, contact loss, mesial drift and the presence of alveolar bone loss were recorded from 190 bite-wing radiographs from 60 boys and 46 girls. Proximal caries was evident in 297 quadrants. In 63.0% of quadrants, both primary molars had proximal caries. Contact loss was evident in 38.4% of the quadrants with proximal caries. Bone loss was found in 12.1% of the quadrants with proximal caries or 31.6% of those with contact loss. Analysis of variance for the presence of bone loss indicated statistically significant values (P less than 0.05) for one or two teeth with caries by quadrant, the presence or absence of contact loss, and age. A second examination was available for 41 children. Among these, eight out of the 36 bone defects disclosed at the first examination were present. At the second examination, after tooth restoration, healing of five bone defects was evident. These findings indicate a connection between the presence and treatment of extensive proximal caries and alveolar bone health in the primary dentition.


Class IV preparation design for microfilled and macrofilled composite resin.

The current standard preparation for Class IV composite restorations is the placement of a bevel on all enamel margins. This study evaluated chamfered and beveled preparations for Class IV restorations of lesions with microfilled and macrofilled composite resin. Forty incisors were obtained and standardized lesions for Class IV restorations were formed. Twenty teeth had a 1.5-mm bevel placed and 20 had a 1.5-mm chamfered preparation placed. Half of the beveled and chamfered preparations were restored with microfilled composite resin; the remaining were restored with macrofilled composite resin. All restorations were fractured with an Instron Testing Machine. The mean force (lbs +/- SD) to fracture the restorations were: (beveled, microfilled composite 16.0 +/- 4.4); (chamfered, microfilled composite 30.6 +/- 20.0); (beveled, macrofilled composite 34.9 +/- 18.6); (chamfered, macrofilled composite 48.8 +/- 14.3). The chamfered preparations provided greater restoration fracture resistance than beveled preparations, for both microfilled and macrofilled composite restorations. Scheffe's test indicated traditional beveled, microfilled Class IV composite resin restorations significantly decreased fracture resistance compared to chamfered, macrofilled composite restorations (P less than 0.001).


Evaluation of the effects of orthodontic pacifiers on the primary dentitions of 24- to 59-month-old children: preliminary study.

This study was designed to compare the occlusions of 24- to 59-month-old children who used orthodontic or conventional pacifiers to the occlusions of a group of controls who had no sucking habits. Information on the habits was collected by parental questionnaires. Ninety-five children were examined for malocclusions involving overbite, overjet, canine, and molar relationships, and posterior crossbites. Users of orthodontic pacifiers had statistically significantly greater overjets, and there was a significantly higher proportion of subjects with open bite in the conventional pacifier group. There was a trend toward a greater number of subjects in the control and orthodontic pacifier group with overbites less than or equal to 50%. These differences were not clinically significant, however. There appeared to be only minor differences between the occlusions of the two pacifier groups.


Influence of vitamins and iron on plasma fluoride levels in rats.

Thirty female Sprague-Dawley rats (250 +/- 10 g) were divided randomly into five equal groups. After overnight fasting, a silastic catheter was placed in the jugular vein of each rat. Each group was intragastrically administered 0.25 mg F/250 g rat weight in 1 ml of one of the following forms of fluoride supplements: Pediaflor (Abbott/Ross, Columbus, OH), Tri-Vi-Flor, Tri-Vi-Flor + iron; Poly-Vi-Flor or Poly-Vi-Flor + iron (Mead Johnson Nutritionals, Bristol-Myers Squibb Co., Evansville, IN). Timed blood samples were collected and plasma fluoride concentration was determined using the microdiffusion method. The presence of iron and vitamins affect the bioavailability of fluoride as measured by the area under the time-plasma fluoride concentration curve.


Use of a bonding agent to reduce sealant sensitivity to moisture contamination: an in vitro study.

Moisture contamination of etched enamel during application of sealant is the most frequently cited reason for sealant failure. In preliminary studies, a dentin bonding agent (Scotchbond Dual Cure, 3M, St. Paul, MN) has been shown to bond adequately to etched enamel after salivary contamination. This study investigated bond strength in vitro, when a bonding agent was used beneath sealants under varied conditions of contamination. Five hundred bovine incisor crowns were separated randomly into eight groups. The enamel samples, etched for 60 sec with a 37% phosphoric acid gel, were contaminated with: 1) fresh whole saliva, air dried, 2) fresh whole saliva, left wet, or 3) moisture from a humidity chamber. All contamination conditions were tested for sealant bond strength with and without the bonding agent as an intermediate layer under the sealant. As controls, both sealant and bonding agent under sealant also were applied to clean etched enamel. Bond strength was measured using a universal testing machine. Data were analyzed using a two-way ANOVA. Under conditions of humidity or intact saliva, sealant alone showed significant reduction in bond strength (P less than 0.001). Bonding agent under sealant on wet contamination yielded bond strengths equivalent to the bond strength obtained when sealant was bonded directly to clean, etched enamel. Bonding agent used without contamination yielded bond strengths significantly greater than the bond strength obtained when using sealant alone without contamination (P less than 0.001). When the saliva was air dried onto the surface, there was no significant difference in bond strengths whether or not a bonding agent was used under the sealant.


Dental caries in relation to nutritional stress in early English child populations.

