[Use of benzoylated DEAE-cellulose for the fractionation of high polymeric DNAs]. A method of fractionation of high polymeric DNAs on a column with benzoylated DEAE-cellulose has been developed. For chromatography it has been used a linear gradient of NaCl in a borate buffer, pH 7.0, concurrent with a gradient in ethanol from 0 to 48% (v/v). It has been shown that DNAs from various sources are eluted by two fractions. For complete elution of the second fraction it has been used supplementary solution of 1 M NaCl in a borate buffer, containing 30% dioxane (v/v). DNAs of every fraction distinguish oneself by base composition. The second fraction, containing DNA more high quantity of GC-pair nucleotides. Some advantages of fractionation on benzoylated DEAE-cellulose are discussed.


[Isolation of sarcolemma acetylcholinesterase fractions by gel-filtration through Sepharose 2B]. Gel-filtration of 0,6 M NaCl and 0,6 M NaCl--0,1% Triton X-100 extracts of freshly isolated sarcolemma through Sepharose 2B (1,5 X 72 cm) has revealed one symmetric peak of acetylcholinesterase activity containing phospholipid and cholesterol, moving faster than fibrinogen and tyreoglobulin. The acetylcholinesterase fraction is substantially separated from other extract proteins. Gel-filtration of extracts from long-store, treated by ultrasound or high concentration of detergent sarcolemma has revealed some peaks of acetylcholinesterase activity, which may be suggested to be degraded forms of the complex high molecular weight structure. All species of acetylcholinesterase are converted by treatment with trypsin to a form moving upon gel-filtration with enzyme-marker catalase. The microsome extracts contain only the form moving with catalase.


[Inhibition of tryptophanyl-tRNA synthetase by modifying ATP analogs]. The interaction between tryptophanyl-tRNA synthetase (EC 6.1.1.2) from beef pancreas and the ATP analogs containing alkylating or phosphorylating groups in the polyphosphate moiety of ATP was studied as an approach to investigate the structure of the enzyme active center. Some of the compounds under study were shown to irreversibly inhibit the enzyme activity; the presence of ATP in the most cases protects the enzyme against inactivation. The kinetic constants Ki and k2 of interaction of the irreversible inhibitors with the enzyme were determined. It was found that the Ki values for a number of irreversible competitive inhibitors are by 1-2 orders of magnitude less than the Km value for ATP; the k2 values were found equal to 0.02-0.04 min-1. this suggests that the compounds may be used as affinity reagents, the most efficient ones being adenosine 5'-(beta-chloroethyl phosphate) and mixed AMP-mesithylene carbonic acid anhydride. The absence of a protective effect of ATP in the case of adenosine 5'-(beta-bromoethane phosphonate) and non-competitive type of reversible inhibition inhibition of the enzyme by adenosine 5'-chloromethane phosphonate indicate that the molecule of tryptophanyl-tRNA synthetase contains sites interacting with adenine nucleotides, other than the ATP binding sites of the active center.


[Accumulation of 14C-nicotinate by membrane preparations]. The incorporation of nicotinic acid (NA) into cells and mitochondria depends on temperature and the presence of glucose, is coupled with the activity of Na, K-ATPase and is disturbed by the energy exchange inhibitors. The addition of a sucrose liver extract or protein-containing fractions of the extract stimulates the 14C-NA accumulation by liver mitochondria. The stimulating effect of the extract on 14C-NA incorporation into mitochondria is retained after dialysis, but is removed upon boiling of the extract. It is assumed that the 14C-NA transport is operated by proteins: NA is incorporated (at least, partially) into liver cells, where it exists both in a free and proteinbound states. It has been found that there exist at least two protein fractions which bind 14C-NA.


[High-yield isolation of chloroplast DNA from Hordeum vulcare barley protoplasts]. A fraction of intact chloroplasts free of other cell components in isolated from barley leaf chloroplasts. Instead of mechanical desintegration of plant tissue, the method described includes the cellulysine treatment, thus increasing the yeild of chloroplast DNA. The mean content of DNA per barley chloroplast is found to be 1.10(-4) g. Base composition of barley chloroplast DNA is 39.8 mol.% of G+C. The treatment of chloroplast DNA with restriction endonuclease EcoRI results in the appearance of 17-19 bands under agarose gel electrophoresis.


