Guanosine 3':5'-monophosphate-dependent protein kinase from bovine lung. Subunit structure and characterization of the purified enzyme. cGMP-dependent protein kinase from bovine lung has been purified to homogeneity using 8-(2-aminoethyl)-amino adenosine 3':5'-monophosphate/Sepharose. Conditions for adsorption of holoenzyme to the affinity chromatography media followed by competitive ligand elution with cGMP have been determined. The holoenzyme of 150,000 molecular weight is composed of two 74,000 molecular weight subunits which are linked in part by disulfide bridges. Two moles of cGMP are bound per mol of holoenzyme compatible with 1 mol of cGMP/monomer. Dissociation of subunits does not occur upon cGMP binding and protein kinase activation. cGMP-dependent protein kinase has an isoelectric point of 5.4 and a Stokes radius of 50 A. The enzyme is asymmetric with an f/f0 of 1.42 and an axial ratio of 7.4. Determination of enzyme activity at varying concentrations of ATP revealed that cGMP increased the Vmax for ATP without significant effect on the Km. The purified enzyme was maximally active at 5 mM Mg2+; other divalent cations could not substitute for Mg2+. In the presence of Mg2+, strong inhibitory effects of other cations were observed with Mn2+, greater than Zn2+, greater than Co2+ greater than Ca2+. Although maximal cGMP-dependence was observed at pH 5.7 to 7.0, basal activity rose at higher pH values to approach activity observed with cGMP. A molecular model comparing cGMP-dependent protein kinase with cAMP-dependnet protein kinase is presented.


Identification, characterization, and quantitative measurement of cyclic AMP receptor proteins in cytosol of various tissues using a photoaffinity ligand. Two protein bands, present in cytosol fractions from each of seven rat tissues examined, specifically incorporated 32P-labeled 8-azidoadenosine 3':5'-monophosphate (8-N3-[32P]cAMP), a photoaffinity label for cAMP-binding sites. These proteins had apparent molecular weights of 47,000 and 54,000 on a sodium dodecyl sulfate-polyacrylamide gel electrophoresis system. These two proteins were characterized in three of the tissues, namely, heart, uterus, and liver, by the total amount of 8-N3-[32P]cAMP incorporation, by the dissociation constant (Kd) for 8-N3-[32P]cAMP, and by the nucleotide specific inhibition of 8-N3-[32P]cAMP incorporation. Several lines of evidence were obtained that the protein with an apparent molecular weight of 47,000 represents the regulatory subunit of a type I cAMP-dependent protein kinase, while the protein with an apparent molecular weight of 54,000 represents the regulatory subunit of a type II cAMP-dependent protein kinase. Almost all of the cAMP receptor protein found in the cytosol of these tissues, as measured by 8-N3-[32P]cAMP incorporation, was associated with these two protein kinases, in agreement with the idea that most effects of cAMP are mediated through protein kinases. The photoaffinity labeling with 8-N3-[32P]cAMP can be used to estimate quantitatively the amounts of regulatory subunit of type I and type II cAMP-dependent protein kinases in various tissues.


Altered absorption spectra of iso-1-cytochromes c from mutants of yeast. Low temperature (-190 degrees) spectrophotometric recordings were made of mutant strains of the yeast Saccharomyces cerevisiae containing various altered sequences of iso-1-cytochromes c. All mutants with replacements of the tryptophan 64 residue had abnormal Calpha-bands, in which the alpha2-peaks were accentuated to various degrees by being more separated from the major alpha1-peaks and by making up a larger portion of the total Calpha-peak. The altered iso-1-cytochromes c included those having the normal tryptophan 64 replaced by phenylalanine, leucine, tyrosine, cysteine, serine, or glycine as well as those having replacements at position 64 and additional replacements at other sites. Tryptophan 64 in iso-1-cytochrome c, which corresponds to tryptophan 59 in vertebrate cytochromes c, appears to be an important residue for preserving the electronic environment of the heme group. It is uncertain, however, whether altered spectra are due specifically to the abnormal residues at position 64 or due to distorted tertiary structures caused by the replacements.


Regulation of lipogenesis in isolated hepatocytes by triglyceride-rich lipoproteins. Very low density lipoproteins, chylomicrons, and remnants caused, within an hour, significant inhibition of fatty acid synthesis but not cholesterol synthesis in hepatocytes isolated from meal-fed rats. In contrast, low density lipoproteins, high density lipoproteins, and the serum fraction of density greater than 1.21 failed to significantly inhibit either fatty acid or cholesterol synthesis within 1 h. The Scatchard plots of specific binding showed that rat and human very low density lipoproteins interact with the high affinity sites on the hepatocytes with the apparent dissociation constants of 64 and 106 nM, respectively. These data also indicated that each hepatocyte was capable of binding 6 X 10(5) molecules of very low density lipoproteins.


Study of autophosphorylation of isoenzymes of cyclic AMP-dependent protein kinases. Type I and type II cyclic AMP-dependent protein kinases, present in the cytosol from each of five rat and two bovine tissues, were separated from one another by DEAE-cellulose column chromatography in order to study their possible autophosphorylation. In each of the tissues studied, autophosphorylation of the regulatory subunit of the cyclic AMP-dependent protein kinase by the catalytic subunit could be demonstrated with the type II enzyme but not with the type I enzyme.


UDP-glucuronate carboxy-lyase in cultured chondrocytes. UDP-glucuronate carboxy-lyase has been demonstrated in chick chondrocytes in tissue culture. It occurs in the particulate fraction, and its activity is stimulated by exogenous NAD. The enzyme is allosterically activated by UDP-glucuronate and inhibited by UDP-xylose, n Values of 2.8 indicate positive cooperativity of at least three interacting sites on the enzyme. These data suggest that UDP-xylose concentration in chondrocytes is regulated by substrate activation and product inhibition of UDP-glucuronate carboxy-lyase. Activity levels of the enzyme during growth of the cells peak towards mid-log phase and decline thereafter, closely paralleling levels of chondroitin sulfate glycosyltransferases determined previously (Schwartz, N. B. (1976) J. Biol. Chem. 251, 3346-3351). Thus, it appears that during chondrocyte development a common mechanism governs induction of glycosyltransferases and of UDP-glucuronate carboxy-lyase.


Liposome oxidation and erythrocyte lysis by enzymically generated superoxide and hydrogen peroxide. Xanthine oxidase, acting on acetaldehyde under aerobic conditions, produces a flux of O2- and H2O2 which attacks artificial liposomes and washed human erythrocytes. The liposomes were peroxidized and the erythrocytes suffered oxidation of hemoglobin followed by lysis. The oxidation of hemoglobin followed by lysis. The oxidation of hemoglobin, within the exposed erythrocytes, could be largely prevented by prior conversion to carbon monoxyhemoglobin, without preventing lysis. Hemolysis thus appeared to be a consequence of direct oxidative attack on the cell stroma. The enzyme-generated flux of O2- and of H2O2 also inactivated the xanthine oxidase. Superoxide dismutase or catalase, present in the suspending medium, protected the liposomes against peroxidation, the erythrocytes against lysis, and the xanthine oxidase against inactivation. Scavengers of O2('deltag), such as histidine or 2,5-dimethylfuran, which do not react with O2- or H2O2, also prevented peroxidation of liposomes and lysis of erythrocytes when present at low concentrations. In contrast a scavenger of OH-, such as mannitol was ineffective at low concentrations and provided significant protection only at much higher concentrations. It is proposed that O2- and H2O2 cooperated in producing OH- and O2('deltag), which were the proximate causes of lipid peroxidation and of hemolysis.


Alterations of rat hepatic cholesterogenesis by heterologous lipoproteins. Heterologous human lipoproteins were infused into rats in order to change acutely the lipoprotein pattern to a predominant kind and the effect on hepatic cholesterogenesis was subsequently observed. A 4-h intravenous infusion of human low density and very low density lipoproteins into rats produced a significant decrease in the incorporation of acetate into cholesterol in both liver slices and homogenates. An infusion of similar concentrations of human high density lipoprotein produced a significant increase in hepatic cholesterol synthesis. These infusions did not change mevalonate conversion to cholesterol in either the homogenates or slices. Concomitant with the changes in hepatic cholesterol synthesis were changes of similar magnitudes in the activity of the enzyme 3-hydroxy-3-methylglutaryl coenzyme A reductase. These alterations in hepatic cholesterol synthesis were associated with significant changes in microsomal cholesterol content. There was a significant increase in hepatic cholesterol synthesis with the infusion of apoproteins of high density lipoprotein. The apoproteins of very low density lipoprotein had no effect on hepatic cholesterogenesis. These studies indicate that circulating lipoproteins modify hepatic cholesterol synthesis and that the apoproteins of these lipoproteins may themselves be important for this action.


Immobilization of urea cycle enzymes. I. Characterization of immobilized carbamoylphosphate synthetase and ornithine carbamoyltransferase. Carbamoylphosphate synthetase (EC 2.7.2.5) and ornithine carbamoyl-transferase (EC 2.1.3.3) extracted from frog liver were successfully immobilized on CNBr-activated Sepharose 4B. The immobilized preparation had a better stability towards heat. The apparent Michaelis constant values for N-acetylglutamate, ammonia, and ATP were not significantly changed by immobilization.


Extraosseous accumulation of 99mTc phosphate tracers. Case report and review of reported causes. A patient with a signet-ring-cell carcinoma of the stomach accumulating 99mTc methylene diphosphonate is described. Other conditions reported in the literature are reviewed.


Hormonal regulation of initiation of DNA synthesis and of differentiated function in Y-1 adrenal cortical cells. ACTH, 8-Br-cAMP, and serum deprivation arrested Y-1 functional mouse adrenal tumor cells in the G1 phase of the cell cycle. Though ACTH and 8-Br-cAMP treated cells were larger with increased macromolecular synthetic rates compared to cells arrested in G1 by serum removal, a similar 8- to 10-hours lag to initiation of DNA synthesis was observed after either ACTH or 8-Br-cAMP removal or after serum addition. After the 8- to 10-hour lag period, cells entered S phase exponentially. ACTH or 8-Br-cAMP opposed serum induced DNA synthesis initiation only when added prior to S. Once commitment to DNA synthesis occurred, ACTH or 8-Br-cAMP addition did not inhibit DNA synthesis although 8-Br-cAMP induced a secondary block in G2. Though ACTH and 8-Br-cAMP inhibited serum induced initiation of DNA synthesis and did not affect serum induced cellular hypertrophy, both substances increased the steroidogenic capacity of the cell. ACTH and 8-Br-cAMP thus appear to specifically oppose the stimulatory effects of serum on initiation of DNA synthesis while inducing the differentiated function of the cell.


