Esophageal function in diabetes mellitus and its relation to peripheral neuropathy. Esophageal function was determined in 50 unselected patients with diabetes mellitus (DM). Fourteen age-matched healthy subjects served as controls. The presence of peripheral neuropathy (PN) was determined by a neurological examination and by nerve conduction studies. An intraluminal transducer assembly placed in the distal esophagus measured pressure in the lower esophageal sphincter and body of the esophagus. Esophageal function was studied both before and after edrophonium chloride, 80 microgram per kg intravenously. There was no significant difference in peristaltic amplitude between the controls and diabetics. There was also no difference in amplitude when DM was divided into presence or absence of PN. However, there was a significant decrease in velocity of peristalsis in DM with PN when compared to DM without PN and to controls. Resting lower esophageal sphincter pressure in DM was similar to controls, with no difference with or without PN. Twenty-eight patients (56%) with DM had abnormal motility, characterized by frequent spontaneous contractions, and decreased prevalence of peristalsis. Abnormal motility in DM was associated with PN and was characterized by a dysfunction of esophageal innervation with intact smooth muscle function.


[Effects of 5-(o-chlorophenyl)-1-methyl-7-nitro-1, 3-dihydro-2HO1, 4-benzodiazepin-2-one (ID-390) as compared to nitrazepam on the sleep-wakefulness cycle in cats (author's transl)]. ID-690 was given at doses over 0.3 mg/kg, and increase in state of arousal and decrease in state of slow wave sleep were observed. Nitrazepam at doses over 1.0 mg/kg revealed similar effects. These agents increased incidences of paradoxical sleep at a low dose of 0.03 mg/kg. When ID-690 was given at doses over 0.3 mg/kg, the number and duration of incidences of paradoxical sleep decreased. Nitrazepam at doses over 0.1 mg/kg had a similar depressive effect on paradoxical sleep. These results suggest that these benzodiazepines have, dose-dependently, both facilitative and depressive actions on paradoxical sleep, and that the dose of these agents which affects arousal is not always the same as that which affects paradoxical sleep.


[Skeletal changes in Down's syndrome. A correlation between radiological and cytogenetic findings (author's transl)]. One hundred and two patients with Down's syndrome aged one day to 17 years were examined radiologically, seven of these repeatedly. Films obtained included the chest in two planes, lateral spine, left hand and pelvis. The following features were noted; the number of paired ribs, ossification centres in the manubrium, height of lumbar vertebral bodies, shortening of the phalanges, bone age and calculation of the acetabula and ilial angle and ilial index. In all patients the chromosomes were examined in order to determine the cytogenetic type. The following features were found to be more common in Down's syndrome than in normals: abnormal ossification of the manubrium (33%), aplasia of the twelfth rib (18%), high lumbar vertebral bodies (50%), brachymesophalangia of the fifth ray (62%) and changes in skeletal maturation (acceleration or retardation in 48%). Changes in the pelvis were typical; with increasing age, the acetabular angle falls and the ilial angle increases. The most striking skeletal changes were found in 84 patients with trisomy 21.


[Problems of corticosteroid-induced adrenal insufficiency in surgery]. Patients previously treated with corticosteroids have a high risk whenever surgery is undertaken. Corticosteroid therapy, especially if prolonged, causes adrenal suppression. Therefore patients under steroid treatment need more, not less corticosteroids during the operative period.


[Dopaminergic Neurotransmission in Parkinson's syndrome (author's transl)]. More recent electrophysiological and biochemical data dealing with the functioning of the central dopaminergic synapse are reviewed in relationship to Parkinsonian symptomatology. Emphasis is put on the influence of feedback mechanisms involving pre and postsynaptic receptor sites, the dopaminergic-cholinergic imbalance and receptor supersentitivity. Stimulation of the dopamine-sensitive adenylatecyclase proved to be a good indicator of central dopaminergic receptor stimulation. Current therapeutic concepts are discussed in view of their limitations and side-effects.


Social work services for radiation therapy patients and their families. A social worker helps meet the social and emotional needs of cancer patients undergoing radiation therapy at the University of Utah Medical Center and the needs of their families. He makes personal contact with the patient and family on their first visit to the radiation division, answers questions, provides information on community services, and generally lets the patient and family know that they can call on him any time during the treatment process. He also follows up patients who are still seriously ill after treatment, and tries to maintain periodic contact with families of patients who have died.


Gestational trophoblastic neoplasms: morphologic considerations. Abnormal trophoblastic proliferation is the hallmark of a spectrum of lesions constituting the gestational trophoblastic neoplasms. Rapid proliferation, infiltration, vascular invasion, hematogenous dissemination, and spontaneous regression are features of both normal and neoplastic trophoblast. Trophoblastic hyperplasia without hydrops, hydatidiform mole, invasive mole, and gestational choriocarcinoma are related lesions, characterized by increasingly aberrant trophoblastic growth and worsening prognosis, if untreated. Difficulties in diagnosis may arise with respect to the normal early implantation site, the hydropic abortus, and postgestational, involuting, residual trophoblast. Histologic grading or hydatidiform moles is relevant to their prognosis and biologic behavior. Trophoblastic neoplasia may begin at any stage of pregnancy or puerperally with immediate or late and local or distant manifestations in the mother or the child. Cognizance of the capricius potential behavior of trophoblast permits successful management of its proliferative lesions, monitored by serial measurement of gonadotropin secretion.


Malignant germ cell tumors of the ovary. This article reviews 281 malignant germ cell tumors of the ovary from the Armed Forces Institute of Pathology and highlights their distinctive clinical and pathologic features. Emphasis is placed on the importance of a combined therapeutic approach utilizing surgery, chemotherapy, and radiation. The rationale for unilateral salpingo-oophorectomy in conjunction with chemotherapy for certain types of neoplasm confined to one ovary (stage 1a) is emphasized, and the role of human chorionic gonadotropin and alpha-fetoprotein as tumor markers in the management of patients with these tumors is discussed.


Functional ovarian tumors of stromal and sex cord origin. Most functional ovarian tumors are of specific stromal or sex cord origin, capable of differentiating in either a female direction or, less commonly, a male direction. Tumors of stromal origin such as thecoma, stromal luteoma, and Leydig cell tumors are for all practical purposes benign, and evolve from mature ovarian stroma, recapitulating common non-neoplastic transformations such as stromal changes associated with follicle development and nodular stromal hyperplasia. Sex cord tumors of granulosa or Sertoli cell types are generally of a low order of malignancy, tending to late recurrence, occasional peritoneal seeding, and only rarely to distant metastasis. Nonfunctional tumors of the ovary may trigger hormone production from adjacent reactive stroma. Massive edema of the ovary due to partial torsion may simulate neoplasm and may produce hormonal syndromes by an unknown mechanism.


The polymorphism of red cell uridine monophosphate kinase in two samples of the Italian population. Red cell uridine monophosphate kinase polymorphism has been studied on a total of 915 individuals from two different areas of Italy (Milan and Rome). The two groups of about the same size show very similar gene frequencies. The UMPK2 allele in the pooled sample has a frequency of 2.8% which is significantly lower than those observed in the two other Caucasian populations so far examined.


An analysis of red cell enzymatic markers in the province of Bologna (Italy). About 280 unrelated individuals living in the province of Bologna (Northern Italy) have been studied for the following red cell enzymatic markers: phosphoglucomutase (PGM), adenylate kinase (AK), adenosine deaminase (ADA) and phosphohexose isomerase (PHI). 116 subjects from the same sample have also been analysed for red cell acid phosphatase (ACP). The observed gene frequencies are PGM21 = 0.280; AK2 = 0.030; ADA2 = 0.091; ACPa = 0.297; ACPb = 0.647; ACPc = 0.056. In the PHI system two individuals with the variant PHI 3-1 phenotype have been found.


Enzyme polymorphisms of Ideles populations (Ahaggar, Algeria) and the Iwellemeden Kel Kummer Twaregs (Menaka, Mali). Surveys dealing with enzyme polymorphisms have recently been conducted in the Sahara. Results from two populations are reported here: 227 inhabitants of Ideles village (Ahaggar, Algeria); 285 nomads of a genetic isolate, the Kel Kummer Twareg tribe (Menaka, Mali). The four classical molecular variants of G6PD:A+, A-, B+, B-, are found in Ideles. The frequency of the G6PD A+ Negroid variant reaches 15% in Ideles and 7.7% among the Kel Kummer. However, gene frequencies will have to be recalculated after a study of the genetic transmission through families. The PGDC gene of 6PGD is especially frequent in the Kel Kummer where 10 'Canning' phenotypes have been observed. The PGM distribution of alleles at locus 1 in Ideles is the same as in the Mediterranean populations. The pa gene of acid phosphatase, relatively frequent in Ideles, has been excluded by drift from the Kel Kummer gene pool. AK and LDH enzymes have also been studied in both samples. The abnormal Ea1 mutation of serum pseudocholinesterase exists in Ideles and in the Kel Kummer as in other populations of the Sahara; the C5 esterase component was revealed by electrophoresis in 5% of the Kel Kummer people.


Adenylate cyclase activity in cultured epithelial cells. The cyclic AMP metabolism of cultured epithelial cells was investigated. Epinephrine or 1-methyl,3-isobutylxanthine (MIX) alone had no effect on cyclic AMP levels in intact cells, whereas the combination of the two agents yielded a 6- to 10-fold increase in cyclic AMP levels. Both basal and stimulated cyclic AMP levels decreased with increasing cell density. Cell-free adenylate cyclase preparations were stimulated markedly by epinephrine or isoproterenol in the absence of MIX. Since the epithelial cells were found to have a relatively small amount of cyclic nucleotide phosphodiesterase (PDE) activity, the requirement for MIX to visualize intact cell responsiveness to epinephrine could be explained only partially by its PDE inhibitory properties.


A further study on the regulation of cyclic nucleotide phosphodiesterase activity in neuroblastoma cells: effect of growth. Adenosine 3',5'-cyclic monophosphate (cyclic AMP) phsophodiesterase activity in mouse neuroblastoma cells in culture markedly increased during exponential growth and reached a maximal level at confluency; whereas guanosine 3'5'-cyclic monophosphate (cyclic GMP) phosphodiesterase activity only slightly but significantly increased under a similar experimental condition. The increase in cyclic AMP phosphodiesterase activity was blocked by both cycloheximide and dactinomycin, whereas the increase in cyclic GMP phosphodiesterase was blocked by only cycloheximide. When the confluent cells were replated at low density, the cyclic nucleotide phosphodiesterase activity decreased; however, when they were plated at high cell density which equaled confluency, the enzyme activity did not decrease. Unlike cyclic AMP phosphodiesterase activity, cyclic GMP phosphodiesterase activity did not change significantly in prostaglandin E1-treated cells, but decreased in cells treated with the inhibitor of phosphodiesterase. Like cyclic AMP phosphodiesterase activity, cyclic GMP phosphodiesterase activity also did not change in cells treated with serum-free medium, X-irradiation, sodium butyrate and 6-thioguanine.


