Induction of epithelial neoplasms by local application of N-nitrosobis(2-hydroxypropyl)amine and N-nitrosobis(2-acetoxypropyl)amine.

The local carcinogenic effect of N-nitrosobis(2-hydroxypropyl)amine (BHP) and N-nitrosobis(2-acetoxypropyl)amine (BAP) in Syrian golden hamsters was elucidated by weekly application to the cheek pouch, lip and vaginal epithelium. The tumor type and incidence in BHP- and BAP-treated hamsters, respectively, was as follows: trichoepitheliomas of the lip, 80 and 90%; cheek pouch papillomas, 10 and 0%, and vaginal papillomas, 80 and 70%. Other lesions were recorded in the perineum, rectum and external urethral ostium and could have been due to a local effect of these nitrosamines. In addition, internal organ tumors were observed with each compound and were possibly caused by absorption of the carcinogens.


Carcinogenic activity of 2-(2-furyl)-3-(5-nitro-2-furyl)-acrylamide, a food additive, in mice and rats.

The carcinogenicity of a food additive, 2-(2-furyl)-3-(5-nitro-2-furyl)acrylamide (AF-2), was studied in Wistar rats and CDF1 mice. The rats developed mammary tumors; the first appearing 9 months after the start of the experiment. In mice fed AF-2 diet, squamous cell carcinoma of the forestomach was observed in the 11th month. Some mice had metastases in the liver, spleen, lymph nodes and peritoneal cavity.


Qualitative and quantitative separation of a series of phorbol-ester tumor promoters by high-pressure liquid chromatography.

A high-pressure liquid chromatographic (HPLC) method using a micro-particulate silica column and gradient elution was developed that separated 12-O-tetradecanoylphorbol-13-acetate (TPA) from 20-oxo-TPA; 12-O-tetradecanoylphorbol (TP); 13-O-acetylphorbol (PA), and from the diterpene alcohol, phorbol (P). A series of other phorbol-ester tumor promoters were also separated via HPLC. Spectrophotometric determination at 232 nm allowed detection sensitivities of 0.05 microgram of TPA. When tritiated TPA was applied to mouse skin, the majority of the tritiated product recovered was TPA, indicating only minimal metabolism of TPA and no need for metabolic activation for tumor promotion.


Membrane glycoprotein changes in primary mammary tumors associated with autonomous growth.

Primary and transplanted mammary tumors of the GR mouse were explanted in tissue culture and grown in the presence of radioactive fucose. Labelled membrane glycopeptides were isolated and compared by cochromatography with differentially labelled glycopeptides from normal mammary gland tissue. Differences with controls in the glycopeptide elution profiles were observed in autonomous, hormone-independent tumors but were absent in histologically similar tumors which required a continuous hormonal stimulus for growth. The results suggest that alterations in membrane glycopeptides are associated with the capacity of autonomous, hormone-independent growth of murine adenocarcinoma cells.


Decrease in the incidence of malignant ileo-caecal immunocytoma in LOU/C rats after surgical removal of the ileo-caecal lymph nodes.

Malignant ileo-caecal immunocytomata, which secrete immunoglobulin, frequently arise spontaneously in LOU/C/Wsl rats. The tumors are confined to the ileo-caecal region and could originate either in the wall of the caecum or in the ileo-caecal lymph nodes. After excision of the ileo-caecal lymph nodes, the incidence of the type of tumor was found to be reduced significantly. The results strongly suggest that the first cells to undergo malignant transformation are located in the ileo-caecal nodes.


A novel class of antitumour agents. III. Morphological findings in treated cells.

The findings of intracellular light-reflecting droplets and clotted mitoses after incubation of Walker and HeLa cells, human lymphocytes and rat fibroblasts with the synthetic amino steroid, 2beta, 16beta-dipiperidino-5alpha-androstane-3-alpha,17beta-diol dipivalate (DAP) were examined in detail. An interaction of DAP with the cellular membranes in all stages of cell life, with the exception of the Go-phase, is postulated.


Effect of split doses of N-methyl-N-nitrosourea on DNA repair synthesis in cultured mammalian cells.

DNA repair was measured in human fibroblasts, mouse C3H 10 T 1/2 fibroblsts and rat hepatocytes by the non-semi-conservative incorporation of [3H]-TdR during DNA repair synthesis using liquid scintillation techniques. Confluent monolayers of these cells grown on cover slips were exposed to split doses (125 or 250 microgram/ml) of the mutagenic and carcinogenic alkylating agent MNU and DNA repair synthesis compared with that produced by a single dose (500 microgram/ml). No significant difference in DNA repair capacity was detected in the three cell lines treated with a single dose or split doses of MNU.


Relationship of aryl hydrocarbon hydroxylase activity to benzo(a)pyrene-metabolizing activity of cells in culture.

The basal level of aryl hydrocarbon hydroxylase (AHH) activity was high in cultures of low passage hamster embryo (HE) cells but AHH inducibility by benz(a)-anthracene (BA) was low; in R72/3 rat liver cells, basal activity was low and inducibility was high. The metabolism of 3H--benzo(a)pyrene was similar in cultures of BA-induced R72/3 cells and uninduced HE cells. Thus, low AHH inducibility may not always be an indication of the cells' absolute hydrocarbonmetabolizing capacity.


Micrometastases formation: a probabilistic model.

A mathematical model of the process of metastases is formulated in which the hematogenous metastatic process from a solid tumor is considered to consist of a series of stages. A mathematical expression is obtained for the probability that no metastases will have been established by a characteristic time interval after tumor initiation. The murine T241 fibrosarcoma that rapidly and reproduceably produces pulmonary metastases was studied. Estimates of parameters required for the expression of probability of metastases formation were derived experimentally. The probability remains close to one for a characteristic time at which point it drops to zero. This indicates that at least in this experimental system there is a predictable critical time period beyond which micrometastases are virtually certain to have been formed.


Effect of excess thymidine on the growth of human melanoma cells transplanted in thymus deficient nude mice.

The effect of thymidine (TdR) on the growth of a human melanoma transplanted in nude mice has been studied. It was found that the injection of 1 g/kg/h of TdR for at least 72 h is sufficient to suppress the growth of the melanoma cells. This inhibition lasts for the duration of the treatment, and causes no apparent toxicity to the host. Nude mice treated for 6--9 days with TdR survived 158 days after melanoma transplant versus 126 days for the controls.


Role of staging in childhood non-Hodgkin's lymphoma.

Childhood non-Hodgkin's lymphoma is characterized by a high incidence of leukemic transformation. A retrospective analysis of this incidence in 227 children is provided. In an attempt to identify factors associated with this phenomenon, the extent of disease in 30 recently diagnosed patients was determined by means of a modified Ann Arbor staging system. Concurrently, the staging system was utilized for the delivery of a new treatment program: chemotherapy was administered to all patients, and those with stage I and II disease also received radiation therapy to sites of bulk tumor. An overall disease-free survival of 75% was achieved. No patient with stage I disease converted to acute leukemia. The data suggest that the major utility of staging is the delineation of anatomic sites of bulk tumor. The chemotherapeutic program utilized in these patients is outlined.


Validity of the Ann Arbor staging classification for the non-Hodgkin's lymphomas.

The Ann Arbor staging classification has proved to be valuable for Hodgkin's disease. When proposed in 1971 it was thought that it could also be applied to the non-Hodgkin's lymphomas. The diversity of the non-Hodgkin's groups, the continued evolution of histopathologic classifications, and the great frequency of advanced disease in the lymphocytic subgroups make the Ann Arbor classification of only limited value for the non-Hodgkin's lymphomas.


Clinical and surgical (laparotomy) evaluation of patients with non-Hodgkin's lymphomas.

A consecutive unselected series of 423 patients with non-Hodgkin's lymphomas was staged prospectively at the Stanford Medical Center between June 1971 and June 1976. The histopathologic classification of Rappaport was used exclusively. Staging laparotomies were performed in 197 of the patients, but another 226 patients were excluded from the staging procedure for a variety of reasons, including stage IV involvement, poor surgical risk, and diagnostic celiotomy before referral to Stanford. Gastrointestinal, splenic, bone marrow, hepatic, and mesenteric lymph node involvement was very common in these patients, whereas systemic symptoms and mediastinal sites of disease were less frequently noted. After staging laparotomy, 15 patients (8%) were downstaged to a lesser extent of involvement while 62 (31%) were upstaged, primarily from clinical stage III to pathologic stage IV. Correlations were made between clinical and pathologic staging and the sites of involvement were compared between those with nodular and those with diffuse lymphomas. The accuracy of diagnostic radiologic procedures was also assessed. Although it is valuable in sequentially determining the extent of subdiaphragmatic involvement by lymphomas, we believe that staging laparotomy should still be regarded as a research procedure which will be undertaken only as indicated in centers of clinical research.


Sequential staging in non-Hodgkin's lymphoma.

The results of staging in 170 patients with non-Hodgkin's lymphoma have been reviewed; all patients were subjected to a series of sequential procedures, including lymphangiography, bone marrow biopsy, and liver biopsy (performed percutaneously, by peritoneoscopy, or by laparotomy). A high incidence of involvement of bone marrow, liver, and abdominal nodes was found in patients with nodular types of lymphoma and in those with diffuse lymphocytic lymphoma, with less than or equal to 20% remaining in stage I or II at the completion of staging in each of these subgroups. Only in patients with histiocytic lymphoma was there an appreciable percentage (31%) remaining in the stage I-II categories after staging. In this study, 75 patients remained in stage III or less after completing the non-surgical phase of staging, and laparotomy was utilized in only 50 patients (30%). At laparotomy, involvement of the liver or the mesenteric or portal lymph nodes was found in 81% of patients with a positive lymphangiogram but in only 18% of those with a negative lymphangiogram.


Determination of the linkages of disaccharides containing a 2-acetamido-2-deoxy sugar unit by solvent effects in circular dichroism.

The circular dichroism spectra of 2-acetamido-2-deoxy sugars in 1,1,1,3,3,3-hexafluoro-2-propanol (F6Pr-2-ol) solutions show a positive band in the n-pi region (209 nm) in contrast to a negative c.d. band in water solution. This difference is interpreted as an indication of a change in the average orientation of the hydroxyl groups adjacent to the amide group. C.d. spectra of 2-acetamido-2-deoxy sugars having a methyl group at O-1 and O-3 confirm this interpretation and suggest that the c.d. spectrum of a disaccharide in F6Pr-2-ol reflects strongly the disaccharide linkage. Large differences in the c.d. spectra of (1 leads to 4) and (1 leads to 6)-linked disaccharides in this solvent lead to rules for distinguishing the linkages of the disaccharides.


Synthesis of beta-L-fucopyranosyl phosphate and L-fucofuranosyl phosphates by the MacDonald procedure.

Fusion of beta-L-fucopyranose tetraacetate with phosphoric acid for 1 min at 50 degrees gives a 9:1 anomeric mixture of the alpha- and beta-pyranosyl phosphates. Longer fusion times give the alpha-anomer exclusively. The L-fucofuranose tetraacetates were synthesized for the first time by acetolysis of methyl-2,3,5-tri-O-acetyl-beta-L-fucofuranoside. Fusion of the furanose tetraacetates with phosphoric acid gave a mixture of the fucofuranosyl phosphates in which the beta-anomer predominated (beta/alpha= 2.4). Anomeric pairs in the fucofuranose series appear to be distinguishable by the chemical shift of the C-6 methyl protons, as already shown by Sinclair and Sleeter in the pyranose series.


Structure of the D-galactan isolated from garlic (Allium sativum) bulbs.

Hot-water extraction of defatted garlic-bulbs yielded a mixture of polysaccharides containing a D-galactan, a D-galacturonan, an L-arabinan, a D-glucan, and a D-fructan. A trace of L-rhamnose was also detected in the polysaccharide hydrolyzate. The pectic acid was partially removed by precipitation with aqueous calcium chloride; from the remaining polysaccharide mixture, a pure D-galactan containing 97.3% of D-galactose was isolated by fractional precipitation and repeated chromatography through a column of DEAE-cellulose. Methanolysis and hydrolysis of the permethylated D-galactan yielded 2,3,4,6-tetra-, 2,3,6-tri-, and 2,3,di-O-methyl-D-galactose in the molar proportions of 1:2:1. On periodate oxidation, the D-galactan reduced 1.18 molar equivalents of the oxidant per D-galactosyl residue, and liberated one molar equivalent of formic acid per 4.13 D-galactosyl residues. Smith degradation of the D-galactan was also conducted. From these results, a structure has been assigned to the repeating unit of the D-galactan.


Structural studies of water-soluble glycoproteins from Cannabis sativa L.

Two carbohydrate-protein fractions, isolated from Cannabis sativa L. by extraction with water and chromatography of DEAE-cellulose, contained arabinose, galactose, glucose, mannose, galacturonic acid, 2-acetamido-2-deoxyglucose, and 2-acetamido-2-deoxygalactose. The structure of the carbohydrate moieties was investigated by methylation analysis and Smith degradation. A high percentage of end-groups indicates a large degree of branching, glucose and galactose being the main branch-points, linked at C-3 and C6. The hexoses are also present as unbranched residues in the chain, largely as (1 leads to 3)- and (1 leads to 4)-linked units and as end-groups. Arabinofuranosyl units constitute the main part of the non-reducing end-groups, and are also present as part of the chain. The polysaccharide chains are probably linked to protein through the hydroxyl group of hydroxyproline.


Characterization of the chondroitin sulfate produced by B16 mouse melanoma cells.

The mucopolysaccharides produced by B16 mouse melanoma cells have been isolated in milligram quantities from the spent media in which the cells were grown in the presence of 2-amino-2-deoxy-D-glucose-t and [35S]-sulfate. The mucopolysaccharides obtained by precipitation with cetylpyridinium chloride from the Pronase digest of the media were further purified by gel filtration, ion-exchange chromatography, and treatment with nucleases. The major components were identified as chondroitin-4-sulfates by identification of the hexosamine as 2-amino-2-deoxy-D-galactose, and by digestibility with hyaluronidases, chondroitinase AC, and chondro-4-sulfatase. The o.r.d. curve and i.r. spectra of these components also confirmed their similarity to chondroitin-4-sulfate from cartilage. The molecular weight of the polysaccharide chains was estimated to be in the range 90,000-120,000 by sedimentation equilibrium analysis.


Structural identification of isomeric O-trimethylsilyl derivatives of some hexuronic acids.

The structures of the isomeric products obtained on trimethylsilylation of naturally occurring hexuronic acids and their sodium salts and lactones have been established by the application of n.m.r. (for anomeric configuration) and mass spectrometry (for ring size). The equilibria of some of the hexuronic acids in methyl sulphoxide involved a larger proportion of furanoid forms than those in water or pyridine. The proportion of furanoses was also increased by the addition of hexamethyldisilazane. Kinetic evidence indicated that two molecules of each hexuronic acid interacted autocatalytically during mutarotation.


An enzyme-P.M.R.-spectroscopic determination of the enantiomers of galactose.

The action of D-galactose oxidase on D-galactose in the presence of oxygen afforded meso-galacto-hexodialdose quantitatively, which allowed p.m.r.-spectroscopic determination of the D enantiomer in a DL mixture. In the spectrum of the products obtained from the enzymic treatment of a mixture of D- and L-galactose, the magnitude of the aldehydrol signals derived solely from the oxidised D enantiomer, relative to those of the anomeric signals, provided the fractional content of the D enantiomer. This simple, accurate, and convenient procedure was applied to the hydrolysate of a seaweed galactan, which was also analyzed, for comparative purposes, by the fermentation technique employing D-galactose-adapted Saccharomyces cerevisiae.


Molecular and metabolic heterogeneity of liver glycogen.

On refeeding after starvation, the resynthesis of rabbit-liver glycogen proceeds inhomogeneously and over-produces material of low molecular weight. The fate of radioactivity incorporated into glycogen from D-glucose-14C can be explained if glycogen of high molecular weight is synthesised on a protein backbone. Confirmation of this view is given by the effect upon glycogen of reagents that break disulphide bonds; these cause loss of the polysaccharide of high molecular weight. Buoyant densities of glycogens are found to be independent of molecular weight and even of extensive degradation. It is concluded that glycogen synthesis proceeds by two routes; one results in the production of polysaccharide of high molecular weight which has a protein backbone capable of forming disulphide bonds, and another results in the production of polysaccharide of low molecular weight which has either no protein backbone or a protein backbone that is incapable of forming disulphide bridges. Apart from size, the two species are physicochemically indistinguishable.


Light-induced alterations in cell shape and pigment displacement in chromatophores of the sea urchin Centrostephanus longispinus.

Alteration in cell shape of the ligh-sensitive chromatophores of Centrostephanus longispinus are described. Upon illumination a centrifugal pigment movement starts within extremely thin filopodia which radiate from the cell body. With continued pigment migration the cellular processes increase in length and diameter and give the cell an irregular stellate appearance. Pigment movement within the cellular processes is discontinuous in space and time and may occur independently in single filopodia. The motion of single granules shows characteristic features of a saltatory movement.


Effect of glucagon on the secretory process in the rat exocrine pancreas.

Glucagon was infused into conscious rats in doses of 10 to 80 microgram/h for periods up to 24 h. The effect on the secretory process of the exocrine pancreas was studied in vitro using isolated pancreatic lobules. A pronounced inhibition of the rate of protein synthesis and discharge of stored and newly synthesized proteins combined with increased enzyme content in the pancreas were observed after 30 min infusion. This effect was absent after longer infusion periods of up to six hours. After 12 to 24 h infusions a marked degranulation and decrease in enzyme content was observed. While the rate of protein synthesis was not significantly enhanced, both the basal and stimulated discharge of enzymes from the pancreas were increased. The results suggest a biphasic response of the pancreas to prolonged glucagon infusion.


The pineal gland of the mole (talpa europaea L.). IV. Effect of pronase on material present in cisternae of the granular endoplasmic reticulum of pinealocytes.

The effect of a proteolytic enzyme, pronase, on material present in cisternae of the granular endoplasmic reticulum of mole pinealocytes demonstrates their proteinaceous nature.


The fine structure of myelinated nerve cell bodies in the bulbus olfactorius of man.

In the bulbus olfactorius of man numerous myelinated nerve cell bodies occur in the stratum plexiforme internum and stratum granulosum internum. In many respects they resemble the neighbouring granule cells: small chromatin clumps border on more than half of the circumference of the nucleus, the thin cytoplasmic rim contains abundant polysomes and sometimes pigment complexes with numerous light vacuoles, the cells often show a process which extends up to the stratum glomerulosum, the perikarya are devoid of synaptic contacts whereas the proximal segment of the peripheral processes display rare contacts. The myelin sheath varies in thickness consisting of 2 to 24 lamellae with distances between the major dense lines ranging from 9.3 to 11.3 nm. The myelin sheath may enclose the cell body completely or partially and accompany the proximal segment of the process arising from the perikaryon. On partially enveloped perikarya, the myelin lamellae end in formations like those of the node of Ranvier, though often less regularly. Within the compact myelin sheath all of its lamellae may be distended for a short distance by glial cytoplasm as in the Schmidt-Lanterman incisures of peripheral nerve fibres. Adjacent to the outermost myelin lamella myelinated axons and cell bodies, tentatively identified as oligodendrocytes, as well as granule cells may be closely joined.


The coxal glands of geophilomorpha (chilopoda): organs of osmoregulation.

The organs terminating at the coxal pores of the tug-legs of Geophilomorpha are not repugnatorial glands, but possess typical transport epithelia with deep apical and basal infolding of the cell membranes, between which numerous large mitochondria are located. Many transport vesicles are found in the basal region but fewer in the apical cytoplasm. The apex is characterized by bundles of longitudinally oriented microtubules, sparse endoplasmic reticulum and free ribosomes. Single neurosecretory axons with synaptoid areas are scattered among the cells. It is suggested that the coxal organs have a diuretic function in moist habitats and an antidiuretic effect in arid environments. The "switch-over" is evidently controlled by a neuroendocrine mechanism.


An ultra-structural study of the gills of Echinus esculentus.

The ultrastructure of the gills of Echinus esculentus is described using transmission electron microscopy. The gills are covered by typical epithelial cells overlying a collagenous basement membrane. The coelomic lumen of the gills is thrown into a series of irregular grooves and ridges which are formed by long narrow cells from each of which projects a single cilium. There is a layer of muscle cells lying underneath these cells adjacent to the basement membrane. They are innervated by axons containing large granular vesicles and the significance of this innervation is discussed in terms of neuromuscular junctions in general within the echinoderms. This study shows that the main function of the gills is excretory and describes three apparent systems whereby excretory products and necrotic coelomocytes are removed.


Ultrastructural evidence for the existence of steroid synthesizing cells in the ovary of the starfish. Asterias rubens (echinodermata).

The ultrastructure of cells showing characteristic features of steroid producing cells in the ovaries of the starfish. Asterias rubens, is described. The correlation between the appearance of these cells and steroid biosynthesis in ovarian tissue of A. rubens is discussed. The importance of the discovery of these cells in view of the biosynthesis of steroids and the phylogeny of Echinodermata is mentioned.


Interommatidial hair receptor axons extending into the ventral nerve cord in the cricket Gryllus campestris.

In the cricket, Gryllus campestris, a branch of the nervus tegumentarius runs to the distal part of the optic lobe. This branch contains the axons of interommatidial hair receptors. The axon terminations extend forward into the trito- and deutocerebrum, and into the subesophageal- and prothoracic ganglia as shown with the cobalt sulfide staining technique. The possible relevance of these connections is discussed.


An amazing sequence arrangement at the 5' ends of adenovirus 2 messenger RNA.

The 5' terminal sequences of several adenovirus 2 (Ad2) mRNAs, isolated late in infection, are complementary to sequences within the Ad2 genome which are remote from the DNA from which the main coding sequence of each mRNA is transcribed. This has been observed by forming RNA displacement loops (R loops) between Ad2 DNA and unfractionated polysomal RNA from infected cells. The 5' terminal sequences of mRNAs in R loops, variously located between positions 36 and 92, form complex secondary hybrids with single-stranded DNA from restriction endonuclease fragments containing sequences to the left of position 36 on the Ad2 genome. The structures visualized in the electron microscope show that short sequences coded at map positions 16.6, 19.6 and 26.6 on the R strand are joined to form a leader sequence of 150-200 nucleotides at the 5' end of many late mRNAs. A late mRNA which maps to the left of position 16.6 shows a different pattern of second site hybridization. It contains sequences from 4.9-6.0 linked directly to those from 9.6-10.9. These findings imply a new mechanism for the biosynthesis of Ad2 mRNA in mammalian cells.


Synthesis of histone messenger RNA of HeLa cells during the cell cycle.

Hybridization of HeLa cell RNA to the DNA of a recombinant phage containing sea urchin histone genes shows that histone mRNA of HeLa cells is synthesized throughout the entire cell cycle. Furthermore, histone messenger is synthesized in cells treated with cytosine arabinoside, an inhibitor of DNA duplication. Although histone RNA is found in the nucleus, the amount of cytoplasmic histone messenger is drastically reduced, in agreement with previous work. The absence of coupling between histone mRNA synthesis and DNA duplication shows that the regulatory process which determines the presence or absence of histone mRNA in the cytoplasm of HeLa cells is not at the transcriptional level.


Discrete sizes of replication intermediates in Drosophila cells.

The size of DNA replication intermediates from Drosophila cells pulse-labeled with 3H-thymidine for 30-120 sec was determined by electrophoresis in formamide-polyacrylamide gels. Replication intermediates were formed in three discrete size classes, with median lengths of 61, 125 and 240 nucleotides. Replication intermediates in the 125 nucleotide size class occurred most frequently. Two of the three size classes may contain discrete species of replication intermediates about 90-400 nucleotides long. The data also suggested that some larger replication intermediates accumulate in pulse-labeled cells. We concluded that 61 nucleotide molecules give rise to 125 and 240 nucleotide molecules, which then form high molecular weight DNA. Mechanisms for forming these replication intermediates are discussed.


Linkage of markers controlling consecutive biochemical steps in CHO cells as demonstrated by chromosome transfer.

Using the technology of metaphase chromosome transfer, evidence has been obtained in CHO cells that genes controlling enzymes in a common pathway in folate metabolism are closely linked. MtxRI and MtxRIII are co-dominant mutations which affect the structure and level of dihydrofolate reductase. Gat- is a glycine-, adenosine- and thymidine-requiring auxotrophic mutant with a lesion in folylpolyglutamate synthetase, an enzyme responsible for addition of glutamates to folate residues. GlyB- is an auxotrophic glycine-requiring mutant whose phenotype may be reversed by folinic acid. Using purified metaphase chromosomes, the MtxR genes were co-transferred into recipient cells with the auxotrophic markers, as demonstrated by the isolation of transferents when two of the phenotypes, either Mtx and Gat, or Mtx and GlyB, were selected at the same time. When recipient cells were selected for Gat+ or GlyB+ alone, the transferents carried the MtxR markers. The GlyA mutation, another glycine-requiring auxotrophic change, is not co-transferred with methotrexate resistance. Because of previous evidence that only a small fragment is involved in  chromosomal transfer experiments, these results seem to provide the first indication that some genes which control enzymes on a common metabolic pathway in eucaryotes are closely linked or are at least syntenic.


Viral DNA sequences from incomplete particles of human adenovirus type 7.

Large pools of empty viral capsids accumulate in cells infected by subgroup B human adenoviruses. Such infected cells also yield DNA-containing incomplete particles in larger quantities than cells infected with serotypes representing other adenovirus subgroups. DNA isolated from carefully purified classes of Ad7 incomplete particles was analyzed by restriction endonuclease cleavage, gel electrophoresis and electron microscopy. At least 90% of the DNA molecules in each sample consisted of sequences that extended from the left end of the viral genome map by variable lengths toward the right end. The average length of DNA is linearly related to the average buoyant density of the  incomplete particles from which the DNA is isolated. The results indicate that each capsid contains one DNA molecule. There is also a specific association of the left end of the viral genome with assembled or assembling capsids. The characteristic distributions of Ad7 incomplete particles may result from intracellular pools of assembly intermediates in which the incompletely packaged DNA has been fragmented in vivo or by shear during preparative procedures.


Decrease in adhesion of cells cultured in polyunsaturated fatty acids.

The addition of long chain unsaturated fatty acids (linoleic, linolenic and arachidonic acids) to BHK cells reduces the cell to substrate adhesion, causes morphological changes and alters the cellular growth properties. The new characteristics are similar to those of transformed cells. The data indicate that the effects are probably due to actual changes in the surface membrane lipids and not due to prostaglandin synthesis.


Continuous protein synthesis is required to maintain the probability of entry into S phase.

"Normal fibroblast" lines such as 3T3 cells arrest in the G0/G1 compartment of the cell cycle when starved of serum. Following readdition of serum and after a lag of 14 hr, the cells enter S phase with first-order kinetics. Cell cycle progress after stimulation is thus consistent with the existence of a single, rate-limiting random event (or transition) in G1 as proposed by Smith and Martin (1973). The addition of low concentrations of cycloheximide (33-100 ng/ml) at any time after the end of the lag phase brings about a rapid reduction (within 1-2 hr) of the rate constant for entry into S phase by an amount that is proportional to the inhibition of leucine incorporation. This suggests that the transition probability depends upon the continuous synthesis of a protein with a short half-life, or on some other unstable substance whose concentration is geared to the rate of translation. More importantly at present, the results indicate that the rate-limiting transition occurs within 2 hr of the start of DNA synthesis. When the same low concentrations of cycloheximide are added at the time of serum stimulation, they also lead to a marked elongation of the lag phase which again is related to the inhibitor concentration. This result is surprising since the lag is independent of serum concentration which itself influences the rate of protein synthesis.


Further mapping of late adenovirus genes by cell-free translation of RNA selected by hybridization to specific DNA fragments.

RNA isolated from the cytoplasm of human cells at late times after infection by adenovirus type 2 (Ad2) has been fractionated by hybridization to fragments of Ad2 DNA which were produced by digestion with the restriction endonucleases Hpa I, Eco RI, Bam HI and Hind III. Cell-free translation of these partially purified mRNAs indicates that the genes for the late Ad2 proteins lie within the following intervals on the conventional Ad2 map: 15K (4.4-17.0 map units), IX and IVa2 (7.5-17.0), IIIa (29.1-40.9), III and V (29.1-57.0), pVIII (40.9-57.0), pVI and II (40.9-70.7), 100K (59.0-83.4), pVIII (70.7-83.4) and IV (85.0-100). In addition to the primary hybridization of the late Ad2 mRNAs to the regions indicated above, most late Ad2 mRNAs (except those for 15K, IX and IVa2) exhibited some hybridization to a secondary site between 17.0 and 29.1 map units.


Two adenovirus mRNAs have a common 5' terminal leader sequence encoded at least 10 kb upstream from their main coding regions.

The messenger RNAs encoding two late adenovirus serotype 2 (Ad2) proteins, fiber and 100K, were purified by hybridization to restriction endonuclease fragments of Ad2 DNA followed by electrophoresis on polyacrylamide gels containing 98% formamide. The 5' terminal oligonucleotides generated by RNAase T1 digestion of the messengers were selected by dihydroxyboryl-cellulose chromatography. Both mRNAs gave an identical 5'-undecanucleotide with the general structure 7mG5'ppp5'AmC(m)U(C4,U3)G. This undecanucleotide could be removed by mild RNAase treatment from the mRNA after hybridization to DNA fragments containing the main coding sequence of the messenger. In contrast, a small region defined by Bal I-E (14.7-21) protects this undecanucleotide from RNase. A second region contained within both Hind III-B (17-31.5) and Hpa I-F (25.5-27.9), although unable to protect the undecanucleotide, hybridizes to both fiber and 100K mRNAs and protects a similar sequence of 100-150 nucleotides. These observations suggest that both mRNAs contain a long common sequence, complementary to at least two different sites on the Ad2 genome remote from the start of these two genes. The implications of these findings are discussed, and a general mechanism is presented for the biosynthesis of mRNAs from larger precursor molecules, based on intramolecular ligation.


The capillary and sarcolemmal barriers in the heart. An exploration of labeled water permeability.

Although the exchange of labeled water between blood and tissue in the heart has usually been assumed to be flow-limited, the outflow patterns of labeled water, relative to intravascular references, in a multiple indicator dilution experiment, have appeared to be anomalous in terms of the models used to explain the transport of less permeable substances. Data showing a change in the shape of the labeled water outflow curve after vasodilation and after the infusion of toxic doses of 2,4-dinitrophenol led us to propose a new model for labeled water permeation which includes barriers at both the capillary wall and the sarcolemmal membrane. This model explains adequately the form of the outflow curve, provides parameters related to the permeability at the two barriers, and gives an estimate of the ratio of the intracellular to interstitial space. Dinitrophenol infused intra-arterially in a dose sufficient to cause S-T elevation in the electrocardiogram is found to reduce the sarcolemmal water permeability by an order of magnitude, but to have no effect on capillary water permeability. We conclude that water transport in the heart is barrier-limited at both the capillary and sarcolemmal membranes and that sarcolemmal water permeability is probably mediated at least in part by a structure sensitive to the effects of dinitrophenol, presumably a protein channel. Since the outflow patterns of inert gases resemble that of labeled water, it is possible that oxygen distribution is also barrier-limited.


