Urinary excretion of adenosine 3',5'-monophosphate and guanosine 3',5'-monophosphate in normal children and those with cystic fibrosis. The urinary excretion of adenosine 3',5'-monophosphate (cyclic AMP) and guanosine 3',5'-monophosphate (cyclic GMP) was examined in 98 normal children and 46 children with cystic fibrosis between the ages of 9 months and 18 yr. Diurnal variations in cyclic AMP and cyclic GMP excretion were observed in subjects from either group, and peak levels of cyclic nucleotide excretion were generally observed during the period of 0700 to 2100 h. Excretion rates (mumol/day) of cyclic AMP and cyclic GMP increased significantly with age. When cyclic AMP and cyclic GMP excretion rates were normalized for urinary creatine, or body weight, the values declined significantly with age in both groups of patients. Cyclic GMP excretion normalized for body surface area also decreased with age, while the value for cyclic AMP (2.86 plus or minus 0.08 mumol/day/m2, mean plus or minus SE) was constant with age in both normals and cystic fibrosis children. With some comparisons of age groups there were significant differences in cyclic nucleotide excretion between normal subjects and children with cystic fibrosis. The differences noted were dependent upon the methods used to normalize excretion rates (urinary creatine, body weight, surface area, and the ratio of cyclic AMP to cyclic GMP excreated). In general patients with cystic fibrosis excreted greater amounts of cyclic GMP than did normals. The most striking comparison was the ratio of cyclic AMP to cyclic GMP excreted which was 9.09 plus or minus 0.50 in all normal children and 4.41 plus or minus 0.32 in children with cystic fibrosis (P smaller than 0.001).


Reversible hypothyroidism in growth hormone-deficient children treated with human growth hormone. Six children with human growth hormone (hGH) deficiency became hypothyroid during the course of their therapy with hGH. This was accompanied by a decreasing growth rate, clinical symptoms of hypothyroidism and decreased serum T4 concentrations. Three of the 6 patients returned to the euthyroid state, both clinically and biochemically, with cessation of hGH therapy, and reinstitution of hGH precipitated hypothyroidism again in 2 of the three. The patients who remained hypothyroid have evidence of multiple pituitary trophic hormone deficiencies while those who reverted to euthyroidism appear to have isolated hGH deficiency. Evaluation of thyroid function while on hGH showed low T4, free T4 and T3 concentrations. The serum thyrotropin (TSH) response to thyrotropin-releasing hormone (TRH) was absent or markedly blunted in 4 of 6 patients while receiving long-term hGH therapy but was normal or exaggerated in all patients when tested before or after only 5 days of hGH therapy. These data indicate that exogenous hGH results in an inhibition of the TSH response to TRH. The mechanism of this inhibition is unclear, but we postulate that it may be mediated by somatostatin secretion in response to pulse doses of hGH.


Radioligand receptor assay for urinary and serum hCG. A radioligand receptor assay system for urinary hCG and serum hCG was developed, using 1100-2000 times g fractions of homogenates of pig testis. This method is specific for hCG and LH, and the limit of detection for hCG was 20 mIU, which could be significantly discriminated from zero at the 95% confidence level. A nonspecific inhibition reaction which exerts its influence on the assay system has been found in the urine and serum, and in performing assays for urinary and serum hCG, it was necessary to remove the inhibition reaction by diluting the urine more than eight fold or subject the serum to an extraction procedure. Since the hCG of normal pregnancy and trophoblastic neoplasia showed a dose response curve similar to that of the International Standard hCG, it became clear that these hCG could be assayed with this method. The potency ratio R/I measured by RRA and RIA of urinary hCG and serum hCG was different from those of normal pregnancy and trophoblastic neoplasia.


Lack of correlation between gonadotropin and adrenal androgen levels in agonadal children. To evaluate the relationship between the secretion of gonadotropins and adrenal androgens, patients with gonadal agenesis were evaluated by (a) administering human luteinizing hormone (hLH) for 5 days with or without estrogen pretreatment to agonadal patients who had prepubertal LH levels; (b) correlating circulating gonadotropin levels with adrenal androgens in 45 patients; and (c) comparing adrenal androgens with gonadotropins after long-term administration of estrogen or androgens. Results are as follows: (a) No alteration in serum concentrations of dehydroepiandrosterone (DHA), dehydroepiandrosterone sulfate (DHAS), estrone (E1), testosterone (T), or in excretion of urinary 17-ketosteroid (17 KS) occurred after the administration of hLH. (b) No clearcut relationship between endogenous level of LH or FSH and DHA OR DHAS was demonstrated although a coincident increase of all hormones with age occurred. (c) Administration of estrogen to patients with gonadal agenesis did not affect their levels of DHA and DHAS although those patients given androgen developed higher DHAS, but not DHA, levels. Hence, increasing gonadotropin concentrations would not appear to be a primary etiologic factor in the maturation of the adrenal.


Relationship of bioassayable and immunoassayable plasma ACTH and cortisol concentrations in normal subjects and in patients with Cushing's disease. Plasma ACTH and cortisol concentrations were determined at 5-min intervals over a 3- or 4-h sampling period in 2 normal subjects. Time spans studied were 10:00 AM-1:00 PM, 4:00 PM-8:00 PM, 8:00 PM-11:00 PM, and 4:00 AM-8:00 AM. Similar sampling for 3 h, (onset 9:00-9:30 AM) was performed on 4 patients with Cushing's disease, 3 untreated and 1 in remission following pituitary irradiation. Two of these patients were studied on 2 separate occasions. Plasma ACTH was determined by both immunoassay (I) and bioassay (B). Although in general, these studies demonstrated significant correlation between I-ACTH or B-ACTH concentrations and those of plasma cortisol, a striking finding in both subject categories was the presence of 30- to 50-min episodes during which marked rises in both I- and B-ACTH concentrations occurred without concomittant, or markedly diminished, increments in plasma cortisol concentrations. This could not be explained by biological inactivity of the ACTH, since a highly significant correlation was present between I- and B-ACTH concentrations at all times; r values ranged between 0.86 and 0.98 for normal subjects, and 0.76 and 0.96 for patients with Cushing's disease. The lack of correlation in these episodes also does not appear to be secondary to an 11-beta-hydroxylase block, differences in the rate of change of plasma ACTH concentrations, lack of adrenal "priming" by prior ACTH or incapacity of the adrenal gland to further increase secretion. I/B ACTH ratios were similar in the normal subjects (1.42-1.64) and in the patients with Cushing's disease (1.27-1.47). "Apparent" ACTH half lives calculated from "peaks" of ACTH secretion were 7-12 min for I-ACTH and 3-9 min for B-ACTH in the normal subjects; and 9-13 min and 7-9 min respectively, in the patients with Cushing's disease. Mean plasma ACTH I-and B-concentrations at comparable time periods were higher in patients with active Cushing's disease than in normal subjects. These studies also indicate that in Cushing's disease, the abnormality present resides in ACTH regulatory mechanisms, not in the nature of the ACTH secreted. Approximation of the total amount of immunoassayable ACTH secreted in one normal subject over a 24-h period yielded a value of 73 mug. Total mug/h secreted in the 2 normal subjects were highest in the hour preceding awakening (6:30-7:30 AM; 12.9 and 12.2 mug/h); were 5.3 and 4.0 mug/h between 10:00-11:00 AM, and 1.4 and 1.7 mug/h between 9:00-10:00 PM. In the 3 patients with clinically active Cushing's disease, apparent ACTH secretion between 10:00-11:00 AM varied from 19.2-34.3 mug/h, the magnitude of such secretion being positively correlated with the extent of increased adrenal cortical activity present.


The effect of melatonin on steroidogenesis by the human ovary in vitro. Melatonin stimulated steroidogenesis in two compartments of the human ovary in vitro. In a corpus luteum of the menstrual cycle, melatonin increased progesterone synthesis in a dose related manner. Both serotonin and N-acetyl serotonin had no effect on progesterone synthesis. In the ovarian stroma, melatonin stimulated the incorporation of acetate-1-14-C into androstenedione. Binding of radioactive hCG by the corpus luteum was unaffected by melatonin. No specific binding of radioactive melatonin of low specific activity could be detected in homogenates of a human corpus luteum. These observations suggest that melatonin may directly modulate steroidogenesis in the human ovary.


Characterization of the glucagon receptor in a pheochromocytoma. Glucagon activated adenylate cyclase in a homogenate of a pheochromocytoma over the concentration range 1 times 10 minus 8M to 1 times 10 minus 6M. Several other hormones including adrenocorticotropin, thyrotropin, parathyroid hormone and histamine were without effect. The tumor glucagon receptor was characterized and found to be similar in several ways to the glucagon receptor previously reported in normal tissue such as liver and heart. One, the receptor specifically bound 125-I-glucagon. Two, solubilization of the pheochromocytoma abolished glucagon-activation of the adenylate cyclase. Three, glucagon-responsiveness of the adenylate cyclase was partially restored by the addition of phosphatidylserine to the incubations. One major difference was observed between the glucagon receptor in tumor tissue and that in liver and heart, namely, a marked lability in 125-I-glucagon binding and adenylate cyclase activity. Within four days, despite storage in liquid nitrogen, 75% of the binding activity and all of the adenylate cyclase activity in the solubilized preparation were lost. The factor(s) responsible for this lability remains unidentified.


Measurement of the estradiol receptor in human breast tissue by the immobilized antibody method. A simple immobilized antibody procedure for estradiol receptor detection has been applied to samples of human breast tissue cytosols previously assayed by the charcoal-dextran method. Good agreement between the two procedures was realized confirming the validity of the immobilized antibody assay.


The effect of alpha-MSH on plasma growth hormone, cortisol and TSH in children. The effect of synthetic alpha-MSH injected intravenously in a uniform dose of 3 mg was studied in 19 prepubertal children. A marked growth hormone (GH) response was seen only in 2 out of 8 constitutionally small children with a normal GH response to insulin and arginine stimulation. Three of of 11 children suffering from hypopituitarism with documented GH and other hormone deficiencies, unexpectedly, showed a significant rise of GH after alpha-MSH: all three had craniopharyngiomas. Alpha-MSH led to an increase of plasma cortisol in all except 3 patients who had secondary adrenal insuffciency. The increase of cortisol after alpha-MSH and after insulin was of the same extent: but the hypoglycemia and stress responsible for the insulin effect were not observed after alpha-MSH. It is possible that alpha-MSH acts by an ACTH-like direct stimulation on the adrenals. There was no effect of alpha-MSH on plasma TSH or on blood glucose.