This research studied an association between dental caries prevalence in the primary dentition and cribra orbitale, an indicator of nutritional stress in archeological populations. Skeletal remains of 221 British children (Pre-Roman to Late Medieval) were examined, of whom 100 had orbital material available. Dental caries, as dmfs, was related to the presence of cribra orbitale (none, slight, moderate, or severe). Thus, data for 50 children showed a statistically significant association (P less than 0.005) between caries prevalence and cribra orbitale using the Fisher Exact Test, suggesting that nutritional stress is an associated factor in caries etiology in children.


Nutritional value of some non-conventional plant foods of India.

Thirteen non-conventional foods including fruits, leaves and grains consumed in various parts of the Indian subcontinent were analysed for their nutritional value. Khejri beans (Prsopsis cineria), Pinju (Capparis decidua) and Kachri (Cucumis species) contained considerable amounts of protein (15-18%). Kachri was rich in fat (13%). Bhakri (Tribulus terristris), Gullar (Ficus glomerata) and Peehl (Salvadora oleoides) were found to be rich sources of calcium; Gullar contained about 15 times the amount of calcium present in wheat. Phosphorus content of Santhi (Boernavia diffusa), Khejri beans, Bhakri, Pinju and Lehsora (Cordia dichotoma) were noticeable. Zinc was present in high amounts in Peepalbanti (Ficus religiosa) and Gullar; as was iron in Santhi and Bhakri and manganese in Santhi. Besides iron, zinc and calcium, Pinju contained appreciable amounts of beta-carotene and vitamin C. However, Santhi contained high amounts of oxalic acid.


Physico-chemical composition and characterisation of the seed and seed oil of Sclerocarya birrea.

The physicochemical composition of Sclerocarya birrea was assessed by standard methods and was found to contain 11.0% Crude oil, 17.2% Carbohydrate, 36.70% Crude protein 3.4% fibre and 0.9% crude saponins. The fatty acid distribution in the seed oil was obtained by fractionating the volatised fatty acid by GC-MS. The oil is made up of nine fatty acids of which palmitic, stearic and arachidonic acids are the most dominant.


The effect of fermentation on the nutrient status and on some toxic components of Icacinia manni.

The effect of fermentation on the nutrient status and on some toxic components of Icacinia manni was investigated. Chemical analysis of both unfermented and fermented products revealed an increase in protein, ash and fibre content while the lipid and carbohydrate content showed a decrease. The results indicated that fermentation resulted in protein enrichment of the fermented Icacinia manni mash. Fermentation was also observed to cause a marked decrease in the level of some toxic components (oxalic acid, phytic acid and hydrocyanic acid) of the product. The possibility of incorporating Icacinia manni among the edible starchy plant tubers is discussed.


Protein quality of developed home made weaning foods.

Home made weaning foods developed from locally available foods like bajra, barley, green gram (Vigna radiata L.), amaranth grain (Amaranthus sp.) and jaggery using household technologies like roasting and malting had a PER ranging from 2.04 to 2.13, BV 79.56 to 80.68, NPU 66.75 to 67.86, NPR 2.13 to 2.76 and PRE 34.18 to 44.18. The values were comparable to that of cerelac--a commercial weaning food.


The nutritive quality of sorghum-commonbean tempe.

The nutritive quality of sorghum-commonbean (40:60) tempe manufactured by Rhizopus oligosporus: Rhizopus oryzae (1:1) mixed culture fermentation was determined. The protein, crude fat and ash content increased slightly, while carbohydrates decreased. The dietary fibre of the tempe increased by 10%. Mould fermentation increased the content of reducing sugars, total acid and aminonitrogen 15.3, 6.7 and 4.6-fold, respectively. It decreased the phytate content by 44% and it increased the tannic acid content by 52%. In vitro iron absorption increased from 2.8 to 12.5%. The protein efficiency ratio of tempe was 1.61 +/- 0.33; the net protein ratio was 2.39 +/- 0.20; the in vitro and in vivo protein digestibility were 88.2 and 80.0 +/- 0.05% respectively, while the protein efficiency ratio, net protein ratio in vivo digestibility of skim milk was 2.96 +/- 0.17, 3.51 +/- 0.17 and 98.0 +/- 1.87, respectively. The sorghum-bean tempe could be used for supplementary feeding.


Certain functional properties of defatted pumpkin seed flour.

Defatted pumpkin (C. pepo and C. maxima) seed flour has potential food uses because of its high protein content, 61.4 +/- 2.56%. The functional and electrophoretic properties of the defatted flour were investigated. Polyacrylamide gel electrophoresis and electrofocusing indicated 14 bands of water-soluble protein subunits with isoelectric points between 3.81-8.08 and apparent molecular weights between 19,200 and 97,000 daltons. Minimum nitrogen solubility was observed at pH values between 3.0-7.0 and exceeded 90% at pH above 9.0. Solubility was a function of ionic strength. It appeared that, even at the pH of minimum solubility, the pumpkin seed proteins could be dissolved up to high concentrations by increasing NaCl molarity. The viscosity of flour-water dispersion was affected by flour and salt concentrations, and temperature. The least gelation concentration was 8% (w/v) and the water and oil absorption 24.8 +/- 2.03 and 84.4 +/- 4.05 g/100 g respectively. Sorption isotherms, BET monolayer moisture and binding energy of sorption were also calculated. Both foam capacity and stability were pH dependent.


In vitro enzymatic hydrolysis of protein and protein pattern change of soya and faba beans during germination.

