Temperature and the progression of HeLa S-3 cells from G2 into mitosis in the presence and absence of the amino acid analogue, para-fluorophenylalanine. HeLa cells grown in suspension at 37.5 degrees C show an immediate reduction in the rate of progression from G2 into mitosis at suboptimal temperatures. The rate of progression becomes slower the longer the cells reside at the lower temperatures. Shifting cells back to 37.5 degrees C from 33.5 degrees C results in an immediate increase in the flow of cells from G2 into mitosis but not to the optimal (37.5 degrees C) rate of traverse found in unshifted controls. The longer cells reside at 33.5 degrees C before being shifted to 37.5 degrees C, the slower the rate of G2 traverse. G2 cells allowed to incorporate p-fluorophenylalanine (in the absence of phenylalanine) are inhibited from entering mitosis at 37.5 and 39.5 degrees C, partially inhibited at 33.5 and 35.5 degrees C, but remain uninhibited at 32 degrees C. Greater incorporation of 3H-p-fluorophenylalanine and of 3H-leucine in the presence of 0.2 mM p-fluorophenylalanine seen at the lower temperatures than at 37.5 and 39.5 degrees C. Although protein synthetic rates and progression of cells to mitosis at different temperatures show a close correlation, they do not appear to be casually related.


Informational molecules in amoebae: an attempt to follow injected fractions using autoradiography. Cytoplasmic fractions from Amoeba discoides contain informational molecules which, when injected into A. proteus, may influence the characters of resulting clones. An attempt was made to follow and locate these molecules by autoradiography. Using either 3H-uridine or 3H-leucine, radioactive fractions of A. discoides were microinjected into A. proteus and cells examined at varying times after injection. A. discoides were injected with the same preparations to serve as controls. Grain counts were compared for equivalent areas of nucleus and cytoplasm, and results expressed in terms of the ratio between nucleus and cytoplasm. When the light microsomal or the supernatant fractions were injected, the level of labelled molecules was higher in the nucleus than the cytoplasm in over 50% of the cells examined after 6 h, except for those cells injected with 3H-leucine-labelled supernatant material when only 15% cells showed a nuclear/cytoplasmic grain count greater than 1.1. After some purification by sucrose density gradient centrifugation, 'information-containing' peaks '13' from the light microsomal fraction, and '16' from the supernatant fraction were injected. A different pattern of results was obtained. The problems of interpreting these results in the light of known migratory molecules and strain incompatibility are discussed.


On the distinction between fear and wariness. The study seeks to illuminate the mechanisms that mediate avoidant reactions in young humans. 40 babies repeatedly were observed throughout their second year in a variety of mildly challenging situations, and then again in a series of follow-up assessments at age 3 1/2. Analysis of the covariance patterns among the repeated measurements supports positing 2 distinct types of avoidant mechanism--an inclination to be "wary" of the unfamilar, and a proclivity to be "fearful" after exposure to a mildly difficult situation. Each type of phenomena constituted an independent source of individual differences. Behaviors that differed as a result of prior experience proved fairly predictive of future social orientations; variations in initial wariness did not.


Infant crying and maternal responsiveness: a rejoinder to Gewirtz and Boyd. The criticisms leveled by Gewirtz and Boyd focus on 1 of 6 analyses upon which Bell and Ainsworth based their interpretation of findings--namely, cross-quarter correlations of measures of infant crying and maternal response. It is maintained that these measures were approximate, and that they were independent of each other in cross-quarter correlations. It is also maintained that the statistical procedures were appropriate to a small sample of subjects in naturalistic, longitudinal research. The three "illustrative" cases offered by Gewirtz and Boyd as ways of testing a contrary hypothesis are considered. These cases assumed maternal behavior patterns which, it is argued, do not occur. Finally, other evidence is cited, both from other studies and from within the Bell-Ainsworth paper itself, that supports the interpretation that Gewirtz and Boyd claim to have been unjustified.


Attachment of kibbutz infants to mother and metapelet. 122 children, born and reared on Israel kibbutzim, were observed in a cross-sectional study of infant attachment behaviors. Their reaction to either mother or metapelet (caretaker) separation and reunion was recorded over a 13-sequence experimental paradigm. Results indicated that children protested equally to either mother or metapelet separation when left with a stranger. Reunion behaviors seemed to be sensitive to the different attachment relationships the infant had with each caretaker, while separation behaviors were not. The  quality of the infant-mother interactions as it is related to the infant's ordinal position had a significant effect on infant attachment behavior in the experimental situation. Firstborn (only) children were more anxious than later-borns during the session. Speculation as to the origin of these significant ordinal differences is discussed.


Sex differences in parent-infant interaction during free play, departure, and separation. 20 18-month-olds were observed with their mothers and 20 with their fathers in laboratory free-play, departure, and separation situations. Parent and infant behaviors were allowed to vary naturally. Although there were no parent sex or infant sex differences in infants' free-play behavior, there were parent sex X infant sex differences in parental free-play behaviors. Parents talked to, sat on the floor more with, and tended to share play more with same-sexed than opposite-sexed infants, and the patterning of free-play behaviors was different for mothers and fathers. During departure, fathers talked to the infants more than mothers. Infants were more distressed in the absence of same-sexed than opposite-sexed parents. There were infant sex X parent sex differences in the relationships between separation distress and parental free-play and departure behaviors. The implications of these findings for understanding differential roles of mothers and fathers, the development of sex differences, and the determinants of separation distress are discussed.


Sex differences in parent and infant behavior in the home. Observations were made in the homes of 32 families, each with a child aged around 12 or 24 months. 16 of the children were boys and 16 girls. Boys showed more active play, play with transportation toys, and play generally forbidden by parents. Girls played more with soft toys and dolls. On questionnaire responses many parents rated these and other behaviors as sex appropriate. However, actual sex differences in immediate parental response to children were few or undetected. Sex differences in children's behavior did not appreciably increase from 12 to 24 months. Also, there was no correlation with how stereotyped parents' questionnaire responses were and the degree to which their children exhibited sex-typed behavior.


Initiating day care at three years of age: effects on attachment. Using the Ainsworth-Wittig strange situation, 12 42-month-old children with approximately 6-months of day-care experience were compared with individually matched children who had not had group child-rearing experience; 8 members of each experience group were female, and 4 members were male. While the day-care children showed less distress than the home-care children in the latter parts of the laboratory situation, the 2 groups were not different with respect to most other behaviors, including those toward the mother. However, day-care experience appeared to have differential effects as a function of sex; large differences were often found between the scores of the male subgroups, whereas the differences between the female subgroups were usually small. The day-care males exhibited the most exploratory manipulation, and the home-care males showed the most approach and proximity seeking toward the mother and the strongest approach toward and avoidance of the stranger. These results did not support the idea that day-care experience impair attachment to the mother.


Aggression and peer acceptance in adolescent boys: two short-term longitudinal studies of ratings. 2 longitudinal studies covering a 1-year and a 3-year interval, respectively were conducted on 2 samples of boys aged 13 years (N1=85, N2=201). On the basis of an adaptation of multimethod-multivariable analysis and other analyses it was found that information contained in the 4 rating dimensions under study to an overwhelming degree reflected characteristics of the boys, not rater biases. The results indicated very high or high degrees of stability over time in the dimensions studied concerning aggression and peer acceptance. The results for the aggressive area were construed as strong evidence for assuming relatively stable, individual-differentiating reaction tendencies or motive systems within the boys. The findings also provided strong evidence that certain aspects of each boy's interpersonal environment were stably different for different boys.


Development of sex-trait stereotypes among young children in the United Staes, England, and Ireland. The Sex Stereotype Measure II (SSM II), a 32-item revision of the Williams, Bennett, and Best Sex Stereotype Measure, was developed to assess children's knowledge of conventional, sex-trait stereotypes defined by American university students. The procedure employed brief stories and human figure silhouettes which were individually administered to 5- and 8-year-old children in the United States, England, and Ireland and group administered to 11-year-olds in the United States. In the United States, knowledge of sex-trait stereotypes was found to develop in a linear fashion between the ages of 5 and 11, with more male traits than female traits being known at each age level. Cross-nationally, there was a high degree of similarity in the nature of the sex stereotypes being learned by the children in the 3 countries, although the rate of learning appeared slower among the Irish children. In all countries there was a clear progression in sex-stereotype learning from age 5 to age 8. English boys had greater knowledge of stereotypes than English girls, but this was not true in Ireland and the United States. Generally, knowledge of male stereotype traits appeared to develop earlier while knowledge of the female traits increased more rapidly between ages 5 and 8. The similarity in sex-stereotype learning in the 3 countries is discussed, and studies in progress in other countries of greater cultural diversity are noted.


Behavioral and fantasy measures of fear of success in children. Fear of success was assessed for 61 fifth-, seventh-, and ninth-grade children using Horner's and Baruch's "fantasy" measures and a behavioral measure developed by the authors. Males and females did not differ significantly on fear of success as measured by the fantasy technique. Developmentally, male fear of success remained unchanged in the three grades tested, while female fear of success decreased from grade 5 to grade 7. The behavioral measure of fear of success tapped a drop in achievement striving in grade 7 for the female subjects. Results are discussed in light of a "situational" versus personality theory of the development of fear of success.


Facial expressions used by children in a conflict situation. 72 pairs of kindergarten children were each given a brief play session involving an object with which only 1 child could play at a time. Facial expressions used by children defending their possession of the object were studied. Results showed a relationship between the facial expressions a child used and both his own subsequent behavior and that of his partner. "Agressive" facial expressions were associated with persistent attempts by the expressers to maintain access to the disputed object. Percipients of aggressive expressions were relatively hesitant about making a new attempt to take the object from the expresser. 1 nonaggressive expression was also followed by percipient hesitancy. The relationship of the aggressive expressions to both primate threat displays and human facial expressions of emotion is discussed.