The structure of syncytia induced by the phytoparasitic nematode Nacobbus aberrans in tomato roots, and the possible role of plasmodesmata in their nutrition. The structure of syncytia induced within galls in tomato roots by the false root-knot nematode Nacobbus aberrans has been examined by light and electron microscopy. A syncytium develops by breakdown or individual cell walls, which allows movement of cytoplasmic contents between transformed cells. The wall breakdown takes place at pit fields, where the plasmodesmata may be protected from digestion until the surrounding wall is removed. Numerous sieve elements differentiate in the cells outside the syncytium. These sieve elements, and also plasmodesmata in pit fields, are demonstrated by fluorescence microscopy. The possibility of a symplastic pathway of solute movement from the phloem to the syncytium is suggested. A massive accumulation of starch occurs in the gall cells and syncytial cells, which may be related to the proliferation of phloem. Wall ingrowths typical of transfer cells are absent, and a comparative survey of the structure and mode of solute entry into nematode-transformed cells in which ingrowths are present or absent is presented.


Method for the separation of organochlorine residues before gas-liquid chromatographic analysis. The adsorption characteristics of alumina and silica for column chromatography have been assessed to develop a method, using a single solvent, for the separation of seventeen organochlorine residues into four eluates prior to gas-liquid chromatographic analysis. The effects on the adsorbents of thermal activation and subsequent deactivation with water, variation of column size and choice of eluents have been critically examined. The lipid capacity of the alumina and the effect of co-extracted materials from animal tissue upon the elution profile of the organochlorines have been determined.


Distribution of antibody to hepatitis A antigen in a population of commerical plasma doners. Plasma samples from 245 regular plasma donors for Plasma Alliance, Inc. (Knoxville, Tenn.) were tested for the presence of antibody to hepatitis A antigen. Antibody was detected at an immune adherence hemagglutination titer of 1:10 or greater in 37% (91 of 245) of the donors. A statistically significant difference in the frequency of anti-hepatitis A was found between the 18- to 29-year-old group (30%) and the 30- to 49-year-old group (57%). No significant differences between whites and blacks or males afemales were observed. Eighty percent of the donors were in the age range of 18 to 29 years, whereas 48% of the higher-titered plasmas (1:1,000 or greater) were in the 30 to 49 age group. By preselection of donors, it would be possible to produce an immune serum globulin with a specific anti-hepatitis A titer.


Pertussis outbreak in Austin and Travis County, Texas, 1975. An outbreak of bacteriologically proven pertussis occurred in Austin and Travis County, Texas, over a 7-month period in 1975. Eighty persons were cultured for pertussis in our laboratory. A total of 62% of specimens from 34 individuals with suspected pertussis was positive for Bordetella pertussis. Diagnosis of acute cases by both culture and fluorescent antibody was attempted, and the correlation of the methods is given. Analyses of cases by age, sex, immunization status, and antibiotic treatment prior to culture are included in this report. Two asymptomatic, culture-positive adults were found.


Elimination of false-positive cytomegalovirus immunoglobulin M-fluorescent-antibody reactions with immunoglobulin M serum fractions. The cytomegalovirus fluorescent-antibody test for immunoglobline M (IgM) antibody was found positive in seven of nine infants with congenital rubella infection, in addition to eight of eight infants with confirmed cytomegalovirus infection. When the test was repeated on IgM fractions of the same sera freed from IgG by ultracentrifugation, only negative reactions were observed in those from the rubella-infected infants, whereas IgM fractions from the cytomegalovirus-infected infants remained positive.


Micro-indirect hemagglutination test for detection of antibody against transmissible gastroenteritis virus of pigs. A micro-indirect hemagglutination (IHA) test was developed for detecting antibody against transmissible gastroenteritis (TGE) virus of pigs. TGE virus propagated in swine kidney cell cultures was highly purified and concentrated by the combination of ammonium sulfate precipitation, treatment with fluorocarbon, and sucrose density gradient centrifugation. Tanned sheep erythrocytes were sensitized with purified virus for use in the IHA test. The results of testing 104 serum samples collected from pigs in the field indicated that the IHA antibody titers were approximately five times higher than those obtained by a serum neutralization test and that there was good correlation between the antibody titers determined by the two tests. High IHA antibody titers developed in pigs experimentally exposed to virulent TGE virus. Sensitized sheep erythrocytes were stable under long-term storage at 4 degrees C (at least for 50 days). The conclusions made are that the IHA test described is more sensitive than the serum neutralization test for the detection of TGE antibody and may be of value for serodiagnosis of TGE.


Concentration of L-thyroxine and L-triiodothyronine specifically bound to nuclear receptors in rat liver and kidney. Quantitative evidence favoring a major role of T3 in thyroid hormone action. To estimate the relative contribution of l-triiodothyronine (T(3)) and l-thyroxine (T(4)) to thyroidal effects, we have measured the concentration of iodothyronine bound to specific hepatic nuclear receptor sites by three different techniques: (a) specific radioimmunoassay after separation of T(3) and T(4) by preparative paper chromatography; (b) in vivo kinetic approaches as reported previously; and (c) isotopic equilibration. By these three methods, receptor concentration of T(3) and T(4) in liver was 0.51+/-0.19 (SD) and 0.08+/-0.06; 0.52+/-0.12 and 0.08+/-0.02; and 0.50+/-0.13 and 0.10+/-0.03 pmol/mg DNA, respectively. The percentage contribution of T(3) and T(4) to total receptor iodothyronine was thus 86.8+/-9.0 and 13.2+/-9.4; 86.3+/-3.5 and 13.7+/-3.5; and 83.7+/-5.6 and 16.3+/-5.6%, respectively. In kidney, specifically bound nuclear T(3) and T(4) were estimated both by isotopic equilibration and by in vivo kinetic techniques to be 0.28+/-0.11 and 0.03+/-0.01 pmol/mg DNA, respectively. Thus, T(3) constituted 89.4+/-3.2% of total receptor iodothyronine in this tissue. No other iodothyronines or analogs were bound to the nuclear sites in either tissue. Kidney and liver nuclear T(3) concentrations also were identical to values previously reported with in vivo kinetic techniques. Other studies from this laboratory have suggested that thyroid effect is related to the molar concentration of iodothyronine bound to specific nuclear sites, that the sites are similar in various tissues, and that iodothyronine in plasma is in equilibrium with nuclear T(3). If these relationships are assumed, T(3) contributes between 85 and 90% of thyroidal effects in the euthyroid rat. The remaining 10-15% of thyroidal effect appears to result from the intrinsic activity of T(4).


The mitogenic effect of the lymphocytosis promoting factor from Bordetella pertussis on human lymphocytes. The purified lymphocytosis promoting factor (LPF) from Bordetella pertussis was found to be a potent mitogen for peripheral blood lymphocytes (PBL) from normal adults as well as for cord blood lymphocytes. Proliferation occurred in autologous plasma or fetal calf serum, regardless of previous exposure to pertussis infection or immunization. Only one adult human serum, from a physician constantly working with B. pertussis, inhibited the mitogenic response to LPF and this serum was shown to contain precipitating antibody against LPF. The proliferative effect of LPF was characteristic of a "nonspecific" mitogen and not of antigen stimulation of sensitized cells.LPF, phytohemagglutinin, and concanavalin A were approximately equal in potency although variation occurred depending upon the cell donor. Experiments with lymphocyte subpopulations obtained by rosetting techniques employing sheep erythrocytes, mouse erythrocytes, and sheep erythrocytes coated with antibody and complement suggested the requirement of a multicellular system for LPF mitogencity.PBL from most patients with chronic lymphatic leukemia and lymphosarcoma cell leukemia were even less responsive to LPF than to phytohemagglutinin, whereas PBL from patients with lymphosarcoma usually responded to both mitogens. It can be inferred from the results of experiments with both normal and leukemic cells that LPF, which is a murine thymus-derived (T)-cell mitogen, is also a T-cell mitogen for human PBL. The exact cell requirement and mode of action, however, are as yet unknown.


Stimulation of surfactant production by oxytocin-induced labor in the rabbit. The respiratory distress syndrome is believed to be due to insufficient surfactant. It is known that there is a greater incidence of the respiratory distress syndrome among infants delivered by cesarean section before labor than among those delivered after labor at the same gestational age. The purpose of this study was to determine the effect of labor on the production of pulmonary surfactant. We measured the phospholipid content of lung lavage in newborn rabbits delivered by cesarean section before labor at 29, 30, and 31 (full-term) days gestation and after oxytocin-induced labor at 31 days. We also measured the activities of pulmonary cholinephosphate cytidylyltransferase and choline-phosphotransferase, enzymes involved in the de novo synthesis of phosphatidylcholine, the major component of surfactant. There was a two- to fourfold increase in the amount of lung lavage phospholipid during the first 6 h after birth. This was not dependent upon gestational age at delivery. There was a further two- to fourfold increase in the next 18 h which was, however, dependent upon gestational age. Labor increased the amount of lavage phospholipid from rabbits delivered at full term by 132%, 177%, and 50% at 3, 6, and 24 h after birth, respectively. There was a postnatal increase in the activity of cholinephosphate cytidylyltransferase. This was almost linear with time during the first 12 h, by which time essentially adult values were attained. Choline-phosphate cytidylyltransferase was not affected by labor. There was also a postnatal increase in the activity of cholinephosphotransferase but this was stimulated 86%, 59%, and 21% by labor at 0, 1, and 24 h after birth, respectively. These studies suggest that labor stimulates both the synthesis and secretion of surfactant in the immediate postnatal period and thus may be an important factor in the prevention of the respiratory distress syndrome of the newborn.


Nephrogenous cyclic adenosine monophosphate as a parathyroid function test. Nephrogenous cyclic AMP (NcAMP), total cyclic AMP excretion (UcAMP), and plasma immunoreactive parathyroid hormone (iPTH), determined with a multivalent antiserum, were prospectively measured in 55 control subjects, 57 patients with primary hyperparathyroidism (1 degrees HPT), and 10 patients with chronic hypoparathyroidism. In the group with 1 degrees HPT, NcAMP was elevated in 52 patients (91%), and similar elevations were noted in subgroups of 26 patients with mild (serum calcium </=10.7 mg/dl) or intermittent hypercalcemia, 19 patients with mild renal insufficiency (mean glomerular filtration rate, 64 ml/min), and 10 patients with moderate renal insufficiency (mean glomerular filtration rate, 43 ml/min). Plasma iPTH was increased in 41 patients (73%). The development of a parametric expression for UcAMP was found to be critically important in the clinical interpretation of results for total cAMP excretion. Because of renal impairment in a large number of patients, the absolute excretion rate of cAMP correlated poorly with the hyperparathyroid state. Expressed as a function of creatinine excretion, UcAMP was elevated in 81% of patients with 1 degrees HPT, but the nonparametric nature of the expression led to a number of interpretive difficulties. The expression of cAMP excretion as a function of glomerular filtration rate was developed on the basis of the unique features of cAMP clearance in man, and this expression, which provided elevated values in 51 (89%) of the patients with 1 degrees HPT, avoided entirely the inadequacies of alternative expressions. Results for NcAMP and UcAMP in nonazotemic and azotemic patients with hypoparathyroidism confirmed the validity of the measurements and the expressions employed.