Interferon inhibition of lymphocyte mitogenesis. Interferon inhibits the early phase of mitogenic action of Concanavalin A on lymphocytes as measured by incorporation of labelled precursors in RNA, DNA or protein. There was no evidence for a role of cyclic AMP in the process of inhibition. Dialysis of interferon against high salt and urea did not separate cell inhibitory activity from antiviral activity. Thus, these results are compatible with the possibility that the antiviral and cell inhibitory activities are in the same molecule. However, further purification of the interferon will be required to verify this assumption.


Effects of whole-body irradiation on antibody affinity. Mice exposed to a sublethal dose of X-rays were immunized with alum-precipitated DNP-KLH (dinitrophenyl-keyhole limpet haemocyanin) and B. pertussis either before or after irradiation. The primary anti-DNP antibody response was evaluated during 8 weeks after immunization by the equilibrium dialysis technique using ammonium sulphate- precipitated serum globulins and the ligand 3H-labelled xi-DNP-L-Lysine. The serum concentrations of antibody sites in mice immunized 1-5 days before or 2 h-8 weeks after 450 rad were below the values in unirradiated controls at all bleeding times. Antibody affinity, however, was found to be up to 20 fold higher in irradiated mice than in control mice when antigen was injected before, or 3-8 weeks after, irradiation. Spleen cells from mice exposed to 450 rad 1-9 weeks before killing were stimulated in vitro with PHA, ConA, or LPS. Recovery profiles of mitotic responsiveness suggest that enhancement of antibody affinity in irradiated mice could result from relative lack of suppressor T Cells.


Action of sphingomyelinase C and other lipid-specific agents as inhibitors of Fc binding and locomotion in human leucocytes. The binding of antibody-coated chicken erythrocytes (EA) to human blood lymphocytes and monocytes is inhibited by pretreatment of the leucocytes with sphingomyelinase C. Inhibition of rosetting of neuraminidase-treated EA with neutrophils is also seen with this enzyme. The cholesterol-binding theta-toxin of Clostridium perfringens and pronase also inhibit EA-rosette formation, but less strongly than sphingomyelinase. The lipid-specific agents also inhibit chemotactic migration of leucocytes to casein and denatured HSA, whereas proteases and glycosidases do not. These results suggest that membrane lipids are important constituents of the binding sites for Fc fragments and for certain chemotactic factors and point to an important role for sphingomyelin in this binding.


The effects of inhaled silica and chrysotile on the elastic properties of rat lungs; physiological, physical and biochemical studies of lung surfactant. When rats breathed air containing approximately 70 mg m-3 of respirable crystalline silica 7 h daily for 10 days (2000 mg m-3 h) the surface tension forces of the alveolar lining film were reduced. This was shown both by surface tension measurements on lung extracts and by pressure-volume studies with air and saline filling of excised lungs. Larger quantities of inhaled silica produced similar effects. Chrysotile inhalation caused an even more marked decrease in the surface tension forces. In the chrysotile studies these findings were supported by biochemical estimations of the quantity of surfactant in the lungs, which was increased 10-fold by an inhalation of 6500 mg m-3 h. Electron microscopy showed an increased number of the type II alveolar cells which produce surfactant and of free phospholipid lattices in the air spaces of the lungs of rats exposed to chrysotile and silica. Both the surface tension and biochemical estimations on a control group of rats suggest that there is an increase in the amount of surfactant in the lungs up to about 12 months of age.


A cell kinetic study of the alveolar wall following dust deposition. The stathmokinetic technique was used to study quantitatively the proliferative response of cells comprising the alveolar wall of inbred mice which had inhaled coal or quartz. The exposures occupied 4 weeks and the observations continued over an extended period thereafter. Control observations suggested that mitotic inhibition was induced solely by residence in the exposure chamber, since cessation of exposure was quickly followed by a temporary rise of proliferative activity. This feature was not apparent when dust was inhaled, though with comparable exposure to both coal and quartz there was an elevation in mitotoic incidence of alveolar wall cells in dust-free areas of lung at a later interval. In dust-laden areas the rise was less in evidence, a situation that persisted throughout the post-exposure survival. The difference in mitotic incidence between dust-free and dust-containing areas was more evident after coal than after quartz inhalation, whilst in higher concentration a quartz aerosol induced a continued depression of mitotic activity. The proliferative response seen in the alveolar walls of control and dusted mice is most likely to be contributed by the interstitial precursors of alveolar macrophages, cells which are recognized to be marrow derived. The changes observed in mitotic incidence are interpreted in terms of demand for alveolar macrophages according to the nature of the dust and the inintensity of the exposure.


The activation of phospholipase A in macrophages after the phagocytosis of silica and other cytotoxic dusts. The phospholipid and lipid metabolism of mouse peritoneal macrophages has been studied in the presence of various silicogenic dusts and asbestos particles. These extensive kinetic studies were possible as a reproducible method was developed to determine these major cellular constituents after labelling macrophages with 1-14C-oleic acid. The following results have been obtained so far: 1. Silicogenic dusts activate a phospholipase A in macrophages leading to a concentration and time-dependent degradation of lecithin and cephalin. 2. Using low doses of SiO2 (less than 1 mg/10(7) cells) the split-off free fatty acid can still be transesterified into the triglycerides. 3. High doses of silica (greater than 2 mg/10(7) cells) induce also a lipolysis. 4. Silica specifically inhibits a plasma membrane-bound acyltransferase 5. Asbestos particles induce in long-term cultures a moderate degradation of diacylphospholipids with transesterification of 1-14C-oleic acid into the triglycerides.


Comparative pulmonary carcinogenicity of inhaled beta-emitting radionuclides in beagle dogs. Beta-emitting radionuclides are important constituents of isotope inventories in light water reactors and may pose an inhalation hazard to industrial workers or the general population if they are released. To study the biological effects of such potential exposures, a series of life span studies was initiated in which beagle dogs were exposed to aerosols of relatively insoluble fused clay particles containing 90Y, 91Y, 144Ce or 90Sr. Groups of dogs exposed to each radionuclide received graded initial lung burdens of radioactivity. When combined with the varied physical half-lives of the four radionuclides, this resulted in a wide variety of radiation doses and dose patterns to the lung. Deaths (greater than 640 days after exposure) were generally associated with pulmonary neoplasia in dogs that inhaled 91Y, 144Ce or 90Sr. These dogs had cumulative lung doses to death greater than 20 000 rads. Exposure to 144Ce or 90Sr with dose rates that decreased slowly induced pulmonary haemangiosarcomas. Pulmonary irradiation from 91Y, with a rapidly decreasing dose rate, resulted in pulmonary epithelial tumours. No malignant lung tumours have been seen within 1540 days after exposure to 90Y. The animals in the main studies have been observed for 1342 to 2756 days after exposure.


The effect of quartz and other non-coal dusts in coalworkers' pneumoconiosis. Part II. Lung autopsy study. Preliminary pathological and mineralogical studies are reported on seventy-four sets of lungs from British coal miners who have been employed at the collieries included in the National Coal Boards's Pneumoconiosis Field Research. The degree of lung damage was considered in relation to the lung dust content and to the known dust exposures of the men concerned. Lungs were classified as having soft macules, fibrotic nodules or PMF. Those with soft macules had the lowest dust content but there was no significant difference between the dust contents of the lungs with fibrotic lesions and those with PMF. The percentage of non-coal minerals in the lung dust appeared to increase with the pathological classification from soft macules to PMF, and comparisons with the exposure data indicated a preferential retention of non-coal minerals, and especially of quartz, in the cases with the more severe lesions. Histological examination of the lesions showed the packing of dust was less close and the cellular response more vigorous with the lungs with the highest quartz content.


Characteristics of lung dusts and their relation to dust exposure and pathological findings in the lungs. Lung dusts were investigated, post-mortem, in twenty-five miners from mixed metal mines, tunnels, and quarries who had exposed to high concentrations of mixed dust containing about 20-25% free crystalline silica. The character of the relations found between the amount of quartz per 100 g dry tissue and the clinical, X-ray and pathological findings is similar to that established in coal miners. The difference lies in the fact that with equal amounts of quartz per 100 g dry tissue, there is less silicosis in coal miners than in our cases; the average residence time of retained dust is longer in coal miners, but its quartz per cent is lower.


Immunoglobulin class-specific antibody response in serum, spleen, lungs, and bronchoalveolar washings after primary and secondary sendai virus infection of germfree mice. Immunoglobulin class-specific antibodies were measured by a solid-phase radioimmunoassay in serum, bronchoalveolar washings (BAW), lung cell lysates, and spleen cell lysates in germfree mice after intranasal (i.n.) and intraperitoneal (i.p.) primary and secondary 10(5), 10(4), and 10(3) mean tissue culture infective doses (TCID(50)) of live parainfluenza 1 (Sendai) virus. The earliest antibody detected in lungs after i.n. virus challenge was immunoglobulin G (IgG), followed by IgM and, lastly, IgA. The local IgA response after both primary and secondary i.n. virus challenge was lowest after the severest infection. It is suggested that the delayed appearance of IgA antibody and the lower response after severe lung damage may be related to a temporary local secretory component-producing cell deficiency. The lungs were a major source of serum IgG antibody after both primary and secondary i.n. virus challenge. Only IgG and IgM antibodies were detectable in lung cell lysates after the i.n. 10(3) TCID(50) secondary response. A secondary response was detected in IgG, IgA, and IgM after secondary i.n. challenge with the other two doses. The lung response to all of primary and secondary i.p. doses of virus was exclusively IgG and IgM. Calculation of radioimmunoassay antibody per microgram of IgG, IgA, and IgM in serum and BAW after both i.n. and i.p. virus challenges showed that, when BAW antibody was present, the ratio in BAW was always higher than that in serum. This finding in the i.n. mice, together with the presence of IgA antibody-containing cells in the lungs, strongly indicates local manufacture and secretion of IgA antibodies in these animals and suggests that the same conclusion could apply to local IgG and IgM antibodies after both i.n. and i.p. challenges.


Antigenic relationship between human and bovine rotaviruses as determined by neutralization, immune adherence hemagglutination, and complement fixation tests. Guinea pig antiserum to bovine rotavirus does not neutralize human rotavirus. Bovine and human rotaviruses were, however, extensively cross-reactive when examined by complement fixation and immune adherence hemagglutination tests with antiserum to either virus. The immune adherence hemagglutination test was 16- to 32-fold more sensitive than the complement fixation test in detecting rotavirus.


Sequential changes in cell-mediated immune responses to herpes simplex virus after recurrent herpetic infection in humans. Lymphocyte responses to herpes simplex virus (HSV) were studied in 23 patients with recurrent herpes labialis and in 19 control subjects. Lymphocytes of seropositive, but not seronegative, controls responded to HSV by thymidine incorporation, and the supernatant fluids inhibited the migration of guinea pig macrophages. Lymphocytes from patients with a recurrent herpetic lesion responded to HSV by significantly greater thymidine incorporation than seropositive controls, but supernatants did not show an increased macrophage migration inhibition response. During the 28 days after the onset of a lesion, the thymidine incorporation to HSV fell to the level of the seropositive controls, and supernatants then induced an increased inhibition of macrophage migration. Lymphocyte responses to Candida albicans, purified protein derivative, or phytohemagglutinin did not fluctuate according to the presence of a lesion and did not differ from those of the controls. Lymphocyte responses to HSV were unaffected by culture in the presence of serum from seronegative or seropositive controls, or from patients with or without a herpetic lesion. It is suggested that in patients with recurrent herpes labialis a periodic defect of the migration inhibition response might have allowed the recurrent infection to develop, and that the increased thymidine incorporation stimulated by HSV in vitro is a result of antigenic stimulation from the lesion.


Transplacental transmission of polyoma virus in mice. When pregnant mice were inoculated on day 1 of gestation with polyoma, some of them exhibited total resorption or reduced litter size, the extent depending on the dose of virus. Virus was detected in 4 out of 11 mouse embryo fibroblast (MEF) cultures made from infected mothers. After maternal infection on day 5 or 10 of gestation, virus titers of up to 10(7) 50% tissue culture infectious doses (TCID50)/g of fetus were found in all pools of fetuses tested 5 days later, with the titers falling by day 6. Hemagglutination-inhibiting antibodies against polyoma appeared in maternal serum by day 6 and rose to a maximum by day 14. Immunoglobulin G class antibodies were detected by day 7, with titers rising rapidly to a maximum at day 14. After maternal infection later in gestation (day 15), one out of three litters of newborn mice was found to have 10(5) TCID50 polyoma virus per g in pooled kidney samples.


Heterogeneity of the sensitivity of vesicular stomatitis virus to interferons. Twelve cloned viruses were randomly isolated from original (uncloned) vesicular stomatitis virus (VSV), and their sensitivities to mouse and human interferons were examined. When the interferon sensitivities of these various VSVs were compared by the plaque reduction method in L cells, virus 3 was found to be sevenfold more sensitive than virus 11, and the interferon sensitivity of the original (uncloned) virus was intermediate. The present study shows that uncloned VSV Indiana strain is a mixture of viruses that have different sensitivities to interferon. The slope of the dose-response curve of original (uncloned) virus to mouse interferon was less steep than those of cloned viruses. Virus 11, which was the least sensitive to mouse interferon, was relatively sensitive to human interferon. There was no correlation between the sensitivities of virus clones to mouse interferon and their sensitivities to human interferon. When the interferon sensitivities were tested by various assay methods (plaque reduction, yield reduction, and cytopathic effect inhibition), the ranking of the interferon sensitivities of different viruses was not changed. These results indicate that the relative sensitivity of a virus to interferon is determined by the host cells in which the tests are performed, but not by assay method used.


Characterization of SA12 as a simian virus 40-related papovavirus of chacma baboons. SA12 virus, originally isolated from an uninoculated South African vervet monkey kidney culture, was identified as a new member of the simian virus 40 (SV40)-polyoma subgroup of papovaviruses. The virus produced a cytopathic effect with nuclear enlargement in primary rhesus kidney cells. The virion had papovavirus morphology and a diameter of 44 to 45 nm. The DNA of the virus was a circular, double-stranded, superhelical molecule with a mean length 101% that of SV40 DNA and an estimated molecular weight of 3.3 X 10(6). The virus was found to be unrelated to other papovaviruses by neutralization, immune electron microscopy, and immunofluorescence tests with antiviral sera. SA12 virus-infected cells exhibited a capsid antigen, which has recently been found to be common to viruses of the SV40-polyoma subgroup. The virus readily transformed kideny cells from 10-day-old hamsters. Inoculation of transformed cells produced tumors in 3- to 4-week-old hamsters. The T antigens of SA12 and SV40 viruses were strongly and reciprocally cross-reactive. A high proportion of the sera of chacma baboons, Papio ursinus, and a comparatively lower proportion of the sera of vervet monkeys, Cercopithecus pygerythrus, had neutralizing antibodies to SA12 virus


Polymorphonuclear leukocyte bactericidal activity and oxidative metabolism during glutathione peroxidase deficiency. Glutathione peroxidase (GPx) deficiency has been proposed as a cause of some instances of chronic granulomatous disease (CGD). GPx activity varies greatly among species, and specific deficiency of this selenium-dependent enzyme can be produced by dietary selenium deficiency in rats. Bactericidal activity of polymorphonuclear (PMN) leukocytes from normal rats, humans, and guinea pigs (GPx high, intermediate, and nearly absent, respectively), selenium-deficient rats (GPx absent), and a patient with CGD were compared. There was no correlation between natural levels of GPx and bactericidal activity; only CGD was associated with inability to kill a Proteus mirabilis strain in vitro (killing known to be dependent on oxidative mechanisms). Postphagocytic metabolism was examined in normal and GPx-deficient rats. Both demonstrated normal iodination and superoxide production during phagocytosis and gave similar histochemical reduction of nitroblue tetrazolium dye under either resting or endotoxin-stimulation conditions. Postphagocytic hexose monophosphate shunt activity was somewhat lower in PMN from GPx-deficient animals as compared with normal but was substantially (10-fold) higher than that observed in resting cells. Thus, postphagocytic oxidative responses and subsequent bactericidal activity of PMN leukocytes were not compromised by complete absence of GPx, even in the species with the highest natural level of this enzyme. These results are not compatible with the hypothesis that CGD can be caused by a deficiency of GPx.


Beta-adrenergic blocking activity of Yersinia pestis murine toxin. Yersinia pestis plague murine toxin has been found to inhibit the mobilization of free fatty acids in mice in a manner similar to that of beta-adrenergic blocking agents. The blockage is detectable 75 min after injection of the toxin (1 to 2 mean lethal doses). The degree of inhibition was directly correlated with the toxicity of a given toxin preparation. Agents such as cholera toxin or glucagon, with apparently distinct receptors from beta-adrenergic receptors, stimulated adenylate cyclase and lipolysis and effectively modified toxicity. Likewise, cyclic adenosine 3',5'-monophosphate bypassed the toxin block and antagonized toxicity. Energy-rich compounds such as fatty acids, organic acids, and glucose effectively modified the intoxication process. The biological activity of plague toxin showed profound temperature sensitivity. Mice placed at 5 degrees C were highly susceptible to the effects of the toxin, whereas mice placed at 37 degrees C were totally resistant to intoxication. Results showed that plague toxin cannot block epinephrine-induced mobilization of free fatty acids in mice placed at 37 degrees C. These studies suggested that plague toxin acts at the receptor level in a manner similar to that of beta-adrenergic blocking agents. A complete, analogous activity was shown between toxin and known beta-adrenergic antagonists in their effect on beta-adrenergic agonist action in stimulating lipolysis. It is hypothesized that, since toxin shows no in vitro activity, it is in some way modified in animals.


Establishment of simian sarcoma virus, type 1 (SSV-1)-transformed non-producer marmoset cell lines. Simian sarcoma virus, type 1 (SSV-1)-transformed non-producer cell lines were established by infection of normal marmoset fibroblast cells (HF) with limiting dilutions of SSV-1. Four focus-derived cell lines were identified as non-producers by assay of culture fluids for focus-forming activity and by the mixed culture cytopathogenicity test with XC cells. Further studies failed to detect production of type-C virus by 3H-uridine labelling, reverse transcriptase assay or electron microscopy. The non-producer cell lines, designated HF/SSV-NPI, IV, V, and VI, appeared morphologically transformed and cloned in soft agar with the same efficiency as HF/SSV-1 virus-producing transformed cells. Expression of either cytoplasmic or cell surface virus-related antigens was not detected by immunofluorescence or serum cytotoxicity tests. The presence of sarcoma genome in the transformed non-producer cell lines was demonstrated by rescue of focus-forming activity following superinfection with non-transforming helper virus or by cocultivation with helper-virus-producing cell lines. The SSV-1-transformed non-producer primate cells provide a useful tool for future studies.


Susceptibility to and escape from complement-mediated lysis of guinea-pig hepatoma line-10. Rabbit xenoantisera produced against diethyl-nitrosamine-induced strain-2 guinea-pig hepatoma line-10 cells (L-10) according to various immunization schedules were compared for their cytotoxicity on L-10 cells in the presence of guinea-pig complement. The highest activity was obtained with antisera (Cx-1) produced by repeated intravenous injections of living L-10 cells at high cell dosage, whereas intramuscular injections of living or glutaraldehyde-treated L-10 cells at similar frequency and cell dosage were less effective for the production of cytotoxic antibodies against L-10 cells. Intravenous injections of smaller cell doses were less effective. The cytotoxic antibody in Cx-1 antiserum was shown to be IgG by various methods including gel filtration on Sephadex G-200, ion exchange chromatography and immunoelectrophoresis of purified tumor-specific antibodies. It was concluded that L-10 cells can be lysed by guinea-pig complement and tumor-specific antibodies (IgG). Some antisera contained IgM antibodies which were not cytotoxic. A decrease in susceptibility of L-10 cells to complement-mediated lysis was observed when the cells were maintained in vivo for a long period (more than 20 passage generations). This was due to a lower density of tumor antigens on the cell surface. When tumor cells were treated with a second antibody directed against rabbit IgG or F(ab')2, cytotoxicity of Cx-1 antisera was completely abolished.


Epstein-Barr herpesvirus infection: inhibition by immunologically induced mediators with interferon-like properties. When sensitized leukocytes were re-exposed to EBV antigen or to tuberculin-purified protein derivatives, they produced lymphokines, including the migration inhibition factor, which inhibited the migration of guinea-pig peritoneal exudate cells, and other lymphokines with the characteristics of interferon, which inhibited EBV-induced transformation and EBV superinfection of target cells. Unsensitized leucocytes from sero-negative adults and from neonates did not produce lymphokines when challenged with the antigens. This indicates that cell-mediated immunity and its associated soluble mediators may be involved in the control of EBV infection and that the interferon release assay is a useful in vitro correlate for the study of cellular immunity to EBV.


Comparative studies on immunity to EBV-associated antigens in NPC patients in North America, Tunisia, France and Hong Kong. This study compared the relative antibody titers to EBV-related antigens in patients with nasopharyngeal carcinoma (NPC) and controls from a high-incidence (Hong Kong), an intermediate incidence (Tunisia), and two low-incidence (France, North America) areas to determine which of several EBV antibodies best differentiated NPC patients from controls. Antibodies measured include anti-virus capsid antigen (VCA), anti-early antigen (EA), anti-soluble antigen by complement-fixation (CF) and antibody-dependent lymphocyte cytotoxicity (ADLC). A matched pair analysis showed that significantly more NPC patients had higher VCA and EA but not CF or ADLC antibody titers than their matched cancer controls. The comparison of geometric mean titers between NPC cases and controls was more than seven-fold (816 vs 11.5) for EA antibody and more than three-fold (359.7 vs 95.4) for VCA anti-body (p less than 0.01). A two-fold difference was seen for CF antibody to soluble antigens (27.3 vs 12.9, p less than 0.01) and a three-fold difference (2657.7 vs 870.9, p less than 0.05) was observed for ADLC. Our finding of significant differences between NPC patients from four countries and their matched controls suggest that if EBV is the etiological agent of NPC in Chinese, it is quite likely to cause the majority of NPC cases in other ethnic groups living in other countries as well.


Membrane receptor stripping confirms the association between EBV receptors and complement receptors on the surface of human B lymphoma lines. Complement (C3) receptors, EBV receptors, Fc receptors, membrane IgM and beta2microglobulin (beta 2m) were individually stripped from the surface of human B lymphoma lines. The cells were subsequently tested for their ability to absorb infectious EB virus. Stripping of Fc receptors, IgM and beta2m did not reduce EBV absorption. Stripping of either EBV receptors or C3 receptors eliminated or drastically reduced EBV-absorptive capacity. The results confirm the distinctive association between EBV receptors and C3 receptors on human lymphoma cells.


Factors interfering with cellular immunological responses to the murine mammary tumor virus in tumor-bearing mice. T-cell-enriched spleen cell fractions from BALB/c mice bearing virally induced mammary tumors become unresponsive with progressive tumor growth in the lymphocyte blastogenesis test, using purified mammary tumor virus (MTV) as an antigen. reactivity can be restored by mild trypsinization or extensive washing of the cells. When reactive cells from mice immunized with MTV were incubated with the wash fluid of the initially unresponsive cells. MTV-specific blastogenesis was inhibited. Washings of normal cells had no such effect. Leukocytes from tumor-bearing mice also become unresponsive to MTV in the leukocyte adherence inhibition (LAI) assay. Trypsinization or extensive washing does not restore reactivity. However, such treatment of reactive cells from immunized animals abolishes a positive reaction in the LAI assay, using MTV as an antigen. The washings of spleen cells from tumor-bearing mice inhibit the reactivity of leukocytes from immunized animals. Sera from these mice could block the reaction in both cellular immunological tests. The inhibiting factors appeared early after the onset of tumor growth. By means of the Sepharose bead immunofluorescence assay it was established that wash fluid and sera with blocking activity contain MTV antigens as well as anti-MTV antibodies.


Dose-response curves of gamma-irradiated purine and pyrimidine powders. E.s.r. spectra have been obtained of gamma-irradiated dry purine and pyrimidine powders. Relative radical yields have been calculated and dose-response curves obtained. Relative recombination rates and recombination rates and formation rates of the radicals have been derived from the dose-response curves. The effect of crystalline structure on the dose-response curves is discussed.


E.s.r. radiation studies of erythrocyte membrane haemoglobin interaction. The dependence of the yield of free radicals in gamma-irradiated, freeze-dried erythrocyte membranes on their haemoglobin content was studied. A non-monotonous relationship was found--different from that observed in mixtures of freeze-dried membranes and haemoglobin, which suggests the existence of radiation-energy transfer between the membranes and bound haemoglobin.


Cytochrome-linked respiration in host grown M. leprae isolated from an Armadillo (Dasypus novemcinctus, L.). The bacilli were isolated from an armadillo (Dasypus novemcinctus, L.) and cytochrome systems as well as oxidation of succinate and NADH by M. leprae were studied. Cell-free extracts of M. leprae contained cytochromes of the a + a3, b, c and o type. Whole cell suspensions catalyzed the oxidation of succinate. The process was unaffected by rotenone but was markedly inhibited by thenoyltrifluoroacetone, antimycin A and cyanide. Cell-free preparations of M. leprae also oxidized NADH with oxygen as the terminal electron acceptor. Although NADH oxidation was completely inhibited by rotenone, the process was inhibited to only 50% by 5 millimols cyanide. The results indicated that complete respiratory system is present in M. leprae isolated from leprous tissues of an armadillo. The effect of inhibitors on succinate and NADH oxidations showed that the respiration in host-grown M. leprae is mediated through the cytochrome system with oxygen as the final electron acceptor.


Cytochemical contributions to differentiating GERL from the Golgi apparatus. Recent studies from our laboratory are described which deal with endocrine cells (insulinoma, beta-cells of the pancreas, thyroid epithelial cells), pancreatic exocrine cells, and hepatocytes. These emphasize the importance of the hydrolase-rich specialized region of endoplasmic reticulum, known as GERL, in secretory cells. Also reviewed in this paper are the varied molecular transformations which apparently occur in GERL in different cell types, as reported from other laboratories as well as our own. Evidence of the continuity of GERL with rough endoplasmic reticulum is presented. Two hydrolytic enzyme activities in GERL, in addition to acid phosphatase activity, are recorded. Finally, the use of cytochemical staining procedures in the study of microperoxisomes is briefly described.


Mechanics of the rib cage and diaphragm during sleep. The pattern of motion of the rib cage and abdomen/diaphragm was studied in three normal subjects during sleep. Sleep state was monitored by electroencephalograph and electrocculograph. Intercostal electromyographs (EMG's) were recorded from the second interspace parasternally. Abdominothoracic motion was monitored with magnetometers and these signals calibrated by isovolume lines either immediately before going to sleep, or if there was movement, on awakening. Respiration was recorded using a jerkin plethysmograph. In the awake subject in the supine position, the rib cage contributed 44% to the tidal volume and had essentially the same contribution in quiet sleep. However, in active or rapid eye movement sleep the rib cage contribution fell to 19% of the tidal volume. This was accompanied by a marked reduction in the intercostal EMG. With the subject in the upright position the rib cage appears to be passively driven by the diaphragm. However, the present data suggest that active contraction of the intercostal muscles is required for normal rib cage expansion in the supine position.


beta-Receptor influence on lung vasoconstrictor responses to hypoxia and humoral agents. The role of the adrenergic receptor in mediating pulmonary vascular responses to gaseous and humoral agents was investigated by use of epinephrine injections in the perfused feline pulmonary circulation. Alteration of the balance between alpha- and beta-adrenergic activity was quantified by measurement of decreasing vasoconstrictor activity to epinephrine and rising lobar tissue 3',5'-adenosine cyclic monophosphate (cAMP) levels. The increased beta-adrenergic activity thus generated was associated with marked reductions in the pulmonary vasoconstrictor responses to hypoxia, hypercapnic acidosis, and histamine, but not to serotonin. Repeated pulmonary vasodilations or increases in blood, but not pulmonary tissue, levels of cAMP induced by theophylline doses, which would not necessarily affect the beta-adrenergic activity, did not alter the pulmonary vasoconstrictor responses to hypoxia, hypercapnia, or histamine. These data support the significant role which the adrenergic system plays in mediating pulmonary vasoconstrictor responses to certain specific gaseous and humoral agents, and the specificity with which this mediation occurs serves to link hypoxia and histamine together so that the latter could serve as a mediator of the former.


Time course of muscular atrophy during immobilization of hindlimbs in rats. The hindlimbs of rats were immobilized, in plaster casts, for varying durations, and the time course for atrophy of muscle and of selected proteins in these muscles was determined. In those muscles whose lengths were at less than resting length during the fixation procedures, exponential decay to a new apparent steady state after atrophy was shown by wet and dry muscle weights and by the amounts of biuret protein, cytochrome c, and citrate synthase. The time taken to decrease to one-half of the final decrease at the new apparent steady state level was about 4-6 days for the above parameters which decayed exponentially. In contrast, the myoglobin concentration increased during atrophy and the amount of myoglobin remain unchanged during atrophy. When fixation procedures on limbs were such that muscles were stretched to lengths greater than resting length, then the onset of atrophy was delayed; indeed, in some cases muscles hypertrophied when fixed in the stretched position.


Purification and properties of poly(adenosine diphosphate ribose) synthetase. Poly(ADP-ribose) synthetase has been purified approximately 5000-fold from rat liver nuclei. The activity of the purified enzyme is absolutely dependent upon the presence of native or synthetic DNA, and the further addition of histone(s) stimulates the activity 3- to 5-fold. When the ADP-ribosylated material synthesized in the absence or presence of various histones is analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, the major product in all cases migrates between histones H1 and H3-H2B with the same RF value of 0.58 relative to the marker dye. No ADP-ribose was found to co-electrophorese with any of thehistones. The addition of histones does not affect the chain number of the poly(ADP-ribose) synthesized but does result in an increase in the average chain length of the polymer. In the presence of histones, the Km for NAD+ decreases from 80 micron to 25 micron and the Vmax doubles. These results indicate that, in the purified poly(ADP-ribose) synthetase system, histones are not ADP-robosylated but act as allosteric activators.


Crystalline alkaline form fructose-1,6-diphosphatase. A simple purification procedure and preliminary x-ray diffraction analysis. A simple and rapid procedure has been found for the preparation of higly pure alkaline form fructose-1,6-diphosphatase from turkey liver. The protein is tetrameric and is composed of subunits comprised of a 29,000 molecular weight core fragment and an associated 7,000-dalton S-peptide. Large single crystals of the protein, suitable for x-ray diffraction analysis, are produced in the course of the preparation. These are of space group R3 and have rhombohedral cell dimensions of a = 153 A and delta = 163 degrees (equivalent hexagonal dimensions a = 303 A, c = 75 A). There is one entire tetramer of 144,000 daltons in the asymmetric unit. The enzymatic properties of this enzyme appear to be very similar to those reported for the protein from other organisms, but its unique propensity to crystallize opens the way for a full three-dimensional structural analysis.


Cyclic AMP and the plasma membrane potential in Neurospora crassa. Diverse treatments, which have been shown by Slayman, C. L. (1977) in Water Relations in Membrane Transport in Plants and Animals (Jungreis, A., Hodges, T. K., Kleinzeller, A., and Schultz, S. G., eds) pp. 69-86, Academic Press, New York, to depolarize the plasma membrane of Neurospora, increase levels of adenosine 3':5'-monophosphate (cyclic AMP) in the organism. The treatments include those producing large transport fluxes of metabolizable or nonmetabolizable compounds, rapid temperature drops, and addition of agents which uncouple oxidative phosphorylation. Severe mechanical stress, which may also act to depolarize the plasma membrane, leads to increases in cyclic AMP. The maximal depolarization appears to precede the maximal cyclic AMP levels. It is proposed that the membrane depolarization produces the increased cyclic AMP levels by stimulating the plasma membrane-bound adenylate cyclase and that cyclic AMP may be important to the maintenance of membrane integrity.


Mast cell binding of neurotensin. II. Molecular conformation of neurotensin involved in the stereospecific binding to mast cell receptor sites. Systematic substitution of the natural L-amino acids in neurotensin by their D isomers reveals that the COOH-terminal portion of this tridecapeptide is required for binding to mast cell receptors: D-amino acid replacements from Pro10 through Leu13 substantially decrease that binding. Either blockage of the COOH-terminal carboxyl group as with N-methylamidation, or formation of a cyclic structure by the inclusion of a disulfide bond, a Cys2,13 substitution, markedly reduces the specific binding to mast cell receptor sites. Modifications in the NH2-terminal portion of neurotensin do not affect the binding to mast cells. However, D-Arg8 and D-Arg9 substitutions increase binding by factors of 5- to 6-fold. The hydroxyl group at position 3 or 11 is not essential for binding since Phe3 or Phe11 is equivalent to Tyr3 or Tyr11. The COOH-terminal penta- and hexapeptides are able to displace approximately 70% 125I-neurotensin relative to the intact peptide. Of 18 other biologically active peptides tested, only xenopsin, a naturally occurring COOH-terminal analog of neurotensin, and bradykinin effectively compete in the binding assay to an extent of 60 and 100%, respectively. Histamine, diphenhydramine, and noradrenaline are ineffective in this regard.


Orthopaedic treatment of hemangiomatous hypertrophy of the lower extremity. The triad of varicose veins, hypertrophy of soft tissue and bone, and cutaneous hemangiomas (The Klippel-Trenaunay syndrome) often results in major malformations. A review of eleven children treated for this condition showed that the overgrowth of the limb was a major problem in management. In three patients the altered hemodynamics resulted in cardiac complications. Venous stripping, excision of the arteriovenous fistulae, and radiation therapy should be avoided.


Abductor digiti quinti opponensplasty.

Abductor digiti quinti opponensplasty. Fifteen neurovascular pedicle transfers of the abductor digiti quinti to the base of the proximal phalanx of the thumb were performed: seven for laceration of the median nerve, seven for neurological disease, and one for congenital absence of the thenar muscles. Fourteen of the transfers were viable, and twelve gave excellent results. The three failures all had had inadequate release of contractures of the thumb. This transfer offers the following advantages over other opponensplasties: (1) an intrinsic muscle with correct amplitude and direction of pull replaces intrinsic muscles; (2) the tension of the muscle transfer is automatically correct; and (3) the appearance of the hand after this procedure is superior to that after other types of opponensplasty.


Cyclic AMP-induced morphological transformation of cells infected by temperature-sensitive mouse sarcoma virus. Expression of transformation-associated markers. Normal rat kidney (NRK) cells infected with a temperature-sensitive (ts) mutant of mouse sarcoma virus (NRK [MSV-1b]) express the transformed phenotype when grown under permissive conditions, but acquire the normal phenotype when grown under restrictive conditions. Addition of 3', 5' cyclic adenosine monophosphate (cAMP) to NRK (MSV-1b) cells grown at the restrictive temperature results in morphological transformation. To determine whether other markers associated with the transformed phenotype were coordinately expressed after cAMP exposure, concanavalin A (Con A) agglutinability, hexose transport rate, and incorporation of radioactively labeled fucose into fucolipid III and fucolipid IV (FL III and FL IV ) of the cells were examined. NRK cells transformed by wild-type MSV or NRK(MSV- 1b) grown under permissive conditions were agglutinated by low concentrations of Con A and exhibited relatively high maximal agglutination levels which were specifically inhibited by alpha-methyl-D-mannoside. In contrast, NRK (MSV-1b) cells grown under restrictive conditions were weakly agglutinated by Con A and exhibited reduced maximal agglutination levels, similar to uninfected NRK cells. Treatment of NRK (MSV-1b) cells at the restrictive temperature with cAMP resulted in morphological transformation and a change in the pattern of incorporation of labeled fucose inot FL III and FL IV to one comparable to that of NRK (MSV-1b) cells at the permissive temperature or to NRK cells transformed by wild-type MSV. In contrast, cAMP treatment resulted in no increase in Con A agglutinability or 2 deoxy-D- [(3)H]glucose transport relative to mock treated cultures. The results demonstrate that cAMP-induced morphological transformation and altered fucolipid composition of NRK (MSV-1b) cells are not correlated with alterations in hexose transport rate or Con A agglutinability.


Iontophoretic release of cyclic AMP and dispersion of melanosomes within a single melanophore. Selective dispersion of melanosomes was often observed after iontophoretic injection of cyclic adenosine monophosphate (AMP) from a glass microelectrode positioned in a target melanophore in frog skin (as viewed from above through a microscope), with other melanophores in the field serving as controls. Because the skin has orderly arrays of several types of closely spaced cells, it is probable that at times the microelectrode also impales cells other than melanophores. When cyclic AMP injection inside a cell resulted in dispersion of melanosomes from a perinuclear position into dendritic processes, the onset of dispersion was relatively rapid, in many cases less than 4 min (mean time of onset, 5.3 +/- 2.9 [SD] min). A much slower dispersion (mean time of onset, 19.0 +/- 5.0 min) of melanosomes was observed when the microelectrode was positioned adjacent to a melanophore, and much larger quantities of cyclic AMP were released. In addition, no changes were observed for injections of 5'-AMP or cyclic guanosine monophosphate (GMP) through electrodes positioned inside or adjacent to melanophores. Potential measurements showed that after impaling a clell, a constant transmembrane potential could often be recorded over many minutes, indicating that the membrane tends to seal around the microelectrode. The results indicate that cyclic AMP acts more rapidly on the inside of a cell than when applied outside a cell and allowed to diffuse through the plasma membrane. This study introduces a model system whereby the properties of the plasma membrane and melanocyte-stimulating hormone (MSH) receptors can be studies within a single target cell.


Characterization of the myosin-phosphorylating system in normal murine astrocytes and derivative sv40 wild-type and A-mutant transformant. Myosin and myosin light-chain kinase have been isolated and characterized from small quantities of normal and SV40-transformed, murine astrocytic neuroglial cells in culture and from intact normal mouse brain. Sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis of the astrocyte myosins revealed a heavy chain of 200,000 daltons and two light chains of 20,000 and 15,000 daltons. These myosins are similar to other cytyplasmic myosins. The astrocyte 20,000-dalton light chain can be phosphorylated by an endogenous myosin light-chain kinase which has properties similar to those of the myosin light-chain kinase found in human platelets. No differences were detected in either the astrocyte myosins or myosin light-chain kinases between (a) the normal and transformed cells, (b) the transformed cells grown at the permissive and nonpermissive temperatures, or (c) the SV40 wild-type and A-mutant transformants.


Locomotory behavior, contact inhibition and pattern formation of 3T3 and polyoma virus-transformed 3T3 cells in culture. The social behavior of 3T3 cells and their polynoma virus-transformed derivative (Py3T3 cells) was examined by time-lapse cinemicrography in order to determine what factors are responsible for the marked differences in the patterns formed by the two cell lines in culture. Contrary to expectations, both cell types have been found to exhibit contact inhibition of cell locomotion. Therefore, the tendency of 3T3 cells to form monolayers and of Py3T3 cells to form crisscrossed multilayers cannot be explained on the basis of the presence versus the absence of contact inhibition. Morevover, with the exception of cell division control, the social behavior of the two cell types is qualitively similar. Both exhibit cell underlapping and, after contact between lamelliopodia, both show inhibition of locomotory activity and adhesion formation. Neither cell type was observed to migrate over the surface of another cell. The two cell types do show quantitative differences in the frequency of underlapping, the frequency with which contact results in inhibition of locomotion, and the proportion of the cell margin that adheres to the substratum. The increased frequency pf Py3T3 underlapping is correlated with the reduced frequency of substratum adhesions, which in turn favors underlapping. On the basis of these observations, it is concluded that the differences in culture patterns are the result of differences in the shapes of the individual cells, such that underlapping, and hence crisscrossing, is favored in Py3T3 cell interactions and discouraged in 3T3 cells.


Intracellular protein degradation in growing, in density-inhibited, and in serum-restricted fibroblast cultures. Exponentially growing Balb/3T3 mouse fibroblasts contain protein populations with slow and fast turnover. These two stability classes were labelled selectively with 3H-leucine. The intracellular degradation of the proteins was then followed as the release into the medium of radioactive leucine. The degradation rate of both stability classes of protein is increased by about 55% in cultures whose growth is inhibited by high cell density. Serum-deprivation, which also halts cell growth, accelerates protein breakdown to a smaller extent, the increases for relatively stable and unstable proteins being 30% and 13%, respectively. The density-dependent increase in protein breakdown is also found in BHK21 cells but not in chick fibroblasts. Protein degradation in Balb/3T3 cells transformed by simian virus 40 is affected by serum-deprivation but not by cell density. The proteins which are relatively stable during growth were shown to become less stable in density-inhibited or serum-deprived cultures, and vice versa. Cycloheximide inhibits degradation to a variable extent. Dibutyryl adenosine-3',5'-cyclic monophosphate has no effect on the protein degradation under the conditions investigated here.


Coexpression of mutant and wild type protein kinase in lymphoma cells resistant to dibutyryl cyclic AMP. A mutant clone resistant to dibutyryl cyclic AMP was isolated from S49 mouse lymphoma cells. The mutant expressed a form of cyclic AMP-dependent protein kinase distinguishable from wild type kinase by its decreased sensitivity to activation by cyclic AMP and its increased thermal lability. Hybrids formed between mutant and wild type cells were resistant to dibutyryl cyclic AMP and expressed both mutant and wild type activities in about equal amount. The parent mutant cells also appeared to express wild type kinase activity, but at a lower level. We conclude that wild type S49 cells have and express two identical alleles for the regulatory subunit of protein kinase, one of which has undergone mutation in the mutant cells.


On the regulation of DNA synthesis in a line of adrenocortical tumor cells: effect of serum, adrenocorticotropin and pituitary factors. Y-1 adrenal cells responded to serum step down by a several fold decrease in DNA synthesis. Serum starved cells resumed DNA synthesis upon serum step up. ACTH and cAMP inhibited DNA synthesis both at low and high serum concentrations, a fact previously known. Pituitary, brain and liver crude extracts stimulated DNA synthesis in serum starved cells. Purified pituitary factors preparations contained two activities: one specific for Y-1 cells and another active with both fibroblasts and Y-1 cells. The kinetics of restimulation of DNA synthesis by serum and pituitary factors was studied. DNA synthesis restimulation occurred after a lag of 11 hours. This lag did not vary irrespective of the type of stimulator or its concentration. Cells entered S phase continuously at a rate which increased with increasing concentrations of the stimulator. Cells became refractory to the inhibitory action of ACTH five hours before entering S phase. The implications of these data to the understanding of cell growth control are considered.


Transformation of human cystinotic fibroblasts by SV40: characteristics of transformed cells with limited and unlimited growth potential. Human skin fibroblasts derived from patients with nephropathic cystinosis were transformed with SV40 virions, cloned and permitted to enter the degenerative stage of growth termed "crisis," characteristic of SV40 transformed human cells. Nephropathic cystinosis is an autosomal recessively inherited metabolic disorder resulting in the intracellular accumulation of the amino acid cystine. A transformed cystinotic cell line which was recovered from the crisis stage was indistinguishable from its transformed precrisis parental cell strain in growth rate in media containing either 1% or 10% serum, cloning efficiency on plastic, in semisolid media, or upon confluent monolayers of normal skin fibroblasts, expression of SV40 T antigen, or production of virus. However, the modal DNA content of the recovered postcrisis transformed cystinotic cell line was different from that of the cloned parental precrisis transformed cell strain, suggesting that the postcrisis line was derived from a small subpopulation of the precrisis strain. The DNA content of the established cystinotic cell line continued to be unstable during subsequent subculturing and gave rise to subclones with both more and less DNA per cell. This line now has an apparently infinite growth potential and still has the hallmark of the cystinotic parental line, the storage of abnormally large amounts of intracellular nonprotein cystine.


Resistance of teratocarcinoma stem cells to infection with simian virus 40: early events. Multipotential stem cells of a murine teratocarcinoma are resistant to typical infection with either polyoma virus (PV) or Simian virus 40 (SV40). Differentiated progeny of the stem cells are susceptible to infection in a manner identical to other mouse somatic cells, i.e., they are permissive for PV and nonpermissive for SV40. The early interactions between the stem cells (embryonal carcinoma or EC cells) and SV40 and PV were studied. Virions adsorbed to and penetrated into the cytoplasm and nucleus of EC cells, but did not induce expression of T antigen in the EC nuclei. Purified SV40 DNA was capable of inducing T antigen in differentiated teratocarcinoma cells but not in EC cells. Virus could not be rescued from EC cells previously exposed to SV40. The resistance of the stem cells to infection apparently involves a block in the infectious cycle after adsorption and penetration but before T antigen induction.


Estriol conjugates in human breast cyst fluid and in serum of premenopausal women. A sensitive method was developed for the assay of the four major estriol (E3) conjugates in human breast cyst fluid and in serum during the menstrual cycle. In the cyst fluid, estriol-3-sulfate (E3-3S) was found in each of ten samples, the concentrations ranging from 240-4310 pg/ml. Estriol-16-glucosiduronate (E3-16G) was detected in six samples at levels of 19-153 pg/ml; estriol-3-glucosiduronate (E3-3G) in five samples, 13-79 pg/ml and estriol-3-sulfate-16-glucosiduronate (E3-3S-16G) in four samples, 28-152 pg/ml. Unconjugated estriol was found in three of the ten cases (12-30 pg/ml). Serum samples obtained in the follicular and luteal phases of the cycle from eight different subjects were assayed in the same way. There was considerable variation between subjects and many values were indistinguishable from zero. But preliminary data suggest that E3-3G is the predominant E3 conjugate in the serum and E3-3S-16G is quantitatively least important. It appears that E3 conjugates in the cyst fluid are not derived from the blood directly, but are produced locally from precursors which have not been identified.


Reversible gonadotropin deficiency in male Cushing's disease. Twelve adult males with documented active Cushing's disease were studied. Mean plasma testosterone (T) was significantly decreased: 1.8 +/- 0.3 (SEM) ng/ml (N=6.8 +/- 0.5); gonadotropin measurements in 8 patients, in basal conditions and under LH-RH iv, showed a significant decrease in both FSH and LH. A further study of 11 patients in remission of Cushing's disease indicated a significant increase in plasma T and gonadotropins up to the normal range. One patient with an initial low T value had a normalized T while in remission, then a dramatic decrease when the disease relapsed. We conclude: a hypogonadotropic hypogonadism is found in male Cushing's disease; it disappears as early as hypercortisolism is suppressed. Some possible mechanisms are discussed.


The variations of plasma corticosterone/cortisol ratios following ACTH stimulation or dexamethasone administration in normal men. A radioimmunoassay for human plasma corticosterone has been developed. Plasma corticosterone increased 4.83 times as much as basal value at 60 min after an im injection of 0.25 mg synthetic beta1-24 ACTH (Cortrosyn) in normal subjects, whereas plasma cortisol increased 2.12 times as much at 60 min. And basal corticosterone/cortisol ratio of 0.053 +/- 0.017 increased to 0.116 +/- 0.022 (P less than 0.001) after ACTH. This might be mainly due to a larger increment of corticosterone than that of cortisol after ACTH. Corticosterone decreased to 36.7% of basal value at 4 h after 1 mg dexamethasone administration in normal subjects, whereas cortisol decreased to 13.9% of basal value at 4 h. The basal corticosterone/cortisol ratio of 0.059 +/- 0.020 increased to 0.137 +/- 0.055 (0.001 less than P less than 0.01) after dexamethasone administration. This may have been mainly due to a more effective suppression of cortisol than that of corticosterone after dexamethasone.


Center for disease control diagnostic immunology proficiency testing program results for 1976. Over 900 laboratories participated in the Diagnostic Immunology portion of the 1976 Proficiency Testing Program, which was provided by the Center of Disease Control under the authority of the Clinical Laboratories Improvement Act of 1967. One hundred specimens prepared by the Center for Disease Control for analysis were distributed on a quarterly schedule or in special surveys. Feedback from participating laboratories included over 37,500 qualitative and 33,000 quantitative responses, which were analyzed to determine individual laboratory proficiency levels. In addition, information supplied by participants in each survey helped to delineate trends in  testing protocols. The specimens chosen for analysis called for a broad range of tests commonly performed in diagnostic immunology laboratories, including those for rubella antibodies, hepatitis B surface antigen, bacterial antibodies, rheumatoid factor, immunoglobulins and other serum-specific proteins, and carcinoembryonic antigen. A summary of the data analysis is provided so that the laboratories can improve their overall performance levels.


Enrichment medium for the isolation of Bordetella. The development of a specimen collection and transport medium outfit for the rapid laboratory diagnosis of whoping cough is described. The transport medium consisted of a semisolid agar containing charcoal, cephalexin, and defibrinated horse blood. It was also found to be an excellent enrichment medium for the selective isolation of Bordetella pertussis and B. parapertussis from scantily populated specimens. The investigation of 3,237 specimens that yielded 1,419 positive isolates of Bordetella, including 86 B. parapertussis, during a 20-month period is presented. A total of 3,076 specimens were processed in the laboratory by using the enrichment medium in addition to the routine procedure. Of these specimens, 757 were submitted in our medium, from which 137 (18%) were positive. Of the 567 specimens received in Amies transport medium, 290 (51%) positive cultures were obtained by the enrichment method only and not by primary culture.


Characterization of high density lipoproteins in patients heterozygous for Tangier disease. In this study a large family group affectd with Tangier disease has been investigated. Besides two homozygous propositi, several heterozygous patients have been identified on the basis of quantitative measurements of high density lipoproteins and their constitutive polypeptides. By a variety of quantitative immunological methods, such as one-dimensional Laurell eletrophoresis, two-dimensional immunoelectrophoresis, and double-antibody radioimmunoassay, the total amount of apoprotein A-I and apoprotein A-I contained in the serum of heterozygous patients and the distribution of these A apoproteins among serum lipoproteins have been determined. The molar ration of apoprotein A-I and apoprotein A-II contained in high density lipoproteins of heterozygous patients did not significantly differ from that of control preparations, although the total mass of high density lipoproteins was reduced by approximately 50%. The elution profile of high density lipoproteins from agarose columns and their morphological appearance, as ascertained by electron microscopy, were similar to control preparations. In addition to the quantitative alterations of serum lipoproteins, lipid storage in histiocytes of the rectal mucosa obtained from heterozygous patients has been documented. It is concluded that patients heterozygous for Tangier disease have normal high density lipoproteins in circulation, the total mass of which is reduced by approximately 50%.


Collagen-mediated platelet aggregation. Evidence for multivalent interactions of intermediate specificity between collagen and platelets. We have shown previously that periodate oxidation of collagen carbohydrate does not affect its ability to aggregate platelets. We now describe an additional characterization of periodate-modified collagen which demonstrates that collagen devoid of intact carbohydrate is fully capable of fibril formation, and we confirm its capacity to initiate platelet aggregation. Furthermore, we demonstrate that the platelet aggregating abilities of Types I, II, and III fibrillar collagen are quite similar despite differences in carbohydrate content and amino acid sequence. We also demonstrate that monomeric, pepsin-solubilized Type I human collagen is ineffective inhibiting aggregation by performed fibrils derived from the same molecule, thus establishing that the affinity of platelets for collagen depends upon prior polymerization of collagen. We interpret these and other findings to demonstrate that the hydroxylysyl glycoside regions of collagen are not highly specific sites involved in platelet-collagen interactions leading to "physiological" aggregation, and that the possibility must be considered that multiple interactions involving collagen sites of comparatively low structural specificity may be the initiating events in release of platelet ADP and the ensuing aggregation.


The biochemical consequences of hypoxia. The various phases of energy production have been described. These include glycolysis which is unique in its ability to produce ATP anaerobically, the tricarboxylic acid cycle with its major contribution to ATP production coming through the generation of NADH, and the cytochrome system at which reducing equivalents are converted to water, the released energy being incorporated into high-energy phosphates. The regulation of these pathways has been briefly described and the importance of the small amount of ATP generated anaerobically emphasized. The adaptation of muscle to periods of hypoxia through the presence of myoglobin, creatine phosphate and large amounts of glycogen is then discussed. The role of pH in limiting anaerobic glycolysis in muscle and the importance of the circulation in providing oxygen for exercising muscle are outlined. The effects of hypoxia on certain other tissues such as liver and brain have been detailed and finally methods for assessment of tissue hypoxia in man such as the measurement of the lactate:pyruvate ratio in blood are presented.