Detection and localization of multiple epicardial electrical generators by a two-dipole ranging technique.

The ability of a numerical procedure to detect and to localize two experimentally induced, epicardial dipolar generators was tested in 24 isolated, perfused rabbit heart preparations, suspended in an electrolyte-filled spherical tank. Electrocardiograms were recorded from 32 electrodes on the surface of the test chamber before and after placement of each of two epicardial burns. The second lesion was located either 180 degrees, 90 degrees, or 45 degrees from the first. Signals were processed by iterative routines that computed the location of one or two independent dipoles that best reconstruced the observed surface potentials. The computed single dipole acounting for 99.68% of root mean sequare (RMS) surface potential recorded after the first burn was located 0.26 +/- 0.10 cm from the centroid of the lesion. Potentials recorded after the second lesions were fit with two dipoles that accounted for 99.36 +/- 1.51% of RMS surface potentials and that were located 0.42 +/- 0.26 cm and 0.57 +/- 0.49 cm from the centers of the corresponding burn. Seventy-one percent of computed dipoles were located within the visible perimeter of the burn. Thus, two simultaneously active dipolar sources can be detected and accurately localized by rigorous study of the generated electrical field.


Purification of high molecular weight forms of renin from hog kidney.

To elucidate the possible regulatory function of high molecular weight renins in blood pressure control, studies were initiated to purify and characterize the high molecular weight enzyme. To avoid the destructive effect of acidification during affinity chromatographic elution, pepstatin-aminobutyl-agarose gel was used from which renin could be eluted at a neutral pH in 2 M urea. Three additional steps of conventional chromatography after the affinity technique produced two types of high molecular weight renins in electrophoretically homogeneous forms. Big renin with a molecular weight of 61,000 has a specific activity approximately 20% of the fully active small molecular weight renin (mol wt 40,000). Big big renins with molecular weights of 140,000 were discovered and purified in two homogeneous forms with specific activities less than 1% of the fully active renin. The possibility of zymogen-enzyme relationship involving the big big, the big, and the fully small molecular weight renins is discussed.


Purification of human angiotensinogen.

A four-step method for the purification of human angiotensinogen has been devised. The four steps are: (1) removal of albumin by affinity chromatography on blue dextran-Sepharose, (2) chromatography on DEAE-Sephadex, (3) chromatography on hydroxylapatite, and (4) chromatography on DEAE-cellulose. This method is capable of producing 8 mg of purified angiotensinogen from 150 ml of plasma with 33% overall recovery of renin-releasable angiotensin I. The angiotensinogen appears homogeneous by immunochemical and ultracentrifugal techniques. The N-terminal amino acids have been determined to be alanine and aspartic acid or asparagine.


A direct radioimmunoassay for human renin substrate and identification of multiple substrate types in plasma.

Plasma renin substrate, a widely measured parameter of the renin reaction, is quantitated indirectly by the measurement of liberated angiotensin I upon exhaustive incubation of plasma with added renin. To overcome methodological problems of this assay system, we have developed a direct radioimmunoassay for this plasma protein using renin substrate purified from pooled plasma of normotensive subjects as the antigen. Comparison of substrate quantitated by the two assay systems (direct and indirect) indicates a 1:1 correlation with the exception of certain subjects with elevated substrate levels induced by estrogen therapy. To study the possibility of multiple substrate forms, we have made a comparison of substrate quantitated by both radioimmunoassays in conjunction with electrophoresis of plasma on polyacrylamide gel. One major form of substrate with a retardation factor (Rf) = 0.60 was found in normotensive and essential hypertensive subjects which gave a 1:1 correspondence on quantitation by the two methods. In contrast, six of 16 women on oral contraceptives demonstrated three forms of substrate (Rf = 0.16, 0.35, and 0.60) on electrophoresis. Substrate with Rf = 0.16 and 0.35 did not cross-react with the antiserum prepared against substrate from normotensive subjects, implying structural differences in these proteins.


Partial purification of human renin.

An affinity gel for the purification of renin was prepared by coupling pepstatin to aminohexylagarose gel. Partially purified human renin (Haas et al., Arch. Biochem. Biophys., 110, 534-543, 1965) was purified further by affinity chromatography on the pepstatin-aminohexyl-agarose gel, gel filtration on a Sephadex G-75 column, and ion exchange chromatography on a DEAE-cellulose column. Separation from a protease permitted further purification without progressive loss of activity. Three peaks of enzymatically active renin were obtained after the DEAE-cellulose chromatography, with specific activities ranging from 206 to 166 and 85 Goldblatt units/mg protein, respectively. The specific activity of 206 GU units/mg represents a 103,000-fold purification compared to the renin present in the first crude extract.


Hormonal control of plasma renin substrate; (angiotensinogen).

The effect of ethinyl estradiol (EE2) and dexamethasone (Dex) upon plasma renin substrate (PRS) was studied in rats in relation to sexual maturation and pituitary function. Normal adult rats had large increases in PRS when given either EE2 or Dex. Prepubescent (25-day-old) rats also had a substantial increase in PRS following Dex, but no increase in PRS after EE2. In adult rats, hypophysectomy prevented the rise in PRS due to EE2 but did not prevent the increase in PRS due to Dex. Hepatic estrogen and glucocorticoid receptors were measured in immature and adult rats. Immature rats had markedly reduced concentration of hepatic estrogen receptors (16%), compared with adults. In contrast, glucocorticoid receptor concentration was not significantly different between immature and adult rats. The results indicate that the effect of estrogens and glucocorticoids upon PRS are mediated by pathways that are separable at the hormone receptor level.


Inhibition of the in vitro renin reaction by circulating neutral lipids.

We have previously reported that the addition of plasma from normotensive human subjects to renin-renin substrate inhibits the in vitro rate of angiotensin generation. The purpose of the present study was to determine whether a renin-inhibiting factor could be identified in human plasma. Neutral lipids and phospholipids were extracted from plasma of seven normotensive control subjects. During 30-, 60-, and 180-minute incubations, neutral lipids inhibited the reaction between human renin and homologous renin substrate (P less than 0.01). Phospholipids did not affect the rate of angiotensin generation (P less than 0.05). The neutral lipid extract was further separated on silica gel chromatographic plates, and several fractions inhibited renin (P less than 0.01) Addition of each of six synthetic neutral lipids to renin-renin substrate did not affect the rate of angiotensin generation. In conclusion, several neutral lipid fractions extracted from human plasma inhibit the in vitro renin reaction.


No evidence for product inhibition of the renin-angiotensinogen reaction in the rat.

The influence of the product of the renin-angiotensinogen reaction, the des-angiotensin I-substrate (des-AI-substrate) on the renin-angiotensinogen reaction has been studied. Des-AI-substrate was prepared from purified rat angiotensinogen by reaction with immobilized renin. The des-AI-substrate had no inhibitory effect on the reaction of partially purified rat renin with rat angiotensinogen at concentrations corresponding to 0.225 or 0.45 micrometer angiotensinogen.


New vistas for the study of structural and functional dynamics of the heart, lungs, and circulation by noninvasive numerical tomographic vivisection.

Major segments of the biologic sciences and the practice of medicine are based on study and knowledge of the relationships of anatomic structure to biologic function. Traditionally, this knowledge has been gained by indirect means, inference, or by direct surgical vivisection or postmortem examination. The revolutionary capability of nondestructive, operator interactive, mathematical vivisection provided by synchronous cylindrical scanning tomography to obtain similar information non-invasively and painlessly will provide these data to the internist for individual patients. Furthermore, this information will be in a computerized format which can be subjected to myriad types of objective measurements and display. These developments promise beneficial effects of clinical diagnosis and health care which may approach those associated with the discoveries of the biomedical investigative and clinical diagnostic value of X-rays and cardiac catheterization.


Comparative hemodynamic effects of inotropic and vasodilator drugs in severe heart failure.

In 12 patients with severe congestive heart failure (CHF) due to ischemic heart disease or nonischemic cardiomyopathy the hemodynamic response to intravenous infusion of sodium nitroprusside (N) was compared to that of dobutamine (D) 10 microgram/kg/min. D and N produced comparable increases in cardiac output (CO) (2.8 to 5.8 L/min and 2.9 to 5.0 L/min, respectively), but, compared to N, D caused a higher arterial pressure (99.3 vs 86.2 mm Hg, P less than 0.01) and heart rate (102.5 vs 95.3, P less than 0.05) and less reduction in pulmonary wedge pressure (PWP) (28.9 to 20.2 mm Hg vs 29.1 to 16.6 mm Hg, P less than 0.05). In five additional patients N and D were studied separately and then were infused together. The combination resulted in a higher CO, lower PWP and greater reduction in systemic and pulmonary vascular resistances than either drug alone. Brachial arterial infusion of nitroprusside produced prominent forearm vasodilation in a dose less than 10% of the systemic dose, whereas vasodilation with dobutamine was only modest even when 50% of the systemic dose was infused. Therefore, potent inotropic and vasodilator drugs produce similar and additive augmentation to left ventricular performance in heart failure. Reduction in vascular resistance with dobutamine probably is largely of reflex origin, but the vasodilation itself may be an important determinant of the rise in cardiac output.


The influence of heart rate on pulmonary arterial-left ventricular pressure relationships at end-diastole.

Increased resistance to blood flow stemming from structural and functional abnormalities of the lungs may cause pressure in the pulmonary artery to exceed that in the left ventricle at the end of ventricular diastole. This study explores the possible contribution of heart rate to the diastolic pressure gradient observed in the presence of acutely induced hypoxia. Pulmonary hemodynamics were examined in mongrel dogs with chronic atrioventricular dissociation with and without hypoxia at two different heart rates and during sequential increments in heart rate while the animals breathed room air. Studies during sequential pacing indicate that heart rate was of greater importance than blood flow in determining the magnitude of the gradient. Heart rate has to be considered when the causes of pulmonary hypertension and the effects of drugs or other agents on the pulmonary circulation are being investigated.


Postextrasystolic compliance of the left ventricle.