Effect of perceptual processing strategies on problem solving of reflective and impulsive children. Several studies employing the "20 questions" test reported that reflective children asked proportionally more constraint-seeking (CS) questions than did impulsive children. The finding was interpreted as supporting the generally accepted view that reflective children are more mature and more efficient problem solvers than impulsive children. Inexplicably, the reflectives' higher proportion of CS questions was not associated with fewer questions to solution, that is, with greater efficiency. The present study examined an alternative hypothesis that differences in performance of impulsive and reflective children on the "20 questions" test are due to individual differences in preferred perceptual processing strategy rather than in cognitive maturity of problem-solving strategy. Efficiency of performance of reflective and impulsive children was shown to be related to type of stimuli and experimental conditions employed, supporting a perceptual rather than cognitive-maturational  interpretation.


Reflection-impulsivity as a predictor of children's academic achievement. To examine the implications of differences in reflection-impulsivity for later academic achievement, 70 children were administered the Matching Familiar Figures test (MFF) in grade 4 and the Comprehensive Tests of Basic Skills (CTBS) in grades 4, 5, and 6. Children identified as reflective based on grade 4 MFF performance scored significantly higher on the CTBS achievement battery at all grade levels than those classified as impulsive. However, the 2 groups did not differ on the grade 5 or grade 6 achievement measures when scores were adjusted for initial differences in grade 4 CTBS. Similarly, while each of the continuous variables MFF error score and MFF response latency was significantly predictive of grade 5 and grade 6 achievement test scores, neither of the MFF variables significantly improved the prediction of academic performance when current level of achievement was statistically accounted for. Sex differences in the relations between the MFF variables and the achievement measures were identified; MFF error score was more strongly related to later academic achievement for boys than for girls, while MFF response latency was a better predictor of academic achievement for girls than for boys.


Hyperactive children's underuse of learning time: correction by stimulant treatment. This study investigates the effects of methylphenidate (Ritalin) on hyperactive children's performance on a paired-associate learning task under 3 presentation rates  (4, 8, and 12 sec per item). The total-time hypothesis states that a fixed amount of time is necessary to learn a fixed amount of material, regardless of the number of trials into which that time is divided. In a double-blind crossover design, the total-time hypothesis was supported by performance in a drug state, but not by performance in a placebo state, where slower presentation rates were not utilized effectively. These results are discussed in terms of inattention in the placebo state and improved attention and use of learning time in the drug state. The methodology presented is suggested as a means of assessing attentional deficits as well as providing a diagnostic procedure for objectively evaluating the appropriateness of stimulant treatment for children referred for symptoms of hyperactivity.


Seeing, hearing, and doing: a developmental study of memory for actions. The ability to recall and organize actions was studied in children from 5 to 11 years in age. 8 different auditory or visual commands were successively presented for 10 trials in each modality in a free-recall task. Younger children performed fewer commands but recalled relatively more recent ones, and they showed the same degree of subjective organization and the same degree and structure of hierarchical clustering as the older children. The hierarchical structure was independent of recall, age, and modality, with the motor actions being organized by the locus of the object or instrument of the verb in the command. The difficulty of the commands was highly correlated with uncertainty of the locus of the action, that is, the number of possible arguments (objects or instruments) a verb could assume, as measured by a subsidiary experiment on 8-year-olds who were asked to name as many parts of the body upon or with which one could perform each action. Developmental differences in recall appear to rise because of primary organization (retrieval) and rehearsal strategies rather than secondary organization.


The development of recognition memory for the left-right orientation of pictures. 3 studies examined the development of recognition memory for the left-right orientation of pictures of common objects. In study 1, 5-year-olds, 9-year-olds, and college students were shown 20 pictures of objects. Subsequently, they were tested on their ability to discriminate between 10 of these pictures and their left-right mirror images and between the other 10 pictures and completely new ones. There were large developmental differences in memory for orientation, although all age groups could discriminate accurately between familiar pictures and completely new ones. In studies 2a and 2b, training concerning the relevance of orientation improved second and fourth graders' long-term memory for this characteristic, but training effects were minimal for kindergartners. However, even kindergarten children showed accurate short-term memory for orientation on the task used for training. In contrast to study 2, study 3 produced accurate long-term memory for orientation in kindergarten children by using a verbal training procedure. Results are discussed in terms of the range of memory factors involved in children's mirror-image confusions and in terms of general implications for the development of recognition memory.


The effects of different modes of verbalization on the recognition of object detail in pictures. The effects of different modes of verbalization on recognition memory of object detail were examined in first- and third-grade children and adults. The results indicated that both age of the subject and the type of verbalization used during initial picture viewing influenced recognition accuracy for object detail. When compared to nonverbal viewing groups, only specific types of verbalization increased recognition accuracy for third-grade and adult subjects. None of the modes of verbalization was effective in increasing first-grade recognition scores. The results indicate that general statements concerning the facilitating or inhibiting influence of verbalization on recognition memory must be qualified. In addition, the results showed that developmental differences in recognition accuracy are contingent upon the type of strategy used during the encoding process.


The development of children's understanding of cyclic aspects of time. Developmental psychological approaches to the study of time have fallen into 3 categories: studies of time perception; studies of logical, reconstructive abilities; and studies of the understanding of conventional time systems. The present work examines problems spanning the latter 2 categories--the development of children's understanding of temporal cycles and the relationship between cyclic concepts and cognitive development. 62 children, ranging in age from 4 to 10 years, were administered Piagetian tests of classification and seriation and a variety of specially designed cyclic tasks. Results show major progress in the representation of cyclic order and recurrence during the age period examined. For a variety of particular cycles, order responses were shown before continuity responses. The ability to produce a correct order is related to seriation performance but not classification performance when the variance attributable to age is partialed out. Continuity responses appear to be unrelated to performance on either of the Piagetian tasks tested when age is controlled.


Following their companions as a form of exploration for human infants. Infants' following of other persons, often considered a measure of attachment, qualified as a form of exploration. The results of the first experiment indicated that following can be a means of investigating novel leaders: 16 9--12-month-old infants were as likely to follow an unfamiliar woman as they were the parent and were reliably more likely to follow a moving toy. The second and third experiments assessed the extent to which the experience of following familiar persons promotes learning about the environment. Infants who followed their mothers to 1 place were more likely later to investigate a similar place than those who initially either locomoted independently (experiment 2) or were transported by their mothers (experiment 3). The findings suggested that infants' transactions with the environment need not be considered antithetical to their social behavior.


Environmental characteristics of a neonatal intensive-care unit. An increasing number of intervention programs are designed to ameliorate the "sensory deprivation" of hospitalized premature infants. However, observation of the premature infant's hospital environment every 15 min for 5 24-hour periods indicated that acoustic stimuli were clearly audible in over 95% of the observations, infants were handled during more than 10% of the times they were observed, and illumination levels were always sufficiently high to permit easy visibility. Analysis of the nature of this stimulation suggests that premature infants may suffer from an inappropriate pattern rather than an inadequate amount of stimulation.


Infant visual scanning of two- and three-dimensional forms. 2- and 3-dimensional forms affected 4- and 8-week-old infant visual behavior differently, 4-week-old infants fixated a recessed 3-dimensional form more than a 2-dimensional form but did not scan it differently. 8-week-old infants did not look more at 1 stimulus than another but scanned a raised 3-dimensional form differently than a 2-dimensional equivalent. For all stimuli and both age groups, there was a relation between attractiveness of stimuli and how they were scanned; babies who were most attracted to stimuli inspected them with the smallest eye movements.


Toys and social interaction between infant peers. Claims that young infants fail to react in a social manner to one another and that toys preempt attention to peers were assessed by comparing the interactions observed between infant peers when they met in the presence of toys versus in their absence. 44 pairs of unacquainted infants (either 10--12 or 22--24 months of age) came with their mothers to an unfamiliar room. Without toys available in the room, infants of both ages more often contacted one another, smiled at and gestured to one another, and duplicated each other's actions. With toys, they showed and exchanged toys and spent more time synchronously manipulating similar play material. The results document that infants as young as 10 months of age are responsive to the person and behavior of an unfamiliar peer and that they are no less responsive than older infants to the social versus nonsocial aspects of a novel setting.


Stranger responses: effects of familiarity, stranger's approach, and sex of infant. Most investigators examining contextual influences on infant responses to a stranger have limited their study to the initial responses of the infant. Few have considered infant reactions beyond the first minutes of the encounter or the enduring effects of contextual variables once mothers have left. The present study examines the effects of stranger's style of approach, familiarity with the stranger, and infant sex on infant response before and after maternal separation. Familiarity was operationalized in 2 ways: the length of time mothers stayed before leaving infants with the stranger and meeting the stranger a second time. Findings suggest that all these factors influence infant behavior. Ifants looked and smiled at the stranger significantly more when approached slowly. Contrary to expectations, infants (particularly girls) who met the familiar stranger after 1 week protested significantly more when left with her at that time. Trends indicate the length of time mothers stayed was least influential since most infants became extremely distressed once mothers departed.


Maternal language to prelinguistic infants: syntactic aspects. Maternal speech to children has been shown to vary by age and language ability of the children. Previous studies have usually involved children over 1 year of age. In this study maternal speech to male and female 4-, 6-, and 8-month-old infants was recorded in the laboratory. Mothers used shorter utterances to 8-month-olds than to 4- or 6-month-olds, presumably in response to the infant's changing level of comprehension. Mothers used more sentences with subjects, verbs, or objects deleted to 8-month-olds and more complex sentences to 4-month-olds.


Infants' perception of temporal grouping in auditory patterns. Infants' processing of temporal parameters of auditory patterns was investigated by assessing the ability of 5-month-old infants to discriminate between multitone patterns with identical component tones but contrasting temporal arrangements of these tones. Following habituation of the cardiac response to a 6-tone stimulus with 2,4 grouping infants were presented with the same tonal sequence with 4,2 grouping. Significant dishabituation was evident, indicating discrimination of different temporal groupings of auditory stimuli.


Sex stereotypes in adult-infant play.

Sex stereotypes in adult-infant play. 24 14-month-old infants played with adult strangers in a laboratory playroom. To examine the effects of sex-role labeling on adult behavior, each infant was introduced as a boy and as a girl in 2 different sessions. After controlling for the adult's scores on the Spence-Helmreich Attitudes toward Women Scale, the adults were found to engage in more Encouragement for Activity and more Male Role Toy Choice when playing with children designated as boys. They showed more Interpersonal Stimulation and more Nurturance Play when playing with children designated as girls. No significant differences were observed between the behavior of the real girls and boys. Men and women showed some differences in their style of play.


Reciprocity in vocal interactions of mothers and infants. Social responses of 24 mothers and 3-month-old infants were continuously observed and coded in the home to examine reciprocal influences in early vocal interactions. One of 4 dyadic vocal states (simultaneous vocalization, mother vocalizing alone, infant vocalizing alone, mutual silence) was assigned to each consecutive 1-sec interval of the 90-min observational records. The sequence of dyadic vocal states was represented by a first-order transition probability matrix. Transitions between states involving maternal and infant vocal onsets and offsets were analyzed. Results indicated that vocal onsets for both mother and infant were more likely when the other dyad member was vocalizing. There was no evidence of reciprocal influences for vocal offsets. It is suggested that differentiating response onset and offset is necessary for understanding vocal reciprocity in mother-infant interaction.


Social class, birth order, and newborn experience. Early home experiences of a group of 32 2-week old infants, equally divided according to social class, birth order, and sex, were studied. Naturalistic observations were taken on 2 consecutive days for a total of 6 hours. Compared with later-borns, firstborns received significantly more caretaker interaction on all variables studied. Compared with lower-SES infants, those from higher-SES homes received significantly more direct verbal interaction although the 2 groups did not differ on total periods of other types of caretaking interaction. The findings are consistent with birth order and social class differences reported in home environments of older infants and children.


Intellectual development in family constellations with adopted and natural children: a test of the Zajonc and Markus model. A confluence model was developed by Zajonc and Markus to account for the effects of birth order, child spacing, and family size on intellectual development. the fit of the model was tested on a sample of families with biological and adopted children. Although the model fits population data very well, it fails to account for intellectual patterns within families for whom both parent and child IQ data are available. The discrepancy is discussed in terms of sources of variance within and between families that are undetected in average values for a population.


Distance distortions in memory for spatial locations. The present study examined developmental differences in the effect of route extensity on the memory for the locations of objects in a spatial array. Kindergarten and adult subjects were trained to remember the locations of 4 objects. During this training, objects were either connected by a combination of indirect, looped train tracks and direct train tracks (experimental subjects) or connected by entirely direct train-track routes (control subjects). Analyses of actual interobject distances, from subjects' reproductions of object locations on a response board (without train tracks), revealed that children, but not adults, distort distance in terms of the nature of travel observed between objects. Further testing revealed that differences in the amount of time taken for travel could not account for the results obtained with children.


An evaluation of the use of spelling error analysis in the diagnosis of reading disabilities. This study examined whether significant differences could be observed in types of errors made by normal versus retarded readers (fifth graders) on the WRAT spelling test. Although researchers have reported high numbers of certain types of errors, such as reversals, this study revealed that the proportion of errors attributed to any particular error type is virtually identical for both normal and retarded readers. Retarded readers were thus shown to be unlike normal readers in the number of errors committed but like normal readers in the types of errors committed.


Caregiver-child interactions and the type A coronary-prone behavior pattern. Type A behavior, 1 of the risk factors for heart disease, is characterized by impatience and competitive achievement striving. An opposing type B pattern is characterized by patience and a lack of competitive achievement striving. Previous observations of mother-child interactions revealed that mothers gave type A and type B children different frequencies of pushing and positive task evaluation. To isolate the effects of type A and B children on caregivers, the present study examined the caregiver behavior of female strangers elicited by boys' type A and B behavior. In addition, it examined whether the child effects differed according to the caregivers' behavior pattern. Results indicated that, relative to type B children, type A children elicited more pushing and positive task evaluation from type B caregivers. However, type A caregivers did not respond to the differences in the children's behavior pattern. The caregiver behavior of type A's and type B's is discussed in light of previous experimental findings of their general responsiveness to variations in their environment.


The modes of the anterior hypothalamic area as the regulatory center for the gonadotropin release. The effects of the anterior hypothalamic area (AHA) implants of gonadal steroid estrogen and progesterone as well as the effects of electrical stimulation and electrolytic lesion confined in this area on the gonadotropin secretion were investigated in ovariectomized estradiol (20 microgram sc)-primed adult Wistar rats housed in light and temperature controlled room. Progesterone implants evoked the rise of serum LH by 6 hr whereas estradiol implants suppressed serum FSH by 24 hr after implantation. Electrical stimulation effectively depleted both gonadotropins with a latency not shorter than 6 hr. The lesion significantly prevented FSH elevation investigated at 72 hr post ovariectomy and potentiated FSH secretion in response to estradiol treatment at 3 week post ovariectomy. The result revealed the involvment of the AHA in LH release mechanism which required progesterone activation while its involvement in FSH regulatory mechanism depended upon estrogen. The area was elucidated as the inhibitory as well as the stimulatory loci for the feedback action of estrogen on FSH release.


Mammary gland differentiation in adult male rat--effect of prenatal exposure to cyproterone acetate. Breast tissue of adult male Holtzman rats exposed to cyproterone acetate during embryonic differentiation showed presence of specific estradiol receptor proteins and C-19 steroid aromatase. We reported similar findings in gynecomastia in man. It is therefore proposed that gynecomastia probably results from failure of adequate testosterone action on the breast primordia during embryonic differentiation.


Plasma basal levels of FSH, LH and testosterone in homosexual men. Plasma basal levels of FSH, LH and total as well as apparent free testosterone were determined in homosexual and heterosexual males. Significantly higher FSH concentrations (mean +/- SD = 6,89+/-5.00 mlU/ml; P less than 0.01) and LH concentrations (28.2+/-30.7 mlU/ml; P less than 0.002) were found in plasma of 50 homosexual males compared with those of 24 or 40 heterosexual males 4.17 +/- 2.34 mlU/ml and 12.6 +/- 7.6 mlU/ml, respectively). Significantly lower free plasma testosterone was observed in 35 homosexual males (10.7 +/- 3.3 ng/100 ml; P less than 0.01) than in 38 heterosexual males (13.3 +/- 4.5 ng/100 ml), whereas total testosterone in plasma of homosexual males (590 +/- 148 ng/100 ml) showed no significant difference in comparison with the heterosexual control group (562 +/- 126 ng/100 ml). The tendency to higher FSH and LH values as well as to lower free testosterone concentrations in plasma compared with the heterosexual control group was more evident for effeminized than for non-effeminized homosexual males. FSH and LH concentrations in plasma of 5 transsexual males were also significantly higher (11.74 +/- 5.06 and 18.3 +/- 3.4 mlU/ml, respectively; P less than 0.02) than those of the heterosexual control group. Our findings may be explained by the possible existence of a prenatal testicular androgen deficiency in homosexual males that is widely compensated by increased gonadotrophin secretion in adult life.


Effect of long-term administration of estrogens on the subcellular distribution of cholesterol and the activity of rate-limiting enzymes of cholesterol biosynthesis and degradation in pigeon liver. To determine the mechanism of the hypocholesteremic effect of estrogens noted in pigeon, studies were done on the subcellular distribution of cholesterol and the activity of beta-hydroxy-beta-methylglutaryl CoA reductase (HMG-CoA reductase) and and cholesterol 7alpha-hydroxylase in pigeon liver after long-term (6 months) estrogen administration. Estrogens significantly reduced free cholesterol concentration in microsomes and mitochondrial fraction. The concentration of cholesteryl ester was reduced in the supernatant fraction. The activity of HMG-CoA reductase was significantly reduced in the estrogen-treated birds, while cholesterol 7alpha-hydroxylase activity showed no changes. Thus, the hypocholesteremic effect of estrogen noted in pigeons could be mainly due to the inhibition of cholesterol biosynthesis in the liver.


Lipid metabolism in rats with isolated medial hypothalamus. Based on determinations of the serum total cholesterolemia, triglyceridemia, free fatty acids and obesity Lee index, performed in rats with isolated medial hypothalamus--VMH included--which had either free food access, or after 24 hours of starvation, or maintained on limited food intake, it has been concluded that the VMH nucleus is involved in the control of lipid metabolism especially through its inhibitory action on the lateral hypothalamic parasympathetic area.


The problem of diabetic angiolopathy in sand rats (Psammomys obesus). No diabetic angiolopathy was found in the retinas, kidneys and skeletal muscles of protodiabetic and overtly diabetic sand rats. The terminal blood vessels were investigated using histological, enzyme histochemical, immunofluorescence microscopic, autoradiographic and electron microscopic methods. There seems to be little or no connection in sand rats between the diabetic metabolism syndrome and the metabolic process leading to angiolopathy.


Variations of glucose utilization in muscle and adipose tissue of sand rats (Psammomys obesus) during adaptation to laboratory holding. Sand rats, captured in Egypt and fed with a low caloric vegetable diet during adaptation, were investigated before and after 2.5 and 8 weeks diet treatment (30 and 40 kcal/100 g body weight daily). In hexobarbital anaesthesia the sand rats were loaded with 1 g glucose/kg body weight in a single dose intravenously. After a rapid increase the content of glucose in blood remained at a level of about 600 mg glucose/100 ml blood. The insulin immunoreactivity in blood did not change uniformly after application of glucose and remained in a physiologic range. In the islets of Langerhans a degranulation was found during diet treatment. The sensitivity of the epididymal adipose tissue towards insulin in vitro decreased to a nearly complete resistance in the course of diet treatment. A diminution of insulin sensitivity was also found in the m. soleus in vitro. The content of glucose-6-phosphate in the m. semimembranosus was found enhanced after the preparation of the animal. It was found progressively increased up to the five-fold at the end of diet treatment. In the corresponding muscle the glucose distribution volume was increased to about double the extracellular volume. An accumulation of free glucose within the muscle cell must be taken into account. In conclusion the treatment of sand rats with a diabetogenic diet results very quickly in a loss of insulin sensitivity of adipose tissue. The progressively increased stress-mediated accumulation of glucose-6-phosphate and free glucose refers to an inhibition of glucose utilization in the phosphorylation step of glucose in skeletal muscle.


The effect of serotonin given into the lateral ventricle of the brain on serum TSH level in normal and thyroxine-blocked rats. The effect of serotonin given into the lateral ventricle of the brain in rats was investigated. Serotonin was solved in artificial cerebrospinal fluid and administered in three doses: 250 microgram, 50 microgram, 10 microgram. Only the highest dose of serotonin caused a statistically significant increase in serum TSH level 0.5 h after the injection. The possible role of serotonin in the blocking mechanism of thyroid hormones on TSH secretion was also taken into consideration. 50 microgram of serotonin significantly strengthened the lowering effect of 1-thyroxine given 0.5 h before on serum TSH level.


Experience of prolonged treatment of hand-Schüller-Christian disease with prednisolone. An account is presented of the experience acquired in the prolonged prednisolone treatment, at 2-day intervals, of 6 patients with Hand-Schüller-Christian disease. A substantial improvement was observed in the basic process, with scarcely any side-effects. On the basis of the literature data and the present findings, glucocorticoid administration every second day is recommended for the treatment of these patients.


Effects of oxytocin and estradiol on volumes of the cell nuclei of the pituitary pars intermedia in male rat. After administration of estradiol, oxytocin and both together to a male rat, investigations of the pituitary pars intermedia were performed. Diminution of the cell nuclei volumes after simultaneous administration of oxytocin and estradiol was observed. The experiment has stressed the role of both hormones in melanotropin release control.


Ventilation-perfusion lung scans for pulmonary emboli. Accuracy of reporting. Reports were made on combined ventilation-perfusion lung scans by three observers on three occasions and by another observer once. Reproducibility for each observer varied between 80 and 88%. There was complete agreement about the areas of scans reported as abnormal. Agreement between observes on whether or not the abnormality represented a pulmonary embolus averaged 77%. There was 86% agreement with the final clinical diagnosis. Our results show that reporting of ventilation perfusion lung scans by eye is reproducible. They support the claims that, under routine clinical conditions, the technique is 91% to 95% accurate for pulmonary emboli.


Value of radioisotope axial tomography in the diagnosis of a cystic malignant glioma. A case is presented in which the patient had a deep tumour in the posterior aspect of the temporal lobe close to midline. The brain scan including a transverse section view at 8 cm from the vertex was negative and the other neuroradiological examinations inconclusive. One month later a follow up brain scan was performed and the conventional views were negative whereas the transverse section scan done at 10 cm from the vertex showed a lesion in the left temporo-parietal region and near midline. Three months later a repeat brain scan was done, and the lesion was visualized in the conventional views and confirmed by angiogram.


The measurement of spleen perfusion in man: a non-invasive method using the ratio between Rubidium-81 and its decay product Krypton-81m. A non-invasive method has been developed for measuring spleen perfusion in man. This involves recording the gamma-ray energy spectra over the spleen following the localisation of Rubidium-81 within the organ by injecting intravenously labelled heat denatured red cells. The spectra are analysed to provide the ratio of Rubidium-81 to its radioactive decay product Krypton-81 m. This ratio is dependent on the rate of perfusion through the organ.  The difficulties encountered in applying this steady state method for monitoring tissue perfusion are illustrated and practical means for their solution presented. The results of applying this method to patients with various splenic disorders are presented and compared with those obtained by other workers using different monitoring techniques.


A computer algorithm for triple radionuclide subtraction studies applied for preoperative localization of enlarged parathyroid glands. This paper presents a computer algorithm for triple radionuclide subtraction studies and its application to preoperative localization of enlarged parathyroid glands. In the clinical examination procedure, 131I-toluidine blue was used as the principal radionuclide, 99mTcO4 and 113mIn were used to obtain additional images of the thyroid and the blood distribution respectively. A scintillation camera with a pinhole collimator and connected to a digital data aquisition system was used to record the images. Subtraction of uniform background, thyroid and blood contributions to the principal image is done automatically in the computer program. The results from a clinical study is used to illustrate the method.


Use of 131I-19-cholesterol in functional dynamic studies of the adrenals in vivo. II. Clinical application. Functional dynamic studies of the adrenal in vivo employing 131I-19-cholesterol were carried out as a further screening test, according to the method previously described (Pavonia and Magrini, 1977), on seven patients in whom hyperadrenocorticism was suspected on the basis of clinical symptoms. The data obtained in basal conditions, under ACTH stimulation, and under dexamethasone suppression, compared with each other and supported by the results of conventional examinations, demonstrate that the quantitative information provided by this radioisotope test is correlated with the functional state of each adrenal, thus confirming the potential value of the method for diagnostic purposes. The functional dynamic test proposed does not require a higher dose of tracer than morphologic studies and the radiation risk (approximately 2.5 rad to the ovaries or gonads) seems acceptable in adult subjects.


Comparison of technetium uptake in small joints with other indices of inflammation in rheumatoid arthritis. The uptake of 99mTc in the small joints of the hand has been compared to various other mesures of inflammation in 22 patients with rheumatoid arthritis. The 99mTc uptake in the finger joints can reflect both disease activity at a given time and change in activity over a period of time. The place of Tc uptake amongst other measures of inflammation, from the point of view of the 'relative efficiency' of the assessment method, is rather inferior. In evaluating the effect of antiphlogistic drugs on local inflammation, techneticum uptake may, however, prove more discriminating than grip-strength or the number of painful joints, since the latter measures depend largely on the degree of destruction.


Fully automatic computer-assisted pancreas imaging. Good pancreas imaging is difficult to achieve even if a computer is applied. The special computer program proposed here was developed to optimize the results and to minimize the operator interaction with the data processing system.


Determination of trace elements in whole blood by photon-induced X-ray fluorescence. Photon-induced X-ray fluorescence is used to determine elements in human whole blood samples. The method is reliable and has a good sensitivity for a wide range of elements. The simplicity of the technique and the associated instruments makes the method suitable for use in hospitals.


A comparison of a new whole body scanner with a large crystal scanning camera in whole body imaging. Physical parameters and the clinical experience obtained in one year with a new whole body scanning system are presented. Advantages and disadvantages of this system are described in comparison with a large crystal scanning camera. The whole body scanner has two detectors each an array of ten crystals with a large area (700 cm2 per head) able to scan a surface of 60 X 193 cm. The information is formed in a 160 by 512 point matrix and stored on a floppy disc. X-ray or polaroid film output is available. Whole body bone scans (both A.P. and P.A.) with excellent resolution are obtained in less than 25 min. The relative sensitivity and resolution of both imaging systems was studied using line and plane source phantoms and clinical material. Taking into account the different physical parameters of the systems an attempt at a practical comparison of scans is made in terms of time, sensitivity, resolution and image quality.


Evaluation and improvement of different methods for the enrichment of antigen-binding cells of chickens. Antigen-binding cells were enriched using bovine serum albumin immunized chickens as an experimental model. A comparison of density centrifugation, adherence to derivatized surfaces (polystyrol and gelatin) and affinity chromatography revealed that optimal enrichment was obtained using the latter procedure. Polyacrylamide was superior to agarose.


Studies using labelled C. parvum preparations in mice. The in vivo distribution of 125I and fluorescein labelled C. parvum, after local and systemic injection mice has been studied. After i.v. injection large amounts were found in the liver, spleen and lungs, with less in the lymph nodes. The distribution after s. c. injection was more restricted; the bulk of inoculum was retained at the injection site and draining lymph node, and only small amounts were found in the liver and spleen. C. parvum was found in bone marrow, but not peritoneal cells after i. v. injection. It was, however, possible to recruit C. parvum-containing cells into the peritoneal cavity by i. p. injection of thioglycollate, and to a lesser extent, i. p. injection of syngeneic tumour cells.


Comparative studies on the cell wall composition of some anaerobic coryneforms of varying lympho-reticular stimulatory activity. Anaerobic diphtheroids possessing lympho-reticular stimulatory properties may differ considerably in their peptidoglycan composition. Spleen weight-increasing activity of strains directly parallels their antitumour properties. P. granulosum strains, inactive in assays for lympho-reticular stimulation, appear to have a higher cell wall alanine content than most of the P. acnes and P. avidum strains tested. Two P. acnes strains, however, had equivalently high alanine ratios and were stimulatory. The presence of galactose does not appear to be required for activity since P. acnes II strains which lack this sugar can be fully stimulatory. The existence of the species P. lymphophilum (Torrey) is further supported by the finding of two more serologically identical strains which do not cross react serologically with the other species in the group. These organisms are fully stimulatory but have lysine rather than DAP as their cell wall diamino acid.


Further studies on the enhancement of the antibody response in vitro by C. parvum. We have shown previously that spleen cells from mice pre-treated with C. parvum give enhanced antibody response when cultured in the presence of thymus-dependent antigens. In order to establish if C. parvum has a similar effect when added in vitro, spleen cells from untreated mice were cultured with C. parvum in Marbrook chambers with sheep erythrocytes (SRBC) as antigen. At concentrations of 35-140 microgram/ml/20 x 10(6) spleen cells C. parvum enhanced, but at higher concentrations suppressed the in vitro response. Cultures containing lymphocytes from C. parvum treated mice and adherent cells from untreated animals, which do not show in vitro the enhancement in the absence of C. parvum, gave higher responses to SRBC when a critical dose of the vaccine was added. Supernatants from cultures of normal spleen cells with added C. parvum could enhance the in vitro response of normal spleen cells. In double Marbrook cultures, cells from mice pre-treated with C. parvum and cultured with this organism failed, however, to enhance the response of untreated cells in the outer vessel. Under these conditions the response was enhanced only when SRBC were also present in the inner vessel.


Antibody responses, antitumour antibodies and Ig class and subclass levels in C. parvum treated mice. This paper summarises the results of extensive studies performed in our laboratory during the past three years aimed at establishing the effect of C. parvum administration on humoral immunity in normal mice and mice previously challenged with viable MC fibrosarcoma cells. This treatment was found to (a) increase the response to thymus dependent and independent antigens, (b) result in the production of circulating immunoglobulins which bound to tumour cells and syngeneic tissues in vitro, and (c) marked increases in the levels of certain immunoglobulins, especially IgG2a and IgG2b. The latter effects were dependent upon the dose and route of injection and were inhibited by the administration of gold salts. Similar though less marked effects were also observed in T cell deprived and nude mice and in normal mice injected with other bacterial adjuvants. The mechanisms by which C. parvum exerts the above serological effects and their relevance to antitumour immunity, will be discussed.