A hypothesis concerning the structure of cAMP-and cGMP-dependent protein kinases. cAMP-and cGMP-dependent protein kinases have been purified. Each enzyme demonstrates high specificity and affinity for the cyclic nucleotide with binding of two moles of nucleotide per holoenzyme and each enzyme is an ATP: phosphotransferase. The holoenzymes have similar molecular weights and demonstrate similar molecular asymmetry. A structural model relating the two enzymes is proposed. cGMP-dependent protein kinase is proposed to be a dimer composed of two identical protomers in isologous association with the chains arranged in anti-parallel fashion. cAMP-dependent protein kinase is proposed to have a similar structure with a dyad axis of symmetry but with a discontinuity in each chain. These structures account for the differing mechanisms of cyclic nucleotide activation of the two enzymes.


Synchronous generation of ovarian hCG binding sites and LH-sensitive adenylate cyclase in immature rats following treatment with pregnant mare serum gonadotropin. A concomitant increase in the activity of LH-senstive adenylate cyclase and in the number of LH/hCG binding sites was induced in ovaries of immature rats upon administration of pregnant mare serum gonadotropin (PMSG), a hormone preparation known to have predominantly follicle stimulation (FSH-like) activity. When an optimal dose of PMSG (15 i.u./rat) was administered to 25-day-old rats, specific activity of LH-dependent adenylate cyclase and the number of binding sites for LH/hCG per mg protein remained unchanged during the first 24h, but 48h after injection a 2-to 4-fold increase in both parameters was observed. By contrast, there was no change in basal adenylate cyclase activity or in the response of the enzyme to the stimulatory action of guanosine-5'-(beta gamma-imino) triphosphate (Gpp (NH)p), GTP, or NaF. Specific activity of succinate cytochrome c reductase, glucose-6-phosphatase and 5'-nucleotidase were found to be unaffected by the hormonal pretreatment, although total protein determined in these homogenates increased 3-fold in the course of this treatment. It is inferred that during follicular maturation, FSH enhances the responsiveness of ovarian adenylate cyclase to LH by stimulating the insertion of LH/hCG-receptors into the cell membrane.


Cyclic AMP-protein-DNA complex formation in mammalian cell-free systems. A cytoplasmic protein fraction from KB and Chinese hamster ovary cells (CHO-Kl) was shown to bind in vitro to cAMP and subsequently to DNA-cellulose. This protein complex was not found in DE-52 purified CHO-K1 cAMP-dependent protein kinases. The complex appeared to exist as a small fraction of the total cAMP binding proteins, preferred native to denatured DNA and exhibited multiple sedimentation coefficients in glycerol gradients. This complex, after elution from the DNA cellulose column, was shown to have bound specifically to [3H]-cAMP which could be displaced by non-radioactive cAMP in competitive binding assays.


Lack of effect on cyclic GMP content of cells treated with mycophenolic acid. Mycophenolic acid, an oncolytic agent and a known inhibitor of guanine ribonucleotide synthesis, has proven to be an effective drug against psoriasis. With reports of greater guantities of c-GMP in psoriatic tissues than in normal tissue, and with the correlation of c-GMP content of cells to proliferation, the effect of mycophenolic acid on cellular c-GMP was investigated. When HeLa, green monkey BSC-1, and mouse L-cells were treated with inhibitory concentrations of mycophenolic acid, no decrease in c-GMP was observed from that of untreated cells. Though mycophenolic acid inhibits guanine ribonucleotide synthesis, this inhibition does not extend to c-GMP synthesis. The inhibition of proliferation of cells by mycophenolic acid then does not include the inhibition of synthesis of c-GMP, but apparently resides solely in limiting the guanylate necessary for nucleic acid synthesis.


Phosphorylation of casein by the lactating mammary gland: a review. The lactating mammary gland synthesizes and secretes large amounts of phosphoproteins that mainly are associated with the casein fraction of milk. The free amino acids and inorganic phosphate of blood serve as building materials for casein, and the final product appears in milk as a colloidal-sized particle, the casein micelle. According to our present concept, the biosynthesis of casein occurs in two steps: synthesis of the polypeptide chain, followed by phosphate addition. Phosphate groups are transferred to the nascent casein by a protein kinase localized in the Golgi apparatus. The enzyme uses adenosine 5'-triphosphate as the phosphate donor and requires divalent cations. Neighboring amino acids may be important in determining which serine residues in casein are phosphorylated. This review discusses historical and current research on the phosphorylation of casein.


Effects of polybrominated biphenyls on health and performance of pregnant Holstein heifers. The toxicity of a commercial blend of polybrominated biphenyls was determined in 24 pregnant Holstein heifers that were allotted randomly to one of four experimental groups given 0, .25, 250, or 25,000 mg/day of fire-Master BP-6. The polybrominated biphenyls were mixed with finely ground corn and given by bolus for 60 days or until the animal became moribund. Average body weight of heifers at onset of experiment was 381 kg. No clinical signs of toxicosis were evident in heifers fed 0, .25 or 250 mg/day. Toxicosis was induced in heifers fed 25,000 mg/day resulting in reduced dry matter intake, body weight, heart rate, and respiration rate. Clinical signs were anorexia, emaciation, dehydration, excessive lacrimation and salivation, diarrhea, depression, and abortion or fetal death. All heifers fed 25,000 mg/day became moribund within 33 to 66 days.


Myocardial uptake of Tc-99m skeletal agents in the rat after experimental induction of microscopic foci of injury. The cardiac uptake of Tc-99m tagged skeletal agents was studied after myocardial injury produced by subcutaneous catecholamine injection and random foot-shock stress. Rats stressed for 2 hr developed microfocal myocardial injury, without gross change, whereas those stressed for 12 hr sustained more confluent and sometimes grossly visible damage. Tc-99m MDP and Tc-99m PPi concentrations in these hearts were significantly above control (undamaged) heart levels, producing positive gamma-camera images. Subcutaneous epinephrine injections resulted in grossly visible lesions, with tracer concentrations higher than those previously reported in vasoocclusive infarcts. We postulate that the stress-induced scattered microfocal lesions may accumulate radiopharmaceutical on a per-gram basis in the same way as the larger catecholamine-induced lesions, since tracer delivery to the injured areas in each case is probably less impeded than in frankly vasoocclusive models. Such microfoci, then, could provide an explanation for some of the "false positive" myocardial scans observed clinically.


Cochlear micromechanics--a mechanism for transforming mechanical to neural tuning within the cochlea. A linear mathematical model is proposed which will account for the differences observed between mechanically measured data of Rhode (1971) for basilar membrane motion, and the responses of neural tuning curves (Kiang et al., 1974). We show that theoretical tuning curves may be derived from mechanical responses by forming the difference between the pressure across the basilar membrane and its displacement. Some ramifications of this proposal are discussed. We then propose a hypothetical physical model which could perform such a function.


The in vitro effect of toluene diisocyanate on lymphocyte cyclic adenosine monophosphate production by isoproterenol, prostaglandin, and histamine: a possible mode of action. Toluene diisocyanate (TDI) significantly inhibits the rise in intracellular cyclic 3',5'-adenosine monophosphate (cAMP) that follows in vitro incubation of human lymphocytes with 6.7 x 10(-3) M isoproterenol and 1 x 10(-6) M prostagladin E1 (p less than 0.05). TDI has no significant effect on th production of lymphocyte cAMP following incubation with histamine (1 x 10(-3) M). The inhibitory action of TDI is greatest at a concentration of 3.3 x 10(-4) M and diminishes as the TDI concentration is increased or decreased. TDI also caused four- to fivefold stimulation of lymphocyte cAMP, an effect that is maximal at 1 x 10(-3) M, a concentration which has no significant inhibitory effect on stimulation of cAMP by isoproterenol or prostagladin E1. Conversely, 3.3 x 10(-4) M TDI, which inhibits cAMP production by isoproterenol and prostaglandin, has little stimulatory effect itself on cAMP production. This evidence suggests that TDI might induce obstructive airways disease through pharmacologic mechanisms and that TDI may be acting as a partial agonist.


Drug-induced changes of adenylate cyclase activity in cells from asthmatic and nonasthmatic subjects. Monolayers of adherent mononuclear leukocytes prepared from normal subjects and asthmatic children whose bronchodilater therapy did not include sympathomimetic drugs, respond to relatively low concentrations of isoproterenol (ISO) and prostaglandin E1 with increased intracellular cyclic adenosine monophosphate (cAMP) levels. The magnitude of in vitro responses to ISO is decreased by previous contact of the cells with ISO or other, orally effective, adrenergic drugs. The desensitization is rapid, concentration- and time-dependent, and is readily reversible after removal of the desensitizing drug. The phenomenon exhibits pharmacologic specificity. Cells in which the response to restimulation with ISO was decreased exhibited full sensitivity to prostaglandin E1. No differences in these behaviors were detected in cells from normal or asthmatic subjects. The results suggest that earlier observations reporting decreased responses to beta adrenergic stimulation in asthmatics may have been due to the treatment of these patients with sympathomimetic agents and not caused by a disease-related beta adrenergic receptor dysfunction.


Depression in the aged: theoretical concepts. The etiology of depression in the aged is discussed from the viewpoint of biologic, psychodynamic, genetic and sociocultural concepts. The chronic neurotic depression of the aged seems to differ from that of younger age groups. The understanding of geriatric depression requires a multidisciplinary approach.


Surgical management of brachial plexus injuries. Exploration of the brachial plexus was done as an elective procedure in 56 patients with complete or partial lesions. The indications were based on clinical findings, a Tinel-Hoffman sign indicating that at least one root was available for direct repair, or a cessation of signs of progressive recovery. In young patients with supraganglionic lesions and evidence of root avulsion, nerve grafts from intercostals to various portions of the plexus were done. Evaluation of the results of motor functions showed that 38 of 54 (70 percent) recovered a useful motor function in at least one important area. There were two postoperative hematomas leading to delayed healing and failure of nerve recovery. Two patients had temporary loss of power in uninvolved muscles but both recovered satisfactory function. Only one patient had a persistent pain syndrome. Two failures were due to the late operation (19 months after injury) and one because of a 15 cm. which was grafted.


Peripheral neuropathy--an unusual complication of meningococcemia. A case of meningococcemia with a persisting peripheral neuropathy and gangrene of the fingers is presented. These unusual complications are thought to be related to the diffuse intravascular coagulation syndrome.