In addition to technological processes like heat treatment, germination can be an alternative process for the improvement of protein quality of legumes. This was demonstrated by enzymatic protein hydrolysis of flour of germinated faba and soya beans, using a pepsin-pancreatin enzyme system. SDS-PAGE was used to study the changes in protein pattern of these legumes during germination. In addition, the effect of germination on the content of condensed tannins in flour from germinated faba beans and trypsin inhibitors in flour from germinated soya beans were studied. Germination for five days resulted in a maximum increase in enzymatic protein hydrolysis by 21.3% in flour from faba beans and by 25.7% in flour from soya beans after 12 hours of germination. Protein patterns, obtained with SDS-PAGE demonstrated a considerable protein breakdown during germination between day 2 and 3 in faba beans and between day 1 and 2 in soya beans. The tannin content in flour from faba beans decreased by 29.7% after seven days of germination, but the tannin content of the hulls of the faba beans did not change during that period of germination. The trypsin inhibitors in flour from soya beans decreased by 25.5% after seven days of germination. We conclude that the increased enzymatic hydrolysis of protein in both legumes cannot be explained by a decrease of tannins or trypsin inhibitors. The possible explanation is that through degradation of proteins during germination of the legumes, the cleaved protein fragments are more susceptible for hydrolysis by pepsin-pancreatin.


Expression of transferrin receptors during differentiation of human placental trophoblast cells in vitro.

In most cell types, transferrin receptor expression is correlated with the proliferation rate, being increased by growth stimulation, or decreased by induction of terminal differentiation. In the human placenta the multinucleated syncytiotrophoblast, in direct contact with maternal blood, is derived by differentiation from mononucleated cytotrophoblast. In this study we examined changes in transferrin receptor expression during in vitro differentiation of trophoblast. Cells cultured in Ham's/Waymouth's medium (HWM) remained primarily mononuclear throughout the study, whereas incubation in keratinocyte growth medium (KGM) led to formation of multinucleate masses within 2-3 days of culture. Cell surface binding of 125I-labelled transferrin increased fivefold between days 1-5 of culture in both media and surface receptors were saturated at 7-14 micrograms/ml (90-200 fM). At saturation, the amount of transferrin bound to syncytiotrophoblast was 37 per cent lower than in cytotrophoblast. Scatchard analysis revealed a reduction in the number of surface transferrin receptors in syncytiotrophoblast compared to cytotrophoblast. A corresponding 29 per cent reduction in the binding of transferrin to intracellular sites was observed in syncytiotrophoblast. Distribution of receptors between surface and intracellular sites was therefore similar in both cytotrophoblast and syncytiotrophoblast. The affinity of transferrin for transferrin receptors was 3.7-fold higher in syncytiotrophoblast when compared to cytotrophoblast. Observed differences between the two cell types were not due to the presence of growth factors or higher iron levels in KGM. Expression of a high number of surface transferrin receptors in syncytiotrophoblast (1.5 x 10(12)/mg protein), along with a high affinity of these receptors for iron-saturated transferrin, could help explain the efficient transport of large amounts of iron from mother to fetus.


Overexpressed full-length human BCL2 extends the survival of baculovirus-infected Sf9 insect cells.

Full-length and truncated human BCL2 lacking the entire C-terminal hydrophobic domain have been overexpressed in Spodoptera frugiperda insect cells with the baculovirus expression system. Immunoblot analysis with BCL2-specific antibodies revealed that both full-length and truncated BCL2 are expressed as multiple immunoreactive species, suggesting posttranslational modifications. The expression of the full-length but not the truncated BCL2 extended the survival of baculovirus-infected cells by preventing virus-induced DNA cleavage. This result is consistent with the reported protective effect of BCL2 against apoptosis in mammalian lymphocytes and suggests a conserved function in evolution. Subcellular fractionation and indirect immunofluorescence studies in intact cells demonstrated that the recombinant full-length and truncated BCL2 proteins were expressed predominantly as nuclear membrane-associated proteins. These results imply that BCL2 must utilize hydrophobic domains other than the deleted domain for its association with the subcellular membranes. Metabolic labeling of insect cells expressing the full-length and the truncated form of BCL2 with 32P(i) demonstrated that BCL2 is a phosphoprotein.


Evolution of consciousness.

The hypothesis of the origin of consciousness is built upon the unique properties of the mammalian neocortex. The apical dendrites of the pyramidal cells bundle together as they ascend to lamina I to form neural receptor units of approximately 100 apical dendrites plus branches receiving hundreds of thousands of excitatory synapses, the collective assemblage being called a dendron. It is proposed that the whole world of consciousness, the mental world, is microgranular, with mental units called psychons, and that in mind-brain interaction one psychon is linked to one dendron through quantum physics. The hypothesis is that in mammalian evolution dendrons evolved for more effective integration of the increased complexity of sensory inputs. These evolved dendrons had the capacity for interacting with psychons that came to exist, so forming the mental world and giving the mammal conscious experiences. In Darwinian evolution, consciousness would have occurred initially some 200 million years ago in relation to the primitive cerebral cortices of evolving mammals. It would give global experiences of a surrounding world for guiding behavior beyond what is given by the unconscious operation of sensory cortical areas per se. So conscious experiences would give mammals evolutionary advantage over the reptiles, which lack a neocortex giving consciousness. The Wulst of the avian brain needs further investigation to discover how it could give birds the consciousness that they seem to have.


Chorionic peptidase inactivates GnRH as a post-proline peptidase.

Recently, we have described a chorionic peptidase (C-ase-1) which inactivates gonadotropin releasing hormone (GnRH), oxytocin, angiotensin II and thyrotropin releasing hormone. Since all these hormones contain a proline residue, we proposed that C-ase-1 may act as a post-proline peptidase. Using HPLC and amino acid analyses, we have defined the products which resulted from enzymatic inactivation of GnRH by C-ase-1. The N-terminal nonapeptide of GnRH was isolated by HPLC and confirmed by amino acid composition analyses. Thus, it was demonstrated that C-ase-1 acts as a post-proline peptidase when inactivating GnRH, yielding the nonapeptide, i.e., des-Gly10-NH2-GnRH, and Gly-NH2. The levels of intrauterine GnRH, angiotensin II, oxytocin and thyrotropin releasing hormone may be affected and integrated by this enzyme. Thus, C-ase-1 may play an important role in the regulation of the paracrine and endocrine function during pregnancy.


Current topic: comparative physiology of placental oxygen transport.

Development of knowledge about placental O2 transport (PO2) is discussed by focusing attention on the factors that determine umbilical venous PO2. In near-term pregnant sheep umbilical venous PO2 is much lower than maternal arterial PO2 and is about 20 torr lower than uterine venous PO2 in ewes who are the homozygous carriers of low O2 affinity ovine hemoglobin. Experimental evidence points to two main reasons for the low umbilical venous PO2 of sheep: (a) the uterine and umbilical circulations form an ineffective venous equilibration exchanger, and (b) a large uterine-umbilical venous PO2 gradient is required to draw O2 across a placental barrier which has a small O2 diffusing capacity relative to placental and fetal O2 demand and relative to the ineffective perfusion pattern. The latter explanation contradicts theoretical models which represent placental O2 transport as virtually 100 per cent blood flow limited. In near-term rabbits and guinea-pigs umbilical venous PO2 is also quite low, but for different reasons. In these species, the uterine and umbilical circulations form a countercurrent exchanger which allows the mother to perfuse the uterus at a very low rate. The effectiveness of countercurrent exchange is exploited to decrease the demand of pregnancy on the maternal circulation, rather than to increase the level of fetal oxygenation. There is suggestive, as yet inconclusive, evidence suggesting that in some species, notably the domestic cat, placental countercurrent exchange is combined with a low O2 affinity maternal hemoglobin and a sufficiently high uterine blood flow to produce a high level of umbilical venous PO2. The striking diversity and complexity of data about placental O2 transport demands great caution in applying comparative knowledge to the human placenta. Experimental evidence seems to indicate that the near-term human placenta is a venous equilibration exchanger, but the information which is presently available is inadequate for a firm conclusion.


Dual role of the nucleolar transcription factor UBF: trans-activator and antirepressor.

In a reconstituted system consisting of partially purified RNA polymerase I (pol I) and the initiation factors TIF-IA, TIF-IB, and TIF-IC, the nucleolar factor UBF (upstream binding factor) stimulates transcription from the rRNA-encoding DNA (rDNA) promoter at least 50-fold. This activation is not observed at high template concentrations or in the presence of highly purified pol I. Template commitment experiments suggest that UBF activates transcription by relieving inhibition exerted by a negative-acting factor(s) in the polymerase fraction that competes for TIF-IB binding to the rDNA promoter and prevents the formation of preinitiation complexes. Using purified histone H1 bound to DNA as a model for the repressed state of the rDNA promoter, we show that UBF counteracts H1-mediated repression of pol I transcription. The implications of these findings are discussed with respect to the protein-protein and protein-DNA interactions at the rDNA promoter and the possible involvement of UBF in control of ribosomal gene transcription.


Multiple forms of rat placental lactogen-II (rPL-II): purification and partial characterization of rPL-II.

The present study was designed to develop a procedure for purifying rPL-II and a homologous radioimmunoassay (RIA) for rPL-II. Molecular profiles of rPL-II were also investigated in tissue and plasma. rPL-II was purified 3,780-fold, based on its radioreceptor assay (RRA) activity compared to ovine prolactin (0PRL), from the placenta of day 18 pregnant rats using ammonium sulfate precipitation and chromatography on phenyl-Sepharose, DEAE-TOYOPEARL 650S, AF-chelate TOYOPEARL 650M and Sephadex G-100. Electrophoretic analysis on SDS gel revealed molecular weight heterogeneity of purified rPL-II, which consisted of three proteins; a major form with a molecular weight of 20.0 K and two minor forms with molecular weights of 20.6 K and 21.0 K under non-reducing conditions. One of the minor forms of rPL-II observed under non-reducing conditions disappeared with 2-mercaptoethanol treatment and the rest of the hormones migrated as 24.5 K and 25.0 K molecular weight species, suggesting that it is a cleaved form of rPL-II. Purified rPL-II displaced 125I-labelled oPRL from binding sites on rabbit mammary gland membranes in a dose-dependent manner. rPL-II and rPRL were, respectively, 21 and 2 per cent as effective as oPRL in the displacement. Antibody to purified rPL-II was raised in rabbits and a homologous RIA for rPL-II was developed. No displacement was observed with rPRL, rGH, oPRL, and other pituitary hormones up to 1,000 ng/ml. Molecular profiles of rPL-II in the placental tissue and plasma from day 18 pregnant rats were examined by gel chromatography on Sepharcryl S-300 HR and by Western blotting. Chromatography of the placental extracts revealed a single peak, which accounted for 86 per cent of the total RIA activity. Anti-rPL-II antiserum detected proteins of at least three molecular sizes as monomeric forms with molecular weights of 20.0, 20.6, and 21.0 K in the non-reducing placental extracts. One of them disappeared with 2-mercaptoethanol treatment and other two proteins had molecular weights of 24.5 and 25.0 K, indicating monomeric heterogeneity of rPL-II in the tissue. The elution profile of day 18 plasma in RIA activity gave two major peaks; the first, eluting just after the void volume (approximate molecular weight of 530 K) accounted for 35 per cent of the total RIA activity, and the second coinciding with the same elution volume as the monomeric form in the placental extract constituted about 26 per cent of the total RIA activity.(ABSTRACT TRUNCATED AT 400 WORDS)


Toward rules relating zinc finger protein sequences and DNA binding site preferences.

Zinc finger proteins of the Cys2-His2 type consist of tandem arrays of domains, where each domain appears to contact three adjacent base pairs of DNA through three key residues. We have designed and prepared a series of variants of the central zinc finger within the DNA binding domain of Sp1 by using information from an analysis of a large data base of zinc finger protein sequences. Through systematic variations at two of the three contact positions (underlined), relatively specific recognition of sequences of the form 5'-GGGGN(G or T)GGG-3' has been achieved. These results provide the basis for rules that may develop into a code that will allow the design of zinc finger proteins with preselected DNA site specificity.


Spatial pattern of cdc2 expression in relation to meristem activity and cell proliferation during plant development.

The p34 protein kinase encoded by the cdc2 gene is a key component of the eukaryotic cell cycle required for the G1- to S-phase transition and entry into mitosis. To study the regulation of plant meristem activity and cell proliferation, we have examined the tissue-specific accumulation of cdc2 transcripts in Arabidopsis thaliana and the related crucifer radish (Raphanus sativus) by in situ hybridization using A. thaliana cdc2 cDNA sequences as a probe. cdc2 transcripts accumulated in leaf primordia and within the vegetative shoot apical meristem. During flower development, high levels of expression were observed in meristems, in the basal regions of developing organs, in the developing vasculature, and associated with rib meristems elaborated late in the development of some floral organs. In root tips, cdc2 transcripts accumulated in the meristematic region and adjacent daughter cells but were not detected in the quiescent center. There was strong hybridization throughout the pericycle, and a further localized accumulation of cdc2 transcripts was observed in the initial stages of the activation of a new meristem at sites of lateral root development. We conclude that cdc2 expression is a critical factor in the regulation of meristem activity and establishment of proliferative competence.


Development of a quasispecies of human immunodeficiency virus type 1 in vivo.

During treatment with one specific batch of blood clotting factor IX, a number of hemophilia B patients in Germany recently became infected with human immunodeficiency virus type 1 (HIV-1). The nucleotide sequences of cloned HIV-1 envelope gene regions including the variable V3 loop and the V4 region derived from short-term virus cultures and directly from peripheral blood cells of these patients were shown to be highly homologous. Based on the assumption that the corresponding consensus sequence (termed HIV-1MBK) was identical to the genotype of the initially infecting virus, we were able to construct phylogenetic trees of the developing quasispecies in two patients studied in detail. True intermediates between input and multiply mutated genotypes were found in individual blood samples. Except for the initially infecting variant HIV-1MBK, variants found at 11 months postinfection had replaced those seen at 5 months postinfection. Variability early after infection was shown to cluster in two small regions located 3' of the V3 loop (i.e., outside the loop) and within the V4 region. This communication therefore describes the evolution of an HIV-1 quasispecies in humans starting from a single genotype.


A pattern of accumulation of a somatic deletion of mitochondrial DNA in aging human tissues.

An assay that selectively amplifies a specific deletion of the mitochondrial genome has been used to study the extent of the deletion's accumulation in a variety of human tissues. The deletion occurs at much higher levels in nervous and muscle tissues than in all other tissues studied. The variation in deletion level between the same tissues in different persons of similar age appears to be less than the variation among tissues within an individual. Tests for artifactual explanations of the level differences were each negative. Three cellular parameters that are correlated with the level of the deletion are identified. The preferential accumulation of deleterious mitochondrial mutations in a restricted subset of aging human tissues may compound deficiencies of function in those tissues that accrue with age.


Involvement of endogenous tumor necrosis factor alpha and transforming growth factor beta during induction of collagen type II arthritis in mice.

Both tumor necrosis factor alpha (TNF-alpha) and transforming growth factor beta (TGF-beta) are found in synovial fluid from arthritic joints of humans and of rodents with experimental arthritis. The role of endogenously produced TGF-beta and TNF in the pathogenesis of collagen type II-induced arthritis (CIA) in DBA/1 mice was examined by determining the effect of neutralizing monoclonal antibodies to these factors on the course of the disease. Endogenously produced as well as systemically administered TGF-beta 1 and TNF-alpha had opposite effects, since TGF-beta 1 and anti-TNF protected against CIA, whereas anti-TGF-beta and TNF-alpha increased CIA incidence and/or severity. Intraperitoneally injected TGF-beta 1 at a dose of 2 micrograms per day for 14 days significantly ameliorated arthritis, even when started at the time of arthritis development, although it did not reverse established disease. The resistance to CIA induction caused by a prior intravenous injection of collagen type II was not significantly influenced by the simultaneous injection of TGF-beta 1, TNF-alpha, or interleukin 1 alpha. It is concluded that the endogenous production of TNF and TGF-beta is important in determining the course of CIA.