Comparative studies on the significance of in vitro data of BCG vaccines. In vitro parameters of BCG vaccine, such as colony-forming units or bacterial weight, are significant with respect to efficacy and occurrence of side effects only for vaccines which are made by one manufacturer according to one method and tested in one laboratory. With respect to colony-forming units from different products, widely varying doses must be applied to gain comparable effects. Similar effects can be expected from the human dose, recommended for each particular product. These conclusions, derived from experiences in vaccination against tuberculosis can also be considered essential for comparative immunotherapy with BCG vaccines.


Distribution of 3H-thymidine-labelled C. parvum in mice. Little is known of the localisation of injected Corynebacterium parvum. We therefore developed a method of radiolabelling this vaccine and determined its distribution in the mouse. Live C. parvum (Wellcome, strain CN 6134) was grown in the presence of 3H-thymidine and subsequently killed by formalin. A high activity, 1-6 x 10(5) cpm/0.1 ml of a 7 mg dry weight/ml concentrated suspension, was obtained. Its biological properties (hepatosplenomegaly and antitumour effects) were similar to those of commercially available vaccine. After intravenous or intraperitoneal injection into normal mice, high activity was recovered in liver and moderate activity in spleen, lungs and small gut. In contrast, after subcutaneous injection, most activity was recorded at the injection site, and little in other tissues. A similar distribution of labelled C. parvum was found in tumour-bearing mice. Only moderate counts were detected in tumour.


Monocytosis-inducing activity (MIA) of serum in Corynebacterium parvum treated mice. Intravenous injection of 548 microgram of C. parvum (Lot 0407, Mérieux Laboratories, France) into C57Bl mice produced rapidly appearing monocytopenia which was followed by marked and prolonged monocytosis after the third day. The serum of these animals, collected during the monocytopenic but not the monocytosis phase, showed monocytosis inducing activity (MIA) as was demonstrated by the intravenous injection of the serum into normal test mice. Serum from normal untreated mice or from mice given an intravenous injection of sterile pyrogen-free saline did not cause monocytosis in the test mice. Monocytosis induced in the test animals presented two interesting peaks. The first was observed 2 h after the injection of serum and the second 5 days later. The former was accompanied by a decrease and the latter by an increase in the number of bone marrow monocytes, suggesting that MIA probably represents a releasing activity. The late increase in marrow monocytes is considered as a phenomenon secondary to the initial reduction.


In vitro anti-tumour properties of peritoneal exudate cells of conventional germ-free and stimulated mice. Adherent cells of peritoneal exudates were obtained from conventional, or from germ-free mice or from mice having received in intraperitoneal infection of a variety of phlogogenic substances such as Corynebacterium parvum (C. parvum) (Mérieux) 500 microgram, thioglycolate (Difco) 3 ml, Bayol (Esso) 0.50 ml + 0.50 ml culture medium, glycogene 1.2 ml. The cytotoxic properties adherent cells were studied in vitro by the chromium release technique (CRT) and their cytostatic properties by the inhibition of the incorporation of tritiated thymidine by YC8 lymphoma cells. C. parvum was found to be the most active agent in enhancing the cytotoxic properties of adherent cells, followed by BCG and Bayol. Glycogen peptone and other substrates were without effect. The unstimulated peritoneal macrophages of conventional mice were found to inhibit thymidine incorporation by tumour cells, whereas those of germ-free mice could not do so. C. parvum markedly increased the cytostatic property adhered cells from both germ-free and conventional mice.


Cell mediated immunity to Corynebacterium parvum and its role in tumor control. The injection of C. parvum provokes a transient cell-mediated immune response to its own bacterial antigens. During this period a subcutaneous challenge dose results in the appearance of a characteristic delayed type hypersensitivity (DTH) skin reaction. If a suspension of syngeneic tumor cells is included in this challenge, they are destroyed in immune but not control animals. Similarly, solid tumors of limited size can be made to regress under these conditions. Adoptive transfer studies revealed that, in keeping with other DTH reactions, this effect was mediated by a population of rapidly dividing, theta-bearing lymphocytes. The ramifications of this response with respect to antitumor therapy are discussed.


Comparison of freeze-dried vaccines prepared from four different strains of BCG. Cancer patients in many centers throughout the world are being treated with a variety of immunological procedures and one of the most widely used requires the administration of BCG vaccine. BCG vaccines made under the same conditions from four different strains of BCG were investigated as to their viable counts and residual virulence. The highest number of culturable particles (CP) was obtained from the vaccines prepared with the Japanese and Connaught strains, whereas the lowest number was obtained from the vaccines prepared from the French and Danish strains. The virulence of the vaccines in Syrian golden hamsters was independent of the number of CP present in each vaccine. The most virulent vaccines were the French and Danish vaccines, the least virulent was the Japanese vaccine and the Connaught vaccine was of intermediate virulence. Although the hamster can be used as a good model to determine the level of residual virulence of BCG, it should be said that, thus far, there is no evidence to show that a BCG vaccine with a high level of virulence for the hamster will necessarily be a better adjuvant or immunostimulant in the treatment of malignancies in man than a vaccine with low virulence.


Effects of surgery and C. parvum on tumour metastases in mice. Surgical operations depress immune responses. The aim of this study was to observe the effects of operation (amputation) on the growth of the Lewis lung tumour and its metastases and to see if C. parvum counteracted them. Anaesthesia alone, or with amputation, did not affect the growth of the primary tumour but C. parvum depressed it. Anaesthesia did not affect the number of pulmonary metastases but amputation caused significant increase. C. parvum inhibited metastases and completely counteracted the effects of operation. Large amounts of cortisone significantly increased metastases but small doses did not. Experiments in adrenalectomized mice suggested the effects of operation were due to non specific stress.


Effects and mode of action of Corynebacterium parvum on murine tumour metastases. The effects of Corynebacterium parvum (C. parvum) on Lewis lung tumour metastases have been studied. I.v. and i.p., but not s.c. C. parvum significantly reduced pulmonary metastases. When combined with excision of the primary tumour, C. parvum given not more than 2 days before caused slight reduction of metastases, but when given earlier, it caused significant reduction and some mice were cured. Metastases were increased by silica or cortisone acetate but were unaffected by trypan blue. The antimetastatic action of C. parvum was not altered by these treatments. Thymectomy and irradiation did not affect tumour metastases, or the antimetastatic action of C. parvum, whereas ALS depressed metastasis and abrogated the protective effects of C. parvum. It appears that the inhibitory effects of C. parvum on tumour metastases are mediated through macrophages in concert with a subpopulation of T2 lymphocytes.


Immuno BCG Pasteur F vaccine.

Immuno BCG Pasteur F vaccine. Immuno BCG Pasteur-F (fresh) vaccine is prepared from young cultures, 7-8 days old. Suspended in Dubos' modified medium it may be preserved for 3 months at 4 degrees C or for many years at--70 degrees C. Tests with the vaccine for colony-forming units during storage, biological controls in animals and allergy in children have demonstrated its good stability.


A comparison of the antitumour activities of glucans and C. parvum. After i.v. injection of glucans (yeast cell walls, lentinan and pseudonigeran) into CBA mice, the peritoneal macrophages became cytostatic to R1 leukemia cells in vitro, and the mice showed resistance to lung nodule formation by i.v. injected T3 fibrosarcoma cells. The glucans were less active than C. parvum. None of the glucans reproducibly acted as an adjuvant when mixed with irradiated tumour cells; nor did they inhibit the growth of a subcutaneous tumour when injected i.v. or directly into the tumour.


Chemo-immunotherapy of mouse tumours using cyclophosphamide and C. parvum. Treatment of a chemically induced mouse solid fibrosarcoma using either non-specific (C. parvum 350 mu i.v.), or specific active (s.c. C. parvum mixed with 5.10(5) irradiated tumour cells) immunotherapy, 4 days after a single dose of cyclophosphamide (200 mg/kg) was synergistically more effective than either C. parvum or drug treatment alone. A contributory factor may be that cyclophosphamide pretreatment has been shown to potentiate the specific antitumour immunity that arises from C. parvum interaction with tumour antigen. Systemic C. parvum before cyclophosphamide will potentiate the antitumour effects of the drug--previously ineffective low doses becoming effective. No similar potentiation of the effects of another alkylating agent, Melphalan, was evident.


Increased sensitivity of C. parvum treated mice to ionizing whole body irradiation. We have investigated whether C. parvum (CP)-induced stimulation of hematopoiesis affects the survival of mice exposed to the whole body irradiation (WBI). C3Hf/Bu mice treated with CP exhibit an increased hematopoietic colony forming activity in their spleens and blood, but not in their bone marrow, as determined by the exogenous spleen colony assay. Also, CP-treated C3Hf/Bu as well as CBA mice show an increase in their endogenous colonies. This increased hematopoietic activity caused by CP treatment did not protect mice from the consequences of the WBI ranging from 650 to 950 rads. In fact, more mice died if they had been treated with CP. A decrease in the number of erythrocytes was more pronounced in mice treated with CP and irradiation than in those given irradiation alone.


In vivo transfer of antitumor activity by peritoneal exudate cells from mice treated with C. parvum. We have investigated whether peritoneal exudate cells (PEC) from C. parvum (CP) treated (C3Hf/Bu mice could transfer in vivo the resistance against a syngeneic fibrosarcoma (FSa). Inhibition of tumour development and prolongation of survival of recipients were observed when CP-activated PEC were admixed with FSa cells before their intraperitoneal (ip) or subcutaneous (sc) injections into normal mice. The antitumour activity increased with the increase of the ratio of effector to target cells. Heat killed CP-PEC were unable to transfer the resistance. Also, pretreatment of recipients with 600 rads whole body irradiation (WBI) substantially reduced the efficacy of CP-PEC. Reconstitution of WBI mice with mixed normal spleen and lymph node cells, or spleen cells alone, or bone marrow cells did not restore the antitumor activity of transferred CP-PEC. In fact, reconstituted mice showed a further reduction of transferred antitumor resistance. CP-PEC activity was also inhibited in sc transfer experiments when normal PEC, spleen cells, T-cells or even fetal fibroblasts were admixed with tumor cells and CP-PEC. Possible reasons for the failure of WBI recipients to be fully protected by transferred CP-PEC are discussed.