[Effect of alcohols on the activity of bacterial agmatinase]. Aliphatic alcohols inhibit the activity of bacterial agmatinase (E. C. 3.5.3.11). A correlation is demonstrated between the inhibition constant and the length of alcohol carbohydrate radical, and the presence of polar groups has decreased the inhibitory effect. The importance of hydrophobic interactions in the formation of agmatinase-substrate and agmatinase-inhibitor complexes is suggested. The increment of free binding energy within the homologous alcohol row is practically constant (0.5-0.7 kcal/mole). Comparison of deltaF amines and their respective alcohols makes possible to evaluate the contribution of electrostatic interaction in the formation of the enzyme-inhibitor complex. A correlation is observed between solubility of alcohols and their inhibitory effect.


[Possible role of ammonium sulfate in inducing invertase biosynthesis in plants]. The induction of invertase from beetroot plants and wheat shoots was studied using the "ageing tissues" technique. It was found that ammonium sulfate at concentration 0.001 M enhances invertase induction. This effect of ammonium sulfate was not revealed when the preparations were pretreated by actidione, puromycin and chloramphenicol. A possibility of activation of invertase induction by ammonium sulfate is discussed.


[Electron microscopic study of the products from dissolving unstabilized fibrin with complex heparin compounds]. Effects of some heparin complex compounds (heparin-urea, adrenaline-heparin, fibrinogen-heparin complexes and secondary complex adrenalin-heparin-fibrinogen) on factor XIIIa unstabilized fibrin were studied using electron microscopy. Fibrillar network of unstabilized fibrin destroys with the formation of globular molecular particles similar to fibrinogen molecule or fibrin monomer ultrastructure. A mechanism of fibrinolytic action of all the complexes mentioned is probably the same, since under dissolving of unstabilized fibrin, structures are found, which are similar to those forming under dissolving of unstabilized fibrin with urea.


[Inhibition of leucyl-tRNA-synthetases by modified ATP analogs]. The interaction between modifying ATP analogs containing alkylating or phosphorylating groups in the polyphosphate moiety of the ATP molecule and leucyl-tRNA synthetases from cytoplasm and chloroplasts of Euglena gracilis (strain Z) was studied. It was shown that most of the ATP analogs irreversibly inhibit the cytoplasmic enzyme, having no inhibiting effect on the chloroplast synthetase. The kinetic constants K1 and k2 for the interaction between the most effective irreversible inhibitors and the cytoplasmic enzyme were determined. The data on the protection of the enzyme activity by substrates against irreversible inhibition suggest, that the effect of the adenosine 5'-(beta-chloroethyl phosphate) is directed to the ATP-binding site of the cytoplasmic enzyme, whereas the mixed anhydride of AMP and mesithylene carbonic acid acts predominantly on the binding site of 3'-terminal adenosine of the tRNALeu molecule. ATP analogs may be effectively used for affinity labelling of the cytoplasmic leucyl-tRNA synthetase.


[Study of the role of arginine residues in aspartate transaminase from chicken heart cytosol]. Reaction of 1,2-cyclohexanedione with chicken heart cytosolic aspartate transaminase results in loss of enzyme activity complying to first order kinetics up to 70% inactivation. The inactivation rate is markedly decreased in the presence of alpha-ketoglutarate, glutarate or alpha-methylaspartate. The number of arginine residues modified per subunit was approximately two (in enzyme preparations which retained 30% residual activity). The diketone-modified enzyme nearly completely loses affinity for alpha-methylaspartate and glutarate; in contrast, its ability to bind alpha-alanine and catalyze its transamination half-reaction with the bound coenzyme remains unimpaired. From these data it can be inferred that a functional arginine residue is the cationic binding site for the distal carboxyl group of the substrates. The transaminase apoenzyme was inactivated with cyclohexanedione at the same rate as reconstituted holoenzyme. Measurements of circular dichroism showed that the modified apoenzyme is capable to bind pyridoxal-P. No evidence was obtained for the presence of an arginine residue in the coenzyme binding site.


[Comparison of the catalytic properties of alpha- and beta-forms of thrombin]. 18-25-fold purified alpha-thrombin, having high esterase activity and coagulating ability of 2500 NIH u per 1 mg of protein, was isolated using chromatography of commercial thrombin through SP-Sephadex C-50. Limited proteolysis of alpha-thrombin on the column with immobilized trypsin resulted in the appearance of beta-thrombin with alpha-thrombin-like esterase activity and tracing coagulating activity (2-5 NIH u per 1 mg of protein). Molecular weight analysis of alpha- and beta-thrombin forms suggests that a peptide (or peptides) with Mr of 1100 is splitted off under proteolysis. Some similarity is revealed in kinetic parameters (Km(app) and kkat) of TAME and BAME hydrolysis by alpha- and beta-thrombin, although Km(app) is somewhat low (approximately 2-fold) for alpha-thrombin. Investigation of TAME hydrolysis kinetics by both thrombin forms at a wide range of substