Association of vesicular stomatitis virus glycoprotein with virion membrane: characterization of the lipophilic tail fragment.

The proteolytic enzyme, thermolysin, degraded the external segment of the membrane glycoprotein of intact vesicular stomatitis (VS) virions but left behind a small nonglycosylated fragment, presumably embedded in the virion membrane. Other proteases generated membrane-associated glycoprotein fragments differing somewhat in molecular weight. The thermolysin-resistant, virion-associated fragment, which can be selectively solubilized by either Triton X-100 or chloroform/methanol, has a molecular weight of 5,200. Amino acid analysis of the glycoprotein fragment reveals a preponderance of hydrophobic amino acids (64% of the residues); the amino-terminal amino acid is alanine as determined by dansylation. Cyanogen bromide digestion of the tail fragment generated two peptides, confirming the presence of one methionine residue per thermolysin-resistant glycoprotein fragment. The secondary structure of this glycoprotein tail peptide is maintained by at least one disulfide bridge. Thermolysin treatment is isolated VS viral glycoprotein in the presence of Triton X-100 also generated a hydrophobic peptide fragment which is very similar to the virion-associated glycoprotein fragment. The amino acid terminus of intact glycoprotein was also found to be alanine as was its dansylated Triton-micellar fragment that resisted thermolytic degradation; this finding suggests that the amino-terminal end of the VS viral glycoprotein is embedded in the virion membrane. These results suggest that the VS viral glycoprotein is an amphipathic molecule, the hydrophilic portion of which contains all the carbohydrate and a lipophilic tail segment which forms lipid or detergent micelles, thus rendering it resistant to proteolysis.


Intracellular synthesis of mouse mammary tumor virus polypeptides: indication of a precursor glycoprotein.

Mouse mammary tumor virus polypeptides were detected in the cytoplasm of mouse mammary tumor cell cultures using immunological precipitation techniques. The anti-mouse mammary tumor virus serum precipitated the major virion glycoproteins gp49 and gp37.5/33.5 and a viral-related nonvirion glycoprotein of 76,000 daltons. Subcellular fractionation studies revelaed that the cell-associated virion glycoproteins were present in the membrane fraction. Pulsechase experiments indicated that a viral-related nonvirion glycoprotein of 76,000 daltons may be a precursor to one or more of the virion glycoproteins.


Temperature-sensitive mutants of herpes simplex virus type 1 defective in lysis but not in transformation.

Twelve temperature-sensitive (ts) mutants of herpes simplex virus type 1 (HSV-1), representing seven complementation groups, were isolated subsequent to 5-bromodeoxyuridine mutagenesis. These mutants were identified by their inability to replicate in a line of monkey (CV-1) cells at 39 C. Seven of these mutants, representing six complementation groups, induced thymidine kinase (tk) and transformed Ltk- cells, a line of mouse L cells lacking tk, to a tk+ phenotype at both the permissive (34 C) and nonpermissive (39 C) temperatures. Thus, the defective cistrons in these six complementation groups, although necessary for lysis, have no essential function in this transformation system. Transformation by these 12 mutants was dependent on prior UV irradiation. Infection of cells with unirradiated virus under conditions which did not permit virus replication was not sufficient to allow cell transformation. Five mutants, representing two complementation groups, were tk- and were incapable of causing the tk--to-tk+ transformation at either 34 C of 39 C. The tk defects in these mutants are probably unrelated to the ts defects, since one of these complementation groups contains a tk+ member. Therefore, transformation of Ltk- cells to a tk+ phenotype by HSV-1 requires an active viral tk gene. One complementation group was represented by a single tk- member. The role of this cistron in transformation remains undetermined since the primary block to transformation is presumed to be the tk- phenotype. Mutants representing the seven complementation groups were unable to replicate at 39 C in two lines of HSV-1-transformed cells, indicating that the activities of resident wild-type copies of the defective cistrons, if present, could not be detected by complementation.


Reaction of poliovirus RNAs with antibodies to double-stranded RNA demonstrated by an immunochemical binding assay.

An immunochemical binding assay was used to investigate the reactivity of radioactively labeled viral RNAs from poliovirus-infected cells with antibodies to the synthetic double-stranded RNA, poly(I)-poly(C). A RNase-free antibody-containing serum fraction was employed. Poliovirus replicative form reacted with the antibodies to poly(I)-poly(C) as well as or better than poly(I)-poly(C). Poliovirus replicative intermediate reacted with the antibodies to a greater extent than poliovirus single-stranded RNA, but both were less reactive than replicative form. The use of the immunochemical binding assay with sucrose-gradient fractions demonstrated that for both poliovirus single-stranded RNA and replicative form the peak of reactivity with the antibodies was coincident with the peak of radioactive material precipitated by trichloroacetic acid. The proportion of replicative intermediate that reacted with the antibody increased in sucrose-gradient fractions containing the more slowly sedimenting RI RNA.


Human cytomegalovirus. III. Virus-induced DNA polymerase.

Infection of WI-38 human fibroblasts with human cytomegalovirus (CMV) led to the stimulation of host cell DNA polymerase synthesis and induction of a novel virus-specific DNA polymerase. This cytomegalovirus-induced DNA polymerase was purified and separated from host cell enzymes by DEAE-cellulose and phosphocellulose column chromatographies. It can be distinguished from host cell enzymes by chromatographic behavior, template primer specificity, sedimentation property, and the requirement of salt for maximal activity. This virus-induced enzyme has a sedimentation coefficient of 9.2S and is found in both the nuclei and cytoplasm of virus-infected cells, but not in uninfected cells. This enzyme could efficiently use activated calf-thymus DNA, oly(dA)-oligo(dT)12-18, and poly(dC)-oligo(dG)12-18 as template primers, especially poly(dA)-oligo(dT)12-18, but it could not use poly(rA)-oligo(dT)12-18, poly(rC)-oligo(dG)12-18, or oligo(dT)12-18. The enzyme requires Mg2+ for maximal activity, is sensitive to p-hydroxymercuribenzoate, and is not a zinc metalloenzyme. In addition, the cytomegalovirus-induced DNA polymerase activity can be enhanced by adding 0.06 to 0.12 M NaCl or 0.03 to 0.06 M (NH4)2SO4 to the reaction mixture.


Abortive infection of a rabbit cornea cell line by vesicular stomatitis virus: conversion to productive infection by superinfection with vaccinia virus.

An abortive infection of a rabbit cornea cell line (RC-60) by vesicular stomatitis virus (VSV), yielding less than 1 PFU/cell, was converted to a productive infection, yielding 1,900 PFU/cell, when cells were superinfected with vaccinia. Studies on the synthesis of VSV-directed RNA in RC-60 cells suggest that the abortive infection by VSV alone may be due in part to (i) a limited production of 40S virion RNA and (ii) a markedly reduced activity of virion-bound transcriptase activity in RC-60 cells compared to the activity in mouse L cells, a permissive host for VSV. No recognizable VSV structures, except a small amount of viral core structures, were produced by the abortive infection. In contrast, double infection of RC-60 cells with VSV and vaccinia in the presence of hydroxyurea resulted in the production of infective B particles of VSV. Although the function supplied by vaccinia responsible for the productive replication of VSV in double infected RC-60 cells has not been identified, metabolic inhibitor studies indicate that continuous vaccinia-dependent RNA synthesis is required for maximal production of infective VSV. The possibility is considered that vaccinia may supply a product or function required for VSV replication which is ordinarily supplied by the host but which is lacking in RC-60 cells.


Synthesis of functional bacteriophage T4-delayed early mRNA in the absence of protein synthesis.

When Escherichia coli B207 is grown either aerobically or under limited aerobic conditions, pretreated with chloramphenicol to block protein synthesis, and then infected with bacteriophage T4, the phage RNA which accumulates, termed "immediately early" (IE), contains the transcripts of a limited number of prereplicative genes. Among the transcripts which accumulate is the mRNA which serves as a template for deoxycytidylate hydroxymethylase (HMase) synthesis. Among the prereplicative gene transcripts which do not accumulate under these conditions are deoxycytidine triphosphatase (dCTPase), alpha-glucosyl transferase (alphg-gt), and deoxynucleotide kinase (kinase); these genes have been termed "delayed early" (DE). In contrast, when protein synthesis is inhibited by  depleting aerobically grown E. coli B207 of K+, both IE and DE T4 RNA accumulate, but these transcripts do not contain functional HMase, dCTPase, alpha-gt, or kinase mRNA's. However, if E. coli is grown under conditions of limited aeration and then depleted of K+ prior to T4 infection, the T4 RNA which accumulates contains both IE and DE transcripts and functional HMase, dCTPase, and alpha-gt mRNA's. Functional kinase mRNA does not accumulate under these conditions. The results of these experiments indicate that the synthesis of functional DE RNA in the absence of simultaneous protein synthesis, depends on the physiological condition of the cells and the way in which protein synthesis is inhibited. In addition, data is presented which suggests that extensive transcription of DE genes in the absence of protein synthesis results in the inhibition of transcription of certain IE genes.


Demyelination and cytopathic effects in cultures of mammalian dorsal root ganglia infected with encephalomyocarditis virus.

Replication of encephalomyocarditis virus and its cytopathic effects were studied in myelinated cultures of dorsal root ganglia obtained from newborn mice. Six hours after infection virus progeny was detected in the culture. At 24 h the virus titer reached 2 times 10(6) PFU per culture and remained at this level until 48 h. The first cytopathic alterations began at 24 h and consisted of rounding of Schwann and satellites cells and their detachment from neurons. Later, bead-like swellings of the myelin appeared along the axons followed by splitting and degeneration of lamellae. The cytopathic effect in the neurons started 29 h after infection, reaching complete neuronolysis at 48 h. Virus particles, scattered or arranged in crystal-like aggregates, were first seen in the cytoplasm of glial cells and then in neurons and axons.


Structure, subunit composition, and molecular weight of RD-114 RNA.

The properties and subunit composition of the RNA extracted from RD-114 virions have been studied. The RNA extracted from the virion has a sedimentation coefficient of 52S in a nondenaturing aqueous electrolyte. The estimated molecular weight by sedimentation in nondenaturing and weakly denaturing media is in the range 5.7 X 10(6) to 7.0 X 10(6). By electron microscopy, under moderately denaturing conditions, the 52S molecule is seen to be an extended single strand with a contour length of about 4.0 mum corresponding to a molecular weight of 5.74 X 10(6). It contains two characteristic secondary structure features: (i) a central Y- or T-shaped structure (the rabbit ears) with a molecular weight of 0.3 X 10(6), (ii) two symmetreically disposed loops on each side of and at equal distance from the center. The 52S molecule consists of two half-size molecules, with molecular weight 2.8 X 10(6), joined together within the central rabbit ears feature. Melting of the rabbit ears with concomitant dissociation of the 52S molecule into subunits, has been caused by either one of two strongly denaturing treatments: incubation in a mixture of CH3HgOH and glyoxal at room temperature, or thermal dissociation in a urea-formamide solvent. When half-size molecules are quenched from denaturing temperatures, a new off-center secondary structure feature termed the branch-like structure is seen. The dissociation behavior of the 52S complex and the molecular weight of the subunits have been confirmed by gel electrophoresis studies. The loop structures melt at fairly low temperatures; the dissociation of the 52S molecule into its two subunits occurs at a higher temperature corresponding to a base composition of about 63% guanosine plus cytosine. Polyadenylic acid mapping by electron microscopy shows that the 52S molecule contains two polyadenylic acid segments, one at each end. It thus appears that 52S RD-114 RNA consists of two 2.8 X 10(6) dalton subunits, each with a characteristic secondary structure loop, and joined at the 5' ends to form the rabbit ears secondary structure feature. The observations are consistent with but do not require the conclusion that the two 2.8 X 10(6) dalton subunits of 52S RD-114 RNA are identical.