The effects of a single premature contraction (PC) on left ventricular diastolic distensibility was studied in man, in the conscious chronically instrumented dog, and in the isolated (isovolumic) blood perfused dog heart. In the isolated dog heart at a constant volume, there was no difference in end-diastolic pressure when control diastolic pressure was compared to that following a PC. In man and conscious dogs, there was no difference between the overall pressure-volume or pressure-length plots when control diastolic data were compared to data from the diastole immediately following a PC (no change in the modulus of chamber stiffness). In the intact circulation of man dog, increased filling during the post PC pause causes the ventricle to operate higher on the steep (stiffer) portion of its pressure-volume or pressure-length curve (decrease in preload-dependent compliance). Thus, although a PC does not alter the modulus of chamber stiffness, a preload-dependent change in compliance may occur during post PC diastole.


Individualized values for the disappearance rate parameter (Kd) in the enzymatic estimation of infarct size. A critique.

A mathematical model to measure infarct size from serial estimates of CPK has been developed by Shell et al. Norris et al. have recently introduced a modification of this method by calculating an individualized disappearance rate parameter (Kd) from the uniformly monoexponential portion of the CPK vs time curve. This modification is based on the assumption that when CPK is decaying monoexponentially, the amount of CPK appearing in the circulation is zero. This paper presents a mathematical analysis which shows that the method of Norris et al. is theoretically unsound, as the CPK appearance function may be significantly different from zero and yet CPK vs time may still be monoexponential. Their method may lead to erroneous results in the calculation of infarct size.


Estimation of the probability of exercise-induced ischemia by quantitative ECG analysis.

In order to improve the value of exercise tests for the detection of coronary artery disease (CAD) a system for on-line computer processing of the Frank lead exercise ECG was developed. Data were analyzed from 95 patients with CAD and 129 ostensibly healthy men. All subjects had a normal ECG at rest. Visual ECG interpretation during exercise yielded a sensitivity of 50% and a specificity of 95%. A large number of QRS and ST measurements were compared by discriminant function analysis in a group of 86 normal subjects and 52 patients (designated training group). Best results were obtained with a combination of two ST amplitudes from lead X: sensitivity, 85%, specificity, 90%. This was confirmed in a test group of 43 patients and 43 normal subjects. The results of the discriminant function were expressed as the likelihood ratio for an abnormal or normal ST segment at a given heart rate, a figure which provides a quantitative assessment of the degree of exercise-induces ischemia. This is a more realistic approach than classification into normal or abnormal since persons with and without CAD fall along the same continuous spectrum.


Sudden death after repair of tetralogy of Fallot. Electrocardiographic and electrophysiologic abnormalities.

In order to try to determine the mechanism of sudded death in patients after surgical repair of tetralogy of Fallot, electrocardiographic, intracardiac electrophysiologic, and clinical data of 51 children who had postoperative intracardiac electrophysiologic studies were reviewed. Ninety-four percent had developed right bundle branch block (RBBB) and 16 percent had additional left anterior hemiblock (LAH). Two had had transient complete A-V block (CAVB) and one had permanent CAVB. Six had a first degree A-V block and nine had premature ventricular contractions (PVC). Nine patients were found to have prolonged intra-atrial conduction times, four prolonged A-V nodal conduction, four prolonged His-Purkinje conduction, and five prolonged corrected sinus node recovery times. Patients with first degree A-V block or LAH did not have an increased incidence of abnormalities on electrophysiologic study. No patient with RBBB and LAH developed complete A-V block or died. Three of the nine patients with PVCs died, one of intractable ventricular fibrillation and two suddenly, presumably of dysrhythmia. All three had significant congestive heart failure. Although late complete A-V block occurs and should be watched for, ventricular dysrhythmias in patients with PVCs may be the cause of most sudden deaths after tetralogy repair. We currently are treating all of our postoperative tetralogy patients who have PVCs with quinidine or propranolol.


Doppler echocardiography. Use of a graphical display system.

Pulsed Doppler echocardiography (PDE) is a technique for evaluating blood flow characteristics at specific locations within the heart and great vessels. Because this method assesses blood flow rather than cardiac structures, PDE complements the findings of M-mode echocardiography. A new on-line graphical method for displaying pulsed Doppler information provides 1) a printed, permanent record of flow information, 2) precise timing of blood flow events, and 3) information on the direction of flow in the heart and great vessels.


A new, noninvasive technique for inducing post-extrasystolic potentiation during echocardiography.

Left ventricular function was evaluated in 34 patients with the echocardiogram, and an external mechanical cardiac stimulator was used to induce a ventricular premature contraction (VPC) noninvasively. Extent of post-extrasystolic potentiation (PESP) was determined by comparing systolic dimensional shortening and ejection fraction of the sinus beat preceding the VPC to that of the potentiated beat which followed it. Using this technique, a VPC could be introduced into the cardiac cycle of 30 of the 34 patients, six of whom were free of obvious cardiac disease and 24 of whom had valvular, coronary or myopathic heart disease. The only complication observed was mild breast ecchymosis in a female patient. Systolic dimensional shortening and ejection fraction increased from control values by an average of 21% and 17% respectively, with a range of 0-100%. The degree of PESP was very reproducible in repeat studies and when the same patients were subsequently evaluated during a spontaneously occurring or catheter-induced VPC. The technique can safely and reliably induce post-extrasystolic potentiation during echocardiography and is a potentially important adjunct to the noninvasive evaluation of left ventricular function.


Prediction of the normal blood volume. Relation of blood volume to body habitus.

Predictions of blood volume (BV) assume the existence of a constant ratio between BV and body weight or surface area (SA). We examined the validity of this assumption by calculating BV from plasma volume and body hematocrit in 160 normal volunteers whose weights ranged from -38.7 to 210.8% of desirable weight (assessed by a modification of the Metropolitan Life Insurance Company Desirable Weight tables). BV is not a constant fraction of body weight or SA in this population. Its prediction from such constant ratios results in a large error of estimate which is systematically biased with respect to height and weight. BV prediction from the observed regressions of the parameter on weight and SA reduces the error substantially but remains biased with respect to height. BV prediction from the subject's degree of deviation from desirable weight affords a smaller error of estimate which is apparently free from systematic bias.


The Starr-Edwards model 6000 valve. A fifteen-year follow-up of the first successful mitral prosthesis.

The Starr-Edwards model 6000 mitral valve was the first successful mitral prosthesis. A fifteen year follow-up of 110 patients undergoing isolated mitral valve replacement with this prosthesis from 1960 to 1966 is presented. There were 22 (20%) operative and 37 (42%) late deaths at a mean of 8.6 years postoperatively. Preoperative variables leading to late death and thromboembolism are analyzed. Seventeen of 25 operative survivors had improvement in their NYHA Functional Class status and eight other remained unchanged an average of 13.1 years after surgery. There are 5000 patients world-wide currently relying on this prosthesis. Despite a high rate of thromboembolism in the past, elective replacement is not recommended except in selected patients.


Inhalation imaging with oxygen-15 labeled carbon dioxide for detection and quantitation of left-to-right shunts.

Quantitation of left-to-right shunts was determined noninvasively from the pulmonary clearance pattern of inhaled 15oxygen-labeled carbon dioxide (C15O2). After a single breath inhalation of C15O2, counts over the lungs were obtained from sequential 0.5 sec positron camera images. In 21 patients without left-to-right shunts, counts declined exponentially due to the washout of C15O2 by the pulmonary blood flow. In 22 patients with left-to-right shunts, this monoexponential pulmonary clearance pattern was interrupted by an abnormal upward deviation, indicating tracer recirculation through the shunt to the lungs. Following surgical shung closure in 10 patients, pulmonary C15O2 clearance patterns became normal in nine and showed a small residual left-to-right shunt in one. Shunt size was derived from the ratio of the height of the recirculation curve to the height of the initial inhalation peak. These values significantly correlated with shunt size as determined by oximetry (r = 0.83).


Lung hypoplasia in congenital pulmonary valve stenosis.

The pulmonary function of ten adult patients with congenital pulmonary valvular stenosis was investigated. The patients clearly showed smaller lungs than healthy control subjects of equivalent age and height; lung elastic recoil pressure was normal at any given percentage of measured total lung capacity, indicating that postnatal parenchymal damage is not the cause of the small lungs. The lung diffusing capacity for carbon monoxide was reduced, reflecting the anatomical alterations of the pulmonary vascular bed. Finally, the maximal flow-static recoil curves showed a fixed (not dynamic) reduction of airway dimensions: the critical transmural pressure in the collapsible flow-limiting segment (Ptm') was normal, but the conductance of the S segment was lowered. These abnormalities most likely reflect inadequate development of the lung and suggest that pulmonary blood pressure may be an important determinant of lung growth in the postnatal period.


A study of instruments in preparation for a blood pressure survey of children.

In preparation for the measurement of blood pressure in children of a total geographic community, several preliminary studies of the validity and reliability of various methods and instruments for indirect blood pressure measurements were performed. These studies included Graeco-Latin Square designs, examination of children in a field setting, and assessments of the replicability of reading automatically recorded blood pressures. Each of the studies was designed to monitor the validity and replicability of instruments, methods, and observers. Controlling for subject, we compute biases due to instrument, method and observer and, where possible, eliminated them in the enusing studies. One automatic instrument, the Physiometrics recorder, was selected and used in conducting epidemiologic studies where it complements the measurements by the mercury sphygmomanometer.


Coronary neovascularity and fistula formation: a sign of mural thrombus.

Fifteen patients have shown neovascularity in the left heart during selective coronary arteriography, with associated fistulous communication to a cardiac chamber in ten. In nine the abnormalities were adjacent to and within ventricular mural thrombi, associated with ventricular aneurysms in seven and congestive cardiomyopathy in two. In six patients with mitral stenosis and atrial fibrillation the angiographic changes related to left atrial thrombi. Histologic study of two atrial thrombi showed revascularization patterns in keeping both with the angiographic findings and with an intermediate stage in the evolution of mural thrombus. Though myxomas can produce a similar appearance, clinical and additional angiographic features should permit differentiation in most instances between this infrequent tumor and the far more common mural thrombus.


De subitaneis mortibus. XXVI. Fatal electrical instabiltiy of the heart associated with benign congenital polycystic tumor of the atrioventricular node.