Apolipoprotein AI mutation Arg-60 causes autosomal dominant amyloidosis.

A mutation in the gene for apolipoprotein AI (apoAI) was identified in an English family with autosomal dominant non-neuropathic systemic amyloidosis. The plasma of all affected individuals contained a variant apoAI with one additional charge, as well as normal apoAI. The propositus was heterozygous; the coding region of his apoAI gene contained both the normal sequence and a single-base substitution changing the codon for residue 60 of the mature protein from CTG (leucine) to CGG (arginine). Allele-specific oligonucleotide hybridization showed that the other affected individuals were also heterozygotes and that there was concordance of the mutant allele with the presence of variant plasma apoAI. Amyloid fibrils isolated from the spleen of the propositus consisted of proteins that ran as a doublet with an apparent mass of approximately 10 kDa in SDS/PAGE and a trace band at 28 kDa. Electrospray mass spectrometry of the purified 10-kDa material revealed components with mass corresponding to the N-terminal 88, 92, 93, and 94 residues of apoAI each with substitution of arginine for leucine. These observations were confirmed by direct protein sequencing and laser desorption time-of-flight mass analysis. No material with the normal apoAI sequence was detected. The trace band at 28 kDa yielded the N-terminal sequence of mature apoAI, indicating that intact or minimally degraded apoAI was also present in the fibril preparation. Discovery of this mutation and the detailed characterization of the apoAI fragments that form the amyloid fibrils open additional avenues for investigation of amyloidogenesis.


Somatotopic organization in rat striatum: evidence for a combinational map.

2-Deoxy-D-[14C]glucose autoradiography was used in awake rats to map neural activity in the sensorimotor sector of striatum. Stimulation of hindlimb, trunk, or forelimb activated primary sensory cortex in a localized columnar pattern, indicating activation of somatosensory receptors and a discrete cortical functional unit. In sensorimotor striatum, an image analysis detection technique revealed regions of maximal activity, or features, that formed a patchy pattern of activation reminiscent of the known anatomic patterns of cortico-striate terminals. Ipsilateral as well as contralateral activation was observed. The activated areas revealed a body map in striatum that was organized in a manner consistent with cortical topography (dorsoventrally: hindlimb, trunk, forelimb) at most anteroposterior levels, similar to that found in other species. However, at other levels, a different organization (e.g., trunk, hindlimb, forelimb) was observed. Furthermore, the arrangements of body region and side were also unique at different anteroposterior levels. Thus, functional activity showed multiple, different juxtapositions of body elements--i.e., a combinational map. The data suggest that striatum may provide an anatomic substrate for different combinations of inputs necessary to select and integrate movement.


Serine tRNA complementary to the nonuniversal serine codon CUG in Candida cylindracea: evolutionary implications.

In the asporogenic yeast Candida cylindracea, the codon CUG is read as serine instead of leucine. This is an unusual instance in which the amino acid assignment of a codon deviates from the universal code. To infer the evolutionary process of this change, the tRNA with the anticodon sequence CAG, which is complementary to and thus responsible for translation of the codon CUG, has been identified. Indeed, this tRNA translates an in-frame CUG codon in a synthetic mRNA as serine in an in vitro translation system. The gene for the tRNA is interrupted by an intron in the anticodon loop. Sequence comparisons of the tRNA and its gene suggest that a single cytidine was inserted into the anticodon loop of the gene for tRNA(Ser)IGA during evolution to produce tRNA(Ser)CAG. The tRNA(Ser)CAG may be produced from its precursor molecule containing the cytidine insertion by splicing.


Purification of the heteromeric protein binding to the URS1 transcriptional repression site in Saccharomyces cerevisiae.

The protein that binds to the URS1 site situated upstream of many genes in Saccharomyces cerevisiae is a central element responsible for global negative control of transcription in this organism. Among the genes whose expression is regulated by this protein are those that participate in nitrogen metabolism, carbon metabolism, electron transport, inositol metabolism, heat shock response, meiosis, and sporulation. This factor, binding URS1 factor (BUF), has been purified and shown to be a heteromeric protein composed of 37.5- and 73.5-kDa monomers. The heteromeric form of BUF is stably maintained both in solution and bound to its DNA target site.


Complexes of tissue-type plasminogen activator and its serpin inhibitor plasminogen-activator inhibitor type 1 are internalized by means of the low density lipoprotein receptor-related protein/alpha 2-macroglobulin receptor.

Tissue-type plasminogen activator and urokinase are serine proteases secreted by many cell types that participate in biological processes, such as tissue restructuring, cell migration, and tumor metastasis. Clinically, these proteases are used to dissolve coronary fibrin clots that are the proximal causes of acute myocardial infarction. In vivo, the activity of these enzymes is controlled by plasminogen-activator inhibitors, members of the serpin family of protease inhibitors. This study shows that tissue-type plasminogen activator-inhibitor complexes bind in solution to low density lipoprotein receptor-related protein (LRP), a large heterodimeric ubiquitous membrane receptor. In cultured cells, endocytosis and degradation of these complexes is reduced by polyclonal antibodies directed against LRP and inhibited by a M(r) 39,000 protein that binds to LRP and inhibits its interaction with previously known ligands, including apolipoprotein E and alpha 2-macroglobulin. We propose a role for LRP in the clearance of plasminogen activator-inhibitor complexes that is analogous to its function in the endocytosis of alpha 2-macroglobulin-protease complexes.


