Peptides with NH2-terminal tryptophan in adrenocorticotrophic hormone and melanocyte-stimulating hormone granules of adenohypophysis. Fluorescence microscopy has demonstrated formaldehyde-ozone-induced fluorescence in the pars intermedia cells (melanocyte-stimulating hormone cells) and in certain cells of the pars distalis of the mammalian pituitary. From histochemical and chemical evidence the fluorescence is believed to reflect the presence of peptides with NH2-terminal tryptophan. In the pars distalis of hamster, cat and pig pituitary, the cells that exhibit formaldehyde-ozone-induced fluorescence have now been identified as adrenocorticotrophic hormone (ACTH) cells by immunohistochemistry. Granules from pig pituitaries were purified by passage through a succession of Millipore filters followed by centrifugation on a continuous sucrose gradient. Two granular fractions were identified by electron microscopy and found to contain high concentrations of peptides with NH2-terminal tryptophan as well as high ACTH bioactivity. These fractions, when pelleted and analyzed histochemically, displayed formaldehyde-ozone-induced fluorescence and ACTH-like immunoreactivity.


Macrophages activated in vitro with lymphocyte mediators kill neoplastic but not normal cells. Normal guinea pig macrophages incubated for 3 days in vitro with mediator-rich lymphocyte supernatants become cytotoxic for the syngeneic tumors, line 1 hepatoma and MCA-25 fibrosarcoma. Under identical experimental conditions the survival of two normal syngeneic cell types, fibroblasts and kidney cells, was not affected. The activating supernatants were prepared by stimulating sensitized lymphocyte cultures with an antigen unrelated to the target cells. Therefore, this type of macrophage-mediated cytotoxicity appears to be nonspecific but restricted to cells with malignant growth capacities.


Complement biosynthesis in vitro by rat hepatoma cell strains. Four separate rat hepatoma strains were examined for their capacity to synthesize complement (C). None of the strains synthesized detectable amounts of the first components (C-1), and only one strain (7800C-1) produced the fourth component (C4). However, each of the strains synthesized significant amounts of biologically active C-2 and C-3. Three of the four strains also produced C-5 and the natural inhibitor of C-1 (C1 INH). Two control rat cell strains (fibroblast and pituitary) did not synthesize any detectable C components. Production of C, studied extensively in 7800C-1 and H-4, was reversibly inhibited by cycloheximide (2 mug/ml) and [ 14-C ] amino acids were incorporated into C-2, C-3, and C-1 INH. As assessed by gel filtration, the elution positions of the C components synthesized by the cells in culture were similar to those of the corresponding proteins in normal rat serum. Hydrocortisone (10-6 to 10-7 M) stimulated the production of C-3 by H-4 but C-2 and C-5 production were not affected. These C-producing hepatoma cells may prove useful for studies of the control of C biosynthesis.


Regulation of the in vitro early anamnestic antibody response by exogenous cholera enterotoxin and cyclic AMP. Keyhole limpet hemocyanin was injected into the hind foot pads of rabbits. Six days later cell suspensions were prepared from the popliteal lymph nodes. Various amounts of hemocyanin (1 ng to 100 mug) were added to 1 times 10-7 cells to induce an anamnestic antibody response. Various amounts of cholera enterotoxin, which stimulates the enzyme adenylate cyclase, or dibutyryl cyclic adenosine 3'5'-monophosphate (AMP), were added to the cultures with or without hemocyanin. De novo synthesis of antibody, protein, deoxyribonucleic acid (DNA) and ribonucleic acid (RNA) from radioactive precursors was assayed. The addition  of cholera toxin or dibutyryl cyclic AMP for the first 24 hr with optimal (1 mug) or supraoptimal (100 mug) amounts of hemocyanin enhanced antibody synthesis by at least 100 to 200%. Addition of the toxin or dibutyryl cyclic AMP for the same period to cells minus hemocyanin or with suboptimal amounts (1 to 100 ng) of antigen failed to enhance antibody synthesis. Addition of these agents for 72 to 120 hr to hemocyanin-induced cultures consistently inhibited antibody synthesis. These agents slightly inhibited DNA and RNA synthesis. The increase in protein synthesis caused by the toxin or dibutyryl cyclic AMP was almost totally accounted for by the increase in antibody synthesis. Neither toxin nor cyclic nucleotide promoted the antibody response in the presence of antibody to rabbit thymus-derived lymphocytes; these lymphocytes as well as bursa-equivalent lymphocytes were required for potentiation of the response. Macrophages were not required either for induction of the anamnestic response or for enhancement of this synthesis by cyclic nucleotide or cholera toxin. Both IgM and IgG antibody synthesis were regulated by exogenous cholera toxin and dibutyryl cyclic AMP. A number of possible cellular mechanisms of regulation of the antibody response through the cyclic AMP pathway were discussed. These included the effects of modifications of this pathway on the activities of T lymphocytes early (0 to 24 hr) and B lymphocytes late (72 to 120 hr) in the response and on the apparent reversal of high zone tolerance.


Spleen cell-mediated cytotoxicity of hamster cells transformed by Herpes simplex virus: evidence for virus-specific membrane antigen. The nature of the host's immune response to isografts of hamster embryo fibroblasts (HEF) transformed by herpes simplex virus type 1 (HSV-1) was investigated by the microcytotoxicity assay. It was found that spleen cells from tumor-bearing hamsters killed homologous tumor cells but not HEF transformed by cytomegalovirus or PARA-(defective SV40)-adenovirus 7 (PARA-7). Cytotoxicity was lost as the tumor increased in size. Spleen cells from animals bearing isografts of HSV type 2 (HSV-2) transformed cells also killed HSV-1 target cells whereas spleen cells from PARA-7 tumor bearers did not. Further studies showed that animals immunized with HSV-1 or HSV-1-infected rabbit kidney cells produced spleen cells specifically cytotoxic for HSV-transformed cells. On the other hand, sera from virus-immunized hosts or tumor bearers had no effect on the cells in the presence of guinea pig complement. However, in blocking experiments such sera could significantly reduce spleen cell cytotoxicity. These experiments established that cells transformed by HSV could elicit a cellular immune response in the syngeneic host, and provided evidence that the immunity was directed against virus-specific antigens on the cell surface.


Tumor development after polyoma infection in athymic nude mice. Nude (nu/nu) mice in a CBA/H background show an age-dependent ssuceptibility to tumor development after polyoma virus infection (strain LID-1) when compared with nu/ + or CBA/H mice, which is apparent when 15- or 30-day-old mice are used: tumor incidence was 83 to 90% in nudes and 0 to 10% in controls. Latent perids for tumor development were also shortened in nudes. However, with increasing age nude mice become partially resistant and only 25% develop tumors when infected at 120 days of age. This partial resistance could be transferred with spleen cells to newborn mice. The cells in spleen responsible for this transfer can be eliminated by lysis with anti-Ig and complement or by pre-treatment of the donor with 100 mg/kg of cyclophosphamide and were not affected by treatment in vitro with anti-Thy.1.2 or procedures that remove adherent cells and/or macrophages. When the cells in 15-day-old nu/ + spleen were studied, both anti-Ig or anti-Thy.1.2 treatment eliminated tranfer of resistance to newborn. Virus replication in tissues of nude mice was increased 5 days after infection when compared with nu/ + but became comparable by day 15 after infection. Hemagglutination-inhibition antibodies in serum of nude and nu/ + had comparable titers when measured early after infection but higher titers were observed in nu/ + later after infection.


Lysis of tumor cells by antibody and complement. II. Lack of correlation between amount of C4 and C3 fixed and cell lysis. Two antigenically distinct diethylnitrosamine-induced guinea pig hepatoma cell lines, line-1 and line-10, sensitized with rabbit anti-Forssman or with tumor-specific antibody, were more susceptible to killing by human complement (HuC) than by guinea pig complement (GPC). This difference could not be ascribed to differences in the amount of C1, C4, and C3 fixed: millions of C4 and hundreds of thousands of C3 were detected on cells whether they were killed or not killed by the C sources. Tumor cells sensitized with anti-Forssman IgM antibody generally had more GP C4 and C3  than Hu C4 and C3 bound to their surfaces. Cells sensitized with anti-tumor antibody generally had more Hu C4 and C3 than GP C4 and C3 bound to their surfaces. The resistance to killing of nucleated cells by antibody and C may be due in part to intrinisic properties of the cell.


Mechanism of the suppressive effect of interferon on antibody synthesis in vivo. Mouse interferon preparations significantly suppress the in vivo antibody response to sheep red blood cells (SRBC), a thymus-dependent antigen, and to Salmonella typhimurium lipopolysaccharide (LPS), a thymus-independent antigen. It is also possible to effect the late responses of antigen sensitive "memory" cells observed during secondary immunization by administration of interferon prior to primary immunization. The immunosuppressive activity of interferon was time- and dose-dependent. Maximum suppression was produced when animals were given 1.5 times 10-5 units of interferon between 4 and 48 hr before antigenic stimulation. These findings suggested that interferon affects some early event(s) in the process of antibody synthesis which might be related to the general inhibitory effect of interferon on rapidly dividing cells and viral m-RNA translation. In addition, the use of nonadherent spleen cell cultures from interferon-treated mice, immunized in vitro with a thymus-independent antigen, indicated that in this situation the inhibitory effect of interferon was due to an action on B lymphocytes. A variety of soluble "suppressive" factors are secreted by T cells as a consequence of activation by mitogens or specific antigens in vitro. Since T cells are recognized as one of the sources of interferon, it is suggested that interferon should be investigated as a suppressor T cell-produced lymphokine which can regulate B cell expression.


Release of cyclic AMP from macrophages by stimulation with prostaglandins. The effect of various prostaglandins (PG)on the generation of cyclic AMP in rat peritoneal macrophages has been studied in vitro. PGE1 produced a rapid intracellular accumulation of cyclic AMP which was followed by its release into the extracellular space. More cyclic AMP was released with prostaglandins of the E-type than with A- and F-types. It is suggested that release of cyclic AMP from macrophages may participate in the modulation of leukocyte function.


Lattice and rodlet nuclear inclusions in Merkel cells in rabbit epidermis. A latticee or rodlet inclusion is occasionally observed in the nucleus of a Merkel cell.  The lattice has the external form of a plaque or band and internally consists of alternating parallel wide and narrow layers.  Filaments in wide layers run in at least two directions across the short dimension of the inclusion.  In contrast, the rodlet is a bundle of parallel filaments running the long dimension of the inclusion.  Filaments in lattices and rodlets possibly contract and agitate the highly folded nucleus of the Merkel cell.