Benign congenital polycystic tumors of the atrioventricular (A-V) node are an unusual but not very rare cause of heart block. Two such cases are presented and discussed in conjunction with the reported experience of other. The tumor is always within and only very near the A-V node, seldom involves more than the proximal end of the His bundle, and has not been reported to occur in the sinus node. Although sudden death has been reported in conjunction with these tumors, a surprising number of other patients have lived to old age and died of causes unrelated to the A-V nodal tumor. As a corollary it is important to consider a diagnosis of A-V nodal tumor in any patient of any age who presents with otherwise unexplained heart block or syncope. From accumulated experience it appears that such patients do not tolerate electronic pacing safely and some possible explanations for this are discussed. Escape rhythm in all reported cases has been characterized by QRS complexes which are narrow and a ventricular rate which is from half to two-thirds of the sinus rate. Reasons why an A-V junctional rhythm which is 66% of sinus rate may be especially stable are discussed.


Inhibition of bipolar demand pacemaker by diaphragmatic myopotentials.

This report describes inhibition of a normally functioning bipolar demand pulse generator by diaphragmatic myopotentials. Transient pacemaker suppression occurred repeatedly with deep respiration, straining, the Valsalva maneuver, coughing, sneezing and laughing. When the magnet was applied, none of these maneuvers inhibited the pacemaker. Extensive investigations ruled out an intermittent electrode problem such as a wire fracture or insulation break. Sensing of diaphragmatic myopotentials should be considered in the differential diagnosis of unexplained pacemaker pauses.


Zinc, iron, copper, and magnesium concentrations in tissues of rats fed various amounts of zinc.

An experiment was conducted with rats to determine the effects on the tissue concentrations of Zn, Fe, Cu, and Mg of feeding various amounts of zinc. The rats were pair-fed one of the following diets for four weeks: Diet A, a zinc-deficient diet; a diet containing the recommended amoun of zinc (diet A plus 55 microgram of zinc per gram of diet), or diet A plus 550 microgram zinc per gram of diet. Concentrations of these elements in various tissues were determined by atomic absorption spectrometry after wet digestion. Feeding the rats zinc-supplemented diets resulted in increased zinc in blood, heart, kidney, and liver, and a marked decrease of iron in kidney and liver. Concentrations of the other two elements were unchanged in all tissues. Thus, the effect of zinc in decreasing iron concentrations in liver, observed by several investigators when rats were fed toxic amounts of zinc, also occurs when zinc is administered in normal or subtoxic amounts.


High-pressure liquid-chromatographic method for determination of gentamicin in plasma.

A rapid, specific method for measuring gentamicin in plasma by high-pressure liquid chromatography was developed. After deproteinization, gentamicin in the supernate was dansylated and extracted into ethyl acetate. The organic extract was chromatographed on a microparticulate reversed-phase column, which was eluted with aqueous acetonitrile. Use of the dansyl derivative enables fluorometry, for more sensitive quantitation. Various factors that could affect the assay sensitivity were investigated. With 0.2-ml plasma samples, the method can accurately measure as little as 1 mg of gentamicin per liter. We encountered no interferences from plasma supplemented with various drugs or plasma of patients who were on therapy with other drugs. This method can also separate gentamicin C1 from C1a and C2, all of which are present in various ratios in commercial dosage forms. This method is also applicable to gentamicin determination in urine.


Application of a method for correcting an observed regression between change and initial value for the bias caused by random errors in the initial value.

When studying the correlation between the change of a variable during treatment and the value before treatment, random errors (errors of measurement as well as intra-individual biological variation) may yield seriously biased results. In the present work a method to adjust for this bias is presented. The method was applied to data from blood coagulation factor and plasma protein analyses in chronic alcoholics before and after one week of abstinence. It was found that many of the significant correlations were lost when the data were adjusted.


Estimation of maprotiline in serum by gas-chromatography, with use of a nitrogen-specific detector.

We describe a gas-chromatographic procedure for estimating therapeutic concentrations of maprotiline, a new tetracyclic antidepressant, in serum by use of a nitrogen-specific detector. Desmethyldoxepin, a secondary amine similar in structure to maprotiline, is added as a mass internal standard to the specimen before extraction, to obviate the need for accurate measurements of volumes during extraction and analysis. Both maprotiline and the internal standard are converted to acetyl derivatives, to avoid the adsorption of secondary amines on the column. A highly selective liquid phase is used, which allows a very good separation of desmethyldoxepin, maprotiline, desmethylmaprotiline, and the interferences from serum and reagents.


Calorimetry of cholinesterase activity in serum.

Cholinesterase activity in human serum was measured calorimetrically and compared to values obtained with the spectrophotometric method of Rappaport et al. [Clin. Chim. Acta 4, 227 (1959)]. The response of the calorimeter upon mixing serum with acetylcholine corresponded to the cholinesterase activity of the serum, the thermogram peak height being linearly related to enzyme activity at serum dilution volumes at which interference heat was negligible. The interference heat contributed by the dilution mixing characteristics of serum, studied separately, was found to be similar for heat-denatured, aged, or lyophilized samples. In all cases, heat of mixing was negligible when 1 ml of serum in buffer (2/3 by vol) was mixed with 1 ml of buffer. When this amount of serum was assayed with acetylcholine, cholinesterase activities of 2 and 3 kU/liter were recorded as thermograms with peak heights of 2.3 and 3.5 cm, respectively.


Performance characteristics of rules for internal quality control: probabilities for false rejection and error detection.

When assessing the performance of an internal quality control system, it is useful to determine the probability for false rejections (pfr) and the probability for error detection (ped). These performance characteristics are estimated here by use of a computer stimulation procedure. The control rules studied include those commonly employed with Shewhart-type control charts, a cumulative sum rule, and rules applicable when a series of control measurements are treated as a single control observation. The error situations studied include an increase in random error, a systematic shift, a systematic drift, and mixtures of these. The probability for error detection is very dependent on the number of control observations and the choice of control rules. No one rule is best for detecting all errors, thus combinations of rules are desirable. Some appropriate combinations are suggested and their performance characteristics are presented.


Radioimmunoassay for digoxin with a fully automated continuous-flow system.

We describe a fully automated continuous-flow radioimmunoassay for digoxin with use of a Technicon AutoAnalyzer system. It makes use of antibodies linked to magnetizable particles; a magnet separates the bound and free fractions. The precise dispensing of reagents by the AutoAnalyzer system permits a small incubation volume (about 160 microliter) and the reproducible timing enables non-equilibrium conditions to be used. Thus a single sample is assayed in 15 min, with a sample throughput of 30/h. The standard curve ranges from 0.5 to 8.0 microgram/liter and is most precise between 1 and 3 microgram/liter, the between-assay coefficient of variation being less than 3%. There is no significant carryover between samples of high and low concentration, and results by the method correlate closely (r = 0.969) with those by an established manual assay in which charcoal separation is used.


Protective effect of glycerol against the increase in alkaline phosphatase activity of lyophilized quality-control serum.

We studied the effectiveness of glycerol or ethylene glycol in preventing the increase in alkaline phosphatase activity of lyophilized or refrigerated quality-control serum after reconstitution or repeated freezing and thawing. Control serum reconstituted completely from the lyophilized state with subsequent storage at -20 degrees C showed a considerable decrease in alkaline phosphatase activity immediately after thawing, and a gradual increase in activity on allowing it to stand at room temperature. Adding glycerol or ethylene glycol to the reconstituted serum obviated these changes in activity, glycerol being more effective than ethylene glycol. Reconstituted serum with added glycerol maintained maximum activity before refrigeration during either storage for 30 days or on repeated freezing and thawing. Practical applications of this glycerol-supplemented control serum are discussed.


Creatine kinase and its isoenzymes in the serum of women during pregnancy and the peripartum period.

In serum obtained from 28 women before, during, and after normal labor and delivery, creatine kinase activity was seen to be distinctly elevated immediately after labor and 24 h later, but had returned to normal six weeks later. In most cases the increase was due to the MM isoenzyme and was attributed to skeletal-muscle damage associated with labor. In 15 cases, the BB isoenzyme was observed, and in three patients the MB isoenzyme. Cord blood, which contains all three isoenzymes, may be the source of the MB and BB isoenzymes. Uterine muscle contains exclusively BB isoenzyme, and therefore uterine muscle damage is a likely source of the serum BB isoenzyme. The relationship among these isoenzymes in serum after delivery must be recognized, to avoid misdiagnosing myocardial infarction at this critical time.


Combined Shewhart-cusum control chart for improved quality control in clinical chemistry.

We describe the adaptation of the decision limit cumulative sum method (cusum) to internal quality control in clinical chemistry. With the decision limit method, the cusum is interpreted against a numerical limit, rather than by use of a V-mask. The method can be readily implemented in computerized quality-control systems or manually on controls charts. We emphasize the manual application here and demonstrate how the technique can be implemented on existing Shewhart or Levey-Jennings control charts. This permits both cusum and Shewhart control rules to be used simultaneously on a single control chart and also minimizes the data calculations necessary for the cusum method. Computer simulation studies are used to determine the performance characteristics of several different cusum rules, alone and in combination with a Shewhart rule. These studies indicate that improvements in existing quality-control systems should be possible by addition of this simple cusum method and by use of a combined Shewhart-cusum control chart. This should be particularly advantageous when  introducing the cusum method in laboratories with manual quality-control systems.


Radioimmunoassay for MM and BB isoenzymes of creatine kinase substantiated by clinical application.

We have developed a radioimmunoassay technique that is highly specific for measuring the MM isoenzyme of creatine kinase. The specificity of the radioimmunoassay for the BB isoenzyme was poor. In patients with treated Duchenne-type muscular dystrophy or untreated hypothyroidism, the MM isoenzyme, but not the BB isoenzyme value, was consistently above above normal. In the radioimmunoassay for the BB isoenzyme the antisera might cross react with other materials and the inactivated isoenzyme, but not with MM or MB isoenzymes.


Effect of incubation with Mg2+ on the measurement of alkaline phosphatase activity.

Human serum incubated with 2-amino-2-methyl-1-propanol (860 mmol/liter) and magnesium ion (270 mumol/liter) at pH 10.35 showed greater alkaline phosphatase (EC 3.1.3.1) activity than if an equal amount of magnesium ion was added at the time of measurement. The apparent increase is due in part to a slight lability of serum alkaline phosphatase in 2-amino-2-methyl-1-propanol, which is prevented by the inclusion of magnesium. For some sera, however, a portion of this increased activity is real rather than artifactual. Use of serum rather than 4-nitrophenylphosphate to initiate the reaction produced relatively low activities and, in some cases, markedly nonlinear (increasing) rate progress curves. The behavior of some commercial lyophilized control sera differed significantly from that of patients' sera, in particular exhibiting a marked lability in the presence of 2-amino-2-methyl-1-propanol. Incubation of these labile materials with Mg2+ slightly improved their stability; addition of Zn2+ plus Mg2+ markedly stimulated and completely protected their alkaline phosphatase activity.