Relationships among the secretion of ACTH, GH, and cortisol during the insulin-induced hypoglycemia test in the normal and obese child. The behavior of the secretion of ACTH, GH, and plasma cortisol during the insulin-induced hypoglycemia test on normal and obese children was studied. The secretion of the above-mentioned hormones was determined by calculating the integrals of the curves. The mean values of the integrals of the plasma cortisol and ACTH curves do not show any significant differences between the two groups of children. The mean values of the ratios between the integrals of the plasma cortisol and ACTH curves show a significant difference between the two groups (P smaller than 0.01). Since the mean values of the integrals of the plasma cortisol curves are practically the same in both groups, the difference in the above-mentioned ratios refers to the lower values of the integrals of the ACTH curves found in the obese children. This enables us to make the hypothesis that in the obese child the function of the pituitary-adrenal axis, at least during the insulin test, does not differ from the norm thanks to the adaptation of the ACTH secretion to the greater sensitivity of the adrenal glands to this hormone. In both the groups examined there was no correlation between the secretion of ACTH and plasma cortisol, between ACTH and GH, and between plasma cortisol and GH.


Plasma growth hormone and slow wave sleep increase after interruption of sleep. A comparison was made of plasma hGH and of sleep stages during one night of undistrubed sleep and one night in which sleep was interupted by an hour of enforced wakefulness folowing the end of the second NREM-REM sleep cycle in 8 normal subjects. Plasma was sampled at 15-min intervals. HGH and slow wave sleep were both significantly increased in the two cycles immediately following the period awake compared with the same two cycles during nights of uninterruped sleep. The difference arose predominantly in the fourth cycle of the night, i.e., in the second cycle after the sleep interuption. The findings are consistent with the belief that extra wakefulness brings additional sleep of high RESTORATIVE properties.


The effect of lergotrile on galactorrhea and gonadotropin secretion. Lergotrile (Compound 83636), a specific inhibitor of prolactin release, was administered to 5 women with galactorrhea. Only 1 of the 5 had an elevated serum prolactin (hPRL) value, but Compound 83636 induced a decrease in serum hPRL in all cases. All patients had cessation of breast secretion within 72 days after initiation of lergotrile. LRH administration (100 pg) evoked a variable gonadotropin response before and during treatment with the ergot alkaloid. By contrast, serum prolactin values remained essentially unaltered after injection of LRH under the same conditions.


A specific, high-affinity, limited-capacity estrogen binding component in the cytosol of human fetal pituitary and brain tissues. A macromoleclar component which binds 3H-estradiol with high affinity and limited capacity is present in the 150,000 x g supernatant fraction of pituitary, hypothalamus, cortex, and limbic system of both male and female human fetwses. 17 beta-estradiol and 17alpha-ethinylestradiol effectively compete with 3H-estradiol for binding sites while testosterone, dihydrotes-tosterone, and cortisol do not. The binding component is different from sexhormone-binding-globulin. It is present in these tissues in relatively large amounts and it is tentatively identified as an estrogen recptor.


The effect of serotonin (5-hydroxytryptamine) and derivatives on guanosine 3',5'-monophosphate in human monocytes. The effects of several putative mediators of inflammatory responses on the cyclic nucleotide content of mononuclear leukocytes from human peripheral blood were investigated. Incubation of mononuclear cells with 100 muM serotonin (a maximally effective concentration) for 5 min caused a five- to eightfold increase in their content of guanosine 3',5'-monophosphate (cAMP)...


Effect of adrenalectomy and insulin insufficiency upon the adenosine 3', 5' cyclic monophosphate system of the rat mammary glands. Effects of adrenalectomy and acute insulin insufficiency upon tissue adenosine 3', 5' cyclic monophosphate concentrations, and adenyl cyclase, phosphodiesterase, and protein kinase activities were investigated. Adrenalectomy decreased intracellular adenosine 3', 5' cyclic monophosphate 53% and increased the activities of both adenylcyclase and phosphodiesterase. Cortisol therapy returned these to normal. During insulin insufficiency caused by anti-insulin serum, mammary adenosine 3', 5' cyclic monophosphate concentrations increased. The acute effects of insulin insufficiency and chronic effects of adrenelectomy suggest that insulin acts upon rat mammary glands to decrease and glucocorticoids, acting over longer term, to increase adenosine 3', 5' cyclic monophosphate concentrations.


Comparative ultrahistochemistry of the adepidermal granules of Salmo irideus, Lebistis reticulatus and Hynobiuo tokyoensis. Enzyme digestive experiment for the epoxy-embedded sections;. A comparative ultrahistochemical investigation of the adepidermal granules of Salmo irideus, Lebistis reticulatus and Hynobius tokyoensis was carried out using enzyme digestion methods on epoxy-embedded sections. The granules of S. irideus larvae were decomposed by periodic acid, and digested by lipase without periodic acid pretreatmenetection of the granules. The secondary postosmificated granules were digested by lipase as in S; irideus, but complete decomposition by periodic acid was not observed in this experiment; Both periodic acid and lipase changed the shape of the adepidermal granules of H. tokyoensis to suggest partial digestion, but it appeared that these granules show rather stronger resistance to periodic acid and lipase than those of S. irideus. The granules of H. tokyoensis were completely digested when the sections were treated sequentially with phospholipase C, neuraminidase and lipase.


Inhibition studies of alkaline phosphatase in hard tissue-forming cells. The effects of the alkaline phosphatase inhibitors levamisole and R 8231 on p-nitro-phenylphosphatase, inorganic pyrophosphatase and adenosine triphosphatase (ATPase) activities in dentingenically active odontoblasts were studied. The p-nitrophenylphosphatase and inorganic pyrophosphatase activities were inhibited, while 40% of the ATP-splitting enzyme activity remained under the assay condition used. This finding, togeather with earlier studies, indicates that at least two different phosphatase are active at alkaline pH in hard tissue-forming cells; on nonspecific alkaline phosphatase and one specific ATPase. The ATPase activity is uninfluenced by ouabain and ruthenium red and is activated by Ca-2+ ions.


Immunization interview survey in the Mongolian People's Republic. An immunization interview survey was carried out in the urban and rural areas of Mongolia. The population sample interrogated comprised 1359 households with 6994 household members, 3634 of them younger than 15 years. The data collection unit was a private household, where information on administrative, demographic and socio-economic items was obtained in addition to detailed information on vaccination histories. A fair number of individuals with a positive history was found among children aged 1 to 5 years; the highest rates were determined among preschool and school-children aged 5 to 9. Significant differences in vaccination rates were found between urban and rural dwellers, urban areas displaying a higher proportion of vaccinated children in the younger age groups and rural areas among school-age children. A significantly higher proportion of vaccinated individuals was found among children living in somons than those living in aimac centres. No significant differences were found between children attending child collective institutions and those educated solely at home. The immunization interview survey was specially directed toward obtaining an estimate of the vaccinated population segment with reference to age in demarcated areas and thus detecting "weak spots" requiring remedial action.


Study of Pseudomonas haemolysin. II. Characteristics of Pseudomonas haemolysin. Haemolysins of 4 strains of Pseudomonas aeruginosa producing different fractions [II, 2 + II, I + II + III] were studied. The production of specific antibodies was demonstrated making possible the immunoelectrophoretic record of the whole lysin and of the individual fractions. Tests on laboratory animals [mice, rabbits] showed a high degree lethal and dermonecrotic property of the lysin containing the fractions II + III and I + II + III.


Reproduction of vaccinia virus (strain neurolapina) in rabbit blood leucocytes in vitro. The determining factor in vaccinia virus reproduction is the initial pattern and subsequent development of the given cell populations, in relation to the infection multiplicity, interferon production and other factors. The sensitive cells in fresh suspensions may be the blood monocytes and the small lymphocytes are virus-resistant. In cultures, transformed lymphocytes (blast cells) and macrophages play the major role in virus reproduction. The macrophages are probably of greater significance for the production of infectious virus. Replication of virus antigen was initiated in the polymorphonuclear cells in exceptional cases only, and only in the presence of a larger infection dose.


Immunochemical and chemical studies on streptococcal group-specific carbohydrates. Structural studies on the carbohydrates of Groups A, C,  and A-variant (AV) streptococci have utilized periodate oxidation, permethylation analysis, and immunochemical comparison of intact and periodate-oxidized polysaccharides. The data indicate that a similar 1,2- and 1,3-linked rhamnose chain is present in both the A and AV carbohydrates. The group A carbohydrate contains in addition N-acetylglucosamine residues at nonreducing terminals, whereas the AV is a homopolymer of rhamnose. There is some evidence that Group Ccarbohydrate contains the same rhamnose chain, but structural comparisons to the A and AV carbohydrates are complicated by the presence of intrachain N-acetylgalactosamine residues. Periodate oxidation and permethylation analysis show that while approximately 50% of the N-acetylgalactosamine of the Group C carbohydrate occupies terminal positions, the remainder is present as 1,3-linked units. Removal of the nonreducing terminal hexosamine units from the Group A carbohydrate by periodate treatment significantly enhanced its cross-reactivity with AV antiserum, whereas no enhancement was observed after similar treatment of the Group C carbohydrate. The data indicate the presence of an alpha-1,3-linked N-acetylgalactosamine disaccharide at the nonreducing terminal of the Group C carbohydrate.


Control of proliferation in mitogen responsive human peripheral blood lymphocytes: restimulation of cells stimulated by sodium periodate. Human peripheral blood lymphocytes are stimulated to a greater extent by sodium periodate when cells are incubated in medium containing human serum than when incubated in medium with fetal calf serum. NaIO4 STIMULATION CAN BE REVERSED BY TREATMENT WITH SODIUM BOROHYDRIDE BUT CELLS ALREADY COMMITTED TO DIVISION ARE NOT AFFECTED BY BOROHYDRATE TREATMENT. Maximal commitment to DNA synthesis of a NaIO4 oxidized cell suspens-on occurs after about 28 hr of incubation in medium. The committal time after periodate stimulation is identical to that after stimulation with concanavalin A. Cells treated with periodate and then reduced with borohydride immediately after oxidation are refractory to further per-odate stimulation. Cells stimulated with periodate and then incubated for 6 hr before treatment with borohydride can be restimulated with periodate, indicating a turnover of membrane sites in the 6 hr period. Periodate-stimulated cells divide only once in response to the stimulation. The progeny of cells which were stimulated with periodate can be restimulated by treatment with either periodate or concanavalin A.


The isolation and partial characterization of neutrophil chemotactic factors from Escherichia coli. Heat-stable, chemotactically active peptides have been obtained from Escherichia coli culture filtrates. They range in size between 150 and 1500 daltons and are anionic at neutral pH. Free carboxyl groups but not free amino groups appear to be required for activity. The N-terminal group may be blocked. There do not appear to be internal aromatic or basic residues in the chemotactically active fractions. A highly purified, not completely characterized, fraction was found to contain aspartic acid, serine, glutamic acid, alanine, and glycine.