Further evidence for the existence of a viral envelope protein defect in the Bryan high-titer strain of Rous sarcoma virus.

Electron microscopy observations of purified Bryan high-titer Rous sarcoma virus (BH RSV) using the freeze-drying technique showed that progeny made in the absence of a helper virus lacked visible surface projections or spikes. Phenotypic mixing experiments employing BH RSV and a thermolabile mutant of vesicular stomatitis virus, tl 17, yielded no evidence of pseudotype formation. Since tl 17 is known to be defective for an envelope glycoprotein, the lack of successful phenotypic mixing with BH RSV is consistent with the observed absence of viral spikes.


Association of ribosomes with intracellular vesicular stomatitis virus particles.

Ribosomes are observed intimately associated with nucleocapsids of vesicular, stomatitis virus, especially those that line structures that are either cytoplasmic vesicles or invaginations of the plasma membrane.


Poxvirus infection in the white-backed magpie (Gymnorhina hypoleuca) and pox-like conditions in other birds in Australia.

Lesions, grossly and histologically typical of pox infection, occurred in a white-backed magpie from Melbourne, Australia. Electron microscopic examination revealed typical poxvirus particles in lesion material. The disease was experimentally transmitted to other magpies, but chickens, turkeys, pigeons, and canaries were refractory to experimental infection with magpie poxvirus. The epidemiology of magpie pox and the probable occurrence of pox-like disease in other native Australian birds are discussed.


Epithelioid granulomas in Hodgkin disease. A favorable prognostic sign?

Histologic sections of spleen and liver prepared from tissue obtained during exploratory laparotomy and splenectomy in 91 untreated patients with Hodgkin disease were reviewed to assess the incidence and possible implications of noncaseating, epitheliod, sarcoid-like granulomas. The 17 patients with granulomas and the 74 patients without granulomas did not differ appreciably with respect to pretreatment indexes. All patients were observed for at least 15 months following laparotomy or until the time of death (range, 15 to 67 months; median, 25 months). The 17 patients with granulomas tended to have fewer relapses, longer survival, and lower incidence of subsequent herpes zoster infection compared to the 74 without granulomas. The presence of epithelioid granulomas in association with Hodgkin disease may reflect a host response to the tumor with favorable prognostic implications.


Hepatitis types B and non-B. Epidemiologic background.

Two episodes of acute viral hepatitis occurred in each of 34 patients. One episode in each patient was serologically diagnosable as type B hepatitis on the basis of tests for hepatitis B surface antigen or antibody. The other episode was classified as "non-B" on the basis of seronegativity, reinforced by seropositivity in an alternate bout. An epidemiologic background appropriate to "serum" hepatitis, either transfusion (one bout) or illicit self-injection (46 bouts), was associated just as frequently with serologically non-B episodes as with identified type B disease. The diagnosis of type B hepatitis, therefore, should be made only on the basis of serologic tests specific for hepatitis B virus infection. Other cases of sporadic diseases in adults must be labeled "viral hepatitis, type unspecifiable."


Requirement of newly synthesized protein for the priming activity of interferon.

Pretreatment of mouse L cells with interferon (IF) enhanced IF production in response to polyinosinic-polycytidylic acid (poly I-poly C). Post-treatment of cells with IF caused no significant enhancement of IF production. The enhancing effect of IF pretreatment (priming) reached a maximum after incubation with IF (10 or 100 units/ml) for 4-6 hr at 37 C, but this effect was absent when the incubation was done at 4 C. Cells which were incubated for additional several hours at 37 C after IF pretreatment at 4 C did not develop the primed state nor the antiviral state. The presence of protein synthesis inhibitors during the IF pretreatment depressed, though not completely, the development of the primed state. The residual priming effect was lost when the cells were incubated with the inhibitors at 37 C for 2 hr before they were exposed to poly I-poly C. There was no significant difference in the binding rate of poly I-poly C to cells between IF-treated and untreated cells. The degradation rate of cell-bound poly I-poly C and its sensitivity to exogenous pancreatic ribonuclease in the pretreated cells were also similar to those in the untreated cells.


Participation of mono- and polysynaptic transmission during tonic activation of the stretch reflex arcs.

Through intracellular observations of the cat spinal motoneuron both mono- and polysynaptic reflex arcs were shown to participate in repetitive activation of stretch reflex. Experiments were performed on 25 cats anesthetized by intraperitoneal (i.p.) injection of 3 cc/kg of a mixture of 1% chloralose and 10% urethane. Driving of the motoneuronal discharges accomplished by electric stimulation (16-100 Hz) of the gastrocnemius lateralis and medialis nerves. Repeated electric stimulation of muscle nerves elicited monosynaptic "vibratory" EPSPs and a polysynaptic "augmenting" EPSP in the motoneuron. The firing of the motoneuron occurred when a temporal summation of the "vibratory" EPSPs was sufficiently obtained. The discharge frequency of motoneuron, MNf, was expressed as a product of electrostimulus frequency, Sf, and reciprocal of an integer, n, i.e. MNf=(1/n)Sf. An increase in the stimulus intensity resulted in a dimimution of the vibratory EPSP and a remarkable increase of n. This was assumed to be due to an inhibitory process produced by the Group II fibers. The minimum value of integer n was always a determinant factor for the maximum value of MNf which was referred to as the "preferred" frequency of the motoneuron. Long-lasting electric stimulation of muscle nerves was followed by a more remarkable recruitment of the augmenting EPSP which made the motoneuron attain its critical threshold of firing. Motoneuronal spikes elicited by such a large augmenting EPSP were not locked to the Ia impulses. Unlocked spikes were observed only in a later stage of electric stimulation of Ia fibers.


Antiphosphodiesterase activity and nonspecific smooth muscle relaxation tested on intestinal smooth muscles.

Mitochondrial, microsomal and soluble fractions separated from the guinea pig taenia and from the dog longitudinal smooth muscle were used as phosphodiesterase preparation. Each preparation had low and high Km values, indicating the existence of at least two kinds of phosphodiesterase. Papaverine and Aspaminol (1, 1-diphenyl-3-piperidinobutanol hydrochloride), hydralazine, caffeine Na benzoate and aminophylline were used at test drugs. Aspaminol had little inhibitory effect on phosphodiesterase. Ki value of papaverine almost equalled the concentration (M) which was necessary to produce 50% relaxation. Relaxation of the guinea pig taenia by papaverine was preceded by an increase of intracellular cyclic AMP,. Therefore, the action of papaverine is likely to be mediated by an increase in cyclic AMP, which is caused by inhibition of the phosphodiesterase-catalyzed breakdown of cyclic AMP. There was little correlation between relaxing activities of the drugs used and their antiphosphodiesterase activities. Relaxation of the smooth muscle induced by the smooth muscle relaxants excepting papaverine is not due to inhibition of phosphodiesterase.


Murine virus susceptibility of cell cultures of mouse, rat, hamster, monkey, and human origin.

Studies were initiated to determine the practicality of using various tissue cultures for the propagation of murine viruses isolated from laboratory animals. The cytopathogenic effects of 10 murine viruses known to cause disease in laboratory rodents were compared in monolayer cultures of L929, BHK-21, WI-38, BSC-1, and Vero cells. The susceptibility of primary hamster embryo, hamster kidney, mouse embryo, mouse kidney, and rat embryo cell cultures was also tested. Seven of the viruses produced effects in at least 1 of the cell substrates. The remaining 3 viruses, namely H-1, K, and mouse hepatitis, produced no effects in the cell cultures tested.


Application of agar-gel precipitin test for the detection of Sendai virus antibody in rat sera.

A simplified agar-gel precipitin test was performed for the detection of Sendai virus antibody in rat sera. A close correlation was observed between detection of antibodies by complement-fixation test and agar-gel precipitin test. No correlation was found between results obtained by hemagglutination-inhibition test and agar-gel precipitin test in sera with HI titer of less than 1:8.


Interaction of the 5'-ends of 28S RNA in dimerization of hamster ribosomes.

Free ribosomes extracted from hamster cells and 28S RNA purified from these ribosomes are known to form dimers. We find that spleen phosphodiesterase inhibits ribosomal dimer formation, but only when a free 5'-hydroxyl end group, produced by the action of alkaline phosphatase, is present. Hence, formation of dimer ribosomes probably involves interaction at or near the phosphorylated 5'-ends of 28S RNA. Dimer RNA molecules show a modal length, when measured on electrom micrographs, of 2.1 mum, which is about double the length of 28S RNA. Electron micrographs of 115S dimer ribosomes often show profiles consistent with our interpretation that in dimers the 28S RNA chains are loosely linked by their 5'-ends.


Duodenoscopy and endoscopic retrograde choledochopancreatography: present position in relation to periampullary and pancreatic cancer.

Duodenoscopy and endoscopic retrograde choledochopancreatography represent a major advance in the diagnosis of periampullary and pancreatic lesions. Instruments, techniques and complications are reviewed. A combination of endoscopy and ERCP will yield diagnosis in a high proportion of patients suspected of pancreatic or ampullary carcinomata. Information is obtained which is not only of diagnostic value but is important in the surgical treatment of such patients.


Radiologic diagnosis of pancreatic cancer.

Modern radiology makes possible an accurate diagnosis of pancreatic cancer in about 95 percent of cases. Conventional upper gastrointestinal study, isotopic scanning, hypotonic duodenography, and ultrasonography are suitable as screening procedures. Special methods of high accuracy - pancreatic arteriography, endoscopic pancreatography, and transhepatic cholangiography are used as needed for definitive diagnosis.


Carcinoma of the exocrine pancreas: pathologic aspects.

This discussion is based upon 205 autopsied cases of carcinoma of the pancreas. Sixty-six percent were male, eighty percent white, mean age 61.5 years. Fifty-three percent of tumors occurred in the head of the pancreas. Most lesions were moderately well-differentiated adenocarcinomas. Pathologic aspects of carcinoma of the pancreas are discussed.


The relationship of histology to the spread of cancer.