Susceptibility of anaerobic bacteria to metronidazole: relative resistance of non-spore-forming gram-positive baccilli. Susceptibility of 358 clinical isolates of obligate anaerobes to metronidazole was determined by an agar-dilution technique. Only 66% of all isolates were inhibited by 6.25 mug/ml, whereas 30% required larger than or equal to 50 mug/ml. Considerable variation in susceptibility was observed among different genera and species of bacteria. Fusobacterium was most senstitive, followed by Clostridium, Bacteroides and Peptococcus, Peptostreptococcus, Veillonella and Acidaminococcus, and non-spore-forming gram-positive bacilli. Bacteroides fragilis was more sensitive than other species of Bacteroides. Similarly, Clostridium perfringens was more susceptible than other species of Clostriduim. While metronidazole appears to be a promising antimicrobial agent for infections caused by Fusobacterium, Clostrididium, and B.fragilis, therapy for infections with other anaerobic bacteria should be guided by in vitro tests of sensitiivity.


Psittacine herpesvirus infection resembling pacheco's parrot disease. Herpesvirus was detected by electron microscopy in hepatocytes of psittacine birds that died at a Florida aviary. The virus was identified in hepatocytes of chick embryos and budgerigars that were given injections of liver suspensions from naturally infected psittacines. Infected hepatocytes had prominent intranuclear inclusions that contained naked nucleocapsids. Both naked and enveloped virions were seen in the cytoplasm of hepatocytes, and enveloped nucleocapsids occasionally were seen in the perinuclear cisternae and extracellularly. The experimental and spontaneous disease mimicked a condition in parrots described previously by Pacheco. The viral agent was classified as Herpesvirus on the bases of the character of the intranuclear inclusions, the size and conformation of the agent as determined by electron microscopy and filtration, and the sensitivity of the agent to ether.


Seroepidemiologic study of Epstein-Barr virus infections in a rural community. The prevalence and titers of antibodies to capsid antigens of Epstein-Barr virus ad to the diffuse and restricted components of the Epstein-Barr virus-induced early antigen complex were determined in 109 families of a semirural community in Louisiana. Titers of antibody to the capsid antigens larger than or equal to 10 were found in 84 percent of children aged two to five years, and the prevalence increased with age to nearly 100 percent. There was a positive but variable correlation of the prevalence of anti-capsid antigen reactivity with low socioeconomic status and crowding. An overrepresentation of high titers of antibody to capsid antigens was present in individuals with a past history of pneumonia and urinary tract infections. The geometric mean titers of antibody to capsid antigens were highest in early childhood, lowest in adolescence and young adulthood, and high in the elderly. Females in all age groups and tonsillectomized children showed a higher geometric mean titer than their male and nontonsillectomized counterparts, respectively. Antibodies to the early antigen complex were found rarely (8.2 percent) and only in sera with relatively high titers of antibody to capsid antigens.


Herpes simplex virus microneutralization: a simplification of the test. The herpes simplex virus (HSV) microneutralization test has been simplified; its use has been demonstrated for the identification of HSV isolates as type 1 (HSV-1) or type 2 (HSV-2) and for the measurement of antibodies to HSV-1 and HSV-2. In this test, the relation between the neutralization titer and virus input is linear, and thus test results can be expressed as corrected neutralization titers, rather than as the more complex neutralizing potency values previously proposed. By means of this test, 45 of 46 clinical isolates of HSV were unequivocally identified as either HSV-1 or HSV-2. Evaluation of neutralizing antibody to HSV infection was more difficult because some neutralizing antibody to the heterotypic HSV is produced after primary infection with HSV-1 or HSV-2, because patients previously infected with one HSV type may show a variety of serological responses to subsequent heterotypic infection, and because human sera obtained early after primary HSV infection may not yet exhibit a type-specific response.


Differential action of deoxynucleosides on mammalian cell cultures infected with herpes simplex virus types 1 and 2. Cytopathic effects induced by eight serologically defined isolates of herpes simplex virus type 2 (tested on human amnion cells) were markedly inhibited by thymidine at a concentration of 5 mM; eight serologically defined isolates of herpes simplex virus type 1, however, were not significantly inhibited. A similar effect was seen with thymidine, deoxyguanosine, and deoxycytidine at 1-mM concentrations in tests with rabbit kidney cultures. The inhibitory effect of thymidine was not blocked by the simultaneous presence of deoxycytidine, which has been shown by others to release mammalian cells from thymidine suppression. Differential suppression of herpes simplex cytopathic effects by these deoxynucleosides provides further evidence of biochemical differences between herpes simplex types 1 and 2. This phenomenon offers a basis for rapid differentiation of type 1 from type 2.


Virus-induced diabetes mellitus. III. Influence of the sex and strain of the host. Nineteen inbred strains of male mice were infected with encephalomyocarditis virus. Five strains became hyperglycemic and had abnormal values in glucose tolerance tests; three strains remained normoglycemic but had abnormal values in glucose tolerance tests; and the remaining strains showed no abnormalities in blood glucose levels or glucose tolerance tests. Female mice from three of five strains tested also developed hyperglycemia, but in one strain (DBA/2) the hyperglycemia was less severe in females than in males. Castrated DBA/2 males developed less severe hyperglycemia than uncastrated males, even though the degree of damage to beta cells appeared to be similar in the two groups. Host factors apparently influence both the development and expression of virus-induced diabetes mellitus.


Temperature-sensitive viruses and the etiology of chronic and inapparent infections. Several mechanisms have been put forward to explain how persistently infected virus-carrier cell cultures are maintained. In this report, information is summarized which suggests that selection of temperature-sensitive mutants may be involved in either establishment or maintenance of persistent infections both in cell cultures and in animals. The possible relation of this mechanism to other mechanisms of regulation of persistent infections is also discussed. Finally, a practical suggestion is offered to researchers and clinicians seeking endogenous viruses from cultured cells and tissues that may harbor temperature-sensitive variants.


Connective tissue activation. VII. Evidence supporting a role for prostaglandins and cyclic nucleotides. Prostaglandins added to synovial cultures stimulated hyaluronic acid (HA) synthesis and glycolysis. The order of potency of the prostaglandins was: PGE1 greater than PGE2 greater than PGF2alpha greater than PGF1alpha, PGE1 and PGE2, 1.0 mug per milliliter, stimulated synovial cells, whereas F-series prostaglandins required 5 mug per milliliter for stimulation. Connective tissue-activating peptide (CTAP) activation of synovial cells was markedly potentiated by all four prostaglandins, and by PGE1 in concentrations as low as 0.01 mug per milliliter. Exogenous prostaglandins caused a prompt and marked increment in synovial cell cyclic-AMP, while CTAP caused a delayed peak of cyclic-AMP of lesser magnitude. Treatment of synovial cultures with cortisol (1.0 mug per milliliter), cycloheximide (10 mug per milliliter), or indomethacin (15.0 mug per milliliter) failed to block stimulation by PGE1, 7-OXA-13-Prostynoic acid, a prostaglandin antagonist, substantially inhibited the action of PGE1 and suppressed the effect of CTAP on synovial cells. It is possible that both exogenous and endogenous (synovial prostaglandins are involved in the connective tissue activation sequence.


Anion excretion pattern following infusion of hyperoncotic human serum albumin into dogs. The effects of hyperoncotic human serum albumin on the excretion rates of several electrolytes and cyclic-AMP were measured in mongrel dogs and the results were compared with those obtained after the infusion of saline. Hyperoncotic albumin increased the excretion rates of sodium, potassium, bicarbonate, and phosphate. There was a small increase in chloride excretion after albumin, while there was a significant decrease in the rate of excretion of cyclic-AMP. Saline, on the other hand, caused a marked increase in the rate of of excretion of all the measured ions. Like albumin, saline was associated with a decrease in the rate of excretion of cyclic-AMP. The marked increase in bicarbonate and phosphate excretion suggests that the response to albumin which includes decreased isotonic reabsorption in the proximal tubule and increased urinary sodium excretion could result from decreased reabsorption in the proximal tubule accompanied by distal sodium and chloride reabsorption. Saline, on the other hand, caused a greater increase in sodium excretion and, although phosphate and bicarbonate excretion also increased, a much greater effect on the excretion of chloride was observed, suggesting that saline may also decrease sodium and chloride reabsorption in the distal nephron.


The large-scale isolation of renin-containing granules from rabbit renal cortex by zonal centrifugation. A new method for the isolation of renin-containing granules from rabbit renal cortex by sequential two-dimensional centrifugation is presented. A mitochondrial fraction produced by preparative differential centrifugation was subjected to isopycnic centrifugation on combined discontinuous and linear density gradients in the same SZ-14 reorienting gradient zonal centrifuge rotor. Renin activity, as assayed by radioimmunoassay for angiotensin I, was localized in a region of the density gradient with a corresponding density of 1.196 Gm. per cubic centimeter or 1.53 M sucrose. The renin-containing granules were enriched 5.0-fold over whole homogenate levels, while peroxisomes were enriched 17.4-fold, lysosomes 23.4-fold, and mitochondria 4.3-fold. Electron microscopic examination of the fraction with highest renin activity showed a granular fraction almost free of contamination from other subcellular material. The data supports the view that renin-containing granules are a distinct subcellular particle. This new method for the large-scale isolation of renin-containing granules makes it possible to obtain the quantity of material necessary to study the release renin at the subcellular level and permit the further biochemical purification and characterization of the enzyme.


Studies on the binding of 125I-labelled corticotrophin to isolated rat adrenocortical cells. Isolated adrenocortical cells, prepared by collagenase disaggregation of normal rat adrenal glands, have been used to study the binding characteristics of 125I-labelled corticotrophin (ACTH) of established biological activity. The binding of the labelled hormone to these whole cells was highly specific, only peptides possessing steroidogenic activity displaced the labelled hormone. Binding was rapid, being complete within 5 min of adding the hormone, and the amount of hormone bound remained constant for up to 20 min thereafter. Over the range 160-10000pg ACTH/ml, increased binding of the hormone was observed at all concentrations of hormone which stimulated steroidogenesis. However at levels of ACTH which stimulated maximal steroidogenesis there was no saturation of binding. This provides the first direct evidence for the existence of 'spare receptors' for ACTH on whole adrenocortical cells. Scatchard analysis of the binding data suggests that there are two types of receptor for ACTH in this preparation of cells. One receptor is of high affinity (dissociation constant equals 2-5 times 10-10 mol/l) with about 3000 sites/cell and the other is of lower affinity (dissociation constant equals 1 times 10-8 mol/l) with about 30000 sites/cell.


Adrenocorticotrophin and the development of insulin secretion in the rabbit foetus. Foetal rabbits were injected with adrenocorticotrophin (ACTH), decapitated, or decapitated and injected simultaneously with ACTH or cortisol in utero on day 24 of gestation. The foetuses were killed after Caesarian section on day 29, and blood was collected for measurement of plasma insulin concentration and pancreatic tissue was obtained for incubation in physiological buffer. Insulin release from the pancreatic tissue of decapitated foetuses was significantly greater than that from the pancreas of control litter-mates when incubated in media containing 3-3mM-glucose, 16-5mM-glucose or 16-5mM-glucose plus 5 mug glucagon/ml, but was similar when the incubation medium contained 3-3 or 16-5 mM-glucose plus 1 mM-theophylline or 3-3mM-glucose plus 60 mM-potassium. The pancreata of decapitated or intact foetuses injected with ACTH did not differ significantly from control foetuses in terms of insulin release in response to glucose in vitro. The plasma insulin concentration of decapitated foetuses and decapitated foetuses injected with ACTH was raised, whereas that of intact foetuses injected with ACTH was similar to that of the control foetuses. Cortisol injection at the time of decapitation resulted in a high rate of foetal mortality. The results indicate that foetal ACTH or foetal adrenocortical secretion influences the normal development of glucose-mediated insulin secretion in the rabbit and that exogenous ACTH corrects the effect of decapitation on beta cell function in vitro but not on plasma insulin concentration.


Effects of hypothalamic extract and dibutyryl cyclic adenosine monophosphate on follicle-stimulating hormone production in the rat adenohypophysis in vitro. An assay system, using the characteristic migration of follicle-stimulating hormone (FSH) on polyacrylamide gels, was developed to evaluate the effects of hypothalamic factors on amino acid incorporation into FSH in the rat adenohypophysis incubated in vitro. Follicle-stimulating hormone extracted from rat pituitary secretory granules was found to migrate on 7-5% polyacrylamide gels, pH 9-5, as a protein with an Rf of 0-614. The characteristic Rf was used to measure [14C]amino acid incorpotation into FSH in rat adenohypophyses in vitro. Follicle-stimulating hormone synthesis increased after addition of crude hypothalamic extract to the incubation medium. Hormone synthesis was also increased three times after the addition of dibutyryl cyclic AMP.


Comparative effect of insulin and explanted adrenal medullary tissue and rat adrenal medulla in situ. The adrenal medulla and explanted medullary cells in the anterior chamber of the eye were examined 3 h after the administration of insulin to rats. No alterations were seen in the explanted cells. In the adrenal medulla, light and electron microscopy indicated depletion of adrenaline cells; noradrenaline cells were not affected. Levels of catecholamines were three times higher in the explanted tissue after insulin treatment; in the medulla in situ they declined by 70%. These results confirm that insulin has no direct releasing effect on chromaffin cells.


Biochemical evidence linking the GIX thymocyte surface antigen to the gp69/71 envelope glycoprotein of murine leukemia virus. It is known that the thymocyte surface antigen GIX is found in some strains of mice and not others, and that its expression in mice of strain 129, in which most extensive genetic studies have been made, is controlled by two unlinked cellular chromosomal loci. We have now isolated a protein with a mol wt of approximately 70,000 daltons from the surface of thymocytes from 129 mice, which have antigenic and biochemical properties characteristic of the gp69/71 envelope component of murine leukemia virus. Our evidence is compatible with the conclusion that it carries the GIX antigen.


The influence of intracellular levels of cyclic nucleotides on cell proliferation and the induction of antibody synthesis. The intracellular ratio of adenosine 3',5'-cyclic monophosphate (cyclic AMP) to guanosine 3',5'-cyclic monophosphate (cyclic GMP) may control the developmental pathway followed by antibody-forming cell (AFC) precursors. The evidence for this is derived from several different types of experiments. First lipopolysaccharide (LPS) which is mitogenic for B lymphocytes, stimulates rapid, transient changes in intracellular levels of cyclic GMP but not cyclic AMP when added to mouse spleen cultures. Cyclic GMP itself stimulates DNA synthesis in these cultures, suggesting that the intracellular changes in cyclic GMP levels are involved in the mitogenic signal delivered by LPS to cells. The absolute amounts of cyclic nucleotides may vary widely in different cells under various conditions, however, the intracellular ratio of cyclic AMP to cyclic GMP is always high in nondividing cells and low in dividing cells. AFC precursors appear to respond to antigen in the absence of T-cell activity by inactivation (1-7). In the response to antigen in the presence of specific T cells, precursor cells proliferate and mature to AFC. Raising intracellular levels of cyclic AMP inhibits cell proliferation and leads to precursor cell inactivation (14, 15). It is suggested that the interaction of antigen with immunoglobulin receptors on the surface of precursors cells leads to the stimulation of adenylate cyclase activity and initiates the inactivation pathway. Since cyclic GMP stimulates immune responses in T-cell-depleted cultures (14, 15) and increasing cyclic GMP levels appear to be involved in the delivery of a mitogenic signal to cells, it is suggested that T-helper cells deliver a signal to precursor cells via the stimulation of guanylate cyclase to initiate the inductive pathway. It is suggested that it is the intracellular ratio of cyclic AMP to cyclic GMP that regulates the fate of precursor cells, not the absolute level of one cyclic nucleotide.


Pathogenesis of of cytomegalovirus infection. I. Activation of virus from bone marrow-derived lymphocytes by in vitro allogenic reaction. After infection in utero or at birth with a cell culture adapted strain of mouse cytomegalovirus (MCMV), several mouse strains developed a latent virus infection in the presence of specific antiviral antibodies. Up to 5 mo after infection, MCMV could be activated and recovered from spleen lymphocytes of the infected animals that were co-cultivated with histoincompatible (H-2 foreign) mouse embryo cells from uninfected animals. In contrast, co-cultivation of lymphoid cells from infected mice with mouse embryo cells from syngeneic, histocompatible (H-2 similar) donors did not activate MCMV. Similarly, MCMV was not recovered from sonicated lymphoid cells. Virus was activated by treating viable lymphoid cells with lipopolysaccharide, a B-cell mitogen, but was not activated by a variety of other mitogens such as phytohemagglutinin, concanavalin A, or pokeweed mitogen. Subsequent purification of lymphoid cells from the infected mice by a variety of techniques indicated that MCMV was harbored in the B-lymphocyte population.


Isotope kinetics of human skin cholesterol secretion. Specific radioactivity (SA) time curves of plasma and skin surface cholesterol collected at several skin areas were recorded in 10 patients on formula diets after single intravenous injections of radioactive cholesterol. Earliest detectable radioactivity on skin surface was found in 4-6 days; depending on the skin site, SA's peaked in 13-75 days. SA's of free cholesterol were almost always higher than those of esterified cholesterol. The general forms of the SA time curves were in keeping with the idea that plasma cholesterol is carried to the skin surface in association with the epidermal and sebaceous cells, whereby (a) cholesterol synthesized de novo is mixed with derived from plasma and (b) appearances of plasma cholesterol on the skin surface is delayed by a time factor that corresponds to the movement of epidermal and sebaceous cells from the basal layer to the skin surface. The shorter mean transit times of plasma cholesterol on skin areas rich in sebaceous glands (22-24 days on the head) than on those poor in these glands (38 days on forearms and 72 days on feet) suggest that cholesterol passes faster through the sebaceous glands than through the epidermis, and faster through thin than thick epidermis. The fraction of skin surface cholesterol (f) that is derived from plasma cholesterol was estimated by three independent methods, and comparable results were obtained. Values of f were lower on skin areas rich in sebaceous glands (0.29-0.46 on forehead) than on areas poor in these glands (0.41-0.70 for forearms; 0.60 on feet) and lower for esterified (0.27-0.33) than for free (0.39-0.48) cholesterol. These data suggest that higher proportions of sebaceous gland and of esterified cholesterol, respectively, are synthesized de novo than epidermal and of free cholesterol. In two patients it was possible to calculate that f of total skin surface cholesterol was 0.49 and 0.37, respectively, and that the maximum amount of plasma cholesterol lost through the skin was 29 and 22 mg/day, respectively. Knowing the total daily excretion of total neutral and acidic steroids in feces in these patients, and assuming a total daily urinary steroid excretion 50 mg, we estimated that no more than 3.2% of total steroid excretion occurred via the skin.


Regulation of protein phosphorylation and sodium transport in toad bladder. It is well established that active sodium-ion transport and water flow across isolated toad bladder are increased by antidiuretic hormone (ADH) and by cAMP. These agents were also observed in previous studies to cause changes in the amount of radioactive phosphate in a specific protein in the toad bladder. This protein, found by SDS-polyacrylamide gel electrophoresis of toad bladder epithelial preparations, had an apparent molecular weight of 49,000 daltons. In the present study, a correlation was found between the ability of a variety of substances to affect the amount of radioactive phosphate in this 40,000-dalton protein and their ability to alter the rate of sodium transport. Thus several agents (ADH, cAMP, theophylline, adenine, prostaglandin E1, and Mn Cl-2) caused a decrease in the amount of radioactive phosphate in the 49,000-dalton protein and also stimulated active sodium transport across the bladder. Conversely, ZnCl-2 produced an increase in the amount of radioactive phosphate in this protein and an inhibition of sodium transport. With each of these agents, the time-course of change in phosphorylation of this protein was, in general, similar to that for sodium transport. A second phosphoprotein, with an apparent molecular weight of about 42,000 daltons, showed changes in parallel with, but less extensive than, those observed in the 49,000 dalton protein. There was no consistent relationship between changes in level of phosphorylation of either in the 49,000- or 42,000- dalton protein and changes in osmotic water permeability. The results are compatible with the possibility that regulation by ADH and by cAMP of sodium transport in the toad bladder epithelium may be mediated through regulation of the amount of phosphate in a specific protein.


Threshold phenomena in chemoreception and taxis in slime mold Physarum polycephalum. The plasmodium of Physarum polycephalum reacts to various kinds of chemicals substances and moves towards or away from them. Threshold concentration of recognition of chemicals was examined in terms of membrane potential and of the averaged motive force of tactic movement by using a double-chamber method, i.e., a single plasmodium was placed between two compartments through a narrow ditch, and differences in membrane potential and in pressure between two compartments were measured. Results are summarized as follows: (a) By increasing the concentration of various substances in one compartment, the membrane potential started to change at a certain threshold concentration, C-th, for each chemical. Chemotactic movement of the plasmodium took place at the same threshold concentration. These results held both for attractants (glucose, galactose, phosphates, pyrophosphates, ATP, c-AMP, etc) and for repellents (various inorganic salts, sucrose, fructose, etc.). (b) The threshold concentration, Cth, for inorganic salts decreased remarkably with increase of the valences of cations, zeta, and was proportional to Z-6, I.E., THE Shultze-Hardy rule known in the field of colloid chemistry was found to be applicable. (c) The plasmodium distinguished the species of monovalent cations in the following order: H(Li(K(Na(Rb(Cs(NH-4 Plots of log Cth against the lyotropic number of anion fell on different straight lines for each monovalent cation species. (d) Plots of log Cth, against the reciprocal of the absolute tempe lines were almost the same and gave a value of 12 kcal/mol for the enthalpy change. These results suggest that the recognition of chemical substances appears as the result of a structural change of the membrane at the threshold point, and that the change in membrane structure is transmitted simultaneously to the motile system of the plasmodium.