Specific immunity and nonspecific resistance to infection: listeria, protozoa, and viruses in mice and hamsters,. Specific immunity developed by mice against protozoan (Toxoplasma gondii and Besnoitia jellisoni) and bacterial (Listeria monocytogenes) infections was compared with nonspecific protection conferred by prior infections. The results indicated that homologous immunity protected mice from more than 10-5 LD50 of T. gondii or B. jellisoni, but from only 10-2 LD50 of L. monocytogenes. Heterospecific protection among these organisms was for 10-0.4 minus 10-1.2 LD50. In studies in hamsters specific immunity to protozoan (T. gondii and B. jellisoni) and viral (equine Herpesvirus type 1 and Oriboca virus) infections was compared with nonspecific protection conferred by prior infections with several heterospecific agents: T. gondii; B. jellison; equine Herpesvirus type 1; Oriboca, Ossa, vesicular stomatitis, yellow fever, and Newcastle disease viruses; L. monocytogenes; and the bacillus Calmette-Guerin strain of Mycobacterium tuberculosis. The results indicated that homologous immunity in hamsters was effective against 10-6 minus 10-7 LD50 of T. gondii, B. jellisoni, equine Herpesvirus type 1, or Oriboca virus. Prior infection with Newcastle disease virus protected (probably by interferon induction) against 10-3 LD50 of equine Herpesvirus type 1. Heterospecific protection among other agents was for less than 10 LD50. This insignificant heterospecific protection in infections in which cellular immunity plays a role suggests that both the induction phase and the expression phase are specific.


Treatment of varicella-zoster virus infections with adenine arabinoside. Twenty-three patients with complicated varicella-zoster virus infections were treated with adenine arabinoside. Of 14 patients with herpes zoster, 13 had malignancy treated with irradiation and cytotoxic agents or steroids. Although the duration of active vesicle formation in these patients ranged from two to 14 days before therapy, no new lesions appeared after the fourth day of treatment with adenine arabinoside. Zoster encephalitis developed in one patient on the third day of treatment, and severe postherpetic neuralgia was seen in three patients. Of nine treated patients with primary varicella, six improved, including five with evidence of varicella pneumonia. Two of the three patients with varicella who died were immunosuppressed and had progressive viral pneumonia with persistently high titers of virus in vesicular fluid; the third pateint was a child with Reye's syndrome. Double-blind controlled studies will be necessary to demonstrate the efficacy of adenine arabinoside in the treatment of infections with varicella-zoster virus.


Cellular events in zoster vesicles: relation to clinical course and immune parameters. The cellular responses in zoster vesicles was studied chronologically in 30 patients, some of whom were compromised hosts with disseminated disease. Cell counts were low initially but rose abuptly later. Polymorphonuclear leukocytes predominated at all times in the vesicular fluid and in the cells adherent to the vesicle base. The abrupt rise in the number of cells generally coincided with an abrupt rise in the titer of vesicular interferon; both increases preceded resolution of local infection and cessation of cutaneous dissemination in disseminated disease, but the sequence of the increases was variable. The peak interferon titer correlated with cessation of dissemination better than did peak cell counts, and the timing of the local events contrasted with appearance of complement-fixing antibody, which commonly occurred after the resolution of disease. Possible interpretations are that a few sensitized lymphocytes may initiate the defensive response, produce interferon, and/or produce chemotactic factors that augment the polymorphonuclear response. The appearance of polymorphonuclear cells may be a nonspecific inflammatory response, or these cells or the deeper mononuclear infiltrate seen in biopsies may contribute to the rise in local interferon.


Cervical cytomegalovirus excretion in pregnant and nonpregnant women: suppression in early gestation. Comparison of 659 pregnant and 202 nonpregnant women with similar demographic characteristics showed overall rates of cervical cytomegalovirus excretion that were identical (9.5% vs. 9.4%) and were surprisingly high, especially since 89% of the pregnant group possessed antibody to cytomegalovirus when admitted to the study. Prevalence of cytomegalovirus among gravidas was significantly lower during the first (1.6%) than during the third (11.3%) trimester. Thus, early pregnancy appeared to exert a suppressive effect on viral excretion that waned with advancing gestation. A similar but less significant occurrence was observed in the two groups with respect to viuria. Increasing age also appeared to suppress the virologic expression of cervical and urinary tract infection, whereas multiparity seemingly produced such an effect only in the cervix. Among both cervical and urinary excreters, a few shed virus thoughout pregnancy, and others shed virus intermittently; however, viral shedding most commonly began in late gestation and frequently continued into the postpartum period. Primary infection was not documented, and antibody status remained unchanged with the advent of viral excretion in most cases. Thus, reactivation of endogenous virus seems the most likely explanation for viral shedding in our population. Similar rates of isolation of Neisseria gonorrhoeae in excreters and nonexcreters further argue against the other major possibility, venereal reinfection.


Cellular immune responses to Herpes simplex virus type 1 in recurrent herpes labialis: in vitro blastogenesis and cytotoxicity to infected cell line. Cellular immunity to herpes simplex virus type 1 (HSV-1) in 12 volunteers with recurrent herpes labialis was evaluated by means of two microassays. In the blastogenesis assay, lymphocytes were incubated with tissue culture cells persistently infected with HSV-1. Uninfected cells were used as controls, and a blastogenic index was calculated. The mean blastogenic index (plus or minus SD) for subjects with recurrent herpes labialis was 26.9 (plus or minus 8.3); the mean blastogenic index (plus or minus SD) in control donors with antibody to HSV-1 was 13.4 (plus or minus 7.2). The difference between these values was statistically significant (t equals 4.154; P smaller than 0.001). In the cytotoxicity assay, cells of the same persistently infected line were used as target cells, and release of 51-Cr from these cells or from control cells served as the index of lymphocyte reactivity. Specific immune release attributable to HSV-1 averages 3.7% (plus or minus 1.8%) in subjects with recurrent herpes labialis, compared with 23.1% (plus or minus 9.8%) in controls (t equals 6.135; P smaller than 0.001). These data suggest a dissociation between mechanisms of cellular immunity, with enhanced lymphocyte blastogenesis but decreased cytotoxicity. Recurrent herpes labialis may thus result from subtle cellular immune deficiency involving at least one of the efferent mechanisms.


Organ culture of human aorta: prolonged survival with support of viral replication. Organ cultures were established with use of human fetal aorta obtained after death; cellular elements were preserved in the cultures for periods of up to eight weeks, with maintenance of their in vivo tissue relationships. Histologically, these cultures preserve the three coats of the aorta, including an intact endothelial layer cells. The aorta organ cultures supported the replication of echovirus 11, herpes simplex virus type 1, adenovirus 2, and coxsackievirus B5 without the production of detectable gross cytoarchitectural degenerative changes. Coxsackievirus B5 and adenovirus 2 were excreted by infected cultures for prolonged periods of up to 12 weeks. This human fetal aorta organ culture system, with its long-term viability and capacity to support viral replication, provides a useful model for in vitro study of virus-aortic vessel interactions.


Enterotoxins of Escherichia coli and vibriocholerae: tools for the molecular biologist. With the independent discovery in several different laboratories that Vibrio cholerae and heat-labile Escherichia coli enterotoxins activated an enzyme (adenylate cyclase) in small intestinal epithealial cells to cause an enhanced intestinal secretion mediated by cyclic adenosine 3',5'-monophosphate (cyclic AMP), a molecular mechanism was provided for these disease states. As agents that also elevate intracellular concentrations of cyclic AMP in virtually every mammalian tissue tested, the enterotoxins are potentially invaluable tools in investigation of the molecular sequence of events of all cyclic AMP-related cellular phenomena. Cyclic AMP has been known for some time to be the central regulator in the cellular expression of the effects of hormones and has become known more recently as an agent that controls cellular growth by corrdinately influencing several biochemical processes related to rate of cell division. Successful application of the enterotoxins as cellular probes in the areas of regulation of cell division, determination of the reaction mechanism of adenylate cyclase, and elucidation of the relationship between prostaglandin and adenylate cyclase, both in this laboratory and in those of others, is reviewed.


Tumours of the parotid region.

Tumours of the parotid region. Fifty-eight operatively treated tumours of the parotid region are presented. Of the tumours 85 per cent were benign and 15 per cent malignant. The percentage of pleomorphic adenomas was 36 per cent. During five years of observation there were no recurrences of benign tumours. Operative complications consisted of one paresis of the mandibular branch of the facial nerve and one death due to bleeding and aspiration during the first postoperative day. Frey's syndrome occurred postoperatively in 26 per cent of the cases. It is emphasized that parotid tumours should be concentrated to centres where surgeons have sufficient experience with these cases.


Studies of rat liver microsomal diglyceride acyltransferase and cholinephosphotransferase using microsomal-bound substrate: effects of high fructose intake. Radiolabeled phosphatidate and diglyceride were prepared bound to rat liver microsomes. These compounds were used as substrates in studies of diglyceride acyltransferase, cholinephosphotransferase, and CTP:phosphatidic acid cytidylyltransferase. Optimum incubation conditions for these reactions in microsomes from normal male rats are described. High fructose diets were fed to rats for 11 days; this resulted in an increased rate of neutral lipid formation from sn-glycerol-3-phosphate by liver microsomal preparations. This was attributed, in part, to a previously reported increase in liver phosphatidate phosphatase activity. The significance of this increase is supported by the finding of a fall in microsomal phosphatidate content and a doubling in microsomal diglyceride. In addition, diglyceride acyltransferase measured with microsomal-bound diglyceride was increased twofold with no equivalent change in cholinephosphotransferase activity. Such a change should result in preferential triglyceride formation from the increased microsomal diglyceride pool. CTP:phosphatidic acid cytidylytransferase activity was depressed by the high fructose diet. These combined alterations would lead to an accelerated hepatic triglyceride formation, a result found in vivo during high fructose feeding. The high fructose diet decreased slightly the total microsomal phospholipid content and markedly depressed phosphatidylethanolamine levels.


Mechanism of avian estrogen-induced hypertriglyceridemia: evidence for overproduction of triglyceride. Relying on methods other than the determination of turnover rate of triglyceride from the curve of plasma triglyceride radioactivity after administration of labeled precursor, we have confirmed that the endogenous hypertriglyceridemia induced by estrogenization of the chick is accompanied by increased production of triglyceride. Chicks estrogenized with diethylstilbestrol became grossly hypertriglyceridemic and had elevated levels of plasma free fatty acid. Within 5 min of administration of labeled palmitate, estrogenized hypertriglyceridemic birds converted approximately 10 times more plasma free fatty acid to hepatic triglyceride than did controls. In addition, 2 hr after intraperitoneal injection of [14-C]acetate or [U-14-C]glucose, the specific activity of very low density lipoprotein triglyceride (VLDL-TG) of estrogenized birds reached or exceeded that of the untreated controls, and the rapid enrichment of the vastly expanded plasma VLDL-TG pool with labeled triglyceride further indicated that increased production of triglyceride occurs with estrogenization. Furthermore, [14-C]acetate incorporation into VLDL-TG was calculated to be 1.6 and 6.6% of the injected dose in estrogenized birds compared with 0.1 and 0.2% in untreated birds. Increased production of plasma VLDL-TG was confirmed by a kinetic study of VLDL-TG metabolism, employing reinjected, endogenously prepared [14-C]triglyceride-labeled VLDL. The fractional turnover rate of VLDL-TG in estrogenized hypertriglyceridemic birds was substantially less than that in untreated controls (0.32 plus or minus 0.03 vs 0.71 plus or minus 0.03/hr), but the total turnover rate was nearly 50 times greater (244 plus or minus 52 vs. 5 plus or minus 1 mg/hr).


Lipid composition of plasma membranes from human leukemic lymphocytes. The specific activity of adenosine 5'-monophosphatase and the concentrations of cholesterol, glucosylceramide, lactosylceramide, and phospholipids were compared in the whole homogenates and in plasma membrane fractions in four preparations of human leukemic lymphocytes taken over a 1-yr period from a patient with chronic lymphocytic leukemia. There was a 69.5-fold enrichment of the specific activity of adenosine 5'-monophosphatase in plasma membrane fractions. This enzyme appeared to be the best plasma membrane marker of all compounds studied. The increase in lactosylceramide concentration in the plasma membranes was 34.4-fold. It was significantly higher than that of glucosylceramide. The enrichment of glucosylceramide in the plasma membranes was similar to that of cholesterol and total phospholipids. The pattern of individual phospholipids in the plasma membrane fraction, as compared with the whole homogenate, was characterized by a decrease in phosphatidylcholine and an increase in sphingomyelin.


Correlation of early murine leukemia virus titer and H-2 type with spontaneous leukemia in mice of the BALB/c times AKR cross: a genetic analysis. Tissue extracts from 6-wk old mice of the AKR strain (H-K) show high levels of infectious murine leukemia virus, and these mice show a near 100% incidence of spontaneous leukemia. In F1 mice of the cross, BALB/c times AKR (H-2K/H-2K), both the occurrence of virus and the incidence of spontaneous leukemia are suppressed to very low values, due largely to the presence of the FV-1b allele inherited from the BALB/c parent. Mice of the (BALB/c times AKR) F-1 times AKR backcross generation were observed for possible correlations between virus expressions at 6 wk of age, H-2 type and leukemia incidence. H-2 type showed at most a weak influence on the occurrence of infectious virus, but there was a very strong correlation between the level of virus expression and the occurrence of leukemia and a strong correlation between H-2 type and leukemia. In addition, there was a highly significant nonrandom distribution of virus-negative mice among the backcross litters, suggesting a maternal effect on virus expression.


Separation of antigen-specific lymphocytes. II. Enrichment of hapten-specific antibody-forming cell precursors. Normal spleen cells were separated in dishes coated with thin layers of DNP-gelatin or NIP-gelatin into binding and nonbinding cells and stimulated in vitro with DNP- and/or NIP-conjugated polymerized flagellin (POL). Hapten-specific unresponsiveness was induced in the binding cell population by melting the gel at 37 degrees C or in unfractionated cells by pretreatment with soluble hapten-gelatin and could be reversed by treatment with collagenase. A specific enrichment of anti-DNP and anti-NIP antibody-forming cell precursors (AFCP) could be demonstrated in the binding cell populations after treatment with collagenase in cultures with or without "feeder" cells. However, the response of small numbers of unfractionated and purified hapten-specific spleen cells was suboptimal even in the presence of mitomycin-treated or irradiated feeder cells. Optimal numbers of anti-DNP (anti-NIP) antibody-forming cells were generated by small numbers of normal or purified spleen cells in the presence of spleen cells depleted of anti-DNP (anti-NIP) AFCP. In this system the response of only 2 times 10-4 purified hapten-specific cells was higher than the response of 10-6 unfractionated cells. Purified DNP-specific cells responded only to DNP-POL but not to NIP-POL and purified NIP-specific cells responded only to NIP-POL but not to DNP-POL. The degree of enrichment of anti-DNP AFCP decreased with increasing numbers of binding cells. NIP3-gelatin layers bound four to five times less spleen cells than DNP2-gelatin layers and the enrichment of anti-NIP AFCP (about 300-fold) was three times greater than the enrichment of anti-DNP AFCP (about 100-fold). The immunological significance of hapten-gelatin binding cells which apparently failed to respond to antigen is discussed.


Neutrophil-mediated tumor cell cytotoxicity: role of the peroxidase system. A cytotoxic effect of human neutrophils on mammalian tumor cells is demonstrated. Cytotoxicity depends on the presence of intact neutrophils, phagocytosable particles, and a halide cofactor and is inhibited by azide, cyanide, and catalase. Neutrophils from patients with myeloperoxidase (MPO) deficiency or defective H1O2 production are not cytotoxic, but activity is resotred by addition of purified MPO or H2O2 respectively. The findings support a mechanism involving the phagocytosis-induced extracellular release of MPO and H2O2 and their reation with a halide cofactor to damage the target cells.


Dissociation between mitogenicity and immunogenicity of TNP-lipopolysaccharide, a T-independent antigen. Two models have been proposed to explain triggering of B cells by so-called "T-independent antigen." Feldmann and Basten (1) proposed that the interaction of multiple repeating determinants on polymeric antigens with specific Ig receptors on the B-cell surface is sufficient to provide the signals for division of these cells and differentiation to antibody-forming cells. In contrast, coutinho et al. (2, and see review, 3) have claimed that there is only one signal, a mitogenic signal, receptors acting merely as passive focusing devices to localize the antigen on specific cells where it delivers a mitogenic signal resulting in differentiation to an antibody-producing cell. This model rests primarily on the demonstration that at high concentration all T-independent antigens they have tested are mitogenic for B cells (4-6). Compatible with this hypothesis are the observations that hydrolysis of lipopolysaccharide (LPS) to remove the ester- linked fatty acids of the mitogenic lipid A component abrogates its mitogenic (7,8) activity as well as its ability, when substituted with the TNP hapten, to induce a T-independent anti- TNP response (9). However, alkali treatment of LPS, although not changing its antigenic component (8), may also modify the molecule physically or chemically which could account for loss of immunogenic properties (10). We therefore investigated other reagents which interact with LPS in a more chemically defined manner in an effort to clarify the relationship between the mitogenic and immunogenic properties of this molecule. Polymyxin B (PB) is one of a family of cyclic peptide antibiotics which are bactericidal for most gram-negative bacteria. It prevents the lethal endotoxic activity of LPS (11, 12) and changes the physical structure of LPS (13). We report here that low doses of PB added to cultures of mouse spleen cells inhibit the mitogenic activity of TNP-LPS, a T- independent antigen, and native LPS, but do not suppress the immune response to TNP-LPS. PB interacts with TNP-LPS and LPS causing a physical change in the molecule. In addition, polymyxin-treated LPS is no longer mitogenic. These results suggest a dissociation between the mitogenic and immunogenic properties of TNP-LPS.


Control by Preynaptic Correlation: a mechanism affecting information transmission from Ia fibers to motoneurons. 1. In the unanesthetized spinal cord of the cat, simultaneous intracellular recordings were made from two motoneurons belonging to the gastronemius motor nucleus. 2. Supramaximal iterative stimulation of small branches of the gastrocnemius nerve produced monosynaptic EPSPs (Ia EPSPs) of varying amplitude superimposed on a fluctuating base line. 3. In most cases the variance of the motoneuron membrane potential was increased above base-line levels with a time course approximately matching the Ia EPSP. This suggests that Ia EPSP fluctuations are greater than can be accounted for by the base-line fluctuations alone. 4. For a given series of Ia EPSPs, the smaller responses in the series had about the same decay phase as the larger EPSPs, suggesting that most of the Ia EPSP fluctuations were not due to systematic changes in postsynaptic conductances produced by ongoing activity, but rather to a presynaptic mechanism. 5. Simultaneous recording from two motoneurons showed that base-line fluctuations were positively correlated. In most cases, however, there was an additional increased correlation above base-line levels resembling the time course of the Ia EPSPs, indicating positive correlation between EPSP fluctuations which is attributed to a presynaptic mechanism. 6. Conditioning volleys to group I muscle afferents or to low-threshold cutaneous afferents reduced the variance of the Ia EPSPs and also their correlation in motoneuron pairs, often without changing the mean Ia EPSPs. 7. It is concluded that, in the unanesthetized spinal cord, in addition to the random process which governs transmitter release intrinsic to a given synaptic terminal, there is another stochastic process affecting, in a correlated manner, transmitter release in large sets of Ia synaptic terminals. Most likely, the correlation in transmitter release is achieved by membrane potential fluctuations imposed on the Ia terminal arborizations by ongoing activity of the segmental mechanism mediating primary afferent depolarization. 8. The effects of such a correlating influence on cell firing behavior have been analyzed. The results suggest that this mechanism, referred to as control by presynaptic correlation, is able to modulate the information transmitted from Ia fibers to motoneurons.


Receptive fields of cerebellar cells receiving exteroceptive input in a Gymnotid fish. Single neurons in the caudal lobe of the cerebellum of the weakly electric fish Apteronotus albifrons respond to distortions in the normal electric field produced by the animal. Moving plastic or metal objects as well as a simpler stimulus, a moving electrical dipole, produce adequate distortions of the fish's field to cause the cerebellar cells to respond. The moving dipole stimulated small enough areas of the fish's skin, as determined by the responses of single electroreceptors, to allow maps of the receptive fields of single cerebellar cells to be produced. The receptive fields seen varied widely in complexity from relatively small excitatory or inhibitory areas to larger fields containing multiple excitatory and inhibitory areas usually bordering one another. Most cells studied displayed directional responses. Usually qualitatively different responses resulted from opposite directions of movement, and less frequently units were seen in which no response resulted from movement opposite the direction which caused responses; Varying the rate of stimulus movement caused only small changes in the responses of cerebellar cells; however, motionless stimuli applied over areas of skin known to respond to moving stimuli produced weaker responses of the appropriate sign for that area. Movement seems to be an important component of the stimulus for these cells. Cells were also seen which responded to visual as well as to electroreceptive input. Responses to each of these two modalities presented above were quite different. The cells recorded from frequently displayed burst discharges similar to those produced by Purkinje cells in other lower vertebrates, and most of the cells studied are believed to be Purkinje cells. A somatotopic relationship was found between the position of the center of a receptive field on the fish's body and the position of the cell in the brain. All of the results obtained are compatible with the hypothesis that the caudal lobe of the cerebellum is processing electroreceptive information related to object detection.


Analysis of postural motoneuron activity in crayfish abdomen. I. Coordination by premotoneuron connections. The activity of the crayfish abdominal postural motoneurons and their associated neurons (the accessory neuron(s) and the MRO(1)) were examined with the aid of techniques for the analysis of simultaneously recorded spike trains. A means of reliably identifying the spikes of the individual motoneurons based on their relative axon conduction velocities is presented. The analyses show that: 1) the large, phasically active synergist motoneurons innervating muscles producing the same movement show a marked similarity in their average responses, which is independent of the input source; 2) the small, tonically active and the middle-sized, tonicphasic synergist motoneurons innervating the same muscle and similar synergist motoneurons innervating antagonistic muscles are coordinated entirely by premotoneuron connections; 3) the accessory neuron is coordinated in its activity with the phasically active flexor excitor motoneurons and the extensor inhibitor motoneuron and thereby functions as a flexor synergist; and 4) the simultaneous presentation of flexion-producing and extension-producing inputs to the postural system results in a reciprocal oscillation in flexor-extensor motoneuron output. The functional significance of these results with respect to the operation of the postural system are discussed.


Analysis of postural motoneuron activity in crayfish abdomen. II. Coordination by excitatory and inhibitory connections between motoneurons. The identified spike trains of the crayfish abdominal postural efferent neurons were recorded simultaneously from one or more segments. The efferent activity was analyzed using cross-correlation histograms, peristimulus time scatter diagrams, and specialized antidromic techniques. The analyses show that the larger, phasically active motoneurons are coordinated in their activity by cross connections made at the motoneuron level. The cross connections are both excitatory and inhibitor in nature and result in significant alterations in spike output. Further, the accessory neuron receives an inhibitory cross connection from a middle-sized extensor excitor motoneuron or motoneurons. In each case, the cross connections appear to be appropriate to the function of the postural system.


Experimental optimization of current source-density technique for anuran cerebellum. This paper represents a systematic, semirigorous attempt to optimize the technique of current source-density (CSD) analysis experimentally. We compared different spatial differentiation formulas in terms of accuracy, aliasing, and smoothing, and provide experimental and theoretical rationale for their use. Sources of error have also been investigated. Expressions were derived to enable one to estimate the relative magnitude of errors due to electrical noise, uncertainty in tip position of recording electrodes, and error in the conductivity tensor. Corresponding experiments illlustrating the validity of such estimates are also presented. Methods to determine the optimum interelectrode spacing are given, based on computations of spatial energy-density spectra in the anuran cerebellum. The application of the technique of CSD analysis developed in this, and the accompanying paper, to the vestibulocerebellar input in the toad cerebellum provided significantly better temporal and spatial resolution of neuronal events than conventional field-potential analysis. Considerations germane to the optimum application of this technique to other neural structures are also discussed.


NADH fluorescence and [K+]o changes during hippocampal electrical stimulation. 1. Short (2 s) trains of stimuli were applied to the dorsal hippocampal surface of cats, producing an increase in [K+]o and a decrease in NADH fluorescence (the latter being indicative of an increase in tissue oxygen utilization). 2. The [K+]o rose rapidly during stimulation (delta[K+]o values from 1 to 6 mM) with larger stimulus currents producing larger changes. The time course of the poststimulus decline of [K+]o was an exponential decay function, with T 1/2 values varying from 1.3 to 6.9s, and independent of the magnitude of the delta[K+]o. Consistent undershoots of [K+]o occurred following stimuli causing less than 1 mM change in [K+]o. 3. The maximum depression of fluorescence and the time integral of the fluorescence changes following each stimulus train were both highly correlated with the total increase of [K+]o occurring during the stimulus train. 4. Application of several stimulus trains in close succession resulted in more rapid potassium reuptake following the later trains and an unusually large undershoot after the last train. Concomitantly, there was a progressive decrease in the fluorescence level. 5. When afterdischarges were induced by prolonged (less than 2 s) stimulation, larger and more sustained increases in [K+]o and decreases of fluorescence were observed, and there was some indication that afterdischarges were followed by accelerated reuptake of extracellular potassium.


Vestibular nystagmus and teleost oculomotor neurons: functions of electrotonic coupling and dendritic impulse initiation. 1. Nystagmus in the horizontal plane is evoked in fish by mechanical stimulation of the ampulla of the horizontal semicircular canal or by electrical stimulation of the nerve from this canal. The movements are conjugate and the slow phase is away from the side of stimulation. 2. Medial rectus motoneurons were recorded from intracellularly, during nystagmus. During the slow phase (induced by ipsilateral stimulation), impulses arise abruptly from the base line and appear to arise at a distance from the cell body. During the fast phase (evoked by contralateral stimulation), impulses appear to arise from large PSPs that must be generated at or near the cell body. 3. In the curarized fish, stimulation of the nerve from the contralateral horizontal canal evokes spikes that arise from large EPSPs and that are blocked relatively easily by hyperpolarizing currents. Stimulation of the nerve from the ipsilateral horizontal canal evokes spikes that arise abruptly from the base line and that are much more difficult to block by hyperpolarizing currents. Little if any underlying PSP is observed when these impulses are delayed or blocked. Thus impulses evoked by stimulation of contralateral and ipsilateral side are initiated near to and far from the cell soma, respectively. 4. If impulses evoked by contralateral stimulation fail to excite the cell body due to injury, antidromic spikes are not occluded. Thus contralateral stimulation initiates impulses in the dendrites. 5. Cell bodies of neighboring motoneurons are coupled electrotonically, and graded antidromic stimulation evokes graded depolarizing potentials which result from electrotonic spread of spike activity from adjacent neurons. These depolarizing potentials are adequate to excite the cells in the presence of a background EPSP evoked by contralateral canal stimulation. In this manner coupling tends to synchronize cells during the fast phase of the nystagmus. 6. Antidromic responses of neighboring cells fail to interact with dendritic inputs to a particular cell, although indirect evidence indicates antidromic spikes invade the impulse-initiating regions in the dendrites. Thus coupling between dendrites is negligible and dendritic inputs can mediate the smoothly graded movements of the slow nystagmic phase. Coupling between somata is too weak to cause significant interaction between dendritically evoked impulses (unless the cell bodies are depolarized by EPSPs). 7. Rhythmic firing can be recorded in a single presynaptic fiber corresponding to either the slow or the fast phase of nystagmus, but not to both. Oculomotor neurons appear to be "relay cells" that, during the fast phase of the nystagmus receive a synchronized synaptic input which is initiated in a higher level command nucleus.


Effect of exercise on tissue lipids and serum lipoproteins of rats fed two levels of fat. Adult rats were fed a low fat (4%) or a high fat (40%) diet, and groups of other rats selected for good running were fed these two diets and were treadmill exercised 5 days a week for 6 weeks. The serum triglycerides were significantly lower in exercised rats fed the low fat diet, but not in those fed the high fat diet, as compared with corresponding controls. The liver lipids were vastly elevated in sedentary rats fed the high fat diet as compared with those fed the low fat diet. An interesting effect of exercise was reflected by the near normal liver lipid levels in rats fed the high fat diet and exercised. However, in these animals, the muscle concentration of cholesterol was significantly higher than that in all other groups. The very low density lipoproteins tended to be lower while the low density lipoproteins tended to be greater in exercised as compared with sedentary rats. It was suggested that the decline in very low density lipoproteins was due to decreased synthesis resulting from adaptive changes in the exercised animal. It was further postulated that the increase in low density lipoproteins was possibly due to increased synthesis necessitated by the need to enhance the catabolism and excretion of cholesterol in exercised animals.


Vitamin E deficiency in the rat. Cytoplasmic factors required for suppression of mitochondrial respiratory decline. Liver cytoplasm from either vitamin E-deficient or normal rats has been shown to suppress mitochondrial respiratory decline induced by microsomes in vitro. The present study is an attempt to identify cytoplasmic factor(s) responsible for this suppression. The effect of cytoplasm from both groups of animals could be removed by trichloroacetate precipitation, boiling, and dialysis against 0.3 M mannitol-o.1 mM Tris buffer, pH 7.4, but not by aging or extraction with isooctane. The addition of oxalacetate of alpha-ketoglutarate plus asparate to cytolplasm potentiated the suppressant effect. Dialyzed cytoplasm was not effective in depressing the respiratory decline. However, the addition of NAD, osalacetate, or alpha-ketoglutarate plus aspartate to dialyzed cytoplasm restored its ability to depress the respiratory decline. When oxalacetate or alpha-ketoglutarate plus aspartate was added to fresh cytoplasm, followed by dialysis, the effect of these compounds was not seen. These results suggest that cytoplasmic factor(s) required for suppression of the respiratory decline were enzymes of the malate shuttle and transamination, which are heat labile and trichloroacetate precipitable, plus the dialyzable metabolites that are associated with these enzymes in the generation of NAD in mitochondria.


Tall girls: a survey of 15 years of management and treatment. In the past 15 years 450 girls have been seen because of concern about tall stature: 168 of them have been treated with stilbestrol to control the growth rate. Of the latter group 87 have been followed after treatment for a sufficiently long time to be sure growth had ceased and to provide the data for this report. At the onset of treatment the following data (mean values) were recorded: chronologic age, 13.0 years; skeletal age, 13.2 years; height, 172.9 cm; estimated mature height 180.2 cm; and growth potential, 7.3 cm. At the end of treatment data of interest (mean values) were: reduction in final height, 3.5 cm; duration of therapy, 2.1 years; amount of stilbestrol given, 2.3 gm; age of last visit, 17.6 years. Side effects were minimal, though two girls developed parovarian cysts that required surgery during treatment. Not included in the series of 87, one additional girl was seen with a serous cystadenoma of the ovary, and one girl developed superficial venous thrombosis of the calf. Long-term follow-up has revealed no late complications. It is concluded that estrogen can reduce significantly the growth rate of almost all tall girls, but treatment must be carried out under very careful supervision, bearing in mind possible side effects.


Combined immunodeficiency and vaccine-related poliomyelitis in a child with cartilage-hair hypoplasia. Patients previously described with cartilage-hair hypoplasia, a distinctive form of short-limbed dwarfism, have been found to have deficient cell-mediated immunity with intact antibody-mediated immunity. The patient with cartilage-hair hypoplasia described in the present report is unusual in that she had both deficient antibody-mediated immunity and deficient cell-mediated immunity. In addition, she developed severe, vaccine-related paralytic poliomyelitis. This complication suggests that live viral vaccines should not be administered to children with short-limbed dwarfism until the form of short-limbed dwarfism is established and immunologic evaluation is performed when indicated.