Low density lipoprotein receptor-related protein/alpha 2-macroglobulin receptor is an hepatic receptor for tissue-type plasminogen activator.

Tissue-type plasminogen activator (t-PA), a serine protease that catalyzes the initial and rate-limiting step in the fibrinolytic cascade, is cleared rapidly in vivo by the liver. Using chemical crosslinking, we have recently identified a plasminogen-activator inhibitor type 1 (PAI-1)-independent t-PA clearance receptor on rat hepatoma MH1C1 cells with a relative molecular mass of approximately 500 kDa. Another recently identified membrane receptor, low density lipoprotein receptor-related protein/alpha 2-macroglobulin receptor (LRP/alpha 2MR), was also detected on MH1C1 hepatoma cells by using immunoprecipitation with anti-LRP/alpha 2MR antibody. When analyzed by SDS/PAGE, we found the t-PA receptor identified on MH1C1 cells comigrated with the large subunit of LRP/alpha 2MR. The t-PA receptor was immunoprecipitated by an anti-LRP/alpha 2MR antibody after chemical crosslinking of specifically bound 125I-labeled t-PA to its receptor. Through chemical crosslinking studies, we found that t-PA and methylamine-activated alpha 2-macroglobulin could bind to LRP/alpha 2MR simultaneously without competing with one another for binding, suggesting that the two ligands bound to two independent sites on the LRP/alpha 2MR molecule. Furthermore, a 39-kDa protein, which modulates ligand binding to LRP/alpha 2MR, was also found to inhibit t-PA binding to its receptor. These data thus show that the t-PA clearance receptor identified on MH1C1 hepatoma cells is LRP/alpha 2MR.


Intracellular accumulation and resistance to degradation of the Alzheimer amyloid A4/beta protein.

The A4 or beta protein is a peptide that constitutes the major protein component of senile plaques in Alzheimer disease. The A4/beta protein is derived from a larger, transmembrane amyloid precursor protein (APP). The putative abnormal processing events leading to amyloid accumulation are largely unknown. Here we report that a 42-residue synthetic peptide, beta 1-42, corresponding to one of the longer forms of the A4/beta protein, accumulates in cultured human skin fibroblasts and is stable for at least 3 days. The peptide appears to accumulate intracellularly, since it does not accumulate under conditions that prevent endocytosis and accumulation is correlated with the acquisition of resistance to removal by trypsin digestion. This intracellular accumulation is also correlated with the ability of the peptide to aggregate as determined by SDS/polyacrylamide gel electrophoresis. At low concentrations of the beta 1-42 peptide, which favor the nonaggregated state, no accumulation is observed. Shorter peptide analogs (28 or 39 residues) that are truncated at the C terminus, which lack the ability to aggregate in SDS gels, fail to accumulate. The accumulated intracellular beta 1-42 peptide is in an aggregated state and is contained in a dense organellar compartment that overlaps the distribution of late endosomes or secondary lysosomes. Immunofluorescence of the internalized peptide in permeabilized cells reveals that it is contained in granular deposits, consistent with localization in late endosomes or secondary lysosomes. Sequence analysis indicates that some of the internalized peptide is subject to N-terminal trimming. These results suggest that the aggregated A4/beta protein may be resistant to degradation and suggest that the A4/beta protein may arise, at least in part, by endosomal or lysosomal processing of APP. Our results also suggest that relatively nonspecific proteolysis may be sufficient to generate the A4/beta protein if this part of APP is selectively resistant to proteolysis.


The T-DNA-linked VirD2 protein contains two distinct functional nuclear localization signals.

Agrobacterium tumefaciens causes neoplastic growth in plants by transferring a piece of DNA, called T-DNA, into the nucleus of the plant cell. The virulence protein VirD2 of A. tumefaciens is tightly linked to the T-DNA and is thought to direct it to the plant genome. Here we show that the VirD2 protein contains two nuclear localization signals that are functional both in yeast and in plant cells. One signal is located in the N-terminal part of the protein and resembles a single-cluster-type nuclear localization signal. The second signal is near the C terminus and is a bipartite-type nuclear localization signal. The involvement of these sequences in the entry of the T-DNA into the nucleus is discussed.


MAZ, a zinc finger protein, binds to c-MYC and C2 gene sequences regulating transcriptional initiation and termination.

ME1a1, a 16-base-pair nuclear factor binding site residing between the c-MYC P1 and P2 transcription initiation sites, is required for P2 activity. A cDNA encoding a 477-amino acid zinc finger protein designated MAZ (MYC-associated zinc finger protein) was cloned from a HeLa lambda gt11 library by screening with a concatamerized ME1a1 binding site probe. In addition to six potential zinc fingers of the Cys2His2 type, MAZ contains an amino-terminal proline-rich domain and several polyalanine tracts. Its mRNA was present in all human tissues tested except for kidney, as a doublet of approximately 2.5 and 2.7 kilobases, along with differentially expressed minor species. MAZ bound specifically to the wild-type ME1a1 sequence but not to a ME1a1 mutant that also failed to yield P2 activity. When expressed as a fusion protein in a pMAL-c vector, MAZ binds with specificity to a GA box sequence (GGGAGGG) found in the c-MYC P2 promoter, to the P2 attenuator region within the gene's first exon, and to a related sequence involved in the transcriptional termination of the C2 gene. MAZ may encode a transcription factor with dual roles in transcription initiation and termination.