This paper investigates the question of whether different types of histology at the same site can affect the spread of metastases - that is, producing a greater or fewer number of metastases depending on the histological diagnosis. Autopsy records were the source of the data analyzed in this paper. The metastases were subdivided into four categories - central nervous metastases, endocrine metastases, metastases at various lymphatic areas, and a miscellaneous group of metastases called the "remainder" group. It was found that adenocarcinoma and oat-cell carcinoma of the lungs are more aggressive in their metastatic spread than squamous cell carcinoma at the same site. Adenocarcinoma of the cervix uteri had more widespread metastases than squamous cell carcinoma at the same site. No difference at all was detectable in the number of metastases, in men or in women, between transitional cell carcinoma and adenocarcinoma of the bladder and kidney. Cystadenocarcinoma of the ovaries appeared to be less widespread than adenocarcinoma (not otherwise specified) of the same site.


Malignant melanoma and carcinoma of the breast.

Five patients with breast cancer and malignant melanoma are reported. Two patients had a third primary malignancy. In 4 out of 5 patients the breast tumor was the initial tumor discovered, and in 4 out of 5 the second tumor evolved metachronously. No specific carcinogenic factor could be established. The low malignancy potential of the melanoma by pathologic criteria may explain the lack of previous reports of this association.


Oat cell carcinoma as a malignant apudoma.

The neural crest origin of cells secreting amine and polypeptide hormones (APUD cells) is a unifying concept. The relationship of the classical endocrine system to that of tumor hormone secretion can be explained by the diffuse migration and later neoplasia of these unique cells. This paper describes the APUD cell origins of oat cell carcinoma.


Cyclic AMP and adenylate cyclase in the inner ear.

The activity of adenylate cyclase and the steady state levels of cyclic AMP (cAMP) were determined in stria vascularis (SV) and organ of Corti (OC) of the guinea pig cochlea. The activities are 12 and 19 pmoles/mg dry weight/minute for OC and SV, respectively. The activity was increased two to four-fold by NaF. The base level of cAMP is 4.2 and 4.4 nmoles/g dry weight in OC and SV, respectively. In contrast to brain, neither ischemia nor barbiturates produced major changes of the steady state levels of cAMP. No in vitro effect of cAMP upon the state of activation of glycogen phosphorylase was noticeable in either tissue. cAMP did not exert a significant in vitro inhibition of strial Na+K+-ATPase. Perilymphatic perfusion of cAMP (10-3 M) and of theophylline (5 times 10-3 M) did not produce changes in the endolymphatic potential (EP), but dibutyryl cAMP (10-3 M) led to a significant increase of EP. The alpha adrenergic blocking agent, phentolamine, produced very complex changes of the cochlear potentials. A possible role of catecholamines and cAMP in the secretory phenomena of the SV and in the transduction and/or transmission processes of the auditory sense organ are discussed.


Phosphodiesterase and adenyl-cyclase activities in the cerebral cotex of the aging rat.

In an effort to determine the factors responsible for the four-fold decrease in cyclic-AMP content of the rat cerebral cortex, observed to occur between the ages of 3 and 6 months, studies were performed on adenyl-cyclase and phosphodiesterase, the cyclic-AMP synthesizing and hydrolyzing enzymes. The activities and kinetic characteristics were determined for both enzymes as obtained from the cerebral cortex of rats ranging in age from one to 24 months. No age dependence was observed either in adenyl-cyclase activity, assayed with or without fluoride ion, or in phosphodiesterase activity. It was concluded that age related changes in factors other than the direc levels of these enzymes underlie the age related decline in cortical cyclic-AMP levels.


Insulin and proinsulin release during calcium infusion in a patient with islet-cell tumor.

The infusion of calcium results in the release of gastrin, calcitonin, and serotonin from certain nonbeta islet cell tumors of the pancreas, medullary carcinomas of the thyroid, and carcinoid tumors, respectively. In this study, intravenous infusion of either calcium chloride or calcium aluconate in a patient with an islet-cell carcinoma resulted in a simultaneous rise in plasma immunoreactive insulin and proinsulin, and concurrent hypoglycemia. After resection of the tumor, calcium infusion caused no change in these parameters. Similarly, calcium infusion caused no change in plasma insulin or glucose in normal volunteers. The response of this tumor suggests that calcium infusion may be a useful provocative test to detect insulin-secreting neoplasia. A derangement of the stimulus-secretion coupling mechanism for insulin in the tumor cells may be responsible for their abnormal sensitivity to calcium ion.


Lp(a) lipoprotein: relationship to sinking pre-beta lipoprotein hyperlipoproteinemia, and apolipoprotein B.

To assess the relationship between the Lp(a) and the "sinking pre-beta" (d smaller than 1.006) lipoprotein, the concentration of Lp(a) was quantified by radial immunodiffusion and the presence or absence of sinking pre-beta was assessed by agarose electrophoresis in overnight fasting plasma samples from 485 adults, comprised of 320 with normal lipid levels, 48 with type IIa, 40 with type IIb, and 77 with type IV lipoprotein phenotypes. The median Lp(a) level was 7.6 mg/100 ml, 89% (433 of 485) having detectable Lp(a) levels. Twenty-two per cent (107 of 485) had detectable pre-beta lipoprotein in the d greater than 1.006 plasma fraction (sinking pre-beta). Of the sinking pre-beta positive plasma samples, 96% (102 and 107) exceeded the median Lp(a) level, and sinking pre-beta was detected in all 44 samples with an Lp(a) concentration exceeding 40 mg/100 ml. The relationship of Lp(a) and sinking pre-beta to lipoprotein phenotype was assessed. Compared to the normolipidemic group, the type IIa group had higher Lp(a) percentile values (p smaller than 0.02), whereas the IIb and type IV groups had significantly lower Lp(a) values than the normolipidemic group. Ninety-two per cent (296 of 320) of the normolipidemic subjects had detectable levels of Lp(a) and 22% (70 of 320) had detectable sinking pre-beta lipoprotein. Ninety-four per cent (45 of 48) of the type IIa plasmas had detectable Lp(a) levels and 27% (13 of 48) had sinking pre-beta lipoproteins. Contrasted with the IIa group, only 80% (32 of 40) of the IIb plasmas had detectable Lp(a) levels and 18% (7 of 40) had sinking pre-beta lipoprotein. In the type IV plasmas 78% (60 of 77) had detectable Lp(a) and 22% (17 of 77) had sinking pre-beta lipoprotein. Lp(a) or log Lp(a) levels were not correlated with apolipoprotein B levels (n = 485, r = 0.002 or 0.037, respectively). Furthermore, Lp(a) levels remained essentially constant in three subjects whose aprptein B levels were altered in response to pharmacological and/or dietary manipulation. A fourth subject had a 50% increase in Lp(a) but this change did not correlate with apoprotein B changes. Thus, these findings suggest that Lp(a) is metabolically independnet of low density lipoprotein even though it shares the same structural protein, apoprotein B.


Selective proteolysis of the receptor for parathyroid hormone in skeletal tissue.

Parathyroid hormone, calcitonin, and prostaglandin E2 activate the adenylate cyclase-cyclic AMP system in fetal-rat calvaria. These agents presumably interact with the tissue at separate receptor sites. When calvaria were preincubated with trypsin, 500 mug/ml for 45 min, the subsequent increase in 3',5'-AMP in response to parathyroid hormone was markedly diminished, whereas the response to calcitonin and prostaglandin E2 were not altered significantly. The effect was attributable to an action of the enzyme on the tissue and not to hydrolysis of the hormone. Similarily, preincubation of calvaria with trypsin prior to homogenization and preparation of a crude plasma membrane fraction decreased PTH-sensitive adenylate-cyclase activity by 58% but did not alter the degree of stimulation of the enzyme in response to calcitonin, prostaglandin E2, or sodium fluoride. These studies support the hypothesis that the actions of parathyroid hormone and calcitonin on bone are mediated through distinct receptor sites, and the receptors for parathyroid hormone can be altered selectively with trypsin.


The importance of geographical pathology.

Three different forms of cancer--the Burkitt lymphoma, nasopharyngeal carcinoma and skin cancer--are discussed in their relation to geographical distribution, the possible role of viruses, and genetic predisposition. Certain general conclusions in regard to cancer follow from these discussions. It is stated firmly that cancer is not a disease but a tissue manifestation which has not only many different presentations but also many different complex causes which consequently will need many different forms of treatment. A comparison of geographical differences is of immense value.


Counter immunoelectrophoresis in rapid diagnosis of pox virus infections.

The use of the counter immunoelectrophoresis technique enhanced the reaction between vaccinia virus and its antibody. By means of 0.7% and 1% solutions of agarose, sharp, clear lines of precipitation were obtained within 30 minutes at 200 volts.


[Spectacillin-a new-developed multi-effective penicillin in treating chronic bronchoulmonary diseases author's transl].

50 patients with a chronic disease of the bronchopulmonary system, within the therapy of an acute exacerbation received in addition to bronchosecretolytics, spasmolytic substances, and corticosterones the substance 'Spectacillin' in a dosage ranging from 2 to 6 grams per day during the period of 2 weeks. The acute exacerbation has been stopped in 34 patients and there-by the recurrence could be prevented. Although in 9 patients a clinical improvement has been obtained, no germ elimination could be ascertained; 7 cases of disease remained uninfluenced. The evalution is documentated by the clinical and laboratory findings, the germ analysis and the test of pulmonary functions. The compatibality can be evaluated as well-effective; especially the low rate of exanthema (2 patients) is to be emphasized.


A mammalian spot test: induction of genetic alterations in pigment cells of mouse embryos with x-rays and chemical mutagens.

Embroys heterozygous for five recessive coat-color genes from the cross C 57 BL/6 J Han x T-stock were x-irradiated with 100/r o r treated in utero with 50 mg/3 kg methyl methanesulfonate (MMS) and ethyl methanesulfonate (EMS), respectively. Controls consisted of irradiated embryos of C 57 BL x C 57 BL matings homozygous wild-type for the genes under study, and non-treated offspring of both types of mating. The colors of the spots were observed in the adult fur were either due to expression of the recessive coat genes or were white. I. Irradiated and mutagen-treated offspring of C 57 BL x T-stock matings had almost exclusively nonwhite spots, distributed randomly over the mouse surface. 2. Irraidated offspring of C 57 BL x C 57 BL matings had only white spots which were always midventral. 3. In non-treated offspring of both types of mating no spot could be observed. After correcting for white midventral spots observed in the one type of control, the frequency of expression of one or the other of the recessive color genes is calculated to be about 11% for embryos irradiated with 100r or 101/2 days postconception, about 1% for embryos irradiated with 100r at 9 days postconception, about &% for embryos treated with 50 mg MMS/kg at 101/2 days postconception, and about 8% for embryos treated with 50 mg EMS/2 days postconception. It is discussed that the white midventral spots are preferentially the result of pigment cell killing, while the nonwhite spots are preferentially the result of gene mutations or recombinational processes like mitotic crossing over and mitotic gene conversion. Of numerical and structural chromosome aberrations only those come into question which are able to pass the filter of several mitoses. Therefore, the test system described is supposted to cover not only heitable DNA-alterations, but the whole spectrum of them.