High density lipoprotein metabolism in man. The turnover of (125)I-high density lipoprotein (HDL) was examined in a total of 14 studies in eight normal volunteers in an attempt to determine the metabolic relationship between apolipoproteins A-I (apoA-I) and A-II (apoA-II) of HDL and to define further some of the determinants of HDL metabolism. All subjects were first studied under conditions of an isocaloric balanced diet (40% fat, 40% carbohydrate). Four were then studied with an 80% carbohydrate diet, and two were studied while receiving nicotinic acid (1 g three times daily) and ingesting the same isocaloric balanced diet. The decay of autologous (125)I-HDL and the appearance of urinary radioactivity were followed for at least 2 wk in each study. ApoA-I and apoA-II were isolated by Sephadex G-200 chromatography from serial plasma samples in each study. The specific activities of these peptides were then measured directly. It was found that the decay of specific activity of apoA-I and apoA-II were parallel to one another in all studies. The mean half-life of the terminal portion of decay was 5.8 days during the studies with a balanced diet.Mathematical modeling of the decay of plasma radioactivity and appearance of urinary radioactivity was most consistent with a two-compartment model. One compartment is within the plasma and exchanges with a nonplasma component. Catabolism occurs from both of these compartments. With a balanced isocaloric diet, the mean synthetic rate for HDL protein was 8.51 mg/kg per day. HDL synthesis was not altered by the high carbohydrate diet and was only slightly decreased by nicotinic acid treatment. These perturbations had effects on HDL catabolic pathways that were reciprocal in many respects. With an 80% carbohydrate diet, the rate of catabolism from the plasma compartment rose by a mean of 39.1%; with nicotinic acid treatment, it fell by 42.2%. Changes in the rate of catabolism from the second compartment were generally opposite those in the rate of catabolism from the plasma compartment, suggesting that these two catabolic pathways may be reciprocally regulated.


Regulation of rabbit myometrial alpha adrenergic receptors by estrogen and progesterone. The effects of estrogen and progesterone on uterine alpha-adrenergic receptors were investigated by direct receptor-binding studies. Immature female rabbits were primed with estrogen by intramuscular injections for 4 days. Other rabbits were primed with progesterone by injections of estrogen for 4 days followed by injections of progesterone for 4 days. The alpha adrenergic antagonist, [3H]dihydroergocryptine, was used to directly assess the number and affinity of alpha adrenergic receptors in membranes derived from estrogen-and progesterone-primed uteri. Membranes from estrogen-primed uteri contained 257 +/- 52 fmol of [3H]dihydroergocryptine-binding sites per mg protein whereas membranes from progesterone-primed uteri contained 83 +/- 11 fmol of of binding sites per mg protein. This reduction of alpha adrenergic receptor-binding sites by progesterone was statistically significant (P less than 0.02). In contrast, no significant difference in the binding site affinity was observed between the estrogen- and progesterone-primed groups. The progesterone-induced decrease in the number of uterine alpha adrenergic receptors provides a potential explanation for the reduced alpha adrenergic contractile response to epinephrine in the progesterone-primed myometrium.


Immune complexes in congenital and natal cytomegalovirus infections of man. The occurrence of circulating immune complexes was investigated in 31 patients with cytomegalovirus infection (29 infected in utero and 2 with natal infection) and 34 uninfected controls. Anti-complementary activity above 1:20 occurred in 34% (29/86) of the sera tested from the infected group in contrast to 7.5% (3/40) in the controls (P < 0.005). When assayed by means of a lymphoblastoid cell line (Raji cell test), the reactivity in these groups was 45 (39/86) and 2.7% (1/36), respectively (P < 0.001). Correlation of results between these two complement-dependent assays occurred in 75% of samples collected from the infected group. Frequency of reactivity was higher in severe intrauterine infection and during the 1st yr of life paralleling the patterns of viral excretion and humoral immune responses. Physicochemical characterization demonstrated that reactive substances in sera were acid-dissociable and, in one sample tested, contained 7S IgG antibodies with cytomegalovirus (CMV) specificity. Circulating immune complexes were heavier (18-22S) in sick, as opposed to subclinically CMV-infected patients, in whom intermediate size complexes (12-16S) were found. In three of four symptomatic patients whose demise was due to severe congenital infection, granular deposits of immunoglobulins and C3 were detected in a pattern typical of immune complexes along the glomerular basal membrane of the glomeruli. Whether or not circulation and deposition of heavier immune complexes contributed to the adverse clinical outcome is unresolved. Because of the high incidence of both congenital and natal CMV infections, definition of the pathogenetic potentials of both heavy and intermediate size immune complexes is required to design more effective therapeutic measures.


The interaction of heparin with an apoprotein of human very low density lipoprotein. An arginine-rich apoprotein obtained from human triglyceride-rich lipoprotein was isolated on a heparin affinity column when either the aqueousor urea-soluble apoproteins were applied to the column. Of all the aqueous- or urea-soluble apoproteins, only this arginine-rich protein exhibited a binding affinity to heparin. This protein was eluted from the column at sodium chloride concentrations above 0.35 M in the absence of urea and between 0.17-0.2 M when isolated in urea. The protein has been characterized by amino acid analysis, immunoelectrophoresis, dodecyl sulfate polyacrylamide electrophoresis, isoelectric focusing, and NH(2)-terminal analysis. It has the same amino acid composition, NH(2)-terminal, and molecular weight as previously described for human arginine-rich apoprotein. The triglyceride-rich lipoproteins of fasting normal humans were eluted as two fractions when applied to the heparin affinity column. A small amount was eluted in the unbound fraction and this species contained virtually no arginine-rich apoprotein. The bulk of the triglyceride-rich lipoproteins eluted in the bound fraction and contained appreciable amounts of arginine-rich apoprotein. The bound lipoproteins had more cholesterol and cholesterol ester and less triglyceride than the unbound. The isolated arginine-rich apoprotein was derivatized with phenylglyoxal with a resulting alteration of 75% of the arginine residues. This modified apoprotein did not bind to the heparin affinity column. Similar treatment of the whole triglyceride-rich lipoprotein produced a lipoprotein that was totally eluted in the unbound fraction.


The alcoholic's perception of the ward as a predictor of aftercare attendance. The present study attempted to determine whether attendance in aftercare services could be predicted from the alcoholic's perception of the inpatient ward environment. Thirty-five Ss who completed inpatient treatment and were eligible for outpatient group therapy were followed up 3 months after discharge. One factor of the Ward Atmosphere Scale, autonomy, significantly differentiated attenders (N = 13) from nonattenders (N = 22); attenders perceived more autonomy on the ward than did nonattenders. Two additional factors, aggression and insight, also differentiated the groups, although not to a statistically significant degree. Attenders were found to perceive more encouragement by the staff to express openly their angry feelings and to share their personal selves with others than were nonattenders.


Human lung tissue and anaphylaxis. Evidence that cyclic nucleotides modulate the immunologic release of mediators through effects on microtubular assembly. The addition of specific antigen to IgE-sensitized human lung tissue causes the secretion of the mediators histamine and slow-reacting substance of anaphylaxis. The mechanisms by which increased levels of cyclic AMP suppress and increased levels of cyclic GMP enhance this secretory process were studied. Colchicine, an agent which disrupts many secretory reactions by binding to microtubules in their disassembled 6S form, was a relatively ineffective inhibitor of the antigen-induced release of mediators unless lung fragments were incubated at 4 degrees C for 60 min to induce microtubular disassembly. As colchicine appeared to inhibit the immunologic secretion of mediators from human lung tissue most effectively after microtubular disassembly, the capacity of colchicine to modulate the release reaction indicated the state of microtubular assembly; inhibition by colchicine signaled a shift to the colchicine-sensitive 6S subunits whereas failure to inhibit suggested maintenance in the colchicine-resistant polymerized state.Exogenously added 8-Bromo-cyclic GMP prevented low temperature-facilitated colchicine suppression of mediator release suggesting that increased levels of cyclic GMP stabilize polymerized microtubules. Transiently increased cyclic AMP concentrations, either exogenously added as 8-Bromo-cyclic AMP or endogenously produced by isoproterenol, promoted colchicine suppression of mediator release suggesting that microtubular disassembly was produced. Direct measurement of cyclic AMP levels revealed parallel kinetics after isoproterenol stimulation between control and colchicine-treated lung fragments. The requirement for functional microtubules in the release reaction may occur after the antigen IgE-stimulated activation of a serine esterase, energy utilization, and an intracellular calcium requirement. The mechanism by which cyclic nucleotides influence microtubular assembly is postulated to involve the degree of phosphorylation-dephosphorylation of microtubules.


Neural substrate for brain stimulation reward in the rat: cathodal and anodal strength-duration properties. The trade-off between current strength and duration of a stimulating pulse was studied for the rewarding and priming effects of brain stimulation reward (BSR). With cathodal pulses, strenght-duration functions for BSR had chronaxies of .8-3 msec. No differences were observed between the results for rewarding and priming effects. With anodal pulses. strength-duration curves were parallel to the cathodal curves at pulse durations of .1-5 msec, but at pulse durations greater than 5 msec the anodal curves showed a greater drop in required current intensity than did the cathodal curves. The parallel portion of the anodal curves was interpreted as due to anode-make excitation, and the drop at longer pulse durations was interpreted as due to anode-break excitation. Cathodal strength-duration functions for the motor effect elicited through the BSR electrodes had chronaxies of .15-.48 msec. Measurements of the latency of the muscle twitch confirmed that anode-make and anode-break excitation occurred, the latter becoming evident at pulse durations as brief as .3-.4 msec. The results provide quantitative characterization of cathodal and anodal strength-duration properties of the neural substrate for BSR and are discussed in terms of their value in guiding electrophysiological investigation of that substrate.


Effects of polyphosphates on the solubility and mineralization of HA: relevance to a rationale for anticaries activity. Preexposure of hydroxyapatite (HA) to polyphosphate reduced the near-equilibrium acid solubility of HA, the mineralization of HA, and the exchange of PO4 between medium and HA. Appreciably longer exposure times were required for maximal effects of trimetaphosphate (TMP) than of pyrophosphate (PP), tripolyphosphate (TPP), and hexametaphosphate (HMP). Calcifying solution solubilized small amounts of the HA-bound polyphosphates. This occurred to the smallest extent in the case of TMP, a fact which could have relevance for the superior anticaries effect of TMP.