Refolding and oriented insertion of a membrane protein into a lipid bilayer.

We have studied the refolding and membrane insertion of the outer membrane protein OmpA of Escherichia coli. The protein was extracted from its native membrane by sonication in the presence of urea and dissolved in the urea/water mixture in unfolded form. In this form it was purified. Upon addition of preformed lipid vesicles, the protein spontaneously refolded and inserted into the vesicle membranes. The vesicles had to be small and the lipids had to be in the fluid state. The insertion occurred in an oriented manner.


Sensitivity of transformation to small differences in population density during serial passage of NIH 3T3 cells.

Early passages of the NIH 3T3 mouse cell line undergo spontaneous neoplastic transformation leading to the development of transformed foci if grown to confluence in 2% (vol/vol) calf serum (CS) and left there for more than a week. Transfer of the postconfluent cultures results in the appearance of large numbers of transformed foci; many of them are larger and denser than those in the original culture. If the cells are continually kept at low population densities by frequent passages in 10% CS, they lose the capacity to undergo spontaneous transformation. If however the low-density passages are made in 2% CS or in 10% (vol/vol) fetal bovine serum, both of which support lower growth rates and saturation densities than does 10% CS, they gain the capacities to grow to high saturation densities and produce more foci when grown to confluence in 2% CS. These increases are proportional to the population densities used in the frequent passages, although the densities are all kept well below confluence. We conclude that the combined constraints of submaximal serum plus those of the limited cell contacts of the low cell densities used here elicit an adaptive response that endows the entire population with increased growth capacity. The increased growth capacity of the heterogeneous population in turn increases the capacity of a fraction of the population to initiate distinctive transformed foci. Similar studies have indicated that the capacity of cells to produce tumors and metastases in mice and rats is enhanced by prior maintenance at high density in culture. We propose the concept of progressive state selection to account for the general increase in the growth capacity of cells that is elicited by moderate constraints on their growth and metabolism.


Association of titin and myosin heavy chain in developing skeletal muscle.

To understand molecular interactions that organize developing myofibrils, we examined the biosynthesis and interaction of titin and myosin heavy chain in cultures of developing muscle. Use of pulse-labeling, immunoprecipitation, and a reversible cross-linking procedure demonstrates that within minutes of synthesis, titin and myosin heavy chain can be chemically cross-linked into very large, detergent-resistant complexes retaining many features of intact myotubes. These complexes, predominantly of titin and myosin, occur very early in myofibrillogenesis as well as later. These data suggest that synthesis and assembly of titin and myosin are temporally and spatially coordinated in nascent myofibrils and support the hypothesis that titin molecules help to organize sarcomere formation.


The evolutionary selection of DNA base pairs in gene-regulatory binding sites.

The DNA base-pair sequences that serve as gene-regulatory sites have been selected during evolution to provide an appropriate functional binding for a particular protein. In most cases, the function depends on the binding probability, which can be influenced both by the binding strength and by the abundance of the protein in the cell. As a consequence, the same function can be achieved with strong binding sites and a small amount of protein as with weak binding sites and a large amount of protein. However, increasing the protein burden will decrease the growth rate of the cells, even when all functions remain the same. Thus, for maximal growth, the protein levels should be as low as possible and the binding correspondingly strong. On the other hand, sequences with a weaker binding can be formed in many more ways and are, therefore, more probable, and random mutations are more likely to produce them. Thus, the selection pressure against an increased protein burden can be balanced against the random mutational drift in the recognition sequences, thereby tying together the statistics of base-pair choice, the binding strength, and the protein burden. In terms of this model, the selection pressure can be estimated from the properties of a gene-regulatory protein and its recognition sites. A key feature is the mutational randomization pressure that appears as a fundamental force shaping the optimal solutions that provide maximal growth. The model is tested on a number of gene-regulatory systems in Escherichia coli. The same principles should hold for all proteins for which overall activity in the cell is proportional to abundance; then the selective pressure to increase the efficiency of an individual protein cannot be larger than the selective pressure to decrease the total protein burden.


Pyrimidine dimers in DNA initiate systemic immunosuppression in UV-irradiated mice.

Exposing the skin of mice to UV radiation interferes with the induction of delayed and contact hypersensitivity immune responses initiated at nonirradiated sites. The identity of the molecular target in the skin for these immunosuppressive effects of UV radiation remains controversial. To test the hypothesis that DNA is the target for UV-induced systemic immunosuppression, we exposed C3H mice to UV radiation and then used liposomes to deliver a dimer-specific excision repair enzyme into the epidermis in situ. The application of T4 endonuclease V encapsulated in liposomes to UV-irradiated mouse skin decreased the number of cyclobutane pyrimidine dimers in the epidermis and prevented suppression of both delayed and contact hypersensitivity responses. Moreover, the formation of suppressor lymphoid cells was inhibited. Control, heat-inactivated endonuclease encapsulated in liposomes had no effect. These studies demonstrate that DNA is the major target of UV radiation in the generation of systemic immunosuppression and suggest that the primary molecular event mediating these types of immunosuppression by UV radiation is the formation of pyrimidine dimers. Furthermore, they illustrate that the delivery of lesion-specific DNA repair enzymes to living skin after UV irradiation is an effective tool for restoring immune function and suggest that this approach may be broadly applicable to preventing other alterations caused by DNA damage.