Direct potentiometric measurement of sodium and potassium in whole blood.

I compared results for sodium and potassium in whole blood and plasma as measured with a newly available potentiometric analyzer, the "Orion SS-30". No significant difference was found for either sodium or potassium in 207 such comparisons. With use of the flowing, high mixing-velocity liquid junction of the Orion SS-30, the residual liquid junction potential due to blood cells was found to be less than 0.1 mV and to be independent of the hematocrit. This is in contrast to the hematocrit-dependent residual liquid junction potential of approximately 0.6 mV noted by others at normal hematocrit values with the open capillary liquid junctions now commonly used in pH instruments. I also found that the potassium concentration can increase significantly during the mixing of whole blood, and such samples should be mixed gently, if at all. Evidently sodium and potassium can be accutately and easily measured directly in heparinized blood.


Adaptation to the centrifugal analyzer of an enzymatic method for the measurement of lactate in plasma and cerebrospinal fluid.

A fully enzymatic method for measurement of l(+)-lactate has been adapted for use with a centrifugal analyzer. The procedure is based on a two-point fixed-time kinetic approach for elimination of static interferences, and commercially available reagents are used. Specimen collection and preservation are simplified by the use of standard evacuated tubes containing sodium fluoride. Under these conditions lactate is stable for as long as seven days at 4 degrees C, and deproteinization is unnecessary. The method is extremely simple and ideally suited for emergency and routine use. Determinations may be performed on plasma or cerebrospinal fluid. Sensitivity and precision are excellent, linearity extends to 8.0 mmol/liter, and results compare well with those obtained using the manual enzymatic procedure.


Simplified radioimmunoassay of urinary drugs of abuse adsorbed on ion-exchange papers.

A convenient screening procedure for presence of drugs of abuse in urine consists of two steps: adsorption of the drugs from urine onto a paper loaded with cation-exchange resin and detection of the adsorbed drugs by direct radioimmunoassay. The first step can be performed in the field, the second in a central laboratory. Storage and transport to the laboratory are simplified because specimens adsorbed on dried paper are stable and can be sent in letter-mail. In the laboratory, a small disc of the ion-exchange paper is exposed to antigen and antibody, rinsed, and tested for radioactivity. Discs treated with positive urines are more radioactive than discs from negative urines.


Presence of creatine kinase BB isoenzyme in some patients with prostatic carcinoma.

We have observed creatine kinase isoenzyme BB in the sera of nine of 19 patients with Stage D carcinoma of the prostate. Its presence does not seem to correlate with acid phosphatase activity in serum or the presence of bone metastases as indicated by increased alkaline phosphatase activity in serum. Only three of the nine patients with BB isoenzyme activity detectable in their serum had abnormal values for total creatine kinase activity, but all had abnormal values for alkaline and acid phosphatase activity. All 10 patients with no BB isoenzyme detectable had abnormal acid phosphatase values; however, only seven had increased alkaline phosphatase values and only one had increased creatine kinase activity.


Pharmacokinetics of bendroflumethiazide.

Bendroflumethiazide (bft), 10 mg, was administered orally to 9 healthy volunteers. The concentrations of the diuretic in plasma and urine were determined by gas-liquid chromatography (GLC). Peak plasma levels (86 +/- 18 ng/ml) of bft were reached at 2 +/- 0.4 hr. The concentration declined with a mean t1/2 of 3.0 hr. The apparent volume of distribution averaged 1.48 L/kg. The major part of the drug was eliminated via nonrenal mechanisms, the nonrenal clearance being estimated to 269 +/- 77 ml/min and renal clearance to 105 +/- 24 ml/min. Urinary recovery of the thiazide averaged 30%.


Effects of furosemide on glomerular filtration rate and clearance of practolol, digoxin, cephaloridine, and gentamicin.

Furosemide was shown to decrease inulin clearance in 20 of 27 normal subjects. The depression in inulin clearance occurred in both water-loaded and non-water-loaded subjects. The renal clearance of practolol, but not digoxin, was reduced when furosemide was given. The average total plasma clearances of gentamicin and of cephaloridine over a 6-hr period were decreased after furosemide. The reduced clearances of the antibiotics were associated with higher plasma levels, the increase in antibiotic concentration being as much as 100% at 1 hr after an intravenous bolus injection.


Effects of probenecid on furosemide kinetics and natriuresis in man.

Furosemide kinetics were studied in 4 normal subjects after single intravenous injections (1 mg/kg). One experiment was done after pretreatment with probenecid. The apparent volume of furosemide distribution was unchanged after probenecid (10.9 L). The mean plasma clearance fell from 155 to 85 ml/min and the mean plasma t1/2 rose from 36 to 61 min. Renal clearance of furosemide fell below 50% of control after probenecid, but the kidney remained the main route of its excretion (75% of the dose appeared in the urine). In another experiment in 4 subjects an infusion of furosemide was sustained following a loading dose to maintain a constant plasma level. After a control period, probenecid was given orally. This resulted in a decrease in renal excretion of furosemide with a simultaneous rise in its plasma concentration. Despite the rising plasma furosemide concentration, however, there was a diminution in both urine flow and the excreted fraction of filtered sodium, which suggested some reduction of diuretic action. In doses commonly used, probenecid reduces renal elimination of furosemide in man with only a mild impairment of its diuretic activity. This suggests that furosemide is eliminated predominantly by way of proximal tubular secretion and that tubular rather than plasma concentration is the main determinant of its diuretic effect.


Influence of probenecid and spironolactone on furosemide kinetics and dynamics in man.

The pharmacokinetics and pharmacodynamics following administration of furosemide (40 mg intravenously) have been studied before and after treatment with probenecid (0.5 gm orally every 6 hr for 3 days) and spironolactone (200-mg initial oral dose followed by 50 mg every 6 hr for 3 days) in 6 normal male subjects. Urine losses during each study period were replaced with saline-dextrose-KCl intravenously. The study was performed with the use of a Latin-square design. Probenecid pretreatment induced significant reductions in renal clearance of furosemide by 78%, the extrarenal clearance by 56%, and the volume of distribution by 52%. As a consequence, furosemide half-life was increased by 54%. Probenecid significantly reduced the rate of sodium excretion at all plasma concentrations of furosemide, but the ratio between urinary furosemide concentration and urinary sodium concentration was not altered. Since the proportion of furosemide excreted unchanged in the urine was not markedly changed, total diuretic response was not influenced by probenecid. There was no evidence of any pharmacokinetic interaction between spironolactone and furosemide. The relationship of furosemide kinetics to dynamics observed in these studies confirms that, in man, the diuretic response is determined by drug that reaches the renal tubule rather than the drug level in plasma.


Patterns of plasma concentrations and urinary excretion of salicylate in rheumatoid arthritis.

Intersubject differences in the volume of distribution, whole body clearance, and steady-state plasma concentrations of salicylic acid (SA) were studied in a series of patients with rheumatoid arthritis and healthy control subjects. The measurement of the plasma concentration of SA 12 hr after an oral dose of 1.2 gm aspirin appears predictive of the success of long-term dosage of aspirin. Concentrations below 5 microgram/ml in this single-dose test were associated with failure to achieve therapeutic plasma concentrations of SA (above 150 microgram/ml during long-term therapy with approximately 4.8 gm aspirin per day. Conversely, plasma concentrations above 10 microgram/ml in the single-dose test were associated with levels above 150 microgram/ml during long-term therapy. The volume of distribution of SA correlated poorly with body weight (r = 0.51, p less than 0.01) and did not correlate significantly with plasma albumin levels. Corticosteroids appear to induce the metabolism of SA and most subjects dosed with oral corticosteroids and aspirin 4.8 gm/day did not attain plasma levels of SA above 150 microgram/ml. The clearance of SA was greater in male than in female patients. The difference appears to be of clinical significance since fewer men than women achieved therapeutic plasma concentrations of SA.


The pharmacokinetics of [3H]-vincristine in man.

The pharmacokinetics, metabolism, and excretion of aromatically labeled tritiated vincristine (VCR) was examined in 4 patients. Clearance of radioactivity from the blood was triphasic with half-life t1/2 values of 0.85, 7.4, and 164 min. The initial phases probably represent distribution and binding to formed blood elements which exceeded 50% of the administered dose by 20 min. Excretion of radioactivity was principally fecal, with 33% recovered in the feces by 24 hr and 69% by 72 hr. Considerably less radioactivity (12%) was excreted in the urine over the 72-hr period. Approximately 40% of fecally excreted and 46% of urinary excreted radiolabel represented metabolites, which suggests that at least 34% of the VCR dose was excreted as metabolies. Plasma metabolites represented from less than 1% to 30% or more of radioactivity in plasma. Ultraviolet spectral analysis of all metabolites revealed preservation of the intact VCR dimer, which suggests that metabolism involves alteration of side groups.


Codeine kinetics as determined by radioimmunoassay.

Radioimmunoassay (RIA) was used to determine several pharmacokinetic parameters of codeine in man, including the relative bioavailability after oral and intramuscular administration. The study followed a crossover design in 6 healthy, young (18 to 21 yr), male volunteers. Three subjects received 65 mg codeine phosphate orally in an analgesic mixture which also contained aspirin, phenacetin, and caffeine. At the same time a similar group received an equivalent dose of codeine phosphate in a single intramuscular injection. Two weeks later the study was repeated so that each group received the alternate treatment. Plasma samples were collected at various times after drug administration, and codeine concentrations were determined by a specific RIA procedure. The procedure can detect less than 50 pg of codeine. Following intramuscular administration, peak plasma concentrations (194 to 340 ng/ml) were observed between 0.25 to 1 hr; after oral dosing, peak codeine plasma concentrations (102 to 140 ng/ml) appeared within 0.75 to 1 hr. The mean plasma t1/2 and volume of distribution of codeine following intramuscular injection were 3.32 hr and 5.1 L/kg, respectively. Oral, relative to intramuscular, bioavailability of codeine, based on areas under the codeine plasma curves, was 42% to 71% (mean, 53%).


Procainamide and N-acetylprocainamide kinetics investigated simultaneously with stable isotope methodology.