A dual synaptic effect on hair cells in Hermissenda. Type A photorecptors can produce an initial hyperpolarizing wave followed by a delayed long-lasting increase in firing which is usually accompanied by a small depolarizing wave. The initial hyperpolarizing wave arises from an increase in conductance while the depolarizing wave was shown to arise from a decrease in conductance. The data presented indicate that both effects produced by the type A photoreceptors in ipsilateral hair cells are synaptic.


Study of sensitivity to curare in certain neurological disorders using a regional technique. A regional technique for the study of curare sensitivity has been applied to patients with Duchenne type muscular dystrophy, myotonic disorders, certain lower motor neurone disorders, to patients with weakness in the arm after hemiplegia, to patients with hyper-reflexia and hypertonia without weakness, and to Parkinsonism. In the dystrophy patients, sensitivity to curare differs from normal controls in that the neuromuscular block persists. The possibilities that this latent defect of neuromuscular transmission is the result of acetylcholine deficiency due to a prejunctional defect or the result of alterations in the property of the postjunctional membrane are discussed. In the myotonic and lower motor neurone disorders, curare sensitivity was similar to that of normal controls. After hemiplegia, the affected side shows resistance to curare when compared with the unaffected side. In states of hyper-reflexia and hypertonia, however, the sensitivity to curare is greater than in normal controls. In Parkinsonism, sensitivity is similar to that of the controls. The results in upper motor neurone lesions are discussed in relation to the dependence of neuromuscular transmission upon the motor neurone, which, in turn, is dependent upon descending impulses.


Disturbances in the voluntary recruitment order of anterior tibial motor units in ataxia. The recruitment order of motor units was studied with an electromyographic technique for secure identification of single motor unit potentials. It has been shown in previous studies of normal subjects that the recruitment order in sustained voluntary contraction is predominantly stable, and that motor units which increase slowly in discharge rate with increasing contraction strength and which already attain regular discharge intervals at low frequencies are always recruited before motor units which increase more rapidly in discharge rate and which do not attain regular discharge intervals until at higher frequencies. In this study 15 patients with severe cerebellar ataxia were examined. It was shown that the recruitment order in sustained voluntary contraction in attaxia is unstable and that low- and high-frequency motor units may alternate as the unit of lowest threshold.


Adrenocortical insufficiency and diffuse cerebral sclerosis. Problems of presentation and diagnosis. Three cases of adrenocortical insufficiency with diffuse cerebral sclerosis are described. One, which was confirmed by brain biopsy, had a family history of the combined disorder affecting members of a previous generation; this has not been previously reported; however pathological confirmation of the diagnosis in the earlier cases was not available. Two cases presented as acute neurosurgical emergencies and attention is drawn to this mode of presentation. Problems of aetiology, presentation and diagnosis are discussed and the finding of normal basal steroid levels is stressed.


Establishment of continuous multiple sclerosis brain cultures after transformation with PML-SV40 virus. PML-SV40 virus was used to transform cells from explants and monolayer subcultures of human brain derived from biopsies or autopsies of MS cases, as well as from 1 case of Jakob-Creutzfeldt disease (JC), 1 case of amyotrophic lateral sclerosis (ALS) and several cases without CNS disease. The success of transformation depended on the growth condition of the culture at the time of infection but at present at least 14 cell lines, 10 derived from MS, 2 from JC, 1 from ALS and 1 from a normal non-CNS patient are maintained in continuous culture.


Zebra body myopathy. Clinical, histochemical and ultrastructural studies. Clinical, electrophysiological and histological evidence is presented of a myopathy not previously described. The patient, aged 15 years, was generally weak and wasted. His condition probably began in utero, but abnormalities of the serum CPK and EMG did not become evident until adolescence. Wide variation in fibre diameter, vacuolation, calcification, increased endomysial connective tissue and intra-fibre splitting were prominent in muscle biopsy sections. Electron microscopy showed numerous rod bodies, fibre disorganisation with loss of Z-bands and occasional honeycomb structures. Frequent zebra bodies were present. Because of the large number of zebra bodies seen, the name "zebra body myopathy" is provisionally proposed.


Experimental intracerebral movement of electron microscopic tracers of various molecular sizes. The authors demonstrate that, while the intracerebral movement of proteins and other relatively large molecules is restricted by tight junctions of endothelia and epithelia of the brain, calcium ion penetrates these barriers and moves between the blood and CSF cavities by extracellular pathways. Mechanisms underlying  CSF formation are considered on the basis of this anatomical continuity of the vascular, extracellular, and CSF compartments of the brain for very small molecules.


Differentiation of Entamoeba: a new medium and optimal conditions for axenic encystation of E. invadens. Nutritional and culturing requirements for efficient axenic encystation of Entamoeba invadens have been studied. A simple and reliable axenic encystation medium has been developed. It contains 0.5% tryptic digest of casein, 0.5% yeast extract, and 5% dialyzed serum in 5 mM potassium phosphate, pH 7.0. Mass encystation (avg 70%) occurred within 30 hr when axenically growing trophozoites of E. invadens IP-l were transferred to this medium before they entered stationary growth phase. Mass encystation of E. invadens PZ occurred similarly, but less reproducibly. Two E. histolytica strains did not encyst. Experiments established that differentiation did not depend upon changes in the external environment after amebase were transferred to encystation medium and, therefore, was initiated by the shift from growth to encystation medium.


Cholelithiasis and ileal pathology in childhood. Primary cholelithiasis is rare in childhood. A particular etiology has appeared over the last few years in connection with unusual physiological situations. Three cases of cholelithiasis after ileal atresia or resection are presented in children of 1, 1, and 7 yr. Another case is presented after abdominal irradiation for Wilms' tumor and ileal resection in a child of 7. The pathogenesis is discussed, raising the question of the interruption of the enterohepatic circulation of bile salts.


Islet cell tumors in childhood.

Islet cell tumors in childhood. Islet cell tumors in children are briefly reviewed. Three of the total of 53 children reported in the literature are described in detail. Special emphasis is directed to localization of the tumor by angiography. The difficulties of localizing the tumor in neonates is discussed. Treatment consists in excision of the insulinoma or, if it is not possible to localize the tumor, in subtotal or total pancreatectomy.


Wilms' tumor in seven children with congenital aniridia. This report reviews our experience with 26 infants and children who demonstrate the syndrome of congenital aniridia. Twenty patients exhibited congenital sporadic aniridia, and unilateral Wilms' tumor has developed in seven of them. Six of the total of 26 patients had familial aniridia and none has shown evidence of the development of a Wilms' tumor or any other malignant tumor to date. Fraumeni studied 15 patients with congenital sporadic aniridia. Five developed Wilms' tumors. In the combined series of 35 patients with congenital sporadic aniridia, 12 patients have developed  Wilms' tumor for an incidence of 34%. A brief review of our experience and a suggested protocol for the evaluation of patients with congenital aniridia is presented.


Studies of herpes simplex virus and interferon in human gingival cell cultures. Cultures of human gingival fibroblast cells from patients with a positive or negative history of recurrent herpes simplex virus (HSV) infection were compared with respect to susceptibility to HSV infection and interferon activity. There was a possible inverse relationship between interferon sensitivity of gingival fibroblast cells and a clinical history of recurrent disease. No correlation was found between prior infection and other parameters of interferon activity or susceptibility of the cells to HSV.


Presence of nonoxidative enzymes of the pentose phosphate shunt in Tetrahymena. Tetrahymena pyriformis, strain HSM, do not have glucose-6-phosphate dehydrogenase or 6-phosphogluconate dehydrogenase, but contain transaldolase, transketolase, ribose 5-phosphate isomerase, ribulose-5-phosphate 3-epimerase, and ribokinase.  The nonoxidative enzymes of the pentose phosphate shunt function in metabolism as indicated by the incorporation of label from [1-14C]ribose into CO2 and glycogen and by the increase in total glycogen content of cultures supplemented with ribose.


Effects of ACTH on the uterine weight response to HCG in mice. The effects of ACTH and dexamethasone on the uterine weight of immature mice treated with HCG or oestradiol-17 beta were studied. The animals were treated daily for 3 days and wet and dry uterine weights were measured on the 4th day. Low doses of ACTH (1-25 mug/day) raised the sensitivity of the uterine weight response to threshold doses of HCG (0-05 to 0-1 i.u.). By increasing the doses of ACTH or HCG, the stimulation gradually turned into inhibition. By itself, ACTH was ineffective and it had no influence on the increase in uterine weight induced by oestradiol-17 beta. Dexamethasone failed to stimulate the effect of HCG.


Prolonged antioestrogenic activity of ICI 46, 474 in the ovariectomized mouse. Subcutaneous administration of ICI 46,474 (3 mg) to ovariectomized mice was found to produce vaginal refractoriness to subcutaneously administered oestradiol for up to 6 weeks. Tritiated oestradiol-17 beta accumulated in the uterus and vagina of the ovariectomized mouse, with maximum accumulation at 3 to 4 hr. This property of the target tissues was used to investigate the binding of tritiated oestradiol after the administration of 3 mg ICI 46,474 to ovariectomized mice. Radioactivity in the vagina was found to be comparable to control values 6 weeks after ICI 46,474 administration; uterine levels of radioactivity returned to control values by 10 weeks. Administration of ICI 46,474 had an oestrogenic effect upon the mouse uterus whereas the vagina appeared to be initially stimulated and was then unable to respond to or to bind oestradiol.


Effects of glucose starvation on normal and rous sarcoma virus-transformed chick cells. We studied the effect of glucose starvation on glucose uptake and thymidine uptake and incorporation in cultures of normal chicken embryo cells and those transformed by Rous sarcoma virus. Resting normal fibroblasts increased the rate of glucose transport up to tenfold when they were starved for glucose, whereas fast-growing normal cells doubled the rate of uptake after starvation. Transformed cells did not show any change in the rate of glucose uptake during starvation. Thymidine uptake and incorporation by normal and transformed cells were not affected by glucose starvation. These results showed that a decrease in the glucose concentration of the medium induced a specific increase in the rate of glucose transport by normal chick fibroblasts, but did not change the transport of glucose by transformed cells. Therefore, it is suggested that glucose or one of its metabolic products regulated the hexose uptake of normal chick fibroblasts. Virus-transformed cells were insensitive to this regulation.