Erythrocyte membrane alterations induced by Plasmodium simium infection in Saimiri sciureus: relation to Schüffner's dots. The nature of erythrocyte membrane alterations in Plasmodium simium infections was determined employing light microscopy, carbon replication and transmission electron microscopy. Light microscopy of Giemsa stained preparations shows that infected cells initially acquire a faint stippling (schuffnerization) which becomes pronouced with subsequent parasite development. Enlargement of the host cell usually accompanied stippling. Both phenomena appear to depend on host cell age since infected mature erythrocytes were neither stippled nor enlarged. Carbon replicas show numerous indentations over the outer membrane surface of most infected cells. Their distribution suggests that they account for Schuffner's granules. The surface indentations are manifest as small infundibular which open to the infected cell's surface. Cytoplasmic microvesciles in the infected cell's stroma frequently are observed adjacent or catenated to the surface infundibula. Images suggest their funsion with the surface infundibula thus adding membrane to the cell's surface and accounting for host cell enlargement.


Reduction of blue tetrazolium by corticosteroids. Pseudo-first-order rate constants were observed for the reaction of corticosteroid or corticosteroid esters with blue tetrazolium. The data indicate that the reactivity of corticosteroids is, in part, a function of their geometry in that corticosteroid reactivity toward blue tetrazolium increases with increasing planarity of the steroid molecule and that corticosteroid esters must be hydrolyzed as a necessary prerequisite to reaction with blue tetrazolium. Evidence is presented indicating that free radicals are not involved in the blue tetrazoliun reaction with corticosteroids. Certain pharmaceutically important compounds such as pyrocatechol derivatives and hydroquinone appear to reduce blue tetrazolium by the anion free radical mechanism proposed previously. A spectrophotometric method for determining the number of reduction units transferred to blue tetrazolium per molecule is described.


Cycloalkanones V: synthesis, distribution, and effects on triglyceride metabolism. The 14-C-labeled 2,8-dibenzylcyclooctanone was synthesized to study its absorption, distribution, and excretion in rats. Maximum drug absorption from the GI tract occurred between 12 and 14 hr after administration. The major organs possessed maximum amounts of the drug in 1 hr, with the liver concentrating the most with 6.56% 14-C and the muscle mass reaching a maximum of 41% 14-C after 14 hr. The drug remained in the GI tract over the first 6 hr and was associated with the lipid and glycogen fractions. Eighty-seven percent was eliminated in the feces after 72 hr. 2,8-Dibenzylcyclooctanone caused a significant reduction in vitro of dihydroxyacetone phosphatase acyltransferase and sn-glycerol-3-phosphate acyltransferase, which is the proposed mechanism for the observed in vivo reduction of hepatic, intestinal, and serum triglycerides and total glycerolipids. In vivo administration of the drug resulted in a depression of liver acid phosphatidyl phosphatase, acid phosphatase and lipase, and adipose lipase. The drug increased the rates of excretion of exogenous cholesterol, palmitic acid, and progesterone.


Effects of activity in single sensory fibres on the discharge patterns of dorsal spinocerebellar tract cells. 1. Electrical activity of dorsal spinocerebellar tract (DSCT) was recorded extracellularly and intracellularly in Clarke's column nucleus. 2.Trains of impluses were elicited in DSCT cells by static stretch of the hind limb muscles, and the distributions of intervals between the impulses were computed. These distributions were essentially similar before and after impalement of the cells with the micropipette, suggesting that cell injury caused by penetration was insignificant. 3. Patterns of synaptic activity revealed that the DSCT cells examined were innervated by a limited number of muscle afferents. 4. In some DSCT neurons, all excitatory post-synaptic potentials (e.p.s.p.s) elicited by a single fibre were suprathreshold, and e.p.s.p.s elicited by other afferents were usually subthreshold. These neurons discharge very regularly during muscle stretch, and the spike initiation followed nearly one-to-one relation in response to activity of the sensory fibre eliciting suprathreshold e.p.s.p.s. 5. The discharge pattern was irregular in other DSCT cells. In these cases, single sensory fibres evoked e.p.s.p.s. which did not always reach the firing threshold because of random fluctuation in their amplitude. 6. DSCT neurons often show low frequency discharges (about 10/sec) in the absence of sensory activation. In these conditions only small e.p.s.p.s. were detected (average, 0.53 mV) suggesting that the average membrane potential is very close (1-i mV) to the firing threshold. 7. It is concluded that the statistical properties of firing patterns in DSCT neurons activated by muscle afferents depend on (a) the large amplitude of e.p.s.p.s., (b) their firing threshold and (c) random fluctuations in the e.p.s.p.s. amplitude.


Depression and recovery of transmission at the squid giant synapse. 1. The process of synaptic depression and recovery were studied in the squid (Loligo pealii) giant synapse with intracellular recording and stimulating electrodes in the prescence of tetrodotoxin (10-minus 7 M).  2. When the synapse was stimulated at 50 Hz, depression occurred rapidly.  Recovery after the tetanus was a first-order process with an average recovery time constant of 4-9 sec.  The rate of recovery was independent of the amplitude of the post-synaptic potential (p.s.p.) or the degree of depression.  3. For the first five to seven p.s.p.s in the train there was a linear relationship between depression and the total amount of transmitter previously released.  This may indicate that depression in this preparation was caused by the depletion of the presynaptic store of transmitter (S).  4. Assuming that this interpretation was correct, we could show that recovery from depression during the tetanus (i.e. 'mobilization') proceeded about 10 times faster than after the end of the tetanus.  5. When the amplitude of the p.s.p. was varied by changing the bathing calcium concentration, [Ca], the degree of depression was correlated to the amplitude of the p.s.p.  6. When the amplitude of the p.s.p. was increased by increasing pre-synaptic depolarization, synaptic depression was found to increase as well.  However, synaptic depression increased less than the amplitude of the p.s.p., the relationship between these two measures being non-linear.  7. This finding is interpreted to indicate that the transmitter stores, S, are closely related to the area of the presynaptic membrane which is sufficiently depolarized to release transmitter.


A dual effect of repetitive stimulation on post-tetanic potentiation of transmitter release at the frog neuromuscular junction. 1. End-plate potentials (e.p.p.s) were recorded with a surface electrode from frog neuromuscular junctions blocked with high Mg and low Ca to study post-tetanic potentiation(potentiation).  2. Potentiation is found to decay exponentially over most of its time course.  3. The time constant tsu(H) characterizing this exponential decay is a function of the previous history (frequency and duration) of stimulation.  For example, tsu(H) increased from about 20 sec following a few impulses to over 70 sec following more than 1000 impulses.  4. A new method is presented to obtain estimates of the rise of potentiation (uncontaminated by facilitation or an intermediate facilitatory process) during repetitive stimulation.  It is found that potentiation is present following short trains of impulses and continually increases in magnitude with the duration of the conditioning stimulation.  Potentiation was at a maximum immediately following the conditioning trains.  5. The relationship between P(T), the magnitude of potentiation immediately following repetitive stimulation, and tsu(H), the time constant for the decay of this potentiation, is given by tsu(H)=Ae-P(T)/B, where A=19-8 plus or minus 5-1 sec (mean plus or minus S.D. of an observation) and B increases from 2.2plus or minus 2-1 to 5-7 plus or minus 2-7 as the stimulation rate increases from 5 to 30/sec.  6. The value of A in the above equation can be considered to represent the minimal time constant for the decay of potentiation--that is, the time constant for decay after a simgle impulse.  7. Evidence is presented for a afacilitatory process with a time constant of decay of about 3 sec which is intermediate in duration between facilitation and potentiation.  8. It is suggested that repetitive stimulation has a dual effect on potentiation; each impulse (1) adds an increment of potentiation and (2) increases tsu(H), the time constant for the decay of potentiation.


A quantitative description of tetanic and post-tetanic potentiation of transmitter release at the frog neuromuscular junction. 1. End-plate potential (e.p.p.s) were recorded with a surface electrode from frog neuromuscular junctions blocked with high Mg and low Ca to study post-tetanic potentiation (potentiation). 2. The magnitude of potentiation was not directly related to the number of conditioning impulses, but was a function of the frequency and duration of the conditioning stimulation. 3. Potentiation was always greater following an equal number of impulses delivered at a higher frequency of stimulation. 4. Plots of the magnitude of potentiation against the number of conditioning impulses would sometimes show an upward inflexion depending on the parameters of stimulation. 5. These experimental observations were described by a model based on the assumption (1) that potentiation is linearly related to a residual substance, R(t), which accumulates in the nerve terminal during repetitive stimulation, and (2) that each nerve impulse adds an identical increment, r, of this residual substance. The data were not described by assuming a 4th power relationship between potentiation and R(t). 6. The upward inflexion in potentiation (see paragraph 4) is described by the model as resulting from an increase in the time constant for the decay of potentiation as the magnitude of potentiation increases. 7. The increment of residual substance r added by each impulse was independent of the amount of transmitter released during the conditioning train. This increment typically increased transmitter release by amount 1% of the control level in the absence of potentiation. 8. Suggestions are given to explain why potentiation of transmitter release, which is thought to arise from an accumulation of Ca-2+ in the nerve terminal, can be described assuming a linear relationship between potentiation and R(t), the proposed substance responsible for potentiation, under experimental conditions in which a 3rd to 4th power relationship would be expected to exist between external Ca concentration and evoked transmitter release.


The electrical basis of excitation and inhibition in the rat anoccygeus muscle. 1. The transmembrane potential of smooth muscle cells of the rat anococcygeus muscle was studied with micro-electrodes.  The muscle had a mean resting membrane potential of --61-5 mV and normally lacked spontaneous mechanical and electrical activity.  2. Stimulation of intramural nerves with single pulses produced a small depolarization and contraction.  At frequencies greater than 0.5 Hz a second component occurred which had a maximum value of 35--51 mV with short trains at 30 Hz.  Spike potentials were rarely seen.  Depolarization was accompanied by a decrease in membrane resistance.  3. Noradrenaline (3 times 10-minus 5 M) and guanethidine (3 times 10-minus 5 M) both depolarized the membrane and produced contraction.  Initially, oscillations in the membrane potential were oftern seen.  4. Intramural nerve stimulation in the presence of guanethidine damped the oscillations and produced relaxation.  This was accompanied by neither hyperpolarization nor a change in membrane resistance.  5. TEA (1 mM) depolarized the membrane (--47-6 mV) and initiated spontaneous activity.  Field stimulation evoked spikes with overshoot.