Activity of arginine aminopeptidases and phosphatases in inflamed palatal mucosa in denture stomatitis: a histochemical and biochemical study. Aminopeptidase and phosphatase activity of denture stomatitis mucosa was studied histochemically and biochemically. Stomatitis clearly increased the activity of these enzymes. An enzyme resembling the chloride-activated aminopeptidase B, usually involved in inflammatory processes, was recognized in the inflamed tissue in higher concentrations than in the control tissue.


Tc-99m methylene diphosphonate versus Tc-99m pyrophosphate: biologic and clinical comparison. The biologic and imaging characteristics of Tc-99m MDP and Tc-99m PPi were compared in animals and patients using freeze-dried bone-imaging kits. Biodistribution data in rabbits showed Tc-99m MDP had slightly higher bone uptake, significantly lower blood levels, and faster urinary excretion compared with Tc-99m PPi. Duplicate studies performed on ten patients showed the following: (a) blood clearance of Tc-99m MDP was more prompt and complete, resulting in significantly lower blood levels at 4 hr; (b) urinary excretion was greater with Tc-99m MDP than with Tc-99m PPi; and (c) Tc-99m PPi showed significant red-cell labeling, whereas Tc-99m MDP did not. Image quality was generally better with Tc-99m MDP than with Tc-99 m PPi, although there was no obvious difference in diagnostic sensitivity between the two agents.


A comparison of technetium etidronate and pyrophosphate for acute myocardial infarct imaging. Etidronate and pyrophosphate, labeled with Tc-95m and Tc-99m, were studied in experimentally infarcted mongrel dogs. A distribution study was conducted 2 hr after simultaneous administration of both agents in two groups of dogs. In one group, the injection was made 15 min after release of a 2-hr coronary arterial ligation. Another group was injected 48 hr after release of the ligation. The uptakes for each radiopharmaceutical and the ratio of uptakes for each sample were computed. The data show pyrophosphate to be a superior agent for the imaging of acute myocardial infarcts because of the higher uptake by infarcted myocardium and the greater contrast between infarcted myocardium and neighboring organs.


Correlation of exposure to various respiratory pathogens with farmer's lung disease. Complement-fixing antibodies (CFA) to a panel of microorganisms commonly associated with respiratory disease were measured in a number of agricultural populations. The panel included Mycoplasma pneumoniae, influenza viruses A and B, parainfluenza virus types 1, 2, and 3, adenovirus, and respiratory syncytial virus. The agricultural populations were grouped according to a clinical history of farmer's lung disease (FLD) and the presence of antibodies to the thermophilic actinomycetes (TA). Farmers with precipitating antibody activity to one or more of the TA (groups I and II) demonstrated a greater frequency of CFA to M. pneumoniae and parainfluenza virus types 1, 2, and 3 than those groups without antibody to the TA, but the level of CFA was not higher in the positive subjects. Immunoglobulin levels were also elevated in groups I and II when compared to the control groups. Unlike IgG and IgM, IgA was elevated only in the farmers who had a clinical history of FLD (group I) but not in farmers without a clinical history. The results suggest that farmers who develop FLD are exposed to a wider variety of pathogens than are other farmers, but do not respond in an accelerated manner.


Glomus tumor of the digits.

Glomus tumor of the digits. Eight cases of glomus tumor of the digits seen during an 8-year period are reviewed. This number comprised 1.2% of all hand tumors encountered. Symptoms of cold intolerance and exquisite tenderness were common to all. The average duration of symptoms prior to diagnosis and treatment was 7 years. Five patients gave a history of either frostbite (two) or trauma (three) prior to onset of symptoms. The tumor was subungual in six of the eight patients, with a relatively even distribution among all digits. Roentgenographic changes of erosion of the distal phalanx were present in 50% of the tumors. Surgical excision was curative in all instances.


Nerve tumors of the hand and forearm.

Nerve tumors of the hand and forearm. Peripheral nerve tumors comprise less than 5% of all tumors of the hand. The most common solitary nerve tumor is the neurilemmoma, which arises from the neural sheath, is well encapsulated, minimally symptomatic, and may be surgically enucleated without producing a neurological deficit. Neurofibromas may be solitary, multiple, or associated with von Recklinghausen's disease. They are usually centrally placed with nerve fibers traversing the tumor mass making it more difficult to remove the tumor without producing permanent neurological damage. Malignant tumors include neurofibrosarcomas which often are very aggressive, requiring wide excision or amputation, and the rare neuroepitheliomas. Reported nerve tumors, intraneural in location but nonneural in origin, include fibrofatty infiltration of the median and digital nerves, intraneural lipoma, hemangioma, and ganglion cysts. These lesions may be treated by decompression or excision, depending on the nature of the tumor. Four unusual cases are described.


Malignant fibrous histiocytoma of the forearm: report of a case and review of the literature. The tissue histiocyte can give rise to a variety of lesions, either benign or malignant, and the latter often is confused with other sarcomas. The multipotential nature of the histiocyte results in the presence of at least five different cell types in these tumors, and all may be derived from a common precursor cell. The anaplasia of elements which differentiate as fibroblasts appears to correlate with survival. The tumors may be highly malignant, and the 10 year survival rate approximates 40%. Aggressive surgical management of these tumors is mandatory, with either wide en bloc resection or primary amputation of the involved extremity. Should the tumor recur locally after a wide resection and there be no detectable metastases, prompt amputation is indicated.


The dissociation of transplantable tumors. Four animal transplantable solid tumors, composed of varying morphologic architecture and intercellular specializations, were studied by light and electron microscopy. These tumors were dissociated into viable single cell populations using a combination of mechanical and enzymatic methods. The conditions necessary for optimal dissociation consisted of (a) preparation of the tumor to maximize the tissue surface area, (b) enzymatic digestion with continuous agitation and (c) additional agitation to release loosely attached cells. Other factors that influenced the dissociation were optimized and discussed.


Laser cytophotometric detection of variations in spatial distribution of concanavalin-A cell surface receptors following viral infection. Human cells in culture (H.Ep. 2) were infected with herpes simplex virus type 2 at a multiplicity of infection (m.o.i.) of 3 for 3, 18 and 36 hr. Spatial redistribution of concanavalin (Con-A) cell surface receptor sites was detected by an indirect double-antibody (peroxidase-conjugated goat anti-rabbit to rabbit anti-Con-A) immunoenzymatic staining reaction which rendered the Con-A sites visible to light microscopy and to flow laser cytophotometry. Significant spatial redistribution of cell surface Con-A receptor sites occurred within the first 3 hr after infection with herpes simplex virus type 2. The infected cells must be in the presence of Con-A for the redistribution to occur, and significant redistribution occurs only at 37 degrees C. Fixation of infected cells prior to application of Con-A prevented this spatial redistribution of Con-A receptor sites.


[Action of cyclic N6-monobutyryladenosine-3':5'-monophosphate (mb-cAMP) on cell on cell and explant cultures of the PNS and CNS in vitro]. Chick embryo trigeminal ganglia of nine days and hippocampus excised from fetal rats between the sixteenth and eighteenth day of gestation were cultivated in cell- and explantcultures in Maximow-chambers. The cultures were placed in a medium containing 2 mM, 1 mM, 0.2 mM or 0.02 mM cyclic N6-Monobutyryladenosin-3':5'-monophosphat (mb-cAMP). The cultures of the PNS, the trigeminal ganglai show after 48 h by 1 mM mb-cAMP longer nerve fibers and a increased nerve fiber index. Smaller concentration did not change significant nerve fiber growth. Cell- and explantcultures of the hippocampus show a stimulation of the nerve fiber growth and the nerve fiberindex by concentration of 0.02 mM mb-cAMP. After ten days in vitro the nerve cells degenerated by higher concentrations.


Appearanc of cytotoxic cells within the bronchus after local infection with herpes simplex virus. Cells from rabbit spleens, bronchial washings (BW) and bronchus-associated lymphoid tissues (BALT) were examined for their ability to lyse cells infected with herpes simplex virus (HSV). Specific lysis of HSV-infected cells was mediated by BW cells as early as 4 days after intratracheal infection of the rabbits with the virus whereas lysis by spleen cells and BALT cells was not detected until 7 or more days after infection. Lysis by spleen cells was initially detected 7 days after intraperitoneal injection of the virus but lysis by BW and BALT cells was not observed until 14 days after infection. Although spleen, BW, and BALT cells could lyse antibody-coated target cells, antibodies detectable by antibody-dependent cellular cytotoxicity could not be detected in bronchial washings until 7 or more days after infection. The data suggest that cells capable of direct cytotoxicity of virus-infected cells appear within the bronchus after local infection by the virus.


Thymosin-induced human serum factor increasing cyclic AMP. Thymosin has virtually no in vitro effect on cyclic AMP levels in thymocytes and seems not to react with either the beta-adrenergic receptor or with the putative human serum thymic factor (SF) receptor. However, when thymosin is injected into patients lacking SF activity, it induces the appearance of a serum factor which increases cellular cyclic AMP levels in thymocytes in vitro. This factor appears to act on a membrane site distinct from the beta-adrenergic receptors.


Macrophages are required for the dextran-sulfate induced activation of B lymphocytes. The proliferative response induced by the B cell activating ligand dextran-sulfate (DxS) requires the presence of macrophage-produced factors or the macrophage-substituting compound 2-mercaptoethanol. Thus, the mechanism for activation of mouse B splenic lymphocytes is different for DxS than for other polyclonal B cell activators such as lipopolysaccharides, which can trigger B cells directely in the absence of the helper factors. It is suggested that the disappearance of the DxS-induced response could be studied in a simple functional tests for the efficiency of macrophage depletion obtained by different techniques.


Antibody response to simian virus 40 tumor antigen in nude mice reconstituted with T cells. Athymic BALB/c nude mice (nu/nu) fail to generate circulating antibodies to simian virus 40 (SV40) tumor (T) antigen when immunized with SV40-transformed mouse cells or with T antigen positive somatic cell hybrids derived from SV40-transformed human and normal mouse parental cells. However, normal BALB/c mice readily produce antibodies to SV40 T antigen. When nude mice were reconstituted with normal syngeneic T lymphocytes from spleen or thymus source, the humoral immune responsiveness to SV40 T antigen was restored.


Antibodies to bacterial and tumor-derived antigens in sera from normal guinea pigs. Antibodies that react with radiolabeled antigens derived from guinea pig line-10 tumor cells and Mycobacterium bovis (BCG) were detected in sera from normal tumor-free strain-2 guinea pigs (NGPS). Binding by NGPS to the two antigens was inhibited by extracts of either line-10 cells or BCG. Binding by NGPS to the line-10 antigen was inhibited by a number of other bacterial extracts. NGPS was tested after absorption with a variety of cells including line-10, line-1, normal guinea pig spleen, normal adult and fetal liver cells. Results indicated that some of the antibodies in NGPS were directed to line-10-specific determinants. The specific stimulating antigen for these antibodies was not identified but because of the antigenic relationship between BCG, line-10 cells and other bacteria, antibodies to line-10-associated antigens might have been induced by exposure to environmental microorganisms.


High densitiy lipoprotein (HDL) polymorphisms in rabbit: production of antibody to rabbit allotype (R 67) in sheep. Antiserum to a rabbit HDL allotype has been produced in a sheep. Qualitative and quantitative immunological test of whole rabbit serum and purified HDL polypeptides, as well as association tests, showed that the absorbed sheep immune serum reacts with the HDL allotype referred to as R 67.


Mechanisms of disease and immunity in cholera: a review. The adenyl cyclase-activating enterotoxin of Vibrio cholerae was shown to contain two types of subunit: six smaller units (L) that are responsible for the binding to cell membrane receptors and a larger unit (H) that mediates the toxic action. The receptor was identified as the ganglioside GM1 (galactosyl-N-acetylgalactosaminyl [sialosyl] lactosyl ceramide), and the results suggested that penetration of the toxin molecule into the membrane follows the rapid binding to GM1. The relationship of these findings to the mechanism of protective immunity, which is mediated by antibodies to the enterotoxin as well as those to the cell wall lipopolysaccharide of V. cholerae, was investigated. The antitoxic antibodies were directed mainly against the L subunit and protected by preventing binding of toxin; the antibacterial antibodies probably interfered with adhesion of V. cholerae to the intestine. The finding that the immune responses to toxin and bacteria act synergistically in protection against experimental cholera indicates that an improved cholera vaccine should contain both toxoid and lipopolysaccharide as antigens. In the rabbit, either subcutaneous or enteral immunization gave rise to intestinal synthesis of specific antibodies to V. cholerae.


Pathogenesis of infection with Bordetella pertussis in hamster tracheal organ culture. Hamster tracheal organ culture was employed as a model for study of the pathogenesis of infection due to Bordetella pertussis. Infected tracheal explants were examined with light, immunofluorescence, and electrom microscopy. B. pertussis organisms preferentially attached to the ciliated cells, producing ciliostasis and marked destruction of the subcellular organelles followed by expulsion of these cells from the epithelial layer. Other nonciliated respiratory epithelial cells appeared to be unaffected. Metabolic studies on infected tracheal cultrues indicated that significant deficiencies in syntheiss of host cell protein accompanied early cytopathology. Similarities and differences in host cell and parasite interaction were noted between B. pertussis and other pathogenic agents studied in this system.


Lymphocytosis-promoting factor of Bordetella pertussis: isolation, characterization, and biological activity. The leukocytosis and lymphocytosis-promoting factor (LPF) of Bordetella pertussis has been isolated in apparently pure form. LPF is a protein essentially free of lipid and carbohydrate with an estimated molecular weight of 67,000-73,600 daltons. Purified LPF induced both histamine sensitization and refractoriness to epinephrine-induced hyperglycemia and was a murine thymus-derived (T-) cell mitogen. Adenyl cyclase activity also appeared to be associated with LPF.


Bilateral chemodectoma in the neck.

Bilateral chemodectoma in the neck. A 55-year-old man with bilateral cervical chemodectoma is reported. The differences between tumours of the carotid body and those of the glomus intravagale are described and a brief review of the investigation and management of such tumours is given.


An electron spin resonance study of cholestane spin label in aqueous mixtures of biliary lipids. The effect of cholesterol on the fluidity of the phospholipid matrix in mixed micelles derived from bile salts and lecithin has been determined by the paramagnetic probe technique. It was found that correlation times for the cholestane spin label were discontinuous functions of cholesterol content and that these discontinuities correlate with the equilibrium solubility limit for cholesterol in this quaternary system. The origin of these discontinuities is attributed to the existence of another aggregate in addition to the discshaped mixed micelle in lipid solutions supersaturated with cholesterol.


Differential characteristics of purified hepatic triglyceride lipase and lipoprotein lipase from human postheparin plasma. Evidence is presented that hepatic triglyceride lipase (H-TGL) and lipoprotein lipase (LPL), purified from human postheparin plasma, can each hydrolyze both glyceryl trioleate and palmitoyl-CoA. The average ratio of glyceryl trioleate/palmitoyl-CoA hydrolase activities, obtained with enzyme preparations from 15 human postheparin plasma samples was 1.30 (1.18-1.52) for H-TGL and 8.75 (7.45-10.25) for LPL. Albumin was identified as the serum cofactor required for the hydrolysis of palmitoyl-CoA by H-TGL. It protected this enzyme from inactivation by this substrate. In contrast, palmitoyl-CoA activated and protected LPL from denaturation by dilution and incubation at 25 degrees C. The effects of other detergents were investigated on glyceryl trioleate hydrolase activities of both enzymes. Sodium dodecyl sulfate (0.4 mM) and Trisoleate (0.4 mM), which also effectively activated and protected LPL against inactivation, had only moderate protective effect on H-TGL. Sodium dodecyl sulfate at a higher concentration (1 mM) produced little or no inhibition of LPL, while completely inactivating H-TGL. Conversely, sodium taurodeoxycholate (0.4 mM) protected and activated H-TGL, but had only moderate protective effect on LPL. Triton X-100 (0.1-0.8 mM) and egg lysolecithin (0.05-2 mM) also protected H-TGL, but not LPL. The very dissimilar effects of detergents on preparations on H-TGL and LPL may form the basis for the direct assay of each enzyme in the presence of the other.


Binding, internalization, and degradation of high density lipoprotein by cultured normal human fibroblasts. Comparative studies were made of the metabolism of plasma high density lipoprotein (HDL) and low density lipoprotein (LDL) by cultured normal human fibroblasts. On a molar basis, the surface binding of (125)I-HDL was only slightly less than that of (125)I-LDL, whereas the rates of internalization and degradation of (125)I-HDL were very low relative to those of (125)I-LDL. The relationships of internalization and degradation to binding suggested the presence of a saturable uptake mechanism for LDL functionally related to high-affinity binding. This was confirmed by the finding that the total uptake of (125)I-LDL (internalized plus degraded) at 5 micro g LDL protein/ml was 100-fold greater than that attributable to fluid or bulk pinocytosis, quantified with [(14)C]sucrose, and 10-fold greater than that attributable to the sum of fluid endocytosis and adsorptive endocytosis. In contrast, (125)I-HDL uptake could be almost completely accounted for by the uptake of medium during pinocytosis and by invagination of surface membrane (bearing bound lipoprotein) during pinocytosis. These findings imply that, at most, only a small fraction of bound HDL binds to the high-affinity LDL receptor and/or that HDL binding there is internalized very slowly. The rate of (125)I-HDL degradation by cultured fibroblasts (per unit cell mass) exceeded an estimate of the turnover rate of HDL in vivo, suggesting that peripheral tissues may contribute to HDL catabolism. In accordance with their differing rates of uptake and cholesterol content, LDL increased the cholesterol content of fibroblasts and selectively inhibited sterol biosynthesis, whereas HDL had neither effect.


A comparative study of surface binding of human low density and high density lipoproteins to human fibroblasts: regulation by sterols and susceptibility to proteolytic digestion. Binding of 125I-low density lipoprotein (LDL) and 125I-high density lipoprotein (HDL) was determined in cultured human fibroblasts from a normal subject and two subjects with homozygous familial hypercholesterolemia (HFH). Binding was assayed at 0 degree C to minimize the internalization of labeled lipoproteins. The binding of LDL and of HDL were compared following interventions reported to affect LDL binding in normal fibroblast. LDL binding to normal cells increased two to three fold 24 hours after transfer from medium containing whole fetal calf serum to medium containing lipoprotein-deficient fetal calf serum. This increase was completely blocked in the presence of cycloheximide (200 microgram/ml) or 7-ketocholesterol (2.5 microgram/ml). This increased capacity of normal fibroblasts to bind LDL could be reduced 70-80% by a subsequent 18-hour incubation with cholesterol (50 microgram/ml) or 7-ketocholesterol (2.5 microgram/ml). In contrast, no significant change in HDL binding to normal fibroblasts was observed after any of these interventions. HFH cells to show any significant change in either LDL binding or HDL binding following these interventions. These results suggest that HDL binding sites on normal fibroblasts are for the most part distinct from LDL binding sites. They also support the conclusion that LDL binding sites on HFH cells are for the most part qualitatively different from those on normal cells.


Secretion of the arginine-rich and A-I apolipoproteins by the isolated perfused rat liver. Rates of secretion of the arginine-rich and A-I apolipoproteins into perfusates of rat livers were measured by specific radioimmunoassays. Livers were perfused for 6 hr in a recirculating system in the presence or absence of 5,5'-dithionitrobenzoic acid, an inhibitor of lecithin-cholesterol acyltransferase. Arginine-rich apoprotein (ARP) was secreted at a constant or increasing hourly rate of about 40 micro g/g liver, whereas the rate of accumulation of apoprotein A-I decreased progressively from about 12 to less than 5 micro g/g liver. These rates were not affected by inhibition of lecithin-cholesterol acyltransferase. The distribution of these two apolipoproteins was also measured in ultracentrifugally separated lipoprotein fractions from perfusates and blood plasma. Apoprotein A-I was mainly in high density lipoproteins, with the remainder in proteins of density > 1.21 g/ml. The percent of apoprotein A-I in the latter fraction was lowest in plasma (5%); in perfusates it was greater when the enzyme inhibitor was present (33%) than in its absence (11%). By contrast much less ARP was in proteins of d > 1.21 g/ml in perfusates than in blood plasma. Discoidal high density lipoproteins, recovered from perfusates in which lecithin-cholesterol acyltransferase was inhibited, contained much more arginine-rich apoprotein than apoprotein A-I (ratio = 10:1). The ratio in spherical plasma HDL was 1:7 and that in perfusate high density lipoproteins obtained in the absence of enzyme inhibitor was intermediate (2:1). It is concluded that: 1) the arginine-rich apoprotein is a major apolipoprotein whereas apoprotein A-I is a minor apolipoprotein secreted by the perfused rat liver; 2) the properties of the high density lipoproteins produced in this system are remarkably similar to those found in humans with genetically determined deficiency of lecithin-cholesterol acyltransferase.


Lipid metabolism in cultured cells. XVI. Lipoprotein binding and HMG CoA reductase levels in normal and tumor virus-transformed human fibroblasts. The loss in feedback control of cholesterol biosynthesis in tumor cells was examined in tissue culture. Human fibroblasts from normal subjects, SV40 tumor virus-transformed cell lines, and homozygous familial hypercholesterolemic cells as reference, were grown in tissue culture. Experiments were conducted to relate the regulatory enzyme for cholesterol biosynthesis, HMG CoA reductase, and the membrane-located binding receptors for low density lipoproteins (LDL) that mediate feedback control in normal cells. Monolayers of virus-transformed tumor cells exhibited specific (125)I-labeled LDL binding of 152 +/- 21 ng/mg cell protein, which was essentially the same as that of normal fibroblasts (135 +/- 20 ng/mg). Binding of LDL by familial hypercholesterolemic cells used as controls was only 8 +/- 3 ng/mg under the same test conditions. Basal levels of HMG CoA reductase in tumor cells of 45.2 +/- 6.5 units/mg cell protein were about twice those of normal cells. However, in contrast to the lack of feedback control of this enzyme observed with tumors in vivo, in both the normal and the transformed cells in vitro, activity of the enzyme decreased about fourfold when serum lipids were added. These findings demonstrate that tumor cells growing in vitro contain a normal complement of the membrane-located binding receptors for low density lipoproteins and, although the basal levels are higher than normal, an effective feedback regulation of the enzyme HMG CoA reductase is retained.


Specific binding of prolactin to seminal vesicle, prostate and testicular homogenates of immature, mature and aged rats. Ovine prolactin was iodinated by the lactoperoxidase method and purified by gel filtration on Sephadex G-100. The binding ability of the labelled hormone was determined, by incubation with liver homogenate from rabbits in late pregnancy, to be 8-8% total binding/mg protein, of which 86% was specific. The fraction of 125I-labelled ovine prolactin which bound most strongly was subsequently used to study its binding to rat seminal vesicle, prostate and testicular homogenates. The total binding to the seminal vesicle homogenate taken from mature (80-day-old) rats was the highest (11-69%/mg protein), but the greatest degree of binding specificity (82-6%) was to immature (30-day-old) rat prostate. Both total and specific binding to rat testicular homogenate were consistently very low. The binding specificity was demonstrated by displacement studies: while ovine prolactin caused displacement of specific binding, human chorionic gonadotropin, rat thyrotropin and human follicle-stimulating hormone did not cause any significant displacement of bound 125I-labelled ovine prolactin. Affinity constants (Ka) and binding capacities for the seminal vesicle and prostate homogenates were determined by Scatchard analysis and the effect of age on these parameters was studied. There was no difference in Ka between the aged (220-day-old), immature and mature rat tissue homogenates; however, a significant fall in binding capacity was observed in the mature rat prostate, and a further fall in the aged rat prostate. No such change was observed in the binding capacity of the seminal vesicle, as estimated by Scatchard analysis, although total and specific binding to the mature homogenates was higher than that of the other age groups.


Inhibitory action of porcine follicular fluid upon granulosa cell luteinization in vitro: assay and influence of follicular maturation. Culture medium 199 supplemented with follicular fluid from 1-2 mm antral porcine follicles inhibited spontaneous luteinization of granulosa cells from preovulatory porcine follicles in vitro. Three characteristics of luteinization were inhibited: morphological transformation, progesterone secretion, and accumulation of cyclic AMP in response to LH. The last was inhibited more effectively by culture media containing 50% follicular fluid than by media containing 20% follicular fluid. The inhibitory actions of the follicular fluid were not altered by charcoal or petroleum ether extraction. Follicular fluid from large follicles (6-12 mm) did not exhibit any of these inhibitory actions. These observations may indicate the presence of a luteinization inhibitor in the fluid of small follicles which (1) is lost by the time the follicle reaches the preovulatory stage, or (2) is overcome by a stimulatory agent which may accumulate as the follicle grows.


Regulation of insulin and glucagon secretion from a human islet cell adenoma. The regulation of insulin biosynthesis, and insulin and glucagon secretion have been investigated in a human islet cell adenoma, by incubation of tumour fragments. Both biosynthesis and secretion of insulin were strongly stimulated by incubation of islet tumour cells in the presence of increasing glucose concentrations in the range 2-8 mmol/1. However, 20 mM-glucose or 20 mM-glucose plus isobutyl methylxanthine (IBMX), both of which provide potent secretagogues for normal B cells, failed to stimulate proinsulin biosynthesis and secretion from the tumour cells. Overall rates of secretion, expressed as a proportion of total insulin content, were up to 20-fold higher than those expected for normal pancreatic tissue. Glucagon secretion from the tumour was stimulated by low glucose concentrations; normal A cells also respond in this way under these conditions. However, no stimulation of glucagon secretion occurred in the presence of IBMX. There was therefore a major alteration in the regulation both of insulin and glucagon secretion, in that release of neither hormone was stimulated by cyclic AMP. Ultrastructural examination showed the tumour to be rather heterogeneous. A and B cells with normal storage granule content and structure were seen, as well as a rather larger number of B cells containing some granules of atypical appearance. The insulin content of the tumour (13 i.u./g wet wt) was consistent with 6-8% of the tumour cells being B cells.


Functional activity of the hypothalamo-pituitary complex in the rat after betamethasone treatment. The precise nature of the impairment of hypothalamo-pituitary-adrenocortical (HPA) activity which follows prolonged corticosteroid treatment is not yet understood. To study this problem, hypothalamic corticotrophin releasing hormone (CRH) content, pituitary ACTH content and the functional capacity of adenohypophysial tissue in vitro were measured in rats after treatment with betamethasone. The content of CRH and ACTH in the hypothalamus and pituitary gland respectively were markedly reduced. After stopping the treatment the hormone concentrations in both structures returned to normal with the rise in the hypothalamus preceding that in the pituitary gland. Adenohypophysial tissue from betamethasone-treated rats incubated with hypothalamic extracts from control animals showed a considerable reduction in its ability to synthesize and release ACTH. However, corticotrophin release was impaired in adenohypophyses removed from untreated rats and incubated with betamethasone but synthesis was not affected. The physiological significance and the possible clinical relevance of the results are discussed.


The presence of lactogen but not growth hormone binding sites in the isolated rat hepatocyte. The binding of 125I-labelled human growth hormone (hGH) and bovine growth hormone (bGH) has been studied in hepatocytes isolated from female rats by perfusion with collagenase in situ. The cells appeared to retain normal membrane function, in that amino acid ([14C]alpha-aminoisobutyric acid) transport was both saturable and temperature-dependent. Amino acid ([14C]leucine) incorporation into protein was also linear over 3 h and was inhibited by cycloheximide. Binding of 125I-labelled hGH was dependent on time, temperature, hepatocyte concentration and hGH concentration. At 22 degrees C, binding reached a steady-state after 2-5 h and had a half-life of dissociation of 2-3 h. Hormone specificity studies indicated that binding was specific for hormones with prolactin-like activity (hGH, prolactins) and not for growth hormones themselves (bGH). Scatchard analysis revealed a single class of binding site with a binding capacity of 26-74+/-3-73 fmol/10(6) cells and a binding affinity of 1-24 X 10(9)+/-0-17 X 10(9) (S.E.M.) 1/mol (n=10). There was a significant sex difference in binding (female greater than male) and binding was subject to marked regulation by oestrogens (stimulation of binding) and by androgens (inhibition). The lactogen-binding sites, therefore, were comparable in many respects to those previously reported in rat liver membranes. No distinct GH binding sites were demonstrable as shown by the lack of specific binding by 125I-labelled bGH, purified either by Sephadex chromatography or by binding to and elution from GH receptors in rabbit liver membranes. The value of receptor purification of tracer for use in hormone binding studies was indicated by a substantial lowering of non-specific binding.


Cell surface antigens of chemically induced sarcomas of the mouse. I. Murine leukemia virus-related antigens and alloantigens on cultured fibroblasts and sarcoma cells: description of a unique antigen on BALB/c Meth A sarcoma. As background for a serological definition of the unique antigens of chemically induced sarcomas, we have typed a series of fibroblast and sarcoma cell lines of BALB/c and C57BL/6 origin by cytoxicity and absorption tests for murine leukemia virus (MuLV)-related cell surface antigens and known alloantigens. 7 of the 17 cultured lines expressed the range of cell surface antigens associated with MuLV (GIX, GCSA, gp70, p30), and this was invariably associated with MuLV production. In nonproducer lines of C57BL/6 (but not BALB/c) origin, a MuLV-gp70-like molecule was found on the surface of fibroblasts and sarcoma cells. The alloantigenic phenotype of these MuLV+ and MuLV- cell lines was H-2D+, H-2K+, Thy-1.2+ or -, PC.1+ or -, Lyt-1.2-, Lyt-2.2-, Ia.7-, and TL.2-. A unique antigen was defined on the BALB/c ascites sarcoma Meth A with antisera prepared in BALB/c or (BALB/c X C57BL/6)F1 mice. Tissue culture lines derived from this tumor were MuLV-, which facilitated serological study of the antigen. Absorption analysis indicated that the antigen was restricted to Meth A; it could not be detected in normal or fetal BALB/c tissue MuLV+ or MuLV- fibroblast lines, 12 syngeneic or allogeneic sarcomas, or normal lymphoid cells from 13 different inbred mouse strains.


Cytotoxic T-cell activation by polyribonucleotides: DNA synthesis is not required. We have shown that cytotoxic T cells can be polyclonally activated by a short exposure to complexes of polyadenylic:polyuridylic acid (poly A:U). Activation is optimal at a dose of 100 microgram/ml poly A:U and occurs during a 2 h incubation period in the absence of antigen. Killing of allogeneic, but not syngeneic, target cells can be observed after 12 h in culture and peaks after 21-24 h in the absence of any nonspecific binding ligand. The observed cytotoxicity is mediated by T lymphocytes and dose not require accessory macrophages or DNA synthesis for the activation or expression of effector functions. These results suggest that few requirements exist for the activation of cytotoxic T cells.


Antigenic differences in nuclear proteins of normal liver and hepatoma. Identification of a nuclear protein present in hepatocytes but absent in hepatoma cells. A nuclear antigen was detected in the mouse liver nonhistone protein fraction by using antibodies to whole liver cells. The antigen was purified to homogeneity from perchloric acid extracts of liver tissue. It gave a single band corresponding to tool wt 21,000 in sodium dodecyl sulfate gel electrophoresis. Amino acid and carbohydrate analysis showed predominance of the acidic amino acids, lack of proline, and absence of carbohydrate. Immunofluorescence staining of liver sections confirmed the nuclear localization of the antigen. Its tissue distribution was studied by using radioimmunoassay. Of the various tissues extracted for analysis, the liver contained the highest amounts of the antigen, about 1 mug/mg of solubilized liver protein. Other tissues examined showed 2-4 percent of the amount of antigen present in the liver. Two transplantable hepatomas in C3H/HeJ and C57L/J mice, respectively, and three spontaneous C3H hepatomas showed greatly decreased levels of the antigen compared to normal liver. The amount of antigen in hepatomas varied from nondetectable to 2 percent of the amount of antigen found in the livers of the mice. The antigen was also found in the blood. The antigen was found in high concentrations (up to 13 mg/ml) in the urine of normal mice. This suggests identity with the previously known mouse urinary protein (MUP). In addition to the extremely high urinary output, the properties found to be shared by MUP and the nuclear antigen included similar serum concentrations (2-60 mug/ml), a sex difference with lower values in females, same molecular size as determined by gel filtration, and immunological identity. The nuclear localization of MUP and its disappearance from hepatomas suggest that it may have an important regulatory function.


Genetic control of radiation leukemia virus-induced tumorigenesis. I. Role of the major murine histocompatibility complex, H-2. Resistance to radiation leukemia virus-induced leukemogenesis is associated with the H-2D region of the H-2 complex, or with closely linked loci. The H-2Dd allele confers resistance ot the disease, while the H-2D-Q and H-2Ds alleles are associated with susceptibility. It is not clear whether Ir genes, or an alternative mechanism are responsible for the observed H-2-linked resistance to the disease.


Genetic control of radiation leukemia virus-induced tumorigenesis II. Influence of Srlv-1, a locus not linked to H-2. A single locus, tentatively denoted Srlv-1 (susceptibility to radiation leukemia virus [RadLV]-1), confers dominant susceptibility to RadLV-induced leukemogenesis. Srlv-1 is not linked to H-2, and appears to be distinct from Fv-1 and Fv-2. Preliminary data suggest that Srlv-1 affected virus proliferation. A striking feature of this system is that Srlv-1 overrides the protection afforded by the H2D-associated dominant resistance to RadLV-induced neoplasia.


Resistance genes to murine leukemia in the I immune response gene region of the H-2 complex. A resistance locus to leukemogenesis in mice by A-RadLV (a variant of the radiation leukemia virus) is described. This locus, Rrv-1, was mapped to subregions I-A, I-B, and I-J of the H-2 complex. It is suggested that Rrv-1 may be in complementation with a second locus to the right of it, between Rrv-1 and H-2D. This localization and the complementation of the two loci for resistance are characteristics similar to Ir genes, and indicate a possible relationship between the genetic regulation of immune responsiveness and susceptibility to leukemia.


Exclusive involvement of H-2Db or H-2Kd product in the interaction between T-killer lymphocytes and syngeneic H-2b or H-2d viral lymphomas. It was demonstrated previously that the cytolysis of murine viral lymphoma cells by anti-murine sarcoma virus (MSV) syngeneic T-killer lymphocytes was restricted by some products of the H-2 complex. The respective role of the products of different regions of the H-2 complex were studied with six H-2(b) and three H-2(d) lymphomas induced by five different type C viruses. They were tested in a classical chromium release test against anti-MSV T-killer cells obtained from different inbred strains of mice, including several H-2 recombinants. Tumors o pound the H-2(b) haplotype were lysed only when effectors and target cells have in common the D(b) region. On the contrary an identity limited to the K end of the H-2 complex is necessary and sufficient in the H-2(d) haplotype. An in vitro restimulation of the spleen cells with concanavalin A strongly increased the activity of in vivo-primed T lymphocytes but did not provide any response for in vivo-primed but nonresponder cells. Preincubation of the tumor cells with anti-H-2 sera abolished the lysis by syngeneic anti-MSV effector lymphocytes. The same results were obtained by preincubating the H-2(b) targets with anti-H-2D(b), or the H-2(d) target with anti-H-2K(d). Preincubation with anti-H-2K(b) or anti- H-2D(d) were ineffective. These results show that the T-killer/target cells interaction in the MSV system involved some products of the H-2 complex which might be different with the various H-2 haplotypes and could possibly vary according to the antigenic specificity. A specific association of a viral product with a normal cellular structure, directed by the H-2 region during the viral budding could explain the observed results.


[Differentiation of VLDL subfraction in the diagnosis of type III hyperlipoproteinaemias (author's transl)]. Type III hyperlipoproteinaemias are characterized by "floating" beta-lipoproteins. In lipid electrophoresis these are present as a VLDL subfraction (beta-VLDL). They can also be detected chemically on the basis of the cholesterol/triglyceride ratio of the serum or the corresponding VLDL serum fraction. In the present work, the diagnostic value of these ratios is compared with that of E III peptide loss from VLDL-apolipoproteins, which is thought to be specific for type III hyperlipoproteinaemias. Of 42 persons with a VLDL subfraction, seven showed a loss of the peptide. In these persons, the cholesterol/triglyceride quotient was 0.94+/-0.45 for the whole serum, and 0.59+/-0.25 for the serum VLDL fraction. The quotient for VLDL cholesterol/serum triglyceride was 0.43+/-0,21, and the electrophoretic mobility of the VLDL subfraction was 0.38+/-0.04. Investigation of the VLDL-cholesterol/triglyceride quotient gave four (10%) false positives, while the quotient for VLDL-cholesterol/serum triglyceride gave ten (24%) false positives. This comparison shows that the criteria used for identification of "floating" beta-VLDL subfraction are only indirectly useful in the diagnosis of type III hyperlipoproteinaemia.


The analysis of nonlinear synaptic transmission. In order to characterize synaptic transmission at a unitary facilitating synapse in the lobster cardiac ganglion, a new nonlinear systems analysis technique for discrete-input systems was developed and applied. From the output of the postsynaptic cell in response to randomly occurring presynaptic nerve impulses, a set of kernels, analogous to Wiener kernels, was computed. The kernels up to third order served to characterize, with reasonable accuracy, the input-output properties of the synapse. A mathematical model of the synapse was also tested with a random impulse train and model predictions were compared with experimental synaptic output. Although the model proved to be even more accurate overall than the kernel characterization, there were slight but consistent errors in the model's performance. These were also reflected as differences between model and experimental kernels. It is concluded that a random train analysis provides a comprehensive and objective comparison between model and experiment and automatically provides an arbitrarily accurate characterization of a system's input-output behavior, even in complicated cases where other approaches are impractical.


Isolation and preliminary characterization of temperature-sensitive mutants of the murine sarcoma leukaemia virus complex. Five temperature-sensitive mutants (ts I to 5) were isolated from a stock of the Moloney strain of murine sarcoma leukaemia virus complex which had been mutagenized by ultraviolet irradiation or N-methyl-N'-nitro-N-nitrosoguanidine. In mouse cells at the non-permissive temperature the mutants formed fewer foci of transformed cells than at the permissive temperature. The ts mutants were characterized by testing: (I) murine leukaemia virus (MuLV) clones from the ts complex, (2) the effect of additional wild type MuLV on focus formation, (3) focus formation in rat cells and (4) focus formation with pseudotypes rescued from non-producer cells. Two mutants (ts 1 and ts 3) were found to be ts MuLVs which did not possess heat labile virion proteins and were not ts in post-penetration helper functions necessary for the fixation of sarcoma virus transformation. The remaining three mutants (ts 2, ts 4 and ts 5) were ts murine sarcoma viruses which, however, showed no temperature-sensitive effect on the maintenance of transformed cell morphology nor on colony forming efficiency in soft agar.


Antiviral activity of bovine interferons on primate cells. Potent preparations of bovine leucocyte and fibroblast interferons had substantial antiviral activity on monkey cells and low activity on human cells. Thus, interferon from a 'lower' phylogenetic species can have considerable antiviral activity in primate cells, but not all primate cells are equally sensitive.


Human cytomegalovirus: purification of enveloped virions and dense bodies. Enveloped virions and dense bodies of human cytomegalovirus have been purified by centrifugation, using combination negative viscosity: positive density gradients. Light-scattering bands of each component were obtained, and when these were examined by immune electron microscopy minimal cross contamination was observed.


Protection of mice against encephalomyocarditis virus infection by chemically modified transfer RNAs. Periodate or nitrous acid treatment greatly decreases the ability of unfractionated Escherichia coli transfer RNA (tRNA) to be aminoacylated by tRNA-synthetases but these treatments do not affect their antiviral activity against encephalomyocarditis virus infection of mice. Bisulphite treatment of E. coli tRNA reduces its ability to be aminoacylated by 20% and has no effect on antiviral activity. Bromine water treatment of tRNA under conditions causing extensive base modifications eliminates aminoacylation and the antiviral activity of E. coli tRNA. Periodate treatment of yeast tRNA does not affect its antiviral activity and nitrous acid treatment increases its antiviral activity to that of E. coli tRNA. The ability to be aminoacylated does not therefore appear to be essential for antiviral activity of tRNA but extensive modification (bromine water treatment) does destroy antiviral activity.


Refractory period, conduction of trains of impulses, and effect of temperature on conduction in chronic hypertrophic neuropathy. The refractory period, the ability to transmit trains of impulses, and the effect of temperature on conduction, have been studied in the sciatic-tibial nerve trunks of Trembler mice, which suffer from a dominantly inherited hypertrophic neuropathy. Both the refractory period of transmission and the relative refractory period were increased in Trembler mice when compared with controls. The nerve trunks of Trembler mice were unable to conduct rapid trains of impulses, and conduction block occurred at rates of stimulation as low as 25 Hz. Cold block occurred at temperatures significantly higher in Trembler nerves than in controls. The conduction velocity increased in an approximately linear fashion in both Trembler and control nerves when the temperature was raised from 20 degrees C to 40 degrees C, and the slopes were not significantly different. The Q10 (27 degrees C-37 degrees C) was 1.5 and 1.6 for control and Trembler nerves respectively. Conduction block was regularly observed in Trembler nerves when the temperature was raised within the physiological range. The abnormalities are related to the pathological changes of chronic demyelination.


The motor unit in psychotic patients: a single fibre EMG study. Single fibre EMG recording from extensor digitorum communis muscle is described in patients with psychotic illness. The fibre density, or average number of single muscle fibre action potentials belonging to the same motor unit within the uptake area of the electrode, was higher in the patient group than in normal controls. Increased jitter was occasionally seen. Motor and sensory nerve conduction velocity values were within the normal range. The results indicate that psychosis is associated with denervation and reinnervation by collateral sprouting.