Translation of individual species of vesicular stomatitis viral mRNA. Vesicular stomatitis virus mRNAs from three of the four bands fractionated by polyacrylamide gel electrophoresis in 99% formamide have been eluted from gels and translated in the Krebs II ascites cell-free system. Band 2 mRNA (0.7 times 10-6 daltons) directed the synthesis of the protein moiety of the glycoprotein (G), and band 3 (0.55 times 10-6 daltons) coded for the nucleocapsid (N) protein. Band 4 mRNA (o.28 times 10-6 daltons) directed the synthesis of the NS and matrix (M) proteins. The authenticity of viral proteins synthesized in vitro was shown by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and by analysis of (35-S)metionine-labeled tryptic peptides. These results are consistent with the complexity analysis and coding capacities for the vesicular stomatitis virus mRNA species presented in the accompanying paper.


Translation and identification of the viral mRNA species isolated from subcellular fractions of vesicular stomatitis virus-infected cells. The cytoplasm of vesicular stomatitis virus (VSV)-infected BHK cells has been separated into a fraction containing the membrane-bound polysomes and the remaining supernatant fraction. Total poly(A)-containing RNA was isolated from each fraction and purified. A 17S class of VSV mRNA was found associated almost exclusively with the membrane-bound polysomes, whereas 14,5S and 12S RNAs were found mostly in the postmembrane cytoplasmic supernatant. Poly(A)-containing VSV RNA synthesized in vitro by purified virus was resolved into the same size classes. The individual RNA fractions isolated from VSV-infected cells or synthesized in vitro were translated in cell-free extracts of wheat germ, and their polypeptide products were compared by sodium dodecyl sulfate-polyacrylamide slab gel electrophoresis. The corresponding in vivo and in vitro RNA fractions qualitatively direct the synthesis of the same viral polypeptides and therefore appear to contain the same mRNA species. By tryptic peptide analysis of their translation products, the in vivo VSV mRNA species have been identified. The 17S RNA, which is compartmentalized on membrane-bound polysomes, codes for a protein of molecular weight 63,000 (P-63) which is most probably a nonglycosylated form of the viral glycoprotein, G. Of the viral RNA species present in the remaining cytoplasmic supernatant, the 14.5S RNA codes almost exclusively for the N protein, whereas the 12S RNA codes predominantly for both the NS and M proteins of the virion.


Mosquito cells infected with vesicular stomatitis virus yield unsialylated virions of low infectivity. Vesicular stomatitis virus propagated in and released from Aedes albopictus cells had the normal complement of viral proteins; the glycoprotein contained carbohydrate but no sialic acid. These virions had markedly reduced hemagglutinating activity and exhibited a very high ratio of physical particles to infectious virus. In vitro sialylation of vesicular stomatitis virions grown in mosquito cells resulted in a 100-fold increase in both infectivity and hemagglutination titers to levels approaching those of virus grown in BHK-21 cells. These experiments provide an example of host-controlled modification of viral infectivity.


Host-dependent restriction of mengovirus replication. IV. Effect of some quaternary ammonium ions on the restricted replication of mengovirus in Madin-Darby bovine kidney cells. The quaternary ammonium ions, choline, hexamethonium, and tetraethylammonium, partially reliev the restricted replication of mengovirus in Madin-Darby bovine kidney cells.


Influenza virion RNA-dependent RNA polymerase: stimulation by guanosine and related compounds. The activity of RNA-dependent RNA polymerase of several influenza viruses is stimulated by guanosine. Depending upon the virus strain used, the stimulation of initial reaction rate is up to 10-fold. 5'-GMP, 3',5'-cyclic GMP, and 5'-GDP show lesser stimulation effects. No other nucleosides of 5'-NMPs stimulate, but the dinucleoside monophosphates GpG and GpC show large stimulations. We present evidence that the stimulation represents preferential initiation of genome complementary RNA chains with guanosine: (i) [3-H] guanosine is incorporated specifically at the 5'terminus of RNA in polymerase reaction mixes in vitro. (ii) This incorporation reaction has several properties similar to those of the virion polymerase elongation reaction. (iii) RNA made in the stimulated reaction behaves as complementary RNA in annealing kinetic studies, as does RNA labeled with [3-H]guanosine.


RNA synthesis in cells infected with herpes simplex virus. X. Properties of viral symmetric transcripts and of double-stranded RNA prepared from them. HEp-2 cells infected with herpes simplex 1 virus contain RNA transcripts capable of forming double-stranded (DS) RNA on annealing. The properties of purified DS RNA were as follows (i) DS RNA is resistant to depolymerization by RNase A or T-1 in 2 times 0.15 M NaCl, plus 0.015 M sodium citrate (SSC) but not 0.1 times SSC or following thermal denaturation. (ii) The Tm of the viral DS RNA was 100 C in 0.1 times SSC. (iii) Undenatured DS RNA does not hybridize with viral DNA: upon denaturation, excess unlabeled RNA drove 50 to 55% of labeled DNA into DNA-RNA hybrid. The kinetics of hybridization indicate that the DS RNA consists of at least two populations of transcripts arising from 29 and 26% of viral DNA and differing 40-fold in molar concentration.


Structure and function of herpesvirus genomes. I. comparison of five HSV-1 and two HSV-2 strains by cleavage their DNA with eco R I restriction endonuclease. The restriction endonuclease Eco R I cleaves HSV-1 and HSV-2 DNA into specific fragments that can be resolved by agarose gel electrophoresis. Comparison of HSV-1 strains KOS, 14-012, MP, F, and CI 101, and HSV-2 strains 333 and 186, suggests that the DNAs from type 1 strains are similar but not identical, and that the type 2 strains differ greatly from type 1 strains. The molecular lengths of the fragments have been determined by electron microscopy and can be used to calibrate gel electrophoretic analyses of DNA fragments.


In vitro polyoma DNA synthesis: inhibition by 1-beta-d-arabinofuranosyl CTP. The effects of 1-beta-D-arabinofuranosyl CTP (ara-CTP) on DNA replication were studied in an in vitro system from polyoma-infected BALB/3T3 cells. Ara-CTP concentrations of larger than or equal to 150 muM were found to block in vitro DNA synthesis  completely, and concentrations of smaller than or equal to 0.3 muM had no inhibitory effect. Intermediate concentrations resulted in a concentration-dependent reduction of the in vitro synthesis rate. Long-term labeling with [alpha-32-P]ara-CTP demonstrated the incorporation of the analogue into cellular and viral DNA concomitantly with [3-H]TTP. In pulse-labeling experiments, at noninhibitory concentrations of the analogue, ara-CTP was incorporated into short DNA fragments and long growing strands to relatively the same extent as TTP. Partial venom phosphodiesterase digestion liberated the incoporated are-CTP at essentially the same rate as incorporated TTP, excluding a predominantly terminal incorporation, and after total venom phosphodiesterase digestion greater than 80% of the incorporated ara-CTP was recovered as 5'-ara-CMP. Analysis of the long-term in vitro viral DNA product made in the presence of partially inhibiting ara-CTP concentrations demonstrated that none of the steps leading to mature viral DNA were totally inhibited at the ara-CTP concentrations used. Pulse labeling of replicating viral DNA in the presence of ara-CTP revealed two consistent differences in the pattern found in control pulses: (i) predominant labeling of short chains (5S) with reduced amounts of radioactivity in the longer growing viral DNA strands (smaller than or equal to 16S), and (ii) a one-third to one-half reduction in size for short DNA chains labeled in the presence of ara-CTP. Release of the ara-CTP inhibition with excess dCTP resulted in covalent extension of these smaller short chans to approximately the size of regular short chains labeled in the absence of the inhibitor. Isolated short chains synthesized in the presence of ara-CTP exhibited a slightly lower degree of self-complementarity than regular short chains. The predominant labeling of short chains during pulses is, therefore, not a consequence of discontinuous growth on both sides of the replication fork. Similar results were obtained with ara-ATP and N-ethylmaleimide. The experiments indicate that ara-CTP acts primarily on DNA-polymerizing activities, affecting different DNA polymerases to varying degrees. The results are discussed in terms of the possible number and identity of polymerases involved in viral (and cellular) DNA replication.


Nongenetic complementation of group V temperature-sensitive mutants of vesicular stomatitis virus by UV-irradiated virus. Cells infected with temperature-sensitive (ts) mutants of complementation group V of vesicular stomatitis virus (VSV) give an enhanced yield at nonpermissive temperature when co-infected or superinfected with UV-irradiated virus. Virions produced in these mixed infections are temperature sensitive and do not complement ts V45. Rescue of group V mutants is multiplicity dependent. It can occur in the presence of cycloheximide; kinetics of rescue are similar in the absence or in the presence of the drug. Rescue is due to nongenetic complementation and is interpreted as a trigger effect on maturation of a small quantity of biologically active protein V molecules provided by UV-irradiated virus. These results are comfirmed by rescue of ts V45 by UV-irradiated, defective, interferring T particles.


Temperature-sensitive host range mutants of herpes simplex virus type 2. Herpesviruses are capable of several types of infection of a host cell. To investigate the early events which ultimately determine the nature of the virus-host cell interaction, a system was established utilizing temperature-sensitive mutants of herpes simplex virus type 2. Four mutants have been isolated which fail to induce cytopathic effects and do not replicate at 39 C in hamster embryo fibroblast cells. At least one mutant is virus DNA negative. Since intracellular complementation is detectable between pairs of mutants, a virus function is known to be temperature sensitive. However, all four mutants induce cytopathic effects and replicate to parental virus levels in rabbit kidney cells at 39 C. This suggests that a host cell function, lacking or nonfunctional in HEF cells but present in rabbit kidney cells at 39 C, is required for the replication of these mutants in hamster embryo fibroblasts cells at 39 C. Therefore, we conclude that these mutants are both temperature sensitive and exhibit host range properties.


Virion polypeptide composition of the human papovavirus BK: comparison with simian virus 40 and polyoma virus. The polypeptide composition of labeled BK virus was compared with that of simian virus 40 (SV40) and polyoma virus by co-electrophoresis of disrupted virions in polyacrylamide gels containing approximately 73% of the capsid protein and had a molecular weight of 39,000. It was smaller than VP1 of SV40  and polyoma virus. The other polypeptides of BK virus were similar in molecular weight to those of SV40. A comparison of the proteins of BK virus and SV40 iodinated with chloramine T before and after disruption in alkaline buffer at pH 10.5 revealed differences between the two viruses in the number and distribution of tyrosines available for iodination. The tryptic peptides of VP1, VP3, VP4, and VP5 combined of SV40 were compared with those of the same polypeptides of BK virus. Among the 19 peptides of VP1 resolved, only two were common to both viruses. The analyses of VP4 and VP5, the histone-like proteins, however, showed more similarity between the viruses, with 6 of 15 resolved peptides in common. The tryptic digests of VP3 were completely different.