Detachment of structurally intact nerve endings from chromatolytic neurones of rat superior cervical ganglion during the depression of synaptic transmission induced by post-ganglionic axotomy. 1. Electrophysiological studies showed that injury of post-ganglionic nerve fibres leads to severe and prolonged depression of synaptic transmission through the rat superior cervical ganglion, beginning within 24 h.  This is in line with the results of previous studies in other species and upon other neurones.  2. electron microscopy after post-ganglionic axotomy revealed nerve endings of presynaptic type with all the specialized membrane-related features of a synaptic zone, but which were not apposed to any post-synaptic nervous element.  These umusual profiles were interpreted as detached presynaptic nerve endings.  In normal and control ganglia, such profiles formed at most 0-5% of all vesicle-containing profiles of presynaptic type; in ganglia with all major post-ganglionic branches cut the proportion rose to approximately 7%, between 3 and 7 d post-operatively.  Over this period, the mean incidence of chromatolytic neurones was 74-6%.  3. Concomitantly, the incidence of synapses within the ganglion fell by about 75%, reaching its lowest levels between 3 and 7 d post-operatively.  There was strikingly little evidence of persistence of post-synaptic membrane specializations ('membrane thickenings') following detachment of synapses.  4. At longer survival intervals the incidence of synapses gradually increased, and that of detached nerve endings gradually decreased; recovery was well advanced by 42 d.  5. The fall in the incidence of synapses was closely paralleled by a fall in the incidence of desmosome-like attachments in the ganglion; the incidence of such attachments was found to be correlated to a significant degree with that of synapses.  6. It is concluded that most or all of the synapses upon sympathetic neurones become physically dissociated during the chromatolytic reaction of these neurones to axotomy.  The failure to persist of ultrastructurally specialized post-synaptic sites, and the loss of desmosomes (particularly marked for those involving purely post-ganglionic nervous elements) suggest that the post-ganglionic neurone is losing all its specializations for attachment.  7. Some evidence suggests that the satellite cells may effect the final separation between pre- and post-synaptic structures.


Stimulation and inhibition with LH and other hormones of female mammalian meiosis in vitro. Utilizing some known physiological preovulatory hormone levels and specific cytological end-points, experiments were carried out with two different media to obtain a clearer understanding of meiotic maturation in vitro of ewe, cow, monkey and human oocytes. In the first series of experiments, it was shown that, in McCoy's medium, ovine LH, prostaglandin E1 and ovine prolactin increased the percentage of divisions of ewe oocytes, without cAMP as a demonstrable intermediate. Monkey oocytes incubated in McCoy's medium with fetal calf serum were stimulated by ovine LH, prostaglandin E2 or cAMP. Human oocytes were stimulated to divide in Diploid medium by ovine LH, oestradiol-17 beta with LH added later, or prostaglandin E1. Diploid medium containing added sheep serum markedly inhibited the meiotic activity of ewe oocytes when ovine LH, HCG, cAMP, prostaglandin E1 and E2 of prolactin were added. This effect was studied by separating the contained fetal calf and sheep sera. It was noted that when LH, HCG, or prostaglandins E1 and E2 were added to a system utilizing fetal calf serum, only spontaneous resumption of meiosis occurred: stimulated and spontaneous activity were reduced when cAMP or prolactin were added. The inhibition of ewe oocyte maturation in Diploid medium with sheep serum and additives was also seen with cow oocytes incubated in Diploid medium containing fetal calf serum and LH, HCG, cAMP or prostaglandin E1 and E2. Cow oocytes incubated in McCoy's medium with equivalent fetal calf serum and added LH were sharply inhibited but spontaneous maturation persisted in the presence of added HCG, cAMP or bovine prolactin.


Distribution of delta-5 -3beta-hydroxysteroid dehydrogenase activity in the Graafian follicle of the sheep. The localization of delta-5 -3beta-hydroxysteroid dehydrogenase (3 beta-HSD) has been examined in ovarian follicles in vivo and in vitro, and related to oestrogen and progesterone production. In vivo, during the oestrous cycle, enzyme activity was restricted to the theca interna of the one or two most advanced follicles in each animal, but was present only between Day 2 and 5 and between Day 13 and ovulation. High levels of oestrogen were found in the ovarian venous blood only when follicles containing 3 beta-HSD were present. When sheep were injected with PMSG, the theca interna in a number ofsmall follicles acquired 3 beta-HSD activity and began to secrete oestrogen within 12 hr of the injection. The enzyme was not detected in the membrana granulosa of any follicles before ovulation but within a few hours of ovulation, 3 beta-HSD activity was present in the granulosa lutein cells. In vitro, large activated follicles exhibited 3 beta-HSD activity in the theca interna and secreted high levels of oestrogen into the culture medium. When LH was added to the medium oestrogen secretion was inhibited; within 48 hr, the follicles were secreting high levels of progesterone, and 3 beta-HSD activity was present in both the membrana granulosa and the theca interna. Dibutyryl cyclic adenosine monophosphate mimicked the effect of LH in suppressing oestrogen secretiion, but did not induce production of progesterone; the distribution of 3 beta-HSD activity infollicles treated with this nucleotide was the same as in those cultured in control medium.


Induction of blastocyst implantation in mice by cyclic AMP. Oestrogen administration to female mice with 'delayed' implantation induces changes in the uterus and blastocysts followed by implantation and normal gestation. The second messenger for oestrogen, adenosine 3' :5'-cyclic monophosphate (cAMP), was instilled into the uterine lumen of females with 'delayed implanting' blastocysts to determinewhether it also is able to induce implantation. Implantation of the blastocysts was induced by cAMP but the pregnancies were not maintained to tem. These results are discussed with respect to the changes in blastocysts induced by cAMP and with respect to the reported contraceptive effects of cAMP in normal mice.


Reproductive capacity of the mentally retarded. Ninety-seven mentally retarded females were studied to evaluate parameters of growth and development of secondary sex characteristics and the incidence of ovulation. These patients were selected on the basis of age and etiological classification. Patients were classified as Down's syndrome, brain damaged, or undifferentiated. The following parameters of growth and development were studied: anthropometric, bone age, and sexual maturation. Sexual maturation studies consisted of breast development, pubic hair, axillary hair, menarche, menstrual pattern, and ovulatory patterns. The following observations were made from this study: 1. Physical growth and secondary sexual development were retarded in all parameters. 2. The undifferentiated and brain damaged groups showed almost identical patterns of growth and development of secondary sexual characteristics. 3. The Down's syndrome group was the most retarded in physical growth and in secondary sexual development. 4. Skeletal age was a better guide than chronological age or height and body weight to the prediction of development of secondary sex characteristics and age at onset of menarche. 5. In 26 mentally retarded females who had achieved menarche the incidence of ovulation was approximately 60 percent.


Cell-envelope proteins of Bordetella pertussis. Cell-envelope polypeptides of eight phase-I and five phase-IV strains of Bordetella pertussis were compared by SDS-polyacrylamide gel electrophoresis. All phase-I strains gave a strikingly similar but complex pattern of protein bands, which did not appear to vary with known differences in heat-labile agglutinogens. Phase-IV strains gave the same pattern as phase-I strains, except that one band was missing and another was either much reduced or absent. Envelopes from phase-I strains grown in Hornibrook medium rich in Mg-2+ ions to produce "antigenically-modulated" C-mode cells gave a pattern of bands indistinguishable from phase-IV strains. A phase-IV strain grown in the high-Mg-2+ medium gave the same pattern of bands as when grown in unmodified Hornibrook medium. We suggest that the two polypeptide bands that show changes may be responsible for one or more of the immunological or physiopathological activities that are lost during phase variation and antigenic modulation in B. pertussis.


Testing for possible effects of cedar wood shavings and diet on occurrence of mammary gland tumors and hepatomas in C3H-A-vy and C3H-Avy-fB mice. C3H-A-vy and C3H-A-VY-FB mice transferred from our colony to the laboratory of Dr. J. R. Sabine in Australia had a markedly reduced incidence of both hepatomas and mammary tumors. Sabine and co-workers (J Natl Cancer Inst 50: 1237-1242, 1973) attributed the low incidence of tumors in their laboratory compared to the high incidence in this laboratory to the fact that we routinely use red cedar shavings in the bedding, whereas they use sawdust, predominantly Doublas fir. Results of tests in this laboratory showed that the difference in occurrence of tumors could not be attributed directly to either a difference in diet or bedding. There was no evidence that the cedar shavings were carcinogenic. The animals in Australia on bedding without cedar shavings were not as healthy as our animals, in that they did not grow as well and were infested with ectoparasites; apparently these factors caused the reduced occurrence of tumors.


Carcinogenicity of 5-nitrofurans and related compounds with amino-heterocyclic substituents. Carcinogenicity of eight 5-nitrofurans with heterocyclic substituents at the 2-position of the furan ring was investigated by feeding the chemicals to Sprague-Dawley female rats. N-[5-(5-nitro-2-furyl)-1,3,4-thiadiazol-2-yl]acetamide induced in 30 rats the highest incidence of tumors with the greatest number of tissues involved: forestomach squamous cell tumors (22), kidney pelvis transitional cell carcinomas (15), pulmonary alveolar cell carcinomas (16), hemangioendothelialsarcomas (20) of the intestine, mesentery, liver, lung, and pancreas, and a few tumors of other tissues. 2-Amino-5-(5-nitro-2-furyl)-1,3,4-thiadiazole, 2-amino-5-(5-nitro-2furyl)-1,3,4-oxadiazole, and trans-2-[dimethylamino)methylimino]-5-[2-(5-nitro-2-furyl)vinyl]-1,3,4-oxadiazole produced high incidences of mammary tumors. The other four 5-nitrofurans tested: N-[4-(5-NITRO-2-FURYL)-2-THIAZOLYL]ACETAMIDE;2,3,4-TRIFLUORO-N-[4-(5-NITRO-2-furyl)-2-thiazoly]acetamide;5-(5-nitro-2-furyl)-1,3,4-oxadiazol-2-ol; and N-( [3-(5-nitro-2-furyl)-1,2,4-oxadiazol-5-yl]methyl)acetamide were associated with tumor incidences of 40-60%. Two other chemicals were also tested: 2-Amino-5-nitrothiazole caused a low incidence of breast and kidney pelvis tumors, and 2-amino-4-(p-nitrophenyl)thiazole induced a high incidence of breast and salivary gland adenocarcinomas and lymphomas.


Characterization of spontaneous, chemicak, and viral transformants of a C3H/3T3-type mouse cell line by transplantation into young chick blastoderms. Cells from a C3H/3T3-type cell line were transplanted into defects of the lower layer of stage-4 chick blastoderms before and after "spontaneous," chemical, and viral transformation. To check the validity of inhibition of lower-layer defect closure as a criterion of malignancy, we compared the behavior of the lower layer toward these cells with their in vitro growth pattern, their capacity to invade embryonic chick skin explants that were organotypicaly cultured, and their tumorigenicity in syngeneic mice. No false-positive results were observed with either test. The study of the in vitro growth pattern gave false-negative results during the early phase of spontaneous transformation, whereas these cells, exhibiting an untransformed growth pattern, were shown to be malignant by the other tests. We concluded that the inhibition of lower-layer defect closure is a reliable, sensitive and rapid test for the detection of malignancy in tissue-cultured cells from any source.