Some aspects of the mechanism of action of steroid hormones.

Events related to the very early steps of steroid hormone action are reviewed, after a brief exposé of the methods used for the measurement and calculation of protein-steroid interactions. A list of steroid binding plasma proteins is given, and their physiological variations and possible role(s) are discussed. The mechanism of entry of steroids into target cells is then considered especially the possibility of a protein mediated step. The metabolism of steroid hormone in target cells and its physiological significance are discussed, especially with reference to androgens in prostate, levator ani muscle and hypothalamus. No receptor of steroid hormone has yet been discovered if defined as the last entity with which the active steroid interacts before action is initiated. However, in the target tissues of steroid hormones, specific proteins have been detected (termed receptors) with properties that are compatible with an essential if not an obligatory role in steroid action. The properties ascribed to the cytosol receptors of different steroid hormones in various tissues, their binding properties, and their quantitative variations are reported. The transformation or activation of receptor after binding of the hormone is related to its transfer to the nucleus. The question posed by the recent observations on Non Histone Chromatin estrogen binding protein and of the "insoluble" nuclear receptor are discussed. Finally, a series of experiments are reviewed, dealing more specifically with the increase of protein and RNA synthesis in tissues in which steroids and RNA synthesis in tissues in which steroids promote growth (i.e. estradiol in uterus). The concept of a cascade phenomenon is reported, including the early synthesis of KIP (Key Intermediary Protein(s)) which would responsible for the secondary amplification of the response. It is discussed in connection with the present (and insufficient) knowledge of receptors and transcription machinery.


Aerosol polymyxin and pneumonia in seriously ill patients.

Pneumonia caused by Pseudomonas aeruginosa occurs frequently in critically ill patients and is associated with a mortality rate of 70 per cent. An aerosol of polymyxin B was administered (2.5 mg per kilogram per day) to the upper airways of 292 patients in a respiratory-surgical intensive-care unit during a seven-month period, in an attempt to prevent Ps. aeruginosa pneumonia. Although only one of the patients studied acquired pneumonia due to Ps. aeruginosa, 10 others acquired pneumonia caused by a polymysinx-resistant organism. Seven pneumonias were caused by organisms not frequently pathogenic to man (flavobacteria, serratia and Streptococcus faecalis). The mortality rate for acquired pneumonia in this study, 64 per cent, is greater than that in previous studies in which either no polymyxin or cyclic polymyxin therapy was used. Continuous use of polymyxin B aerosol appears to be a dangerous form of therapy.


Predominantly T-cell infiltrate in rheumatoid synovial membranes.

Lymphocytes infiltrating synovial membranes were characterized in eight patients with proliferative rheumatoid synovitis. Surface immunoglobulins were studied with use of immunofluorescence, and the C3 receptor was detected by adherence of red cells coated with antibody and complement - both are B-cell markers. Spontaneous rosette formation with sheep erythrocytes was used as a T-cell marker. To obtain viable lymphocytes in suspension, the villous synovium of five of these patients was digested with collagenase and deoxyribonuclease. Populations enriched in lymphocytes could be obtained by velocity sedimentation. Whereas only 9 to 35 per cent of lymphocytes bore surface immunoglobulins, the majority (70 to 85 per cent) formed sheep-erythrocyte rosettes. Cells bearing the C3 receptor constituted a distinct minority of synovial lymphocytes in frozen-tissue sections, and were found in follicle-like accumulations. These data indicate that the predominant infiltrating lymphocyte in proliferative rheumatoid synovitis is a T cell.


Characterization of alpha1-antitrypsin in the inclusion bodies from the liver in alpha 1-antitrypsin deficiency.

alpha1-antitrypsin was isolated from periodic acid-Schiff-positive inclusion bodies from the hepatocytes of patients with alpha1-antitrypsin deficiency and further purified to enable more detailed chemical analysis. Amino acid and cyanogen bromide fragmentation studies showed a close similarity between hepatic and serum (PiMM) antitrypsin in contrast to the carbohydrate analysis, which revealed markedly deficient glycosylation of hepatic antitrypsin. A complete lack of sialic acid and a relative deficiency of all other carbohydrate components could fully explain the difference of approximately 6000 daltons in molecular size between the two proteins. The accumulation of hepatic globules is probably related to the physical properties of the defective antitrypsin, which include marked insolubility and tendency toward aggregation. The results strongly suggest an abnormal amino acid sequence in the peptide chain of the deficient antitrypsin. The interference with glycosylation may be related to steric hindrance.


Limitations of the electrocardiographic response to exercise in predicting coronary-artery disease.

The electrocardiographic response to exercise was compared with the results of coronary angiography in 89 patients with Type II hyperlipoproteinemia who had previous myocardial infarction or typical angina or both (43 patients)(Group A), "atypical angina" (16 patients)(Group B)or positive electrocardiographic response to exercise without other evidence of cardiac disease (30 patients)(Group C).  Thirty-nine of 43 in Group A had greater than or equal to 50 per cent stenosis, and 26 (67%) of these 39 had negative exercise tests. In Group B, five of the 16 had greater than or equal to 50% stenosis, and three had positive exercise tests (one patient had a false-positive test).  In Group C, eleven of 30(37%) had greater than or equal to 50% stenosis; however, nine (30%) had minor stenosis (less than or equal to 50%), and 10(33%) normal coronary arteries. The diagnostic usefulness of exercise electrocardiography is limited.  False-negative responses are frequent in patients with clinically suspected coronary disease, and false-positive responses frequent in asymptomatic patients.


Stimulation of adenosine 3',5'-monophosphate formation in guinea-pig cerebral cortical slices in a calcium free medium.

In guinea-pig cerebral cortical slices cyclic AMP concentrations increase during incubation with histamine+noradrenaline. After 10 min of incubation the levels of cyclic AMP start to decline. When calcium ions are omitted from the incubation medium, cyclic AMP levels do increase to a greater extent under the same conditions and do not drop during 30 min incubation. In the presence of calcium ions cyclic AMP synthesis can not be elicited by noradrenaline alone. In calcium-free Krebs-Ringer solution a pronounced effect of noradrenaline on cyclic AMP levels is observed. This effect of noradrenaline is shown to be mediated by a classical alpha-type receptor. 5-Hydroxytryptamine, prostaglandin E1 and dopamine do not significnatly enhance cyclic AMP formation in guinea-pig brain slices in either the presence in, or the absence of calcium ions from the incubation medium. Under depolarizing conditions of incubation the stimulatory effect of ouabain or 125 mM K+ is blocked in a calcium-free medium, while with the depolarizing agent veratridine no significant reduction of cyclic AMP formed during incubation in a calcium-free medium is obtained.


[Electrical reaction of olfactory structure and the amygdaloid complex of the brain of dogs in a paradoxical stage of sleep].

Electrical activity of the olfactory bulb, olfactory tubercle amygdala, hippocampus, hypothalamus and neocortex during physiological sleep was studied in chronic experiments in unanesthetized unrestrained dogs;A synchronous high-frequency sinusoidal rhythm of 36-42 Hz was found in the amygdala and olfactory structures in the paradoxical phase of sleepmthis olfacto-amygdaloid rhythm, unlike that of aroused animals, was not the result of olfactory stimulation and persisted in dogs with trachael breathing; Its generation is probably of central origin; A comparison of the dynamics of this electrical activity with somatic, autonomic and teeg-correlates of sleep gives reasons to regard the olfacto-amygdaloid rhythm of 36-42 Hz as a specific EEG-correlate of the paradoxical sleep stage in dogs


Giant axonal neuropathy. A clinical entity affecting the central as well as the peripheral nervous system.

An 8-year-old girl had progressive muscle weakness and a unique posture of the lower limbs, areflexia, distal sensory impairment, and remarkably kinky hair. Histologic examination of the sural nerve showed giant axons filled with neurofilamentous masses. The clinical and histologic findings resembled those of recent cases reported as "giant axonal neuropathy." Our patient's precocious puberty, Babinski's sign, and electroencephalographic abnormalities suggested central nervous system involvement. Two cases previously reported and the present one appear to represent a new clinical entity that affects the central and the peripheral nervous system.


"Living cytology" in the diagnosis of intraabdominal adenocarcinoma.

Cytologic diagnosis of adenocarcinoma metastatic to the peritoneal surface is often hampered by the difficulty of differentiating between reactive mesothelial cells and macrophages from malignant epithelial cells by light microscopy. With in vitro tissue culture technics, the fibroblastic dedifferentiation of mesothelial cells facilitates the recognition of residual neoplastic epithelial cells. Ascitic fliid from 18 patients who ultimately had histologic documentation of intraabdominal adenocarcinomatosis was studied by tissue culture technics. In 15 of the 18 patients, small islets of replicating tumor cells could be identified after the mesothelial cells had dedifferentiated into fibroblasts. With one exception the average time between collection of the specimen and recognition of adenocarcinoma was 6 days. In 14 cases, a definitive diagnosis of malignancy was made by cytologic analysis.


Cervical carcinogenesis with herpes simplex virus, type 2.

In the past few years there have been a number of reports correlating a high frequency of herpes simplex virus type 2(HSV-2) infection with lesions of the uterine cervix. These studies have used a clinical history of herpetic infection or the demonstration of herpetic antibodies in the cancer patients. The present study was performed to evaluate any possible carcinogenic activity of the formalin-inoculated herpes simplex virus type 2 in the reproductive tract of the female mouse. This approach to the study was selected because of previous experience with a model system of carcinogenesis of the cervix uteri using coal tar hydrocarbons. Cytologic and histologic preparations from experimental animals and controls are presented to demonstrate the mucosal alterations and tumors observed in the animals. Noninvasive lesions of the cervix were identified in 76.8% and invasive adenocarcinoma detected in 30.2% of the mice.


Paget's disease of the vulva and carcinoma of the breast.

Vulvar Paget's disease is an intraepithelial change which is locally recurrent and has minimal if any propensity to invasion. Its major significance lies in the frequency with which concomitant carcinomas are discovered in association with the vulvar skin lesion. Adenocarcinoma of the vulvar apocrine glands is the most commonly associated tumor. Another apocrine adenocarcinoma, carcinoma of the breast, either antecedent or concomitant, is the second most frequently associated tumor, having been noted in 14 cases of vulvar Paget's disease. A case report illustrating this association is presented, and the importance of preoperative breast screening procedures in all patients with vulvar Paget's disease is demonstrated.


Liver tumors in women on contraceptive steroids.

The possible association between oral contraceptives and benign liver tumors has recently been reported. To date the majority of cases have been diagnosed as benign hepatomas (liver cell adenomas). We have had the opportunity to study 13 such cases. Eight have been examples of focal nodular hyperplasia of the liver; however, in addition, there were examples of hepatocellular carcinoma, liver cell adenoma, and possible liver cell hamartoma; all were in women on "the pill." Gynecologists are alerted to the fact that many of the patients present with symptoms of acute abdomen, syncope or shock, and intrahepatic or intraperitoneal bleeding. Prompt diagnosis and treatment may be lifesaving.


Viral replication in human ovarian cell culture.

A method is presented for the preparation and maintenance of human ovarian cell cultures. Sections of normal ovaries removed at surgery were minced, trypsinized, and seeded as cell cultures grown in minimal essential media at 37 C. Long-term, low-passage cultures were grown in quantities sufficient to permit viral studies. Ovarian cells, of passage 23, were challenged with 57 known viruses. Of these viruses, 52 (especially viruses of the picornavirus and adenovirus groups) produced typical cytopathic effects. The significance of these studies is discussed.


Murine tumor induction by cytomegalovirus.

Cytomegalic inclusion disease can be induced in mice with relative ease. In the study reported here, grandular neoplasms were produced in four mice by means of immunosupression together with large doses of cytomegalovirus. The results of this study suggest that cytomegalovirus may well be an etiologic agent in the formation of salivary gland tumors.


Peritonsillolith.

Peritonsillolith appears to be an unknown entity. Two cases of lithiasis in the peritonsillar region following spontaneous rupture of peritonsillar abscesses have been reported. The authors have speculated concerning the cause of stone formation in the peritonsillar region, which has been designated "peritonsillolith," on the basis of lithiasis in other regions of the body, such as gallbladder, kidneys, and salivary glands. Peritonsilloliths most likely form as a result of incomplete evacuation of pus, the dead bacteria and the pus cells providing the nidus for their formation. The salts of these stones are undoubtedly derived from the inflammatory exudates and saliva.


The central odontogenic fibroma. Clinical and morphologic studies.

The clinical, pathologic, and ultrastructural features and pertinent case history of an 11-year-old boy with a rare benign odontogenic neoplasm, the central odontogenic fibroma, are presented. A review of the literature reveals seven other lesions with clinical and pathologic findings similar to those of this case. This study supports the concept that the central odontogenic fibroma is a distinct odontogenic neoplasm of bone which occurs most commonly in the mandible as a multilocular, radiolucent, and slowly growing expansile lesion with no tendency to recur after surgical enucleation. The ultrastructural findings indicate that the central odontogenic fibroma and the odontogenic myxoma share many common morphologic features and have an apparently similar histogenesis.


Congenital granular-cell tumor of alveolar ridge and tongue. Report of two cases.

Granular-cell tumors in the oral cavity occur primarily on the tongue (myoblastoma) in adults and on the gum pads (congenital epulis) in infants. Because of the usual location and age, these histologically and histochemically similar tumors have been separate clinical entities. These case reports, however, report the concurrence of both clinical entities in the same patient. A 7-day-old female infant had a 1 cm. epulis on the right anterior maxilla and two 1.5 mm. masses on the anterior ventrum of the tongue. A 12-day-old girl had a 1 cm. epulis on the anterior mandible and two 1.5 mm. masses on the anterior tongue near the tip. In both cases, all lesions were histologically and histochemically similar and were granular-cell tumors. This suggests a relationship in the origin of these entities and tends to rule out an odontogenic origin for the epulis.


Shock synthesis of amino acids II.

The synthesis of amino acids behind shock waves in methane, ethane, ammonia, and water vapor was further investigated. Aldehydes and HCN are formed separately in the non-homogeneous gas during the high temperature period and recombine with ammonia during the thermal quench period, to form chi-amino nitriles. The chi-amino nitriles are either hydrolyzed by excess water vapor during the quench period or emerge as such after the reaction is completed. A combined gas chromatograph-mass spectrometer analysis of the reaction product showed identical amounts of D and L amino acids, thus confirming the absence of contaminants. Thunder shock waves were shown to be a suitable source of energy for the production of amino acids.


Quantum chemical study of the thermodynamics, kinetics of formation and bonding of H2CN: relevance to prebiotic chemistry.

Using the Iterative Extended Huckel Theory (IEHT), energy-conformation studies have been carried out for H2CN (I), trans-HCNH (IIA), and cis-HCNH (IIB), three possible isomers formed by addition of a hydrogen atom to hydrogen cyanide. Calculations show that the order of decreasing thermodynamic stability is I greater than IIA greater than IIB. Additionally, from calculated energies along simulated reaction pathways, the formation of I from HCN+H appears to be kinetically favored over IIA. Calculated properties of the minimum energy conformers of I and IIA are described and the potential role of H2CN (I) as a reactive intermediate in prebiotic organic synthesis and its possible relevance to interstellar organic chemistry are discussed.


Macromolecules and the origin of life.

From our knowledge of present day organisms, it is hard to imagine a living assembly, even at its most primitive stage, without macromolecules. In order to look for the macromolecules which possibly participated in the assembly of the primitive organisms, the reaction and formation of polymers in HCN under irradiation of ultraviolet ray of 184.9 nm. was studied. As an example of a simple way of producing an assembly of macromolecules, the mechanism of coacrevation was studied by using gelatin as the material.


The origin of proteins: Heteropolypeptides from hydrogen cyanide and water.

Evidence from laboratory and extraterrestrial chemistry is presented consistent with the hypothesis that the original heteropolypeptides on Earth were synthesized spontaneously from hydrogen cyanide and water without the intervening formation of chi-amino acids, a key step being the direct polymerization of atmospheric hydrogen cyanide to polyaminomalononitrile (IV) via dimeric HCN. Molecular orbital calculations (INDO) show that the most probable structure for (HCN)2 is azacyclopropenylidenimine. Successive reactions of hydrogen cyanide with the reactive nitrile side chains of IV then yield heteropolyamidines which are converted by water to heteropolypeptides. To study this postulated modification of a homopolymer to a heteropolymer, poly-chi-cyanoglycine (IX) was prepared from the N-carboxyanhydride of chi-cyanoglycine. Hydrolysis of IX, a polyamide analog of the polyamidine IV, yielded glycine. However, when IX was hydrolysed after being treated with hydrogen cyanide, other chi-amino acids were also obtained including alanine, serine, aspartic acid and glutamic acid, suggesting that the nitrile groups of IX (and therfore of IV) are indeed readily attacked by hydrogen cyanide as predicted. Further theoretical and experimental studies support the view that hydrogen cyanide polymerization along these lines is a universal process that accounts not only for the past formation of primitive proteins on Earth, but also for the yellow-brown-orange colors of Jupiter today and for the presence of water-soluble compounds hydrolyzable to chi-amino acids in materials obtained from environments as diverse as the moon, carbonaceous chondrites and the reaction chambers used to simulate organic synthesis in planetary atmospheres.


Prebiotic nucleotide synthesis-demonstration of a geologically plausible pathway.

Mineral phosphate (apatite) is activated for the synthesis of nucleotides when dilute solutions containing nucleoside and ammonium oxalate are evaporated in its presence. A natural, igneous fluorapatite was found to be even more effective in nucleotide synthesis than the more soluble hydroxylapatite. The phosphorylation is considerably more efficient if urea of cyanamide is also present. Hydrolysis of solutions of cyanogen to form oxalate and urea among other products is a spontaneous process that provides a geologically plausible model for necleotide synthesis on the primitive earth.


Models of prebiological phosphorylation.

The hypothesis that contemporary metabolic pathways envolved from analogous chemical reaction sequences on the primitive Earth leads to a reexamination of models of prebiological phosphorylation. Present-day phosphate uptake by algae and bacteria seems to involve two transport systems: (a) A n active transport process occurring at low external phosphate concentrations (as in umpolluted natural waters), with a transport constant Ks of 10(-7) to 10(-6) M Pi. (b) Another (probably diffusive) process at higher phosphate concentrations (greater than 10-6 M)(as in the interstitial water of reducing sediments). Laboratory model experiments are described for the reation of reducing sugars with orthophosphate in the presence of cyanogen, producing glycosyl phosphates. These reactions proceed with appreciable yields only at high phosphate concentrations (greater than 10-3 M), and may thus possibly serve as simulations of prebiological phosphorylation with diffusive transport, as it may have occurred in the intestial water of reducing sediments.


Evolution of photosystems of photosynthetic organisms.

It is generally accepted that two photosystems function successively in photosynthetic electron transport chain of plants and algae. The interaction of these photosystems results in the enhancement of photosynthesis. It was suggested that only one photosystem is present in purple bacteria, the most primitive photosynthetic organisms. The functioning of this photosystem is accompanied by absorption changes at 890 nm. Recently new spectral changes were found in Chramatium chromatophores under reductive conditions, more favorable for bacterial growth. Some of that spectral changes take place even at liquid nitrogen temperature. It is proposed these absorption changes could be related to other photosystem functioning in low potential region. Such a photosystem is necessary for reduction of NAD in Chromatium, for which the reverse electron transport to NAD was not shown. In contrast to photosystems of plants, the bacterial photosystems appear to function independently because the enhancement of bacterial photosynthesis is not found. Apparently the evolution of photosystems involved interaction between independent photosystmes, one of them functioning under more oxidative conditions.


Hormonal control of glucose metabolism.

Glucose homeostasis is accomplished through two major hormones, glucagon and insulin. These hormones, of course, must work in concert with other permissive hormones such as glucocorticoids and catecholamines to bring about a multiplicity of physiological processes such as gluconeogenesis, glycolysis, lipolysis, and proteolysis that maintain the organism's biochemical integrity.


[The effect of asbestos cement, UICC asbestos samples and quartz on the peritoneum of the mouse].

Aqueous suspensions of asbestos cement powder injected experimentally into the peritoneal cavity of mice act as a fibrogenic agent, as do chrysotile asbestos or chrysotile asbestos-containing soil samples. The fibrotic nodules caused by the dust resemble morphologically silicosis granulomas. In addition, asbestos cement has a characteristically strong cytotoxic effect during the first 2 weeks of the experiment. It is suggested that this is due to the chrysotile asbestos and/or the calcite component of the powder. Amosite and crocidolite, on the other hand, induce a diffuse peritoneal fibrosis with the appearance of numerous foreign body giant cells and asbestos bodies. Dust particles displaced to the regional lymph nodes are frequent in the animals treated with quartz, asbestos cement and asbestos-containing soil samples. A spindle cell type sarcoma arising from the visceral peritoneum is observed in animals injected with crocidolite or asbestos cement. In addition, dusts containing chrysotile asbestos induce considerable amyloidosis of the liver and spleen.


Influence of sex and sex hormones on transplantable hepatocellular carcinoma in the rat.

Palpable subcutaneous transplants of hepatocellular carcinoma-35 appeared slightly earlier in male animals; however, the number of successful growths was no greater than that in female animals. Castration and administration of testosterone or diethylstilbestrol were performed after the transplants reached 1.0-1.5 cm in size. The carcinoma was less well differentiated histologically, had more bile pigment, grew rapidly, mestastasized sooner and killed the host quickly in castrated females given testosterone propionate. Bile was present in lung metastases. There was little difference in the growth rate in intact or castrated male or female animals. Exogenous diethylstilbestrol slowed the growth of the transplants and cause weight loss in castrated males. The weight loss was felt to be related to extensive necrosis of the carcinoma.


Rhinosporidium seeberi: spherules and their significance.

This study of the nature and functional significance of the spherules that develop in sporoblasts show them to participate in the germination of Rhinosporidium seeberi and are the precursors of the trophocytes.


Photochemical synthesis of simple organic free radicals on simulated planteary surfaces-an ESR study.

Electron spin resonance spectroscopy provided evidence for formation of hydroxyl radicals during ultraviolet photolysis (254 nm) at -170C of H2O adsorbed on silica gel or of silica gel alone. The carboxyl radical was observed when CO or CO2 or a mixture of CO and CO2 absorbed on silica gel at -170C was irradiated. The ESR signals of these radicals slowly disappeared when the irradiated samples were warmed to room temperature. However, re-irradiation of CO or CO2, or the mixture CO and CO2 on silica gel at room temperature then produced a new species, the carbon dioxide anion radical, which slowly decayed and which was identical with that produced by direct photolysis of formic acid adsorbed on silica gel. The primary photochemical process may involve formation of hydrogen and hydroxyl radicals by means of (1) photodissociation of H2O physically adsorbed on the silica gel, or (2) absorption of the excitation energy by the silica gel surface with subsequent cleavage of the silanol bonds, or (3) dissociation of H2O molecules throug photosensitization by the surfaces or a combination of (1) to (3). Subsequent reactions of these radicals with adsorbed CO or CO2 or both yield carboxyl radicals, CO2H, the precsursors of formic acid. Our results confirm the formation of formic acid under simulated Martian conditions and provide a mechanistic basis for gauging the potential importance of gas-solid photochemistry for chemical evolution on other extraterrestrial bodies, on the primitive earth and on dust grains in the interstellar medium.


Malakoplakia of the prostate: a morphological and biochemical study.

A case of malakoplakia of the prostate is presented. Electron microscopic appearances support the origin of the Michaelis-Gutmann bodies from phagolysosomes in the histiocytes characteristic of the lesion. Biochemical analysis revealed the presence of muramic acid in the prostate with malakoplakia. This amino sugar is characteristic of bacterial cell walls and despite the absence of demonstrable bacteria in the affected tissues indicates the involvement of bacteria in the disease process.


High frequency discharge experiment. I: Formation of organic compounds from methane and ammonia.

Investigations of the synthesis of organic compounds from methane and ammonia were carried out by using high frequency discharges. The radical of CN was observed in the early phase of the discharge. Therefore it is considered that the CN radical contributes to the successive reaction steps. An important intermediate, a tetramer of hydrogen cyanide was observed to be formed at room temperature. The amounts of this compound progressively increased with time. It was speculated that the purines which were identified in the refluxed sample were synthesized through the above tetramer. Compounds which are thought to be heterocyclic (max:250-265nm) were initially formed in the sample trapped at liquid nitrogen temperature. After hydrolysis with hydrochloric acid, Gly, Asp Ala, etc. were observed in the sample. Overall yield of organic compounds was estimates at 5-6% based on input carbon of methane.


Formation of prebiochemical compounds in models of the primitive earth's atmosphere. I: CH4  NH3 and CH4  N2 atmospheres.

In order to understand the formation of organic compounds in the primitive atmosphere, the first steps of evolution in models of the primitive atmosphere were investigated. Mixtures containing C-H-N elements were subjected to a low pressure silent electric discharge for several seconds, and the resulting effluents were analysed mainly by gas chromatography, infrared spectrometry and chemical analysis. The formation of hydrocarbon (i.e. ethylene, acetylene, methylacetylene) and of nitrogen containing compounds (i.e. hydrogen cyanide, cyanogen, saturated nitriles, acylonitrile, cyanoacetylene) is reported. The influence of the initial mixture composition on the amount of compounds formed was systematically studied. The nature of the nitrogen source (N2 or NH3) in the primitive atmosphere has a great influence on the amount and on the very nature of the synthesized products. It is shown that important precursors such as cyanogen and cyanoacetylene are formed only in very rich N2 mediums. These results show the important role played by the nature of the primitive atmosphere in the determination of the chemical evolution pathways.


The natural history of the Silver-Russell syndrome: a longitudinal study of thirty-nine cases.

The growth of 39 children with Silver-Russell syndrome has been followed for 1-13 years. Pregnancy and labor were normal; none of the 61 sibs had the syndrome. Height at referral (mean age 4.6 years) averaged 3.6 SD below the mean and remained at this level during subsequent growth. Bone age averaged 69 percent of normal at referral but caught up by puberty, which occurred at the normal time. Nineteen cases were treated with human growth hormone without lasting effect. There is no clear-cut distinction between the Silver and Russell syndromes; the name should be Silver-Russell. It is likely that some 10 percent of cases have birth weights in the minus 1.5 to minus 2.0 SD range.


The action of neurotransmitter hormones and analogues and cyclic nucleotides and theophylline on electrolyte transport by the excretory duct of the rabbit mandibular gland.

The effects of autonomic drugs on electrolyte transport by the main duct of the rabbit mandibular gland were studied in vitro. Acetylcholine, in concentrations as low as 10(-10) M, reduced nett Na+ reabsorption and partially depolarized the transepithelial potential difference (P.D.); the effects were blocked by atropine. Acetylcholine was relatively ineffective applied from the luminal rather than the interstitial surface of the duct. Noradrenaline and isoproterenol produced similar effects to acetylcholine but the minimum effective concentrations were much higher (10(-7) M and 10(-3) M, respectively). It was concluded that the duct cells possessed specific muscarinic receptors on the basal cell membrane but that adrenergic receptors, particularly beta receptors, were either scanty or lacking altogether. Theophylline (10(-5) M) could mimic the effects of the autonomic drugs and produced a dramatic potentiation of the action of acetylcholine. Both cAMP and cGMP (4 times 10(-5) M) mimicked the acetylcholine response but cAMP was only effective when applied to the luminal cell membrane. It is proposed that cAMP is the intracellular mediator of the acetylcholine response and that it produces some of its effects by acting on an enzyme system close to the apical cell membrane.


[Biological examination in hyperlipemia].

The authors review the various methods of investigation useful in the classification of hyperlipoproteinaemias. Estimation of cholesterol and triglycerides remain the most important elements but do not suffice alone to reach a conclusion and measurement of phospholipids would seem to be a useful addition. Electrophoresis on agar, isolating a larger quantity of prebetalipoproteins, does not always facilitate classification. Finally, ultracentrifugation is often necessary in those forms which are difficult to classify, these representing almost 20 p.cent of patients.


Catalytic oxidation of two ribonucleosides in the 5'-position.

Guanosine and 7-beta-D-ribofuranosyltheophylline was oxidized to the corresponding nucleoside 5'-carboxylic acids by molecular oxygen and a platinum catalyst.


The 5' end group of tobacco mosaic virus RNA is m7G5' ppp5' Gp.

RNA extracted from CsC1-purified virions of tobacco mosaic virus is shown to give rise to an unusual nucleotide on digestion which RNAase T2, in addition to the four major nucleotides. This minor component has the electrophoretic characteristics of a phosphorylated end group, but is partially resistant to bacterial alkaline phosphatase. It is, however, a substrate for venom phosphodiesterase or nucleotide pyrophosphatase, yielding products which imply the structure m7G5'ppp5'Gp. TMV RNA, like many animal cellular and viral mRNAs recently examined, therefore has a 5' terminus blocked by a methylated nucleotide inverted with respect to the rest of the chain.


Non-random frequencies of short oligonucleotide sequences within poliomyelitis and encephalomyocarditis virus RNAs.

In poliomyelitis and encephalomyocarditis virus RNAs the relative frequencies of several short oligonucleotide sequences differ substantially from those expected on a random basis. The frequencies of CpGp-containing sequences are low, and the frequency of GpUpApGp is considerably lower than that of GpApUpGp. The relationship of these findings to those obtained from host cell RNAs is descussed.


Effect of selected environmental treatments on the incidence of gross Marek's disease lesions in chickens.

The effects of selected environmental treatments on the incidence of gross Marek's disease (MD) lesions in chickens contact exposed to MD virus were examined. Treatments were high levels of airborne decomposition products from chicken litter in the rearing environment, disinfectants applied to chickens and their environments, and formaldehyde fumigation of eggs and newly hatched chicks. There were no significant differences in the incidence of gross MD lesions when treated and untreated groups were compared.


Serological responses of broiler-type chickens, with and without Newcastle disease and infectious Bronchitis vaccine, to experimental infection with Mycoplasma synoviae by foot pad, air sac and aerosol.

Serum plate (SP) and tube agglutination (TA) reactions and geometric mean Mycoplasma synoviae (Ms.) hemagglutination-inhibition (HI) titers were determined on chickens infected at 21 days of age with Ms. by foot pad, air sac and aerosol. One-half had been given Newcastle disease (ND) and infectious bronchitis (IB) vaccine in the water at 20 days of age, and the other half had not been vaccinated with ND and IB vaccine. Blood was taken at 3, 4, 6 and 8 weeks after Ms. infection. Geometric mean HI titers were determined on serums of chickens that had a foot pad injection of Ms. 8 weeks after Ms. infection and were bled at 1, 3 and 5 weeks after foot pad booster. Three weeks after aerosol exposure with Ms., the geometric mean HI titer was significantly higher in the ND- and IB-vaccinated birds than in the nonvaccinated birds. In the Ms. air sac-infected group of nonvaccinated birds the geometric mean HI titer had increased significantly 3 weeks after food pad challenge with Ms. In the Ms. air sac-infected group of vaccinated birds, the geometric mean HI titer had significantly increased 3 and 5 weeks after foot pad challenge with Ms.


Activity of liver fructose diphosphatase from chick embryos treated with aminoguanidine sulfate.

Injection of chick embryos with aminoguanidine sulfate (AGS) on the fourth day of incubation resulted in a decreased specific activity of liver fructose diphosphatase (FDPase) prior to hatch time. This decreased FDPase specific activity was found to be the consequence of increased levels of an enzyme inhibitor (adenosin 5'-monophosphate) rather than a specific repression of enzyme synthesis.


Metabolism of some ethylenediamine derivatives with antiarrhythmic properties. I. Absorption, distribution in tissues and excretion of 14-C labelled N, N'-bis-[3-(p-methoxyphenoxy)-2-hydroxypropyl]-ethylenediamine dimethanesulphonate (MK-142) in rats.

14-C labelled MK-142 (specific activity 2.875 muCi/mg) (1.762 mCi/mM) passes very rapidly from the blood stream to the majority of tissues since 5 min after the iv administration the blood contained only about 5% of the administered dose. The relatively high content of MK-142 in the myocardium as compared with its content in the skeletal muscles indicates a high affinity of the examined substance to the heart muscle. MK-142 is excreted via kidneys, alimentary tract and respiratory ways.


Molecular sizes of egg yolk very low density lipoproteins fractionated by ultracentrifugation.

Egg yolk very low density lipoproteins isolated from normal eggs were separated into six fractions by preparative ultracentrifugation. The sizes of the molecules in three of the fractions were estimated by gel chromatography and by electron microscopy of the lipoproteins using a negative staining technique. The three fractions had average diameters of 39 nm., 32 n., and 25 nm. and average molecular weights of 17 X 10(6), 10 X 10(6), and 5 X 10(6). There was considerable overlap of size of molecules in the three fractions. Size differences were not observed in the ultracentrifuge separated fractions of very low density lipoproteins isolated from eggs laid by hens receiving crude cottonseed oil in their diets. However, the fatty acids from these fractions differed in stearic and oleic acid contents while those from the normal egg lipoproteins did not. The floating very low density lipoproteins contained more oleic acid and less stearic acid then the sedimenting ones did.


Pharmacological studies on 2- and 4-pyridylethylamides of acetyltropic acid (PAT-2 and PAT-4).

Two new compounds: 2- and 4-pyridylmethylamides of acetyltropic acid (PAT 2 and PAT-4) show spasmolytic effects on isolated guinea pig ileum, have mydriatic actevity in rabbits and block restraint produced ulceration in rats. PAT-4 causes a strong inhibition of the salivary secretion in mice. Both of the test compounds inhibit also the effects of ACh and vagus stimulation on the blood pressure and have local anaesthetic activity. PAT-4 inhibits the aggressive behaviour in mice and rats, prolongs the hexobarbital sleeping time, increases the pain treshold and inhibits the oxotremorine induced tremor in mice. Both PAT-2 and PAT-4 had no effect on the convulsant action of pentetrazole. EEG findings show that central cholinolytic activity is present in both compound tested.


Reduction of aflatoxin effects of quail.

A natural occuring outbreak of aflatoxicosis was observed in Bobwhite quail chicks while on a brooding experiment. A slight reduction in effects was noted for birds which received Tylan in the water from 0-3 days and Terramycin from 0-35 days of age. No symptoms of aflatoxicosis were observed in birds which received a product (FloxAid) containing two antibiotics and eight vitamins from 0-14 days. It is postulated that the primary mode of protection was via the water soluble vitamin D content of the product with possibly some protection from vitamins A, E and K.


Isolation and characterization of collagen-synthesizing polysomes from chick embryos.

Collagen-synthesizing polysomes were isolated by low-speed centrifugation of the post-mitochondrial supernatant of chick homogenates. Electron microscopy of the fraction thus isolated shows it to be exclusively composed of ribosomes. Amino acid incorporation in vitro showed that these particles were efficient in the incorporation of proline, but not tryptophan, as opposed to ribosomes obtained from the supernatant of the low-speed centrifugation. The incorporation process was highly dependent on GTP, and exibited an optimal Mg2+concentration of 5.6mM. The reaction was inhibited by RNase, elongation inhibitors as anysomycin, sparsomycin, fusidic acid and GDPCP. It was also moderately inhibited by initiation inhibitors such as aurintricarboxilic acid and pyrocatechol violet. The product of the incorporation was characterized as collagen by its sensitivity towards purified collagenase, lack of tryptophan, chromatography in CM-cellulose and molecular sieve chromatography in Sephadex G-200.


Regulation of sensitivity to beta-adrenergic stimulation in induction of pineal N-acetyltransferase.

Stimulation by isoproterenol causes large increases in the activity of rat pineal N-acetyltransferase (arylamine acetyltransferase or acetyl-CoA:arylamine N-acetyltransferase, EC 2.3.1.5) after a variable lag period. Lengthening periods of exposure to light cause rapid increases in the sensitivity of pineal N-acetyltransferase to induction by isoproterenol. This supersensitivity, which develops gradually over a 12 hr period, is correlated with increasingly longer lag periods in the induction of the enzyme, and with progressively greater maximal response. Repeated administration of isoproterenol to supersensitive animals rapidly reverses the above changes and causes relative subsensitivity. The sensitivity of N-acetyltransferase to induction by dibutyryl-cyclic tamp added to pineals in organ culture was found to change in parallel with the sensitivity to isoproterenol. Stimulation of cyclic AMP levels in the pineal by isoproterenol was also greater in supersensitive rats than in the subsensitive animals, whether the supersensitivity had been caused by denervation or by exposure to light for 12 hr. The above experiments suggest that there are two sites for the regulation of the sensitivity of N-acetyltransferase to induction by catecholamines. The first site regulates the capacity of intracellular processes (i.e., induction of tn-acetyltransferase) to respond to cyclic AMP as a second messenger.


Reversible dissociation of a carbamoyl phosphate synthase-aspartate transcarbamoylase-dihydroorotase complex from ovarian eggs of Rana catesbeiana: effect of uridine triphosphate and other modifiers.

Glutamine-dependent carbanoyl phosphate synthase [ATP6carbamate phosphotransgerase (dephosphorylating), EC 2.7.2.9], aspartate transcarbamoylase (carbamoylphosphate: L-aspartate carbamoyltransferase, EC 2.1.3.2) and dihydroorotase (L-5,6-dihydroorotate amidohydrolase, EC 3.5.2.3), are copurified as a high-molicular-weight complex from extracts of unfertilized eggs of Rana catesbeiana. UTP is required to maintain the integrity of the complex during the last two purification steps. Removal of the nucleotide results in dissociation of the complex. Based on sedimentation behavior in glycerol gradients, the dissociated carbamoyl phosphate synthase has an apparent molecular weight of 260,000 +/- 20,000 and that of dihydroorotase is estimated at 280,000 +/- 20,000. Aspartate transcarbamoylase is broadly distributed over the gradient. The addition of ATP, 5-phosphoribosyl-1-pyrophosphate, Mg++, or inorganic phosphate to the dossociated complex results in the appearance of a peak of aspartate transcarbamoylase activity with an apparent molecular weight of 110,000 +/- 10,000. Icubation of a mixture of the dissociated enzymes with UTP and Mg++ leads to their reassociation into the high-molecular-weight complex.


Motion of subfragment-1 in myosin and its supramolecular complexes: saturation transfer electron paramagnetic resonance.

Molecular dynamics in spin-labeled muscle proteins was studied with a recently developed electron paramagnetic resonance (EPR) technique, saturation transfer spectroscopy, which is uniquely sensitive to rotational motion in the range of 10(-7)-10(-3) sec. Rotational correlation time (tau2) were determined for a spin label analog of iodoacetamide bound to the subfragment-1 (S-1) region of myosin under a variety of conditions likely to shed light on the molecular mechanism of muscle contraction. Results show that (a) the spin labels are rigidly bound to the isolated S-1 (tau2 = 2 x 10(-7) sec) and can be used to estimate values of tau2 for the S-1 region of the myosin molecule; (b) in solutions of intact myosin, S-1 has considerable mobility relative to the rest of the myosin molecule, the value of tau2 for the S-1 segment of myosin being less than twice that for isolated S-1, while the molecular weights differ by a factor of 4 to 5; (c) in myosin filaments, tau2 increases by a factor of only about 10, suggesting motion of the S-1 regions independent of the backbone of the myosin filament, but slower than that in a single molecule; (d) addition of F-actin to solutions of myosin or S-1 increases tau2 by a factor of nearly 10(3), indicating strong immobilization of S-1 upon binding to actin. Saturation transfer spectroscopy promises to provide an extremely useful tool for the study of the motions of the crossbridges and thin filaments in reconstituted systems and in glycerinated muscle fibers.


Inhibition by interferon of delayed-type hypersensitivity in the mouse.

The effect of interferon on delayed-type by persensitivity to picryl chloride and sheep erythrocytes was examined in the mouse. When administered to sensitized animals on the day before or the day of challenge, tissue culture interferon inhibited both the ear swelling induced by pieryl chloride and footpad swelling induced by sheep erythrocytes. Newcastle disease virus, when injected into sensitized If-1l or If-1h congenic mice a few hours before challenge, caused an inhibition of delayed-type hypersensitivity which could be related to the amount of serum interferon induced by the virus. These results indicate that interferon production may represent one of the factors responsible for the depression of cell-mediated immune reactions during virus infection.


Anatomy of herpes simplex virus DNA: strain differences and heterogeneity in the locations of restriction endonuclease cleavage sites.

Digestion of herpes simplex virus DNA by the HinIII or Eco RI restriction endonucleases yielded 11 to 15 fragments with molecular weights between 1 x 10(6) and 28 x10(6). The electrophoretic profiles obtained in 0.3% agarose gels with DNA fragments from none different strains of herpes simplex virus type 1 could be readily differentiated from the patterns exhibited by the corresponding fragments from four separate strains of type 2 virus; however, with each serotype, the laboratory strains differed significantly among themselves and also from isolates passaged a minimum number of times outside the human host. Digestion of all DNAs of herpes simples virus with either enzyme reproducibly generated two classes of fragments (major and minor) which differed in molar ocncentration. Moreover, although the molecular weight of an intact herpes simplex 1(F1) DNA molecule is approximately 98 x 10(6), the summed molecular weights of all major and minor HinIII fragments totalled 160 x 10(6), and the seven major fragments alone accounted for only 60 x 10(6). These unusual features indicate the existence of limited heterogeneity in the positions of cleavage sitet along individual molecules. We have eliminated the possibility that minor fragments arose from contamination with the defective DNA of high byoyant density which appears on serial undiluted passage of the virus. In fact, this latter type of DNA was resistant to cleavage by HinIII and gave large amounts of only two species of EcoRI fragments; suggesting that the defective molecules consist of many tandem repeats of a small segment of viral DNA. The heterogeneity in the viral DNA of normal density appears to be related to the structural organization of the molecules and does not necessarily imply differences in genetic content.


Action of feedback regulator on adenylate cyclase.