RNA metabolism of murine leukemia virus II. Endogenous virus-specific RNA in the uninfected BALB/c cell line JLS-V9. Type C virus-specific RNA sequences of BALB/c endogenous virus were detected in JLS-V9 cells (an uninfected BALB/c derived line) by annealing cell RNA with 3-H-labeled virus-specific DNA. Endogenous viruses used in preparing the 3-H-labeled DNA (mostly xenotropic) was prepared from JLS-V9 cells induced to produce virus with iododeoxyuridine. In whole-cell extracts, two virus-specific RNA species, 38S and 27S, were detected. No 60 to 70S virus-specific RNA was found. The same two species of virus-specific RNA were observed in isolated cytoplasmic RNA and in cytoplasmic RNA selected for polyadenylic acid-containing species by binding and elution from oligo(dT) cellulose. Very little, if any, of the virus-specific RNA was active as messenger RNA on polyribosomes. No virus-specific RNA transcribed from genes coding for the BALB/c endogenous N-tropic virus was detected, since 3-H-labeled DNA prepared from endogenous N-tropic virus did not hybridize measurably with JLS-V9 RNA.


Host-dependent restriction of mengovirus replication. II. Effect of host restriction on late viral RNA synthesis and viral maturation. Restricted mengovirus replication in Mandin-Darby bovine kidney (MDBK) cells is characterized by a 400-fold reduction in infectious virus yield and a 40-fold increase in the production of noninfectious virus. Using conditions which insure that all MDBK cells are infected, virus-specific RNA and protein synthesis were measured in the restrictive host and in a permissive host for mengovirus, HeLa cells. Labeling kinetics and sucrose gradient analysis of mengovirus-specific RNA from MDBK cells show a reduction of 10-fold in virion RNA, 5-fold in double-stranded RNA, and 12.5-fold in single-stranded RNA. The viral RNA biosynthetic processes which occur late in the replicative cycle and result in the production of 90% of the single-stranded viral RNA that is packaged into capsid proteins in the permissive host are absent in restrictive MDBK cells. Viral protein synthesis as measured by labeled viral-specific polysome is decreased, and there is an accumulation of 80S subviral particles in the restricted host. It is suggested that restriction may act at a number of stages of viral replication and maturation.


Cellular adsorption function of the sialoglycoprotein of vesicular stomatitis virus and its neuraminic acid. Exposure of vesicular stomatitis (VS) virions to neuraminidase resulted in loss of their ability to agglutinate goose erythrocytes and to attach to L cells concomitant with hydrolysis of sialic acid. These viral adsorptive functions were also destroyed by tryspsinization. Sialyl transferase resialylation in vitro of neuraminidase-treated VS virions restored their hamagglutinating and adsorptive functions almost to original levels. Erythrocyte and L cell receptors for attachment of VS virions were blocked by fully sialylated fetuin and by VS viral sialoglycopeptides. Smaller VS viral glycopeptides generated by extensive trypsinization were less effective inhibitors of hemagglutination than were larger glycopeptides; neuraminic acid and neuraminosyl lactose had no capacity to inhibit hamagglutination or adsorption of virus to L cells. These data suggest that cellular receptors for viral adsorption recognize sialoglycopeptides of a certain size. Neuraminidase desialylation did not significantly alter the isoelectric point of VS virions. Cells exposed to DEAE-dextran, trypsin, or neuraminidase showed significantly increased capacity to attach fully sialylated but not desialylated VS virions. Neuraminidase desialylation of L cells, Chinese hamster ovary cells, and Madin-Darby bovine kidney cells resulted in enhanced susceptibility to plaque formation by VS virus.


Isolation of two subgroup-specific leukemogenic viruses from standard avian myeloblastosis virus. Two populations of virus having subgroup-specific homogeneity (A and B) were isolated from standard avian myeloblastosis virus stocks by passage in vivo through genetically defined chickens. Each possesses leukemogenic activity in vivo. Other properties and potential usefulness of these agents are discussed.


Identification of a viral protein involved in post-translational maturation of the encephalomyocarditis virus capsid precursor. Translation of encephalomyocarditis virus RNA in a cell-free system from uninfected Krebs ascites cells results in the synthesis of a major polypeptide product with a molecular weight of approximately 112,000. In contrast, when the viral RNA is translated in a cell-free system from virus-infected cells, this polypeptide is absent and the largest polypeptide produced has a molecular weight of about 100,000. This latter polypeptide comigrates on sodium dodecyl sulfate-gels with in vivo virus capsid precursor A, and the two have identical patterns of CNBr-generated peptides. A polypeptide having a molecular weight of 12,500 is also a major translation product in the system from infected cells (but not from uninfected cells). This polypeptide appears to be generated by cleavage of the NH-2-terminal portion of the viral RNA-dependent polypeptides by a proteolytic activity present in the infected cell-free system. This proteolytic activity copurifies with the 23,000-molecular weight viral capsid protein gamma, found in infected cells, through chromatography on DEAE-cellulose and cellulose phosphate. This suggests that gamma is itself a proteolytic enzyme involved in maturation of the viral capsid precursor.


Physical map of polyoma viral DNA fragments produced by cleavage with a restriction enzyme from Haemophilus aegyptius, endonuclease R-HaeIII. Digestion of polyoma viral DNA with a restriction enzyme from Haemophilus aegyptius generates at least 22 unique fragments. The fragments have been characterized with respect to size and physical order on the polyoma genome, and the 5' to 3' orientation of the (+) and (-) strands has been determined. A method for specific radiolabeling of adjacent fragments was employed to establish the fragment order. This technique may be useful for ordering the fragments produced by digestion of complex DNAs.


Nucleotide sequence complexities, molecular weights, and poly(A) content of the vesicular stomatitis virus mRNA species. Poly(A)-containing vesicular stomatitis virus mRNA species synthesized in vesicular stomatitis virus-infected cells have been separated into four bands by electrophoresis on formamide-polyacrylamide gels. Two-dimensional fingerprints of ribonuclease T-1 and ribonuclease A digests of the RNA from each band show that they contain unique oligonucleotide sequences as well as 60 to 125 nucleotides of poly(A). The fingerprints were used to determine the nucleotide sequence complexities of RNA from three of the bands. Two contain nucleotide sequences which account completely for their molecular weights (0.70 times 10-6 and 0.55 times 10-6) determined by gel electrophoresis and sedimentation rate, and, therefore, these are radiochemically pure RNA species. The most rapidly migrating band must contain two ro three different RNA species since it has a molecular weight of 0.28 times 10-6, determined by physical methods, and a nucleotide sequence complexity two to three times that expected for a pure RNA species of this size. These data are in complete accord with translational studies (accompanying paper) which show that each of the two pure RNA species codes for a distinct viral protein, whereas the third codes for two viral proteins. From the molecular weight and sequence complexity determinations on mRNA from the bands, we conclude that most of the vesicular stomatitis virus genome is transcribed into discrete mRNA species.


Correlation of pre-treatment serum 17-hydroxycorticosteroid values with survival in patients with prostatic cancer. Total 17-OHCS values were determined before and after treatment in 715 patients with all stages of prostatic cancer. The pre-treatment 17-OHCS level was an important variable in predicting survival and appeared to be a better prognostic indicator of death in the first year than the patient's age, treatment or stage of disease. Generally, the higher the initial value of serum 17-OHCS the greater the probability of death in the first year of treatment. There was no preponderance of one type of death over another when patients with initial high and low values were compared.


Increased virus shedding with aspirin treatment of rhinovirus infection. In two double-blind trials, volunteers challenged with rhinovirus were treated with aspirin or placebo. Aspirin treatment did not alter the rates of infection or illness but was associated with a moderate reduction in the frequency or severity of some symptoms. The overall benefit in rhinovirus infection was not statistically significant. Aspirin treatment appeared to cause a highly significant increase in the rate of virus shedding in treated subjects. The increase in virus shedding must be considered an adverse event that could influence the course of the disease in the individual and increase the likelihood of the spread of the infection to contacts.


Are chlamydial infections the most prevalent venereal disease? We studied two population groups (more than 1,600 patients) to determine the prevalence and clinical associations of genital tract infections with chlamydiae and herpesviruses. Base-line isolation rates for asymptomatic women having routine pelvic examinations were much lower than rates for symptomatic women. In both groups, chlamydiae were more prevalent than herpesviruses. Chlamydial infection was associated much more commonly with cervicitis (36.6%) than with vaginitis only (4.1%). Among 282 symptomatic men, the cultures of 19.9% yielded chlamydiae, and 4.3%, herpesviruses. Chlamydiae were recovered from 35.5% (27 of 76) of specimens from men with nongonococcal urethritis, and from an even larger proportion, 57% (24 of 42), of the specimens from men with frank discharge.


Hyperlipidemia in cerebrovascular diseases. 1. Hypertriglyceridemia and hyper beta-lipoproteinemia are closely related to the pathophysiologic status of the cerebrovascular patients. 2. The electrophoretic analysis of serum lipoprotein is the usefull method for the study on hyperlipidemia in the cerebrovascular disease. 3. The electrophoretic analysis of the serum lipoprotein gives close relation to the clinical features in the patients with normal serum lipid levels as well as in those with the hyperlipidemia. 4. Hyperlipidemia relates closely to the renal impairment, which is supposed to originate from the sclerotic changes of renal artery.


Ischemic heart disease and hyperlipidemia. Studies on lipid and lipoprotein abnormalities which are associable with ischemic heart disease were presented. None of the subjects studied for this report had clinical signs or symptoms characteristic to "familial or sporadic" hyperlipoproteinemia. Only few showed gross abnormalities in lipid chemistries which are compatible with these clinical entities. Lipid abnormalities characteristic to the majority of ischemic heart patients were modest to moderate increase of serum total and free cholesterol and triglyceride; either independent increase or combined increase of these lipid fractions. Determination of free cholesterol may favor to detect such minute abnormality in modest hyperlipidemia as seen in ischemic heart patients. As expressed by our lipoprotein PAG electrophoresis pattern, B and Bp pattern (Fig. 1) seemed to be important lipoprotein abnormalities because of high incidence of ischemic heart disease (60 to 65%). Another feature of these hyperbeta lipoproteinemic state without (B pattern) or with (Bp pattern) moderate prebeta lipoprotein is highly suggestive of premature onset of ischemic heart suggestive premature onset of ischemic heart disease in the subjects with these lipoprotein patterns. Pb battern (hyperpre-beta lipoproteinemic state) was the next, because of frequent occurrence of this pattern (approx. 30%) among the cardiovascular patients and relatively high incidence (approx. 40%) of ischemic heart disease. PB pattern (combine hyperpre-beta and hyperbeta lipoproteinemic state) occurred rarely but incidence of ischemic heart disease in the subject with this pattern was high (approx. 40%). Midband lipoprotein which is one of the unique lipoprotein species detected by PAG electrophoresis may possibly reflect disordered lipoprotein metabolism. However, its association with ischemic heart disease seemed highly unlikely. However, further studies on this and other unusual lipoproteins detectable with PAG electrophoresis seems productive. Extensive studies on cine coronary angiographically established subjects (well characterized study subjects) with this new method in addition to the others would be highly productive to obtain more reliable conclusion on this subject, and hence, to obtain more effective guide line for early identification or for prevention of coronary atherosclerosis.


Characteristics of serum lipid and lipoprotein patterns in patients with coronary sclerosis and with the slightest hyperlipidemia. 1. Serum lipid and lipoprotein patterns of subjects with/without coronary sclerosis and having less than 300 mg/100 ml of serum total cholesterol were analyzed. Subjects woth coronary sclerosis were classified into 2 groups, myocardial infarction (MI) group and coronary sclerosis (CS) group, in accordance with the criteria for the diagnosis of coronary sclerosis established by the special study group of the Educational Ministry in 1960. 2. While serum total cholesterol was higher in the MI group than in the other groups, there was no difference in total cholesterol concentration between the CS group and the control. However, the concentration of free cholesterol, triglycerides, total phospholipids and total lipids increased in the control, the CS and the MI group in this order. 3. The percent concentration of pre-beta and beta and the ratio of beta/alpha and pre-beta+beta/alpha increased in the control, the CS and the MI group in this order. Type IIb hyperlipoproteinemia was observed more frequently in the MI group than in the control one. Type IIa and IV hyperlipoproteinemia did not change into any other phenotypes, but Type IIb changed occationally into Type IIa. 4. So called extra pre-beta lipoprotein was observed more frequently in the CS and MI groups than in the control one. 5. These results suggested that even slight changes in serum lipid and lipoprotein patterns might be related to atherogenecity.


Lipoproteins and hyperlipidemia; special refference on the LCAT and apo-lipoproteins. 1) HDL3 is the most excellent substrate of LCAT reaction. 2) Significant relationships of LCAT activities with serum concentrations of LDL, VLDL, cholesterol, triglyceride and phospholipid, and body weight were observed. No significant relationships were observed between the activity and serum concentrations of HDL3 and HDL2. 3) LCAT activities increased in the various secondary hyperlipedemic states. However, the activities remained within normal range in the familial hyperlipidemias. 4) Immunological properties of apolipoproteins were consistent between normal and hyperlipidemic subjects. 5) Amino acid compositions of hyperlipidemic subjects were almost consistent with those of normal subjects. However, significant differences of amino acid compositions were observed in the patients with familial type II and type IV hyperlipidemias with genetic factor, as compaired with those of normal and other hyperlipidemic subjects. 6) Discussions were made about the above-mentioned results. Authors greatly acknowledge Drs. T. Shimizu, M. Iijima, E. Sasa, T. Kinoshita and M. Harada for their cooperations.


[Epidemic keratoconjunctivitis: a report on a clinical epidemic (author's transl)]. After description of the clinical and serological features of an epidemic keratoconjunctivitis caused by adeno-virus (Type 8) the author compares results of 4 methods of treatment. The schedule consisted of "Okuzell-Coldan" eye drops as placebo, eye drops of corticosteroid content, intramuscular inoculation with Interferonogen and a combined treatment with 0.3 percent "Neutralrot" eye drops and radiation with daylight lamp HPR 125 W produced by Philips Co. Ltd. The transition from the conjunctival form to the keratic form could be best prevented through the last named therapy.


[Treatment of epidemic keratoconjunctivitis with ABOB. Clinical and laboratory investigations (author's transl)]. In an epidemic of viral keratoconjunctivitis in Bulgaria in 1972, caused by adenovirus 9, we applied ABOB in 1 percent solution topically in 120 of all 151 observed patients. When the treatment could be begun in first 3-5 days of the disease, a very favourable curing effect was noted. In most of the patients occurred a significant subjective improvement. The effect was most pronounced upon the course of ther keratitis stage. When ABOB was applied early, the observed keratitis was lighter, with slight visual impairment (with a duration of more than one month in 14.4 percent of the patients, as opposed to 57.3 percent in a placebo group), with quicker epithelialization of corneal erosions and, as a result, with fewer residual corneal opacities.


Granular cell myoblastoma: another unusual esophageal lesion. The case is presented of a 48-year-old woman with esophageal granular cell myoblastoma who was treated by local excision. A review of the literature is also included. So far as we can determine, this is the thirteenth such case reported.


Fasting serum bile acid levels in the primary hyperlipoproteinemias. A newly developed radioimmunoassay specific for conjugates of cholic acid (CCA) was used to measure the fasting serum levels of these bile acids in 233 patients with primary hyperlipoproteinemia classified as type IIa, IIb, III, IV, or V hyperlipoproteinemia as well as in 80 healthy control subjects. Subjects with type IIa hyperlipoproteinemia had significantly lower levels of CCA (0.9 plus or minus 0.06 muM, mean plus or minus SE) than did healthy controls (0.50 plus or minus 0.08 muM). Patients with type IIb, III, IV, and V hyperlipoproteinemia had values similar to those of the control group. The abnormally low fasting value of CCA in type IIa patients is considered to reflect decreased secretion of bile acid into the intestine in such patients, who are known to have defective bile acid synthesis rates and decreased bile acid pool sizes.


Isolation of Chlamydia from patients with urethritis. From October 1973 through August 1974, 335 genitourinary tract specimens from patients with  urethritis were inoculated into McCoy's cell cultures for the diagnosis of Chlamydia infections.  Of the 45 Chlamydia isolates, 42 were recovered when glass vials rather than plastic microtiter plates were used as cell culture vessels.  Herpes simplex virus was isolated 15 times.  Bacterial overgrowth occurred 42 times; however, in 20 specimens the contamination was not apparent until after the first subculture.  This is the first report of the frequency of Chlamydia infections in patients with urethritis in the Midwest and indicates that these organisms have a significant etiologic role in nongonococcal urethritis in this community.


Recent trends in chemotherapy of solid tumors. A multidisciplinary approach consisting of surgery, radiotherapy, chemotherapy, and immunotherapy is now being used in both early and advanced stages of carcinoma. Positive and constructive approaches available today vary from adjuvant studies for early or primary disease to combinations of radiotherapy and chemotherapy for advanced disease. Methods of treatment considered to hold significant promise are discussed.


Current status of regional arterial infusion chemotherapy. The rationale for this method is that a prolonged, continuous exposure of tumor cells to an antimetabolite is required for effective antimetabolic activity to develop in the tumor cell population. Overall experience in selected sites such as head and neck, liver, and extremities is reviewed.


Syndromes resulting from ectopic hormone-producing tumors. Among the malignant tumors of nonendocrine origin that are capable of producing polypeptide hormones and of manifesting as different endocrine syndromes discussed here are ectopic ACTH syndrome, SIADH, and ectopic gonadotropin-producing tumors.


Cutaneous manifestations of internal malignancy. Skin changes may result from metastases to skin or from chemicals or hormones that are secreted by the tumor, resulting in identifiable cutaneous syndromes, or may be inflammatory dermatoses, benign or malignant proliferative lesions, or nevoid states whose association is recognized but the mechanisms of which are unexplained.


Ifluence of dietary fatty acids on membrane properties and enzyme activities of liver mitochondria of normal and hypophysectomized rats. Studies are reported on the effects of diets containing fatty supplements with (A) a high concentration of arachidonate (46% concentrate of ethyl arachidonate), (B) a high concentration of linoleate (corn oil), and (C) an essential fatty acid deficient, fully saturated fat (hydrogenated coconut oil) upon lipid composition, membrane permeability, and enzyme activities of liver mitochondria of normal and hypophysectomized rats. The fatty supplements produced differences in the fatty acid composition of the liver mitochondria; hypophysectomy, in addition, influenced the neutral and phospholipid composition. Permeability, indicated by swelling properties, correlated generally with the degree of unsaturation and essential fatty acid content of the lipid of the mitochondria of the normal animals. The fatty supplements also influenced the enzyme acitivites of the mitochondria of the normal animals. The mitochondria of the hypophysectomized animals were less responsive to the differences in the dietary fat in both their swelling properties and enzyme activities. Although the relationship was complex, it appeared that the hypophysis was involved in the functions of essential fatty acids in liver mitochondria.


75Se-selenomethionine-labeled lipoproteins in hyperlipidemic and normolipidemic humans. Investigating the feasability and application appropriate to determination of lipoprotein turnover, 75Se-selenomethionine (Se-M) incorporation in lipoproteins was studied in eight hospitalized patients. Three were hyperlipidemic diabetic subjects (HL), and five were normolipidemic but otherwise abnormal subjects (NL). Maximum activity was observed in the VLDL fraction within 2-3 hr, and was followed by a biexponential decay. Incorporation into the LDL-I (density, 1.006-1.019 mug/ml) occurred at a rate which correlated with the rapid decay rate of the VLDL Se-M activity. The decay of the Se-M was single exponential in the LDL-I and LDL-II (density, 1.019-1.063 mug/ml), suggesting undirectional transfer of the label. The HDL fraction showed a rapid initial Se-M uptake which was followed by a slow rise, resulting in a complex time-activity curve. The Se-M activity was significantly higher in the VLDL fraction and slightly lower in the LDL and HDL fractions in the HL as compared to the NL subjects. The VLDL-apoprotein concentration increased by a significantly greater amount in the HL (374 mug/ml) than in the NL subjects (67 mug/ml). There was a significant increase in VLDL-apoprotein turnover in the presence of a fractional turnover rate that was not significantly lower in the three hyperlipidemic subjects compared to the NL controls. In vivo 75Se-selenomethionine labeling allows the estimation of the synthesis and removal rate of apoproteins.


Effects of estrogenic compounds on triglyceride kinetics. Paired studies of triglyceride kinetics were made in 8 women on and off postmenopausal estrogen supplementation. When estrogens were discontinued, mean very-low-density lipoprotein triglyceride (VLD-TG) and VLD-TG turnover rate fell respectively from 555 to 349 mg/100 ml, and from 30-18 mg/kg/hr (p smaller than 0.01). Maximal turnover rate (Vmax) and VLD-TG concentration at 1/2 V max (Km), were 82.4 mg/kg/hr and 704.14 mg/100 ml on estrogen and fell to 41.83 and 347.51 when estrogen was discontinued (p smaller than 0.01). Estrogen supplementation did not lengthen VLD-TG half-life or diminish the fractional turnover rate of VLD-TG. Estrogens apparently increase VLD-TG by augmenting VLD-TG production rates.


[Globular corpuscles in hepatomas of Swiss mice]. In hapatomas formed spontaneously without cirrhosis at Swiss mice numerous globular corpuscules of eosinophil character were found. These corpuscules as revealed by electron microscopy are formed in cyternas of the rough surfaced endoplasmatic reticulum and are of the nature of protein.


[Electron microscopic study of cytoplasmic crystals in liver damages caused by alcoholism]. Liver biopsy of 50 patients with liver disease of alcoholic etiology was examined electronmicroscopically. In every stage of the disease presence of intramitochondrial paracrystalloids was revealed. Hyalin of alcoholic origin of fibrillar structure was seen in cases of alcoholic hepatitis. By the electronmicroscopic examination of three cases of alcoholic fatty liver a special form of cytoplasmatic paracristalloid inclusion could be seen. This inclusion consists of membranless parallely lying fibrills. Changing the plain of the preparats containing inclusions by the aid of a goniometer the structure of the inclusions appeared in a form of "honey-comb". Inclusion in the question can not be regarded identical neither with intramitochondrial inclusions nor with the fibrills of the so called alcoholic hyalin. Formation and function of these inclusions are as yet unknown.


Azaguanine-resistant mutants induced by several mutagens in a neurospora heterokaryon. This paper reports the development of a new system for the detection of forward mutations in a heterokaryon of Neurospora crassa. This system, designed to detect a wide variety of genetic alterations, is based upon the detection of 8-azaguanine-resistant mutants in an adenine-requiring heterokaryon carrying the aza-3 gene. Induction of mutations in the aza-3 gene was detected after treatment of conidia with ultraviolet light, X-rays, ethyl methanesulfonate (EMS), or the acridine mustard, ICR-170. The pattern of appearance of mutant colonies and various factors in mutant selection are discussed.


A simple test for the diagnosis of absorptive, resorptive and renal hypercalciurias. A test was developed to diagnose various forms of hypercalciuria. A two-hour urine sample after an overnight fast and a four-hour urine sample after 1 g of calcium by mouth were tested for calcium, cyclic AMP and creatinine. The 24 patients with absorptive hypercalciuria had normocalcemia and normal fasting urinary calcium (less than 0.11 mg per milligram of urinary creatnine). Urinary calcium was high (greater than or equal to 0.2 mg per milligram of creatinine) after a calcium load. Of the 28 patients with primary hyperparathyroidism (resorptive hypercalciuria), 25 had hypercalcemia and 21 had high fasting urinary calcium. Urinary cyclic AMP, elevated in 30 per cent of fasting patients, was high (greater than 4.60 mu moles per gram of creatinine) in 82 per cent of cases after calcium load. Six patients with renal hypercalciuria had normocalcemia, high fasting urinary calcium, and high (greater than 6.86 mu moles per gram of creatinine) or high-normal fasting urinary cyclic AMP was normal. This simple test should facilitate the differentiation of various causes of hypercalciuria.


Travelers' diarrhea and toxigenic Escherichia coli. In a group of 133 United States students studied for 18 days after arriving in Mexico, diarrhea developed in 38 (29 per cent). Diarrhea rarely began before the fourth day, and the mean onset was 13 days after arrival. Symptoms lasted an average of 3.4 days but persisted in 21 per cent of sick students. Heat-labile enterotoxin-producing Escheria coli was found in the stools of 72 per cent of sick and 15 per cent of healthy students. None had heat-labile Esch. coli when they entered Mexico. The incubation period was short, generally 24 to 48 hours, and the carrier state was five days or less in 82 per cent of students surveyed. Entamoeba histolytica was found in 6 per cent of cases of diarrhea, but not salmonella, shigella or penetrating Esch. coli. These studies suggest that approximately 70 per cent of travelers' diarrhea in Mexico is associated with heat-labile toxigenic strains of Esch. coli.


Pancreatic cholera: benefical effects of treatment with streptozotocin. Two patients with pancreatic cholera and islet-cell carcinoma were treated with intra-arterial streptozotocin. Before therapy, they had stool volumes from 2 to 8 liters per day and required 200 to 800 mEq per day of supplemental potassium. After three to five doses of streptozotocin (1.5 per square meter), both stool volume and number and size of hepatic metastases decreased markedly. One patient has had normally formed stools for 12 months; the other had a 90 per cent reduction in stool volume for 13 months with additional therapy. Both patients' serum potassium returned to normal without need for supplementation. Jejunal adenylate cyclase activity was normal in both, and plasma vasoactive intestinal peptide was detectable in only one. After chemotherapy, these findings showed no consistent change. Pharmacologic studies suggest that arterial administration increased either tumor or hepatic extraction (or both) of streptozotocin by two times and decreased renal exposure to this nephrotoxic drug by one third.


17 Beta-estradiol transport and metabolism in human red blood cells. Attempts to measure the function of red blood cells as steroid metabolizing tissue showed that the membrane plays an important role. Of the 17beta-estradiol associated with red cells, 2/3 was bound to the membrane while 1/3 was in the soluble fraction. The binding appears to be nonspecific and linearly related to the amount incubated. Binding was more extensive at 2 C than at 26 C. Successive washes of red cells with isotonic sodium chloride solution increased their ability to bind 17beta-estradiol. Red cells incubated in isotonic salt media removed 85.1 plus or minus 3.2% (SD) of the total estrogens present and this value remained within narrow limits for incubation times as short as 15 s and as long as 19 h at 37 C. When incubated in homologous plasma, only 8-12% of estrogens in the media became associated with cells. The eight-fold difference in cell binding between salt solution and plasma can be attributed to competition with plasm aproteins. Presence of plasma proteins diminished but does not prevent activity of 17beta-hydroxysteroid dehydrogenase of red cells. Cells produce four times more estrone when incubated in saline media than when incubated in their own plasma. The enzyme is in the soluble portion of the cells and the steroids have to traverse the membrane in both directions. In the transfer of estrogens from plasma of cytoplasm, the membrane needs to be considered as a separate compartment.


Studies of ACTH secretion control in 116 cases of Cushing's syndrome. Plasma ACTH (normal value: 0.16 plus or minus mU/100 ml) was measured in 116 patients with Cushing's syndrome, using a bioassay including dynamic tests and sequential determinations. In 10 patients with adrenal tumors ACTH levels were nondetectable (ND) or low, and usually nonstimulatable. In 10 patients with ectopic ACTH secretion high levels (0.42 plus or minus 0.07 mU/100 ml) were measured. The extracts of 6 tumors yielded an ACTH-like substance. Forty-three patients with Cushing's disease (without pituitary tumor) had, before treatment, a mean ACTH level of 0.18 plus or minus 0.01 mU/100 ml, accompanied by high levels of plasma cortisol (32.1 plus or minus 1.9 mug/100 ml). Irregular nycthemeral variations occurred. ACTH rose to 0.30 mU/100 ml after incomplete adrenalectomy (20 patients) and to 1.14 mU/100 ml after total adrenalectomy (21 patients). Dexamethasone (8 mg per day) suppressed ACTH levels. Metyrapone induced a normal ACTH rise, but at abnormal times. Lysine-vasopressin (LVP) induced an ACTH mean relative increase of 120% before, and of 140% after adrenalectomy (i.e., within the normal range). Six nonadrenalectomized patients with pituitary tumors showed similar abnormalities of ACTH regulation. However, the ACTH rise after LVP was above 500%. When pituitary tumors occurred after adrenalectomy (12 patients) the mean basal ACTH level was 18 mU/100 ml. Dexamethasone induced a 90% decrease, and LVP a 416% increase in ACTH levels. In 6 patients with nodular adrenal hyperplasia, ACTH was undetectable before treatment. After adrenalectomy, ACTH rose to 0.4 mU/100 ml (11 patients) and the increase after LVP was 90%. Five additional patients developed pituitary tumors. These data confirm the abnormalities of ACTH feedback regulation in Cushing's disease. However, even when pituitary tumors occur, ACTH levels can be altered by metyrapone, dexamethasone and LVP. This last test is of particular interest for the detection of pituitary tumors. The follow-up pattern of treated nodular adrenal hyperplasia appears to be very similar to that of Cushing's disease.


Radioreceptor assay for human chorionic gonadotropin. A sensitive radioreceptor assay capable of detecting 100 pg of human chorionic gonadotropin has been developed using ovarian receptors and (125I)hCG. The dose-response curves of a series of urine and serum samples from pregnant women were linear and parallel to standard curves of 2nd International Standard hCG and purified hCG when logit of percent bound was plotted against log dose. Potency estimates of pregnancy sera, measured by radioreceptor assays, were approximately 2 times greater than those determined by radioimmunoassays. Examination of hCG concentrations in the urine of pregnant and nonpregnant women, using a receptor assay, showed good correlation with the result obtained with a standard agglutination pregnancy test. This readioreceptor assay, which can be made as sensitive as radioimmunoassay, provides a rapid and simple method for the measurement of biologically active hCG and LH.


Partial pharmacodynamic model for the circadian-episodic secretion of cortisol in man. A pharmacodynamic model was developed to facilitate computer analysis of the circadian-episodic influx of cortisol into plasma from the adrenal gland. The model consists of a catenary system of a biorhythmic control, the adrenal gland, and a body compartment containing circulating cortisol. Computer nonlinear regression analysis was carried out on data consisting of plasma cortisol concentrations measured at 30-min intervals over 1 day in a group of normal children. Circadian rhythmic synthesis of cortisol by the adrenal gland was described by a sinusoidal function which provides the level (284 plus or minus 50 mugg/h/m2) and amplitude (245 plus or minus 50 mug/h/m2) of the synthesis rate and the period (24 hours) and acrophase (8.14 plus or minus 1.95 h) of the cycle. Cosinor analysis of the data confirmed a highly significant circadian rhythm in the daily synthesis rate of cortisol. Episodic secretion, described by an empirical switch function, is assumed to result from accumulation of small amounts of cortisol precursors in the adrenal gland and intermittent stimulation of cortisol release by ACTH. This was found to take place over 46.6 plus or minus 2.4% of a 24-h day. Circulating cortisol is contained in a single body compartment with an apparent volume of distribution (5.3 plus or minus 0.55 liters/m2) from which elimination occurs by first-order metabolic clearance. The biological half-life averaged 0.96 plus or minus 0.18 h. Upon least-square optimization of selected kinetic parameters, the circadian-episodic model increases the accountable variation (r2 x 100) to 89% in comparison with the 35% obtained by use of only a periodic function.


Ethidium bromide inhibits appearance of closed circular viral DNA and integration of virus-specific DNA in duck cells infected by avian sarcoma virus. The DNA of avian sarcoma virus assumes a closed circular configuration before integration into the host cell chromosomal DNA. Ethidium bromide reduces the formation of superhelical viral DNA and concurrently blocks integration of the viral genome. Inhibition of integration of viral DNA results in the inhibition of virus replication.


Inhibition of release of prostaglandins as an explanation of some of the actions of anti-inflammatory corticosteroids. Corticosteroids as well as non-steroid anti-inflammatory drugs inhibit the prostaglandin-mediated vasodilatation accompanying lipolysis in subcutaneous fat. Whereas the non-steroids produce their effect by inhibition of prostaglandin synthesis, however, corticosteroids inhibit their release. This mechanism may be the basis of some actions of corticosteroids in inflammation, in the gastric mucosa and in the CNS.