Evidence for dopamine-storing interneurons and paraneurons in rhesus monkey sympathetic ganglia. In order to study and quantify the small, intensely fluorescent (SIF) cells and to examine their synaptology, the superior cervical sympathetic ganglia of rhesus monkeys were examined by a sequential glyoxylic acid (GA) monoamine fluorescence and electron microscopic technique. Correlated data concerning the sensitivity of the ganglia to dopamine were gathered by measuring the rise in cyclic AMP levels following incubation in vitro with exogenous dopamine. Cyclic AMP levels after incubation for 12 min with 50 micron dopamine showed marked increases, averaging 288% of control values. This indicates that these ganglia contain a dopamine receptor-adenylate cyclase complex. The number of SIF cells per mg of tissue was about half that found in the cow and rabbit, and only 1% of the number observed in the rat. Morphological data support the view that, in this species, there are almost equal numbers of type I (hypothesized interneuron) and type II (paraneuron) SIF cells. Presumptive SIF cell efferent synapses were observed by electron microscopy, the postsynaptic elements being dendritic spines or very fine nerve processes.


Cyclic AMP and GMP levels in thymocytes and splenic T and B lymphocytes in protein deprived rats. Cyclic nucleotide levels were determined in thymocytes and in splenic T and B lymphocytes separated by nylon wool filtration. In control rats, thymocytes contained less cAMP and cGMP than splenic T cells. The level of cGMP was lower, and consequently cAMP:cGMP ratio was higher in splenic B than in T cells. Protein deprivation increased the nucleotide ratios in both T and B splenic lymphocytes; this effects was due to a decrease in the cGMP levels in the case of the T cells and to an increase in the cAMP levels in the B cells. These data could be related to the reduced mitotic activity of the splenic lymphocytes. However, the nylon-nonadherent thmyocytes of protein deprived rats paradoxically displayed increased cGMP contents without changes in the cAMP levels, despite the drop in their proliferative potential. The impairment of cyclic nucleotide metabolism may intervene in the immunological disturbances due to protein malnutrition.


Biohydrogenation and availability of linoleic acid in lactating cows. Linoleic acid biohydrogenation, absorption and availability for maintenance and milk production in dairy cows fed high grain (60--85% of dry matter) diets were quantitatively estimated by isotope dilution, using two methods of dosing. [1-14C]Linoleic acid-labeled chylomicra and very low density lipoproteins (VLDL) were obtained from lymph of a calf fed [1-14C]linoleic acid and fitted with a thoracic duct-venous shunt. Labeled chylomicra were injected intravenously into two cows: a Jersey (trial 1), and a Holstein (trial 2). Labeled VLDL was injected intravenously into a Holstein cow (trial 3). In trials 4 and 5 the [1-14C]linoleic acid was placed into the omasal canal of two rumen-fistulated Holstein cows. Linoleic acid biohydrogenation (%), absorption (g/day), and availability above requirements for milk production (mg/kg body wt 3/4) were: 68.1 +/- 2.28, 52.1 +/- 2.92, and 244 +/- 19.4 (mean +/- SE), respectively. The biohydrogenation data indicate that both methods of dosing the cows were equally dependable. The estimates of linoleic acid biohydrogenation are consistent with limited data previously reported, indicating that the isotope dilution technique used is a reliable method to estimate linoleic acid absorption in lactating cows. Linoleic acid available to the lactating cow above milk production requirements was more than double the requirement of weanling female rats, when compared on the basis of metabolis body size.


Mannosidosis: pathology of the nervous system. In a girl with mannosidosis, who died at 3 1/2 years of age, histopathological and ultrastructural changes in the nervous system are described. A widespread neuronal storage evidenced by ballooning of the neuronal perikarya is the salient histological feature and the occurrence of electron-lucent storage vacuoloes in neurons, astrocytes, endothelial cells and pericytes is the most striking ultrastructural feature of mannosidosis in the nervous system. By virtue of the deficiency of acidic alpha-mannosidases A and B, the accumulation of mannose-containing oligosaccharides in tissues and the occurrence of storage vacuoles in various cells, mannoisidosis is similar to various neuronal storage diseases associated with lysosomal enzyme deficiencies. In mannosidosis, the storage vacuoles in the neural and visceral tissues are alike with little variation in details and contain chiefly loosely dispersed, finely reticulogranular material. The storage vacuoles in neurons in mannosidosis are, therefore, distinct from those in neurons in other lysosomal storage disease such as Pompe's disease, various lipidoses and mucopolysaccharidoses. However, they resemble closely the storage vacuoles in neurons in fucosidosis and those in liver cells in various mucopolysaccharidoses.


The effects of nortriptyline and mesoridazine on neurons and glia in vitro. The ultrastructural changes produced by Mesoridazine and Nortriptyline in neurons and glia in vitro are reported. Concentrically laminated bodies (CLB) and increased numbers of dense bodies were produced by exposure to these drugs. The number and complexity of the CLB increased with increased dose and longer time in vitro and their formation appeared to be reversible.


Arsenical neuropathy: residual effects following acute industrial exposure. A case report is presented describing a worker who was splashed with arsenic acid in an industrial accident and subsequently developed symptoms of systemic arsenicalism and peripheral neuropathy. This is the only report, to the authors' knowledge, of a single episode of cutaneous absorption of arsenic resulting in peripheral neuropathy. Previous reports of arsenical neuropathy and rationale for BAL therapy early in the treatment of systemic arsenicalism are discussed.


Vaccine-associated poliomyelitis in a child with sex-linked agammaglobulinemia. Paralytic poliomyelitis was observed in a child with a sex-linked defect in immunoglobulin synthesis. Evidence is presented that this was secondary to administration of oral, live poliovaccine. The demonstration of a familial sex-linked gammaglobulin deficiency and the failure to document a defect in cell-mediated immunity in this child extends the risk of vaccine associated poliomyelitis to virtually all forms of immunodeficiency. The critical host factors in the pathogenesis of poliovirus vaccine infection and in particular its unfavorable outcome appear to include either a deficiency in the humoral (B cell) system or in the cell-mediated (T cell) system.


Bronchial carcinoid tumors in children and adolescents. Thirteen histologically proven cases of bronchial carcinoid from the literature and eight cases from the Armed Forces Tumor Registry, all under 20 yr of age, are presented. Hemoptysis, cough, and pneumonia were the most frequent presenting complaints. Bronchoscopy defined the lesion in all 16 cases so studied. Gross and microscopic pathology is briefly described, and treatment in these cases outlined. A good long-term prognosis can be expected, except in those cases with a long delay in diagnosis.


Pleural effusion in Wilms' tumor.

Pleural effusion in Wilms' tumor. The association of pleural effusion and Wilms' tumor is uncommon. We report three patients who developed pleural effusion as a result of different mechanisms, all of which were related to Wilms' tumor or to its treatment.


Cultural characteristics of mesoblastic nephromas. Seven infants and children with renal tumors underwent nephrectomy and the tumors were cultured to investigate the behavior of their cells. In four children over 1 year of age with histologically typical nephroblastoma in the tumor cells were bizarre and disorderly. In one patient 3 months of age where the histology suggested malignancy the tumor culture showed an orderly growth pattern of spindle cells akin to normal kidney and the patient has had no recurrence for 18 months. In one infant seven months of age the histology was consistent with malignancy and this was confirmed on culture showing bizarre cells similar to those of nephroblastoma. The third infant under 1 year of age had a teratoma on histology and an unusual growth pattern on culture characterised by polyhedral cells with long processes and much overlapping. It is suggested that in infants with kidney tumors, tissue culture be used to help in assessing the malignant potential of the tumor and deciding on adjuvant therapy.


A case of EMG (exomphalos, macroglossia, and gigantism) syndrome with associated renal tumor. This is a brief report of a patient with Wiedemann-Beckwith syndrome and nephroblastoma with metastases, apparently cured by surgery, chemotherapy, and radiation.


Wilms' tumor in crossed fused renal ectopia. The occurrence of a nephroblastoma in a crossed ectopic kidney has been reported only one previous time. A second patient with these findings is the subject of this report.


Polyamine metabolism I: Synthesis of dansyl derivatives of N-(monoaminoalkyl)- and N-(polyaminoalkyl)acetamides and elucidation in urine of a cancer patient. The dansyl derivatives of N-(monoaminoalkyl)- and N-(polyaminoalkyl)acetamides were synthesized and unequivocally characterized. TLC of the dansyl derivatives obtained from human urine indicated the presence of N-[3-[(4-aminobutyl)amino]propyl]acetamide (N1-acetylspermidine, N-[4-[(3-aminopropyl)amino]butyl]acetamide (N8-acetylspermidine), and N-(4-aminobutyl)acetamide (N-acetylputrescine) in appreciable amounts. The dansyl derivatives of N1-acetylspermidine, N8-acetylspermidine, and N-acetylputrescine were isolated and purified using various chromatographic methods. The mass spectra of these compounds were similar to those of authentic samples, which confirmed the identity of these compounds and established the presence of N8-acetylspermidine as well as N1-acetylspermidine and N-acetylputrescine in human urine.


Single-dose tolerance to morphine hypothermia in the rat: differentiation of acute from long-term tolerance. The time course for the development and disappearance fo tolerance to the hypothermic effects of morphine was determined after a single subcutaneous injection (10 mg/kg) in rats. Body temperature responses to second injections, given at varying times after the first, were compared with those produced by the initial injections. Tolerance (attenuation of the hypothermic response) induced by a single dose of morphine was found to be biphasic. Acute tolerance was apparent by 4.5 hours and lasted at least 20 hours after morphine administration. Development of long-term tolerance occurred within 24 hours, was maximal at 3 days and persisted up to 11 days. Both acute and long-term tolerance were drug specific since hypothermic responses to pentobarbital were not altered either 4.5 or 72 hours after morphine. Long-term tolerance was attenuated by co-administration of naloxone with initial dose of morphine. Long-term tolerance induced by multiple doses of morphine (300 mg/kg/day) differed from that induced by a single dose in persistence (3-4 weeks) rather than in quality or magnitude. The thermoregulatory response of the rat provides a sensitive measurement system which allows discrimination between acute and long-term opiate effects.


The rapid reduction of disulfiram in blood and plasma. A gas chromatographic assay procedure was developed to quantitate the reduction product of disulfiram, diethyldithiocarbamate (DDC), in blood and plasma. The procedure involved the in situ methylation of DDC prior to the extraction and chromatography of the methyl ester. The minimal sensitivity achieved was 0.2 microgram/ml from 1 ml of blood or plasma. The coefficient of variation about any concentration was 10.5%. Calibration curves having a reproducible nonlinear form were prepared up to 9 microgram/ml. The assay procedure was used to evaluate the stability of disulfiram and DDC in blood. Disulfiram was rapidly and quantitatively reduced to DDC within 4 minutes. The DDC thus formed decomposed in human and dog blood with half-lives of 70 and 100 minutes, respectively. The implications of these findings are discussed with respect to the chemical form of disulfiram responsible for the ethanol-sensitizing effect.