Neuroblastoma and transfer factor.

Neuroblastoma and transfer factor. Some authors have demonstrated the cytotoxic capacity of the mother's lymphocytes against the neuroblastoma cells of the son. It is not known if that is the reason for the better prognosis of these tumors in early infancy, and it was decided to treat some similar patients with transfer factor from the mother. The conditions for the patients were: more than 2 years old, poor response to chemotherapy and/or part of the tumor not resected. 3 to 5 doses of transfer factor were administered to 3 patients 2 1/2, 4 and 6 years old. One dose was taken from 400 ml of blood and prepared as described in the literature. The patients remained without metastasis more than 1 year after the treatment.


Effects of C. parvum on radiation response of murine tumors. The effects of intravenous C. parvum (CP) on the efficacy of fractionated gamma-irradiation in treating a fairly immunogenic fibrosarcoma (FSa) and a weakly immunogenic mammary carcinoma (MDAH-MCa-4) were studied in C3Hf/Bu mice. Tumors were 8 mm in diameter at the start of irradiation. The FSa was exposed to 500 and the MDAH-MCa-4 to 750 rads daily for 3.6 or 10 days. The administration of 0.25 mg CP varied from 4 days before to 14 days after the start of irradiation. CP greatly augmented radiocurability of FSa, especially when applied before irradiation. The effect was evidenced by the increase in the cure rate, and, in mice not cured, by the tumor growth retardation, reduction of pulmonary metastasis incidence and prolongation in survival of mice. This effect of i.v. CP on FSa radiocurability was not further increased by intralesional CP, or by systemic injections of vitamin A or the hypoxic tumor cell radiosensitizer Ro-07-0582. CP was less effective in augmenting radiation response of the MDAH-MCa-4. Here, CP slowed the growth of irradiated tumors and prolonged the survival of mice.


Experimental evaluation of nonspecific immunostimulation and chemotherapy in the intracranial tumor-bearing Fisher rat. One hundred sixty rats bearing intracranial tumors were divided into groups of 10 to evaluate C-I interval +, effect of chemotherapy alone (BCNU), chemotherapy and C. parvum, and C. parvum alone in the rat tumor model. All therapy was administered intraperitoneally: BCNU -5.2 mg/kg (40% of LD10) on the 9th and 16th post tumor implant day, and C. parvum-1.4 mg regardless of the body weight and the 6th or 23rd day and 6th and 23rd post tumor implant day. An additional group received BCNU on the 9th and 16th day and C. parvum every 3rd day beginning with the 11th post implant day. It is concluded that C. parvum given before BCNU yields the greatest increase in survival in this animal system. Two or more injections were detrimental and injections after BCNU were of less value. C. parvum alone was minimally effective. If nonspecific immunostimulation is contemplated in this animal tumor system, it would appear to be best given before chemotherapy. This may relate to the brain as a "privileged immune site", or a peculiarity of the blood brain barrier in an intracranial lesion as it related to combined therapy.


BCG-immunotherapy of L2C guinea pig leukemia. In different experiments 46 guinea pigs of inbred strain 2 were inoculated intradermally with a lethal dose of about 10(5) gelatine purified L2C leukocytes per animal in mixture with 10(7) colony forming units (CFU) of BCG Copenhagen 1331. From these animals 58% showed a significant (p = 0.05) delay of death from leukemia in comparison to 38 animals not treated with BCG all perishing of leukemia with an average life time of 22.7 days (+/- 4.1 S.D.) after tumor cell inoculation. In 15% of the animals BCG caused long term survivors, still alive more than three months after L2C application. Twelve of them were challenged with nearly 10(5) L2C leukocytes; nine animals remained healthy demonstrating strong acquired resistance against L2C leukemia. Fresh submers cultured and lyophilized pellicle grown preparations of BCG Copenhagen 1331 were equally effective in antitumor activity. Using the same CFU dosage the lyophilized Copenhagen 1331 preparation was compared with two other commercial BCG vaccines and was found equivalent or better in efficacy against L2C leukemia.


The effect of pretreatment with Corynebacterium parvum on the growth of a chemical induced transplanted murine tumor. Unspecific immunostimulation with or without specific tumor immunotherapy may well contribute to the control of minimal residual cancer. Corynebacterium parvum (C.p.) has been shown to raise the level of immunocompetence in tumor bearing animals. Experiments are reported, which aim at an evaluation of optimal conditions for an immunostimulation with C.p. prior to the transplantation of BALB/c Meth A ascitis. Using different dosages, routes of injection and schedules of vaccination it was found, that the highest non-toxic dose of C.p. per mouse yielded the optimal inhibition of tumor growth, the longest survival of tumor bearing hosts and the highest number of tumor rejections as compared to untreated controls. If the vaccine was distributed to at least 4 sites of injection, the s.c. route appeared superior to i.p. application. The intravenous route appeared to be most effective. The oral route seemed to inhibit tumor growth, if not less than 4 mg/mouse were fed daily for five days. The mechanism of action of C.p. in the model system used is discussed and correlated with an appropriate timing of both C.p. sensitization and tumor challenge.


Human lymphoblastoid cells as a source of interferon. Interferons have considerable antitumour effects in animals, and have been used with encouraging results in patients with osteocarcomas and certain other tumours. So far only relatively small amounts of material suitable for use in man have been prepared, and almost all of this has come from human white blood cells [buffy coats]. Human fibroblast cell lines are now increasingly being used as an alternative source, but the resultant interferon differs in its chemical and biological properties from leucocyte interferon. Lymphoblastoid cells can also be induced to form an interferon which appears identical to buffy coat interferon. These cells can be grown in suspension in large tanks, and could provide large amounts of relatively inexpensive interferon. The advantages of this type of production system and the problems associated with it will be discussed.


Interference of preceding BCG vaccination with BCG immunotherapy in L2C-guinea pig leukemia. Guinea pigs of inbred strain 2 were vaccinated with BCG 1331 Copenhagen (K-Nr. 240-1) six to seven weeks before intradermal application of about 10(5) leukocytes from a highly leukemic animal in mixture with 10(7) colony forming units (CFU) of BCG. This presensitization with BCG totally abolished or strongly reduced the antitumor activity of L2C-cells adjoining BCG. Increase of bacterial mass applied from 1 X 10(7) to 2 X 10(8) CFU did not restore the antileukemic effect of BCG as observed in unvaccinated guinea pigs by a significant delay of death in a portion of animals and by the occurrence of long-term survivors. Since local reaction to BCG occurred much earlier in presensitized than in unvaccinated animals, we speculate that the destruction of tumor cells and hence the antigenic stimulus was too premature in presensitized animals to build up a BCG-mediated antitumor immunity.


The local effect of intravesical Corynebacterium parvum in the dog. A preliminary study. High doses of C. parvum introduced in the bladder of the dog provoke changes in the submucosa. The significance and importance of these modifications should be investigated.


A vaccine and immune serum for ascites tumor of mice and therapy of skin tumors. Of the 16 dyes studied, methylene blue med (MBM) was found to be the least and the most immunogenic either the dye-prepared vaccine or the dye solution injected simultaneously with ascites tumor (AT) cell suspension. The effective dosages of MBM ranged from 0.2 ml of 0.1% to 0.1 ml of 1%, i.e., 0.2-1.0 mg per mouse, weighed at 15-20 gm. Rabbits immunized with MBM-prepared vaccine produced specific antibody to AT in 2-3 weeks. Antisera produced by intraperitoneal injection were more effective than those produced by subcutaneous injection. Both rabbit immune serum and MBM dye solution were found to have therapeutic value in treatment of skin tumors induced by AT cell suspensions.


Effects of BCG and C. parvum treatment on human melanoma xenografts. CBA mice rendered immunodeficient by thymectomy, potentially lethal gamma-irradiation and reconstitution with bone marrow cells were used to grow a wide variety of human tumours as subcutaneous implants. Samples of human melanoma obtained at surgery were successfully passaged by transplantation and produced rapidly growing tumours, some of which metastasized to lung, lymph nodes and the para-aortic node; this system was used as a model for the study of immunotherapy of melanoma. Preliminary results show that intratumour injections of C. parvum retard or inhibit the growth of melanoma transplants and, therefore, do not require the involvement of T lymphocytes, whilst BCG has no effect on growth rate.


Toxicological studies with therapeutically-applicable Vibrio cholerae neuraminidase. Neuraminidase was submitted to a preclinical toxicological investigation in short--and long--term experiments. The acute toxicity in mice, rats, and guinea pigs is low (LD 50 i.v.: mouse 30,000 units/kg, rat 44,000 units/kg, guinea pig 7,700 units/kg; s.c.: mouse, rat, guinea pig more than 30,000 u/kg). After i.v. injection into dogs, respiration is not at all and blood pressure only slightly altered. Neuraminidase is pyrogenfree in the rabbit test. Assays of antigenicity in guinea pigs and tests on local i.v., i.m., s.c. toxicity in rabbits and guinea pigs showed only moderate toxic signs. Chronicity studies were performed in dogs and rats over 90 days with repeated administration of various amounts of Neuraminidase (dogs 50-2,000 u/kg, rats 500-2,000 u/kg i.v., 26 or 65 injections). Only in very high dose levels (more than 500 u/kg) Neuraminidase was toxic in dogs (damage of liver, kidney and erythrocytes). In lower doses in dogs and in rats the clinical, biochemical, and hematological data as well as urinalysis and anatomopathological results give no evidence of systemic or important alterations which would preclude the clinical investigation of the drug.