The pharmacokinetics of procainamide (PA) and N-acetylprocainamide (NAPA) were compared in 3 normal subjects after simultaneous intraveous injection of PA and NAPA-13C. The distribution kinetics of both compounds were modeled with a 3-compartment mamillary system, and it was found that their steady-state distribution volumes were not significantly different, averaging 1.41 L/kg for PA and 1.46 L/kg for NAPA. However, the intercompartmental clearances of NAPA were slower than those of PA. In these normal subjects, the average elimination t1/2 and total elimination clearance for PA were 2.5 hr and 589.8 ml/min, and for NAPA were 6.2 hr and 233.7 ml/min. Mean renal clearances of PA (346.7 ml/min) and of NAPA (199.5 ml/min) exceeded the usual rate of glomerular filtration, which suggests that both compounds are eliminated in part by renal tubular secretion. All subjects were phenotypic rapid acetylators of isoniazid and converted approximately one fourth of the administered PA dose to NAPA-12C. The fate of 15.4% of the administered PA and 14.5% of the administered NAPA-13C was not determined.


Glutethimide and 4-OH glutethimide: pharmacokinetics and effect on performance in man.

The relationship between the plasma concentration of glutethimide (G) and the change from baseline of the standard error of the mean (deltaSDE) of a tracking test was determined in 7 volunteers. There was excellent positive correlation (r = 0.91) between log G and log deltaSDE and good correlation between log  G and deltaSDE (r = 0.77). The metabolite, 4 hydroxyglutethimide, did not contribute significantly to the effect of G administered in therapeutic doses. No trend in performance versus level was found with 5 other tests (finger tapping, card sorting, digit substitution, subtraction, and subjective perception of drowsiness). Although the numbers were small, when the volunteers were divided into smokers (3) and nonsmokers (4) G decreased tracking ability to a greater extent in smokers than in nonsmokers.


Prediction of the lithium ratio in man by means of an in vitro test.

The movement of the lithium ion (Li+) across the membrane of intact erythrocytes incubated in vitro was assessed under two different experimental conditions in which such transfer occurred primarily due to the activity of a lithium-sodium countertransport system. The 13 subjects on whom the in vitro procedures were done subsequently received lithium carbonate for 14 to 56 days, and the extent of accumulation of Li+ by erythrocytes in vivo was measured. While both in vitro procedures yielded data that correlated with the extent of accumulation of Li+ by erythrocytes in vivo, a system measuring the efflux of Li+ from Li+-loaded cells produced a much higher correlation (0.976). The magnitude of this correlation suggests that this in vitro system can be used for further investigations into the relevance of the erythrocyte accumulation of Li+ to the pathogenesis and treatment of affective disorders.


Activation of monooxygenases in human liver by 7,8-benzoflavone.

Addition of 10(-4) M 7,8-benzoflavone to homogenates of human liver samples obtained by autopsy or surgical biopsy increased the rate of benzo[a]pyrene hydroxylation up to 11-fold. 7,8-Benzoflavone also increased the rates of hydroxylation of zoxazolamine and antipyrine at 10(-4) M but inhibited these reactions at 10(-6) M. The effects of 7,8-benzoflavone on the hydroxylation of benzo[a]pyrene and zoxazolamine in microsomes from human liver were similar to those in homogenates. Addition of 7,8-benzoflavone to homogenates of surgical biopsy samples of human liver had little or no effect on the rates of oxidative metabolism of 7-ethoxycoumarin, coumarin, and hexobarbital. Marked individuality for the activating and inhibiting effects of 7,8-benzoflavone was observed in different liver samples. This individuality may result both from the presence of multiple monooxygenases in varying amounts and proportions in the different liver samples and from a selective effect of 7,8-benzoflavone on certain of the monooxygenases.


Changes in the P-wave terminal segment correlated with left ventricular end-diastolic pressure.

A statistically highly significant correlation exists between the values of left ventricular end-diastolic pressure and the dimensions of the terminal segment of the P wave in the electrocardiographic V 1 lead. Values lower than -0.03 mmsec were found only in patients with pathologically elevated left ventricular end-diastolic pressure. In a group of 94 patients with acute transmural myocardial infarction, pathologically low values were found in 43% of the patients on the first day of the disease; on the fifth day of disease the frequency of pathological values of PTF---V 1 sank statistically significantly. A gradual decrease in the values of PTF---V 1 is a sign of bad prognosis in patients with acute myocardial infarction. Changes in the terminal segment of the P wave, at preserved sinus rhythm, can be read off any electrocardiographic curve. Their evaluation offers valuable information about changes in left ventricular end-diastolic pressure, when facilities for catheterization of the patient are not available. This possibility constitutes the main practical value of the method described.


Parasystole associated with fixed coupling ventricular ectopic activity.

In 5 patients ventricular parasystole and fixed coupling ectopic beats were recorded. In two cases ventricular parasystole and bigeminism acted intermittently. In one case the spontaneous transition from ventricular parasystole into parasystolic tachycardia with "reverse coupling" and finally to ventricular bigeminy could be illustrated. In all these cases ventricular parasystole and fixed coupling extrasystolic activity seems to be a close related mechanism. In other two cases there was a mathematical relationship between the sinus and parasystolic discharge rate which explained the periodic fixed coupling of ectopic beats. In one of these cases His bundle electrography was helpful in documenting the ventricular origin of ectopic beats.


Reproduction of twelve routinely used ECG leads from three orthogonal ones.

The paper presents the results of the reproduction of routinely used leads from three corrected orthogonal leads in the system SVEC III modified by L. I. Titomir. Simultaneously with routine and reproduced leads, the differences among them were recorded at every instant of time. In the reproduction for each proband and each lead an individual selection of vector parameters for the given lead was performed. This method was applied in the examination of 150 probands, including 121 patients with myocardial infarction (MI) and 29 healthy subjects. A comparison of routine with reproduced leads showed that in both healthy subjects and patients with MI the limb leads were well reproducible; reproduction of thoracic leads was poorer, especially in patients with MI. Greatest reproduction errors occurred in leads V2 and V3. In 12.5% of patients with anterior-wall infarct, the diagnostically highly important indicators, pathological Q and informative T waves, were not reproduced. The authors assume that in isolated anterior-wall MI, which manifests itself solely in leads V2 and V3, diagnostic errors occur when only three corrected orthogonal leads are used.


Sex chromatin in cardiovascular diseases in elderly and senile women.

In women aged 60--79 years the author determined the frequency of sex chromatin in epithelial cells of buccal mucosa. A higher frequency was found in women with cardiovascular diseases, especially with arterial hypertension and acute disturbances of coronary of cerebral circulation. The role of hormonal factors in the pathogenesis of cardiovascular diseases in elderly women is discussed.


Brain and kidney renin-angiotensin activity in spontaneously hypertensive rats.

Renin activity in peripheral plasma (PRA), renal cortex and brain tissue (cortex, stem, medulla oblongata) homogenates was studied in a bioassay after Serebrovskaya's modification of the method of Pickens. Fourty-nine spontaneously hypertensive rats (SHR) and 26 normotensive Wistar rats (NWR) were used. The results show that: 1) In spite of very high systolic blood pressure, PRA in SHR is in normal ranges; 2) Renin activity exists not only in the brain of NWR, but in the brain of SHR as well. 3) Renin activity in the different parts of the brain is not equal both in NWR and in SHR. 4) Renin activity in the cortex, the stem and the medulla oblongata of SHR is in normal ranges. 5) Renin activity in the medulla is higher than in other areas of the brain, both in NWR and in SHR, the difference being statistically significant in the latter. 6) No perceptible interrelation between arterial blood pressure, PRA, renal renin activity and renin activity in the cortex, and in the stem of the brain was established. It is suggested that higher renin activity in the medulla oblongata might be considered as an indication of the importance of the brain renin-angiotensin system, and its function in the cerebral (neural) regulation of the cardiovascular system. A causal connection between the renal and the brain renin-angiotensin systems, and the onset and maintenance of hypertension is assumed.


Measurement of systolic time intervals through non-invasive polygraphic techniques in healthy subjects and in patients with ischaemic heart disease. Comparison of results obtained by conventional and computer assisted means.

Computer processing of carotidograms is an incontestable methodical asset. Electronic detection of systolic time intervals excludes human errors, makes possible evaluation of a greater number of systoles, and reduces the effect of nonlinearities of recording. The proposed computerized system saves tedious manual calculations and can yield results with mutual correlations within a short period of time.


Intramitochondrial calcification and myocardial inclusions in the rabbit due to anoxic hypoxia.

Intramitochondrial calcium granules and calcification of the mitochondrial cristae were examined in myocardial cells of rabbits in acute anoxic hypoxia, under normal circulation. The alterations were noted first after 2 hours of hypoxia, and became progressively larger and more ubiquitous with increasing periods of anoxic hypoxia. After 8 hours or longer hypoxia intramitochondrial "cristalloids" were found in some mitochondria.


Left ventricular end-diastolic volume during supine exercise in patients with healed myocardial infarction.

Eighteen men with myocardial infarction in their history and without signs of heart failure were investigated at rest and during standard supine exercise. In nine patients aneurysma or diskinesis of the left ventricular wall were found. The left ventricular end-diastolic volume was determined from the wash-out of 133Xe injected into the left ventricle by means of precordial scintillation counting. During exercise the cardiac index rose owing to acceleration of the heart rate, whereas the stroke index remained unchanged, and the left ventricular work and stroke indices increased. The left ventricular end-diastolic pressure, elevated at rest, reached high values during exercise. The left ventricular end-diastolic and residual volumes decreased during exercise in most patients, and simultaneously the systolic ejection fraction increased. In patients with aneurysma or diskinesis the end-diastolic volume both at rest and during exercise does not differ from EDV of other patients. Six patients developed angina pectoris during exercise, but their haemodynamics did not differ significantly. It is concluded that the left ventricle in patients with advanced coronary heart disease and previous myocardial infarction shows the signs rather of diminished compliance than of heart failure during adequate exercise and still possesses some functional reserves.


Management of Reye's syndrome. A rational approach to a complex problem.

Five patients suffering from progressive Reye's syndrome were aggressively treated in our ICU. Our protocol consisted of endotracheal intubation, curarization, mechanical ventilation, hypothermia to 31 degrees C, and fluid restriction. Intracranial pressure (ICP) was monitored and remained below 10-20 torr in all patients except one in whom ICP rose only once (for a few minutes) to 27 torr. Pulmonary artery wedge pressure was monitored in 4 patients and was maintained at 4-5 torr. Duration of active intervention as outlined above was 3-4 days. All 5 patients started with clinical stage IV and EEG grade IV or worse. Four patients had complete recovery and one died. In this group of patients, high ICP was not a feature of the disease. We recommend that use of mannitol for active dehydration be reserved for cases with proven rise in ICP, in which case hemodynamics should be carefully monitored.