Sarcoma in a hamster inoculated with BK virus, a human papovavirus. An undifferentiated sarcoma occurred in 1 of 52 hamsters inoculated when newborn with BK virus (BKV), A SIMIAN VIRUS 40 (SV40)-related human papovavirus. It was transplantable and grew in tissue culture. Sera of tumor-bearing hamsters were without antibodies reactive to BKV virion antigens in hemagglutination-inhibition and neutralizationtexts, but contained antibodies reactive in immunofluorescence (IF) tests to SV40 T antigen in SV40-TRANSFORMED CELLS AND TO ANTIGENS IN CELLS ACUTELY INFECTED WITH BKV or SV40. The IF reaction between tumor cells and sera of tumor-bearing hamsters was minimal in texts of tumor cells of an early passage but satisfactory in tests of cells from a later passage. Fusion of tumor cells with permissive (monkey and human) cells, induced by inactivated Sendai virus, did not lead to recovery of infectious BKV. Immunization of hamster with BKV or SV40 failed to protect against challenge with tumor cells. There was considerable cross-reactivity between the T antigens of BKV and SV40 viruses


Antibody to herpes simplex virus type 2-induced nonstructural proteins in women with cervical cancer and in control groups. Sera obtained from 15 patients with cervical cancer, 10 patients with breast cancer, and 15 control women, individually matched with the cervical cancer patients, were examined for antibodies to early proteins synthesized in herpes simplex virus type 2 (HSV-2)-infected cells. The method used was an indirect radioimmune precipitation test followed by polyacrylamide gel electrophoretic analysis of immune precipitates. The relative reactivity to a major early nonstructural protein (VP134) was used to compare these selected sera. The results obtained suggest that cervical cancer patients possess sera with a higher reactivity to VP134 than breast cancer patients or matched healthy women,and that serum reactivity is independent of the level of neutralizing antibodies to HSV-2.


Inhibition of embryonic cell aggregation by neoplastic cells. The effects of normal and malignant cells on the aggregation of embryonic cells in gyratory shaker cultures were compared. The addition of 1 times 10-5 simian virus 40 (SV40)-transformed BALB/3T3 (SV40-3T3) cells to 6 times 10-6 embryonic neural retina cells caused a highly significant greater reduct on (22.7 percent) in aggregate diameter than the addition of untransformed BALB/3T3 (3T3) cells. The ratio of the number of single cells to the number of aggregates was significantly higher for cultures containing SV40-3T3 cells than for the cultures containing 3T3 cells. This effect was concentration dependent in the presence of cultured Ehrlich-Lettre hyperdiploid (ELD) ascites cells; however, media from ELD cell cultures or ELD cell sonicates resulted in aggregates of greater diameter and lower ratios of single cells to aggregates. This approach may provide  a sensitive assay system for the interactions of tumor and other cells in vitro.


Microvesicles and vesicles of multivesicular bodies versus "virus-like" particles. Pellets obtained by the centrifugation of unfiltered or filtered fetal bovine serum (FBS) contained, among other things, numerous elongate and spherical microvesicles, ranging from 30 to 60 nm. They possessed a trilaminar (unit membrane) envelope and a moderately electron-dense core. Vesicles of multivesicular bodies and microvesicles associated with secretory epithelial cells and human lymphoblastoid cells in suspension culture possessed the same structure and were in the same size range as the serum microvesicles. These structures were either normal cell components or they represented the breakdown products of normal cell components and should not be confused with viruses. Though microvesicles of FBS may have been involved in the tissue culture material, at least some of the microvesicles associated with, and phagocytosed by, human lymphoblastoid cells in suspension and those in the lumen of acini of the baboon prostate were derived from the breakdown of the cristae of mitochondria in degenerating cells.


Mammary tumor virus particles in the submaxillary gland, seminal vesicle, and nonmammary tumors of wild mice. Type-B mammary tumor virus particles were detected by electron microscopy in the submaxillary glands of 6 of 27 freshly trapped, pregnant wild mice (Mus musculus). Type-B particles were also detected in 3 9f 24 seminal vesicles and 2 pulmonary adenomas from wild mice. Intracytoplasmic type-A virus particles were found in 7 spontaneous nonmammary tumors (lymphoma, hepatoma, lung adenoma) of aging wild mice. Type-C virus particles were also detected in many of these tissues.


Immunization with a lipid-conjugated membrane antigen to suppress growth of a fibrosarcoma induced by simian virus 40. The covalent conjugation of fatty acid to a tumor cell membrane preparation transformed it from an antigen that enhanced tumor growth to one that suppressed it. A crude cell membrane preparation was made by sequential hypertonic and hypotonic salt extraction of tumor cells from a fibrosarcoma induced in hamsters by simian virus 40. The membranes were chemically conjugated with dodecanoic anhydride in 0.5 M carbonate buffer (pH 9.0). Injection of unmodified membranes 10 days before transplantation of live tumor cells produced clear-cut enhancement of the tumor growth rate. In contrast, injection of lipid-conjugated membranes in a similar dose and protocol suppressed tumor growth. The lymphoid proliferative reactions to the tumor cells as demonstrated by the histology of both the tumor and regional lymph nodes were consistent with the hypothesis that unmodified membranes stimulated the production of antibody which participated in the enhancement of tumor growth, and that lipid-conjugated membranes stimulated the production of cell-mediated immunity which suppressed this growth.


Inhalation studies of nickel sulfide in pulmonary carcinogenesis of rats. Two hundred twenty-six specific pathogenfree male and female F344 rats were exposed to nickel sulfide inhalations for 78 weeks (5 days/wk, 6 hr/day) and observed for an adiditional 30-week period. For the same amount of time, 214 rats were exposed to filtered room air and served as controls. Rats exposed to nickel sulfide showed a significantly higher incidence of pulmonary hyperplastic and neoplastic lesions originating from the bronchial and bronchiloo-alveloar segments. The overall incidence of lung tumors in the animals treated with nickel sulfide was 14 percent compared with 1 percent in the controls. Pulmonary inflammatory reactions were also greatly increased. Injection of an agent (hexachlorotetra-fluorobutane) that induced lung infarction did not increase the proportion of animals having lesions, nor did it alter the type of lesions found in animals exposed to nickel sulfide.


Vertical transmission of C-type viruses: their presence in baboon follicular oocytes and tubal ova. C-type viruses were found in baboon follicular oocytes and tubal ova adjacent to the plasma membrane in the perivitelline space or along the inner margin of the zona pellucida. Their presence support the concept of vertical transmission of C-type viruses.


Modification of diethylnitrosamine liver carcinogenesis with phenobarbital but not with immunosuppression. Administration of 40 ppm diethylnitrosamine (DENA) in the drinking water for 10 weeks  to male Fischer rats led to hepatocellular carcinoma in 100 percent with metastasis to the lung in 40 percent, of the animals living for the full experimental period of 20 weeks. Concurrent feeding of phenobarbital and DENA for 10 weeks produced cancer of the liver in 77 percent of the animals, but only 9 percent had metastases in the lung. A brief regimen of DENA for 4 weeks, followed by 16 weeks of observation, induced cancer of the liver in only 13 percent of the rats. Administration of phenobarbital, begun 1 week after cessation of DENA intake and terminated at week 20, led to liver cancer in 64 percent of the rodents. Hydroxyurea had no effect on this enhancement. Treatment with a purified gamma fraction of antilymphocytic serum after the DENA did not influence the outcome. Thus phenobarbital given together with DENA reduced the severity of the carcinogenic process, but when it was given after the hepatocarcinogen, it increased the effect.


Inhibition of bovine leukemia virus release. Sera from 3 cows with the adult form of lymphosarcoma inhibited release of leukemia virus from a cell line of fetal lamb spleen infected with bovine leukemia virus (BLV). Sera from 5 to 7 cattle experimentally infected with BLV also suppressed virus release. The inhibition of virus release was reversible. Sera from cattle with the calf form and the thymic form of lymphosarcoma and normal bovine control sera did not repress virus release.


Studies on a transplantable murine rhabdomyosarcoma. Well-differentiated rhabdomyosarcomas developed in 8 of 10 BALB/c mice inoculated with cell-free extracts of two lesions arising in bats previously inoculated with the Moloney strain of murine sarcoma virus. The tumors could be transplanted in BALB/c mice and other strains of mice compatible at the H-2 locus. A tumorigenic clonal cell culture line (R2) was established from a transplanted tumor. Ultrastructural studies of the primary and transplanted tumors and R2 cells revealed thick and thin myofilaments characteristic of myoblasts. These cells contained numerous cisternal type-A virus-like particles, but no type-C particles were found. Attempts to recover, rescue, or chemically induce transforming or nontransforming viruses from transplantable tumors and the R2 cell line were unsuccessful. The tumor cells lacked the murine type-C virus gs antigen. Hybridization data confirmed the apparent lack of viral RNA in the tumor cells.


Long-term primary culture of mouse mammary tumor cells: production of virus. Long-term primary cultures of mouse mammary tumor cells proved an excellent source of mouse mammary tumor virus (MMTV). Virus purified from these primary cultures had the same morphologic biochemical, immunologic, and biologic characteristics as MMTV. Quantitation of MMTV-protein equivalents released into the medium was measured by the radioimmunoassay for MMTV. Peak production levels were 20-40 mug MMTV protien equivalents/75-cm-2 flask/24 hours. These cultures produced MMTV  for as long as 90 days. MMTV cultivation depended on the initial cell-plating density and hormones. Maximal MMTV release was obtained at a plating density of 1 times 10-6 cells/cm-2 in the presence of insulin and hydrocortisone. Insulin alone gave basal levels of MMTV, and hydrocortisone alone increased MMTV release only three-fold, but insulin and hydrocortisone together effected an eightfold increase in MMTV release. This suggested that hydrocortisone had a primary effect on MMTV release and insulin acted synergistically with hydrocortisone to maximize MMTV release.


Immunologic aspects of 1, 2-dimethylhydrazine-induced colon tumors in rats. Investigation of immunologic aspects of colon tumors induced in rats by 1,2-dimethylhydrazine demonstrated that: 1) An antigen in an extract of colon tumors was not detected in the normal colon. It was related to antigens in rat fetuses and did not cross-react immunologically with carcinoembryonic antigen. 2) Mucinous adenocarcinomas of the colon were significantly associated spatially with lymphoid follicles.