Histological changes of intravascular coagulation in mice resulting from intravenous injection of C. parvum. A histological study was made of liver and other tissues of mice at intervals after injection of C. parvum. As early as 20 min after injection occasional thrombi surrounded by polymorphs were seen in the alveolar capillaries. A general thrombosis rapidly developed in the hepatic vessels with consequent hepatic necrosis. Thrombi also developed in splenic vessels. This intravascular coagulation was prolonged, lasting at least 7 days. There was evidence that fresh thrombi were being formed throughout this time. The thrombosis was mirrored by a fall in platelet counts. The kidney was uninvolved in the reaction. The mechanism of this reaction and its possible relevance to the antitumour action of C. parvum is discussed.


Effects of Corynebacterium parvum vaccine on drug metabolism in the mouse. Mice of the CBA strain were given a single intraperitoneal injection of C. parvum vaccine at 50 mg/m2. There followed a progressive increase in liver weight and in the duration of hexobarbitone sleeping time, accompanied by a marked decrease in liver microsomal protein and in cytochrome P-450/mg microsomal protein. That is to say, content of cytochrome P-450 was reduced out of proportion to the apparent loss of endoplasmic reticulum. These changes were paralleled by decreases in microsomal O-demethylation of p-nitroanisole and glucuronide formation with o-aminophenol measured in vitro; they were most marked at 13-14 days after dosing; they then slowly returned to control levels which they reached 20-30 days later. Changes in the dose of vaccine showed that the increase in liver weight and in hexobarbitone sleeping time were significant at 3.5 mg/m2, but that the decrease in the activities measured in vitro were not significant below 35 mg/m2.


Control of viability, thermostability and residual virulene of BCG vaccines. Multiple comparison of laboratory data to select products for immunostimulation in the treatment of cancer. The aid of the work was to apply the multiple comparison method to select BCG products for immunostimulation. In the IABS-BCG Collaborative Study 1973 program coded BCG products (code) and reference preparation were investigated with 3 laboratory tests to compare differences concerning: (1) viability = viable units E6/ml (VU) of vaccines stored at 4C, (2) 37C thermostability (TS) slope = daily VU decrease of vaccines stored at 37C/0, 7, 14, 21, 28 days, (3) relative persistence capacity (RPC),--residual virulence--of the BCG strains in the mice spleen; regression followed up to 360 days. Culturable particles and recoverable VU were determined on BOAA medium. Means and regression coefficients of VU, TS and RPC were ranked after analysis of variance and evaluated by multiple-comparison-Gabriel method. Figures summarize the results. As to VU the products form 3 subsets; as to TS the products are in two subsets (ie potency of several products could be included in the same subset). Thus, the ranks of VU and TS are not the same. Also there are differences among overall rank of RPC (coefficient concordance of ranks of 2 VU doses, log and sqrt transformation of exp. data) and the VU as well as the TS. parallel VU, TS and RPC investigations and multiple comparison method could ensure the validity and reliability of the estimations in the quality control of products for immunostimulation and could explain the correlations and contradictions between the laboratory data and the results of the clinical trial in neoplasia.


In vitro and in vivo effects of Corynebacterium parvum on lymphocyte transformation. Using a range of doses the effect of C. parvum on the in vitro response of lymphocytes to PHA was studied. C. parvum was not found to be mitogenic itself when added to unfractioned lymphocyte cultures from normal donors and patients with breast cancer, nor did it effect the response to PHA in these cultures except at the highest dose (20 microgram/100,000 cells) when it was inhibitory. 21 patients with breast cancer treated with C. parvum were assessed by their in vitro response to PHA prior to, during and on completion of treatment. We were unable to show consistent changes in PHA response following treatment with C. parvum; 9 patients having shown a fall, 4 having shown an increase and 8 remaining unchanged in their PHA response.


Immunological changes in cancer patients receiving BCG. We employed immunological tests to assess patients with cancer, mainly melanoma, receiving BCG alone or combined with chemotherapy. A comparison of BCG recipients with similar cancer patients not receiving BCG showed differences in the frequency of Mantoux reactivity positivity in the leucocyte migration assay against tumour antigens and positivity in membrane immunofluorescence. Short term sequential studies showed conversion from negativity to positivity or strengthening of reaction to occur during initiation of BCG therapy. Extended sequential studies employing the leucocyte migration assay against tumour antigens and modifications to assess serum effects indicate that the changes are more related to the changing clinical status than to BCG. It may be possible to devise quantitative in vitro techniques to assay sensitivity to tuberculoproteins which may allow assessment of BCG effects in vitro. Current technology permits monitoring for varying clinical stage rather than the effects of immunotherapy.


The effect of intravenous infusion of Corynebacterium parvum on an immune profile of women with breast cancer. An immunological profile has been measured in 21 patients with advanced breast cancer who were treated with C. parvum (Wellcome strain CN6134). Thirteen patients received a single i.v. dose of 15 mg of C. parvum and 8 received 4 mg i.v. on 5 successive days. The "profile" was recorded before and 7-10 days after treatment and included measurement of total white count, absolute lymphocyte and monocyte counts, PHA response, B and T cell percentages. DNCB and Mantoux skin tests, immunoglobulin classes G, A, M and E and spleen size. Most patients showed a rise in white count, due largely to a polymorph leucocytosis, but there was no consistent change in any of the immunological variables recorded. IgG levels increased significantly following the single injection but not after the 5-day course; suggesting the possibility of acquired immunological tolerance. These results fail to demonstrate a consistent effect of C. parvum on either T-lymphocyte dependent function or on the spleen size, properties well documented in the experimental animal.


Effects of BCG and Corynebacterium parvum on immune reactivity in melanoma patients. Sixteen patients with disseminated melanoma were immunised with either BCG (8 cases) or C. parvum (8 cases) on three occasions at 21 day intervals. Blood for assay was taken immediately before the first immunisation and weekly for eight weeks thereafter. Total white count tended to increase but little change was seen in lymphocyte and monocyte counts. Serum IgG increased after BCG BUT NOT WITH C. parvum, serum IgA and IgM did not alter. The 'E' rosette % did show some increase mainly after C. parvum, and 'B' lymphoid cells (sIg staining) increased slightly after BCG; the 'EA' rosette % fell following C. parvum but not after BCG. Lymphocyte PHA blastogenesis increased after immunisation, particularly with BCG. Non-specific lymphocytotoxicity (51 Cr Chang target) demonstrated dramatic increases for 'non T' and 'K' cell function and a smaller increase in 'T' cell cytotoxicity following immunisation. These increases in cytotoxicity were maintained by the 21 day immunisation schedule.


Further studies on the serological effects of C. parvum immunotherapy in cancer patients. Previous serological studies undertaken in our laboratory have shown that the repeated administration of small doses of C. parvum to a small group of cancer patients resulted in a consistant increase in the levels of IgG (especially the IgG2b subclass), the development of antibodies to C. parvum and the transient appearance of 'rheumatoid factor like' substances in the serum (Br. J. Cancer, 32, 310, 1975). In the present paper we report the results of similar studies in small numbers of (a) inoperable bronchogenic carcinoma patients receiving combined C. parvum and cyclophosphamide therapy, and (b) brain glioma patients receiving intracerebral injection of C. parvum. As might be expected some of the serological changes observed in these patients were not as marked as previously noted. Nevertheless a number of the patients developed autoantibodies which were detectable by the latex and Ripley procedures but not by the Rose-Waaler test. There was no evidence of antinuclear factors by the fluorescence procedure.


First impressions of I.V. C. parvum in patients with solid tumours. 6 patients with solid tumours (4 malignant melanomas, one fibrosarcoma, one osteogenic sarcoma) received i.v. C. parvum alone or in combination therapy (radiotherapy, Levamisole and/or vitamin A). The single doses of C. parvum ranged from 5.0-7.5 mg/m2; No. of doses ranged from 1-8; interval between doses ranged from 2-140 days. 2 patients with malignant melanoma had no measurable disease, one of them (stage I) still has no evidence of disease. The patient with fibrosarcoma appeared to have a minor decrease in size of some of the lung metastases for a short time (ca 4 weeks), but soon progressed as well as the other 4 patients. Except for the one patients still having no evidence of disease the other 5 are dead of disease. Survival time did not appear to correlate either with systemic reaction to C. parvum or with No. of doses of C. parvum. The one minor response was observed in a patient receiving a total of 8 doses; but response was seen already after 3 doses, and progression occurred after 6 doses. So this change of lung metastases might have been unrelated to this therapy. Summarizing, there was no evident anti-tumour effect observed after i.v. C. parvum in these patients with solid tumours.


Experience with the quality control testing of C. parvum suspension for clinical trial. The control of C. parvum suspension presents special problems. The intended application of this preparation is in a therapeutic area where clinicians are accustomed to using defined chemical entities the biological activity of which are usually predictable according to weight of active principle administered, the route and number of doses. But the essential character of C. parvum is that of a vaccine in which the character of the active principle or principles is not clearly identified and can vary according to certain conditions. The control of the preparation therefore depends on the nature and relevance of tests for biological activity. These problems are exacerbated by uncertainty as to which of several possible mechanisms of antitumour activity might prove relevant in clinical practice. Experience with these problems as they have affected the quality control of large numbers of batches of C. parvum suspension prepared for clinical trial, will be discussed.


Complications of BCG treatment in patients bearing solid tumors. We treated one hundred patients who had various high risk solid tumors (malignant melanomas, osteosarcomas and lung cancers) by immunostimulation alone or with a sequential and synchronized chemotherapy as a complement treatment. Institut Pasteur BCG (150 mg) was administered either by scarifications (10 X 10 of 5 cm each) or multiple puncture technique (Gun), or in the case of 12 patients, by intra-tumor injections. The following complications were observed: chills and high fever during 1 to 30 days after scarifications or gun technique. In some cases an allergic loco-regional cutaneous reaction was noted after the gun technique. Nevertheless these complications were well tolerated. However, severe reactions were observed after the intra-tumor injections: malaise, chills, sweating, hyperthermia, nausea, vomiting and changes in blood pressure. In 1 case a prolonged high fever (3 weeks) was offset only by the use of corticosteroids. In another case the patient developed hepatitis. A percutaneous liver biopsy revealed noncaseating granulomas and the presence of acid fast organisms in the liver (by means of staining by auramine and observation by fluorescence). In this patient BCG has been replaced by Corynebacterium parvum (2 X 2 mg a week). This type of adjuvant was used in 2 patients and produced the same complications as the BCG. We believe that caution must be exercised in the use of such intra-tumoral treatments. BCG must be given in the hospital and patients must receive antihistaminic preparation before and after immunostimulation.