A factor [the feedback regulator (FR)] formed by adipocytes after the stimulation of a cAMP raising hormone has been found to be a potent inhibitor of membrane-bound adenylate cyclase [EC 4.6.1.1.; ATP pyrophosphate-lyase (cyclizing)]. In a standard assay system using rat adipocyte plasma membrane as the source of adenylate cyclase, the FR inhibited adenylate cyclase by lowering the Vmax without affecting the apparent Km for ATP (0.3-0.6 mM). The apparent Ka for epinephrine (5-6 muM) was also not affected by FR. The inhibitory action of FR was partially countered by Mg2+ ions. An increase in phosphorylation of plasma membrane was observed when FR was present in the incubation system. The concentration required for a 50% inhibition was four times higher when adenosine 5-(beta,gamma-imino) triphosphate [AMP-P(NH)P] replaced ATP as the substrate for adenylate cyclase, implying that adenylate cyclase was inactivated by phosphorylation caused by FR. Increase in FR inhibition obtained by adding low concentrations of adenosine 5-(alpha,beta-methylene) triphosphate or ATP to AMP-(NH)P as the substrate supports this view. The inhibitory action was reversible. These results are consistent with the previously reported phenomena that (1) the undue to the formation of FR, and (2) the recovery of responsiveness of the stimulated cells by washing the cells with regular buffer medium is a result of the removal of FR. The hormone-initiated biphasic curve of cAMP levels in adipocytes is believed to be due to the negative feedback action of FR on adenylate cyclase. The mechanism of action of FR on inhibition of adenylate cyclase appears to be related to the phosphorylation of certain membrane components.


Nuclear receptors for thyroid hormone: evidence for nonrandom distribution within chromatin.

Chromatin receptor proteins appear to mediate some actions of thyroid hormone. In this study, sheared mammalian chromatin containing [125I]triiodothyronine (T3) bound by these receptors was separated using sucrose gradient velocity sedimentation. T3-receptor complexes were distributed throughout the chromatin fractions, but were enriched in the slowly sedimenting fractions. The latter contain most of the template capacity for RNA synthesis and most of the endogenous RNA polymerase activity but a minor portion of the total DNA. Formaldehyde treatment of chromatin containing receptor-bound [125 I ]T3 resulted in fixation of radioactivity, as evidenced by its migration with chromatin after equilibrium density gradient sedimentation in both cesium chloride and Conray. This fixation implies that the T3 receptor protein is closely associated with chromatin. These results suggest that proteins involved in the regulation of gene function may be nonrandomly distributed within chromatin subfractions, and are consistent with a direct role for thyroid hormone in regulating genetic expression.


Structural changes inducted by follicle-stimulating hormone or dibutyryl cyclic AMP on presumptive Sertoli cells in culture.

Cells isolated from testes of 20-day-old rats, maintained in primary culture in a defined medium, are shown to respond to follicle-stimulating hormone or 3:5-cyclic AMP with characteristic morphological changes. No response is observed in cells treated with luteinizing hormone or 5-?AMP. The cells form a mono-layer, and have been identified as presumptive Sertoli cells structurally by identification of unique tight junctions in electron micrographs of the preparations, along with other ultrastructural properties characteristic of Sertoli cells in situ. These cells do not undergo mitosis. The presumptive Sertoli cells are shown to be morphologically and functionally different from peritubular fibroblasts grown in parallel cultures. Fibroblasts have high rates of mitosis, do not respond to follicle-stimulating hormone, and frequently form multilayers. Other information on the biochemical responses of the cells is cited, which supports the conclusion that the cultured cell preparations consist primarily of Sertoli cells.


Folding of the DNA double helix in chromatin-like structures from simian virus 40.

Relaxed circular, covalently closed simian virus 40 DNA molecules were associated with the four histones that are present in virions. In electron micrographs the resulting complexes appear twisted, with globular structures (nucleosomes) along the DNA. Incubation with an untwisting extract converts the twisted complexes to relaxed structures. Extraction of the DNA from the relaxed complexes yields supercoiled molecules. The number of superhelical turns in these molecules corresponds to the number of nucleosomes per DNA molecule in the complexes.


Interrelationship between poly (ADP-Rib) synthesis, intracellular NAD levels, and muscle or cartilage differentiation from mesodermal cells of embryonic chick limb.

Mesodermal cells of embryonic chick limbs have the capacity to differentiate into either muscle or cartilage. Previous reports from this laboratory show a correlation between pyridine nucleotide levels and this differentiation, and thus suggest that fluctuations in the cellular NAD levels play a role in the control of muscle versus cartilage development. Poly(ADP-Rib) is chromatin-associated and forms from the polymerization of NAD with the excision of nicotinamide. The studies reported here show that: (A) the rate of net synthesis of poly(ADP-Rib) is correlated with the differentiation of chondrogenic cells from stage 24 limb mesodermal cells grown in vitro; (B) inhibition of chondrogenic expression caused by exposure to nicotinamide or BrdUrd is correlated with maintenance of basal levels of poly(ADP-Rib) synthesis, and this inhibition is dependent on the concentration of nicotinamide or BrdUrd exogenously supplied; (C) potentiation of chondrogenic expression caused by exposure of limb mesodermal cells in vitro to 3-acetylpyridine is correlated with stimulation of the rate of poly(ADP-Rib) synthesis if corrected for the specific activity of the ATP pool or compared to untreated cultures on a per cell basis; (D) isolated chromatin from mesodermal cells has the enzymatic machinery for synthesizing poly(ADP-Rib); (E) this machinery is inhibited by nicotinamide, thymidine, and 3-acetylpyridine; and (F) newly synthesized poly(ADP-Rib) is either associated with a discrete fraction of chromatin or is completely extracted from chromatin by the high column salts, which result in an aggregation with eventual elution at the exclusion volume position of the agarose column. Taken together, these observations provide a possible explanation for how fluctuations in cellular NAD levels can communicate with or be "sensed" by genomic related machinery and eventually result in differtial phenotypic expression.


Somatic hybrid of thymus leukemia (.

A somatic hybrid of ASL-1 leukemia cells [H-2a, thymus leukemia (TL)1,2,3, Thy-1b] and LM(TK)-cells [H-2-k, TL(-), Thy-1 (-), thymidine kinase deficient] was formed with the aid of inactivated Sendai virus. The selection of hybrid cell clones was facilitated by the inability of ASL-1 cells to grow in vitro and of LM(TK)-cells to survive in hypoxanthine/amethopterin/thymidine medium. The H-2 antigens of both parental cells were formed in approximately equivalent amounts by the hybrid cells, and they possessed a hybrid karyotype. As determined by five independent experimental procedures (antibody and complement-mediated cytotoxicity tests, the reduction of specific antibody activity of antiserum of known titer, immunofluorescent tests, mixed hemagglutination tests, and their direct isolation), TL antigens but not Thy-1 antigens were formed by the hybrid cells. TL antigens of the hybrids failed to undergo modulation under conditions leading to the modulation of TL antigens of parental ASL-1 cells. Modulation was attempted with TL 1,3, TL 2, or TL 1,2,3 antisera of high titer. thybrid cells were incubated for up to 30 hr in medium with TL antisera. Both direct and indirect immune methods were attempted. These results indicate that cellular mechanisms controlling the expression of TL antigens may be distinguished from the capacity of the cells to undergo modulation.


Genetic control of endogenous C-type virus production in pancreatic acinar cells of C57BL/He and C57BL/6J mice.

Electron microscopy revealed very active production of C-type virus particles in the pancreatic acinar cells of untreated normal adult mice of the C57BL/He strain. In C57BL/6J mice, a similar picture was observed after a single intraperitoneal injection of dexamethasone. No viruses were observed in the pancreas of untreated or dexamethasone-treated BALB/c and C3Hf mice. F1 hybrids of both C57BL strains with C3Hf mice produced viruses in the same manner and quantity as the C57BL parents, whereas hybrids with BALB/c mice were entirely negative. Approximately 50% of mice of the first backcross generation of (BALB/c times C57BL/He)F1 hybrids with C57BL/He mice were active producers of C-type particles, while the other 50% were negative. It is suggested that a regulator gene that controls C-type virus production does not function in the pancreatic cells of either C57BL strain, and that BALB/c mice can provide hybrids with an active regulator.


Simian virus 40 DNA directs synthesis of authentic viral polypeptides in a linked transcription-translation cell-free system.

A linked cell-free system has been developed which is capable of transcribing and translating mamalian viral DNA, and its characteristics and requirements are outlined. In this system, simian virus 40 (SV40) DNA Form I (supercoiled) directed the synthesis of discrete polypeptides up to 85,000 daltons in size. One of these products was indistingusihable from authentic major virus capsid protein VPI, as judged by mobility on sodium dodecyl sulfate/polyacrylamide gels, antibody predipitation, and peptide analyses. The cell-free products larger than VPI comprised a number of polypeptides ranging in molecular weight from 50,000 to 85,000. These polypeptides demonstrated no immunological relationship whatsoever to the structural protein VPI. However, two of these products, along with one of approximately 25,000 dlatons, were precipitated with antiserum to SV40 tumor antigen. Linear SV40 DNA generated by the cleavage of Form I DNA with the  restriction endonuclease EcoR1 was an efficient template in this system and also directed the synthesis of a polypeptide migrating with VPI on polyacrylamide gels. The potential of this system for defining a functional map of a DNA genome is discussed.


Biochemically differentiated mouse glial lines carrying a nervous system specific cell surface antigen (NS-1).

Six biochemically differentiated clonal lines have been established from a transplantable glioma (tg26) of the C57BL/6 inbred mouse strain. Antibodies have been previously raised against G26 tumor cells, which define a cell surface component(s), NS-1 (nervous system antigen-1), found exclusively in the nervous system. NS-1 concentrations approximate the levels of the original G26 tumor when the clonal lines are grown as clonal tumors in vivo, but are reduced when the cells are grown in vitro. NS-1 concentrations are further reduced in vitro upon incubation of the cells with 1 mM dibutyryl 3:5-cyclic AMP. H-2 histocompatibility antigen concentration, in contrast, is unaffected by dibutyryl cAMP. In addition to expressing NS-1, the neuroectodermal origin of these cell lines is further confirmed by their synthesis of the nervous system specific acidic protein S-100 and by the high specific activity of the enzyme 2:3-cyclic nucleotide 3-phosphohydrolase. In addition, they respond to catecholamines by the elevation of intracellular 3:5-cyclic AMP levels. Whereas expression of S-100 protein is high under in vitro conditions but negligible after one passage in vivo, 2:3-cyclic nucleotide 3-phosphohydrolase is not detectable in vitro but becomes detectable again in vivo. The two membrane-bound constituents, NS-1 and 2:3-cyclic nucleotide 3-phosphohydrolase, therefore seem to be subjected to different regulatory mechanisms from that of the soluble, intracellular S-100 protein.


In vitro transformation of lymphoid cells by Abelson murine leukemia virus.

Cell cultures prepared from fetal murine liver were infected by Abelson murine leukemia virus. After about 2 weeks, proliferating cells of lymphoid morphology appeared in some of the cultures. Addition of 2-mercaptoethanol to the initial culture medium greatly enhanced the appearance of the lymphoid cells. Immunoglobulin determinants were evident on the cells in some cultures. Continuous passage of the cells in certain cultures was possible and the passaged cells could form tumors after animal inoculation. Because Abelson murine leukemia virus is able to induce in vitro malignant transformation of lymphoid cells, it probably causes leukemia by directly affecting cellular growth control.