Analysis of murine C-type virus structural proteins by rocket and crossed immunoelectrophoresis. We have examined the structural proteins of Rauscher murine leukemia virus (R-MuLV) by means of rocket immunoelectrophoresis and crossed immunoelectrophoresis, using polyspecific antisera to Tween/ether-disrupted purified R-MuLV. Fifteen different precipitation lines were recognized in virus lysates. Using five reference antisera prepared to purified R-MuLV-structural proteins, the precipitation lines of p 10, p 15, p 30 and gp 69/70 were identified. These techniques, although less sensitive than radioimmunoassay, have several advantages, such as simplicity, direct control of precipitation reactions and possibility of using crude antigen preparations.


A rapid microassay for detecting antibodies against poliovirus based on [14C]thymidine uptake of treated cell cultures. DNA synthesis of mammalian cells propagated in microplates can easily be measured if cell cultures incubated with [14C]thymidine are harvested on the glass fibre filters by a semiautomatic harvesting technique. Soon after infection with poliovirus, [14C]thymidine uptake of U cells (established, human amniotic cell line) is inhibited. This inhibition can be prevented by previous virus neutralization with antibody. Based on this effect a rapid, precise assay method was set up to determine neutralizing antibody titres against poliovirus. There was a good correlation between titres obtained by this assay and those obtained by 50% end point titrations in cytopathogenic effect inhibition assays.


Quantitation and immunocytochemical localization of human skin collagenase in basal cell carcinoma. Human skin collagenase was quantitated by radioimmunoassay in 21 basal cell carcinomas. Immunoreactive collagenase protein was found to be approximately 2-fold greater in extracts of these tumors than in extracts of normal skin, suggesting that this enzyme may be important in the pathogenesis of soft tissue destruction in vivo. To further define the role of collagenase in such destruction, immunofluorescent staining with specific antiserum to human skin collagenase was used to localize collagenase in the basal cell carcinomas. The enzyme was found only in the stromal elements surrounding the tumor islands. No staining of the epithelial components of the basal cell carcinomas was found. These findings suggest that the normal connective tissue elements may have been stimulated to produce an increased amount of collagenase and emphasize the importance of epithelial-stromal interaction in soft tissue invasiveness.


Characterization of cryoprecipitates in pemphigus: demonstration of pemphigus antibody activity in cryoprecipitates using the immunofluorescent technique. Cryoproteins observed in patients with pemphigus have been shown to contain IgG, complement components, and other serum proteins. The IgG moiety possessed pemphigus antibody activity and contained kappa and lambda light chains. Cryoproteins appeared in the clinically active stage and disappeared subsequent to clinical improvement. These observations indicate that immune complexes may exist in the sera of patients with pemphigus.


Epidermal adenylate cyclase: stimulation of the histamine (H2) receptor by tolazoline. Tolazoline (2-benzyl-2-imidazoline) activated adenylate cyclase in pig epidermal slices resulting in the accumulation of cyclic AMP. This effect was highly potentiated by the addition of the cyclic AMP-phosphodiesterase inhibitor, theophylline. Specific histamine (H2) receptor inhibitors (metiamide and cimetidine) completely blocked the tolazoline activation of adenylate cylase. At low concentrations (10-100 micrometer), a histamine (H1) receptor inhibitor (diphenhydramine) and a beta-adrenergic blocker (propranolol) did not inhibit this effect. The stimulation of cyclic AMP formation by the combination of tolazoline and histamine was about the same as the stimulation by histamine alone (nonadditive), whereas the stimulatory effects by tolazoline and epinephrine were additive. These data suggest that tolazoline, an alpha-adrenergic blocker, also activates adenylate cyclase at the histamine (H2) receptor site which is distinct from the beta-adrenergic receptor site. Another alpha-adrenergic blocker, phentolamine, did not have this effect.


Murine cytomegalovirus infection of epithelial cells in mouse tracheal ring organ culture. A differentiated tissue model was used to study the course of cytomegalovirus infection in respiratory epithelium in vitro. Mouse tracheal rings in organ culture were infected with murine cytomegalovirus. Infectious virus in the medium reached average titers of 10(5.5) plaque-forming units/ml by day 17. Infected epithelial cells on the rings contained viral antigen as demonstrated by immunofluorescence staining; light microscopy of these cells revealed enlarged nuclei, cytoplasmic vacuolization, and nuclear and cytoplasmic inclusions characteristic of infection with cytomegalo-virus. Development of viral particles, as visualized by electron microscopy, was similar to that observed in other types of cells.


Inhibition of the biologic effects of endotoxin on neutrophils by polymyxin B sulfate. Polymyxin B sulfate diminished the endotoxin-mediated release of human blood neutrophil lysosomal enzymes. Similarly, this antibiotic reduced the endotoxin-induced increase in the neutrophil hexose monophosphate pathway activity as measured by the release of 14CO2 from [l-14C]glucose. Although these effects were seen with therapeutically attainable levels of polymyxin B sulfate, they could not be demonstrated when the cell-endotoxin interaction preceded treatment with polymyxin B sulfate.


Free amino acids in plasma during experimental infection of mice with the MHV-3 strain of mouse hepatitis virus. The concentrations of total free amino acids, single free amino acids, urea, and ammonia were determined in plasma of mice during experimental infection with the MHV-3 strain of mouse hepatitis virus. Analysis of free amino acids was done by ion-exchange resin chromatography under conditions that allowed the use of a single chromatographic column, separation of glutamine and asparagine, and an accelerated rate of chromatography. The results showed that as early as 6 hr after infection there was a decrease in the concentration of several free amino acids as well as in the total concentration of free amino acids in plasma. For most of the amino acids the decrease persisted until 48 hr. Only at 72 hr, during severe cytolysis, did the concentration of amino acids increase significantly. It is suggested that the decrease during the initial phases of the infection may be due to a thermolabile factor that is produced by circulating leukocytes and that effects a flow of free amino acids from the plasma toward the liver. The final increase in concentration of several free amino acids reflects the cytolytic damage to the liver caused by the virus.


Antibody to varicella-zoster virus-induced membrane antigen: immunofluorescence assay using monodisperse glutaraldehyde-fixed target cells. A sensitive and reproducible technique was developed for the detection of antibody to varicella-zoster virus-induced membrane antigen (VZMA) by immunofluorescence. Controlled trypsinization and glutaraldehyde fixation were employed to prepare a monodisperse suspension of noninfectious VZMA-positive target cells that can be stored indefinitely at -72 degrees C. A microtiter immunofluorescence assay utilizing these target cells was shown to provide a sensitive and specific means for the detection and quantitation of antibody to varicella-zoster virus. The properties of the target cell preparation and the characteristics of the assay make practical the rapid assessment of immunity to the varicella-zoster virus.


Prospective study of prevalence, incidence, and source of herpesvirus infections in patients with renal allografts. The prevalence, incidence, and source of infections with different types of herpesviruses were determined prospectively for 25 persons undergoing hemodialysis, 30 allograft recipients, and 16 kidney donors. The prevalence of prior infections with cytomegalovirus (CMV), herpes simplex virus (HSV), and Epstein-Barr virus (EBV) was high (72%-100%) and was similar for healthy persons and those with renal failure. The incidence of infections in patients undergoing hemodialysis was no greater than that before dialysis. In allograft recipients, the incidence of infection with CMV was 73%; HSV, 57%; EBV, 30%; and varicella-zoster virus (clinical), 7%. Ninety-seven percent of the patients developed an infection with one or more herpesviruses. Transfusions, hemodialysis, the allograft, and hospital environment were not significant sources in transmission. Uremia and splenectomy were unimportant in the reactivation of infection. Immunosuppressive drugs possibly algmented by a graft rejection response account for the high incidence of recrudescent infections with CMV and HSV.


Isolation of simian virus 40 from rhesus monkeys (Macaca mulatta) with spontaneous progressive multifocal leukoencephalopathy. Isolates of virus from the brain tissue of two naturally occurring cases of progressive multifocal leukoencephalopathy in rhesus monkeys (Macaca mulatta) have been characterized. Both isolates were demonstrated to be simian virus 40 (SV40) by serological tests and analysis of cleavage fragments of viral deoxyribonucleic acid produced by restriction endonuclease from Haemophilus influenzae. SV40 virions and the nonvirion T antigen were demonstrated in the brain lesions of one monkey by the fluorescent antibody staining technique. SV40 was not demonstrated in the brain of normal rhesus monkeys from the same colony with use of the same methods of viral isolation or demonstration of antigen.


Enzyme immunoassays for viral diseases.

Enzyme immunoassays for viral diseases. Microplate enzyme-linked immunosorbent assays (ELISA) for viral diseases were investigated with special reference to rubella. Standardization of the carrier plates, antigens, conjugates, and substrate was found to be essential. The ELISA results were compared with results of hemagglutination-inhibition tests for rubella, and ELISA was used in an epidemiologic study. Antibodies to cytomegalovirus, measles, adenovirus, coxsackieviruses, and herpesviruses were also assayed by ELISA. ELISA has veterinary applications in detection of respiratory syncytial virus and Newcastle disease virus, and it has been used for assay of plant viruses.


Detection of antibodies specific for herpes simplex virus in human sera by the enzyme-linked immunosorbent assay. An enzyme-linked immunosorbent assay (ELISA) was used for the detection in human sera of antibody to herpes simplex virus antigens. Development and standardization of the assay suggested that antigenic purity, temperature of reactions, concentration of enzyme-conjugated antiglobulin, and concentrations of test sera are all critical parameters of a successful ELISA procedure. When ELISA titers of 30 human sera were compared to micro-complement fixation titers of the same sera, a significant degree of correlation was observed, but quantitatively ELISA was found to be up to 200 times more sensitive. The sensitivity of the assay and its adaptability to automation should provide an additional method for study or diagnosis of infection with herpes simplex virus.


Quantitative, semiautomated, enzyme-linked immunosorbent assay for viral antibodies. A quantitative, semiautomated, enzyme-linked immunosorbent assay for viral antibodies was developed. The quantitation was based on the comparison of the antibody activity of a serum to a standard curve obtained from a reference serum. This comparison was made automatically by a programmable calculator connected with the spectrophotometer. The assay was simplified further by use of specially designed disposable microcuvettes and micropipettes. The reproducibility of the test was good (1 SD = 5%), and the sensitivity for antibodies to influenza A virus and respiratory syncytial virus was 100-200 times greater than that of complement-fixation or hemagglutination-inhibition techniques. The technique seems especially suitable for virus laboratories in hospitals, where the time required for specific diagnosis of viral diseases can be shortened considerably, not only because of the simple performance of the test, but also because the greater sensitivity allows the shortening of the interval between successive samples of serum down to two or three days.