Toxic and immunologic side effects of daily C. parvum-infusion in treatment-resistant cancer patients. Daily increasing intravenous doses of Corynebacterium parvum (C.p.) up to 5 mg/m2 i.v. X 10-14 days were given to 6 patients with widespread metastatic neoplastic diseases resistant to radio- and chemotherapy. The immunotherapy treatment-cycles were evaluated for toxic and immunologic side effects and also for possible clinical benefit to the patients. Immunotherapy with i.v.-C.p. was moderately well tolerated. Subjective discomfort for the patients (headache, chills, nausea) was not better tolerated with ongoing treatment-doses. After the 3rd day the body temperature rose nearly regularly to 40 degrees and more within 3-4 h after i.v.-C.p. and returned to normal levels about 6-10 h after the infusion was stopped. Hematological values were monitored on day 1, 4, 8, 15. WBC counts rose after an initial moderate decrease to normal levels. Monocyte counts rose also after an initial transient fall to pre-treatment levels. The monocytic activity index of Naphthol-AS-D-Chloro-Acetate-Esterase, correlating with the monocyte turnover, did not show a significant change. Granulocyte counts, especially stabs, increased slightly. Lymphocyte counts, the number and relations of B, T and O-cells, did not change in a uniform typical way. Hemoglobin values fell in all patients, reticulocyte counts increased, and the blood sedimentation rate did not change.


Mouse tumour tests for quality control of C. parvum preparations. Description of two mouse tumor tests selected to control the quality of C. parvum pilot productions: transplanted Ehrlich's ascitic tumor in inbred Swiss mice and transplanted YC8 ascitic tumor in isogenic Balb/c mice. The results obtained during three years of control are analyzed, particularly accuracy, long range adequacy and reproducibility of tumor challenges and C, parvum stimulations. The effects of some parameters of the tumor tests, e.g. dose-responses, treatment schedules, tumor-host relationships and standard preparations are discussed.


Prospects for immunotherapy.

Prospects for immunotherapy. The limitations of conventional methods of treating cancer are such that it is worth devoting considerable resources to the search for new methods. The immunotherapeutic approach is based on theoretical grounds which, despite recent criticism, still appear compelling, and on much sound experimental work. Clinical benefits have so far been meagre but this is not surprising in view of the poor design of many of the trials and the large tumour burden in many of the patients. It is suggested that properly controlled trials, designed in the light of existing knowledge to answer a specific question, are much more likely to be rewarding than empirical attempts to win a therapeutic jackpot.


C. parvum skin testing antigen: study on guinea pig model. A skin-testing antigen produced from C. parvum has been developed for exploring cell-mediated immunity and specially C. parvum specific cell-mediated immunity by delayed cutaneous hypersensitivity (DCH) reaction. The DCH antigen and the techniques of intradermal injection and multiple puncture are described. DCH reactions are carried out in C. parvum specifically sensitized guinea pigs: sensitization procedure and adjuvant (IFA and CFA) effects are reported. Measures of DCH reactions for different antigen doses at various times (5-24-48 h) are reported; classical Mantoux and multiple puncture reactions are compared. The specificity of these DCH reactions are explored by comparing C. parvum antigen and tuberculin reactions in corynebacterium and mycobacterium sensitized guinea pigs.


A survey of patients' reactions to intravenous Corynebacterium parvum therapy. A survey was made of patients' reactions to high dose (21 mg) Corynebacterium parvum therapy given intravenously over four hours. The patients completed a pro forma. Rigors were the commonest side effect complained of being present in nearly all patients (24/25) and being very severe in one-third (10/25). Other problems were nausea (15/25), vomiting (17/25), bone pain (12/25) and headaches (12/25). One-third also usffered with other less common side effects. Overall, patients felt the side effects were as disagreeable as those experienced with outpatient chemotherapy.


Proposals for quality control methods of bacterial vaccines for immunostimulation. I. General considerations. Control methods for this type of vaccines include tests on identity, absence of contaminating microorganisms and safety which are required for any vaccine. In addition, for the control of vaccines for immunostimulation in cancer therapy some special methods for safety and potency testing are required. Some assays are proposed, such as stimulation of lymphocytes in vivo and in vitro, stimulation of macrophages (listeria clearance) in mice and the prophylactic and therapeutic tumor model in mice. Details of these tests are given in the companion papers II and III. The influence of the method of preparation of BCG vaccines on its mode of action in some of these models will be emphasized.


Intra-tumour C. parvum therapy in gastric carcinoma; a pilot study. In a preliminary and toxicological study, eight patients with carcinoma of the stomach were treated by Corynebacterium parvum immunotherapy prior to surgical excision. Killed C. parvum suspension was injected at multiple sites directly into the tumour mass with the aid of fibre-optic gastroscope at doses of 1.5 mg, 4 mg or 8 mg. Each patient received two injections of the same dose with an interval of 14 days. The second injections were made 14 to 18 days prior to surgery. No chemotherapy was administered during this treatment. Patients were monitored clinically for signs of toxicity. Immunological parameters were assessed on peripheral blood samples taken immediately before each injection and prior to surgery. Antibody titre to C. parvum was estimated by direct agglutination, while cell-mediated responses to C. parvum and 3M KCl tumour extract and phytohaemagglutinin (PHA) were assessed by lymphocyte transformation. Serum carcinoembryonic antigen (CEA) was measured by radio-immune assay. Tumours removed at surgery were examined histologically for the presence of invading immunocytes. These preliminary data will be discussed with respect to the dose of C. parvum administered.


Proposals for quality control methods of bacterial vaccines for immunostimulation. II. In vivo and in vitro stimulation of lymphocytes by BCG. An in vitro test for measuring the reaction of the draining lymph node after BCG vaccination and an in vitro test based on the stimulation of lymphocytes by concanavalin A were used to demonstrate immunostimulating properties of different BCG vaccines. A marked difference was seen between vaccines grown as a surface pellicle and ground in a ball mill, and those which were prepared from dispersed homogeneous cultures. In both systems the ball mill preparations were the less active.


Rapid turnover serum proteins in fulminant hepatitis. An attempt was made to find reliable indices for early diagnosis of fatal cases of acute viral hepatitis, using the values of serum proteins with rapid turnover. Of the subfractions of serum protein, prealbumin, alpha2-HS-glycoprotein and Normotest were measured simultaneously before the appearance of hepatic coma/or gastrointestinal bleeding in 78 cases of acute viral hepatitis, verified by biopsy or necropsy. The mean value of prealbumin with a very short half-life of one or two days, was 6.0 mg/dl in fatal cases, 7.4 mg/dl in surviving ones of subacute form of fulminant hepatitis. The difference between fatal and surviving cases was not statistically significant. In contrast to this, the values alpha2-HS-glycoprotein with a comparatively short halflife of four to six days, showed statistically significant difference between fatal (21.9 mg/dl) and surviving cases (37.4 mg/dl). Normotest was also depressed in fatal (10.7%) and surviving cases (45.3%). The difference was statistically significant. The present results indicate the possibility of differentiating fatal cases from surviving ones at an early stage, using the reduction of alpha2-HS-glycoprotein and the value of Normotest.


Clinical significance of e-antigen/anti-e, with special reference to HBc-antigen in the liver. e-antigen and anti-e were assayed in sera of asymptomatic HBs-Ag carriers and of patients with liver diseases. Thirteen out of 34 (38.2%) asymptomatic carriers were positive for e-antigen, which was in sharp contrast to the reports from USA and Europe. e-antigen was detected to a greater extent in patients with chronic active hepatitis, reversely anti-e in patients with chronic persistent hepatitis. However, e-antigen was found rarely in patients with cirrhosis and never in 23 cases with hepatoma positive for HBs-Ag. HBc-Ag in the liver was detected in 4 out of 8 e-antigen positive asymptomatic carriers and in 4 out of 5 patients with chronic liver diseases with e-antigen respectively, and moreover in 3 out of 14 anti-e positive cases, so that the presence of anti-e did not necessarily mean the negativity of HBc-Ag in the liver. Anti-HBc titer, however, was lower in anti-e positive sera than in e-antigen positive ones. This may implicate the decreased replication of HBV in cases with anti-e. These results emphasize that the investigation of e-antigen/anti-e is mandatory for the evaluation of the prognosis of asymptomatic carriers and of patients with chronic hepatitis.


Sulfobromophthalein and indocyanine green binding by the hepatic cytoplasmic proteins. The elution patterns of human and rat liver supernate mixed with BSP or ICG have been demonstrated using G-100 Sephadex gel filtration. The elution pattern in the rat liver remained the same as that shown by Levi et al. In the human liver BSP localized to Y protein could hardly be demonstrated, whereas ICG was detected. This pattern was quite similar in Rotor's hyperbilirubinemia and in Crigler-Najjar syndrome Type II. These observations suggest that the impaired hepatic transportation of organic anions in congenital hyperbilirubinemias may not be related to cytoplasmic organic anion-binding proteins.


Clinical studies on dissolution of gallstones using ursodeoxycholic acid. Ursodeoxycholic acid (UDCA), 7beta hydroxy epimer of chenodeoxycholic acid (CDCA), has been used as a choleretica for 20 years in Japan. Recent report showing increased excretion of UDCA in bile after CDCA administration may suggest the possibility that UDCA has similar effects to CDCA on bile cholesterol unsaturation and on gallstone dissolution. The present paper describes the clinical usefulness of UDCA for gallstone patients during the past two years. Seventy-four gallstone patients with functioning gall-bladders, 19 men and 55 women with a mean age of 48 years, have been treated for 6 months or more. UDCA, supplied in tablets (Ursosan), was given 450 mg per day. The disappearance or the reduction of stone size or number, or both (dissolution effect) was recognized in 32 out of 74 patients (43%). In case of radiolucent stones, the overall effective rate was estimated for 24 of 46 patients (52%). There may be no significant difference in dissolution effect between CDCA and UDCA treatment, however, the merit of UDCA treatment seems to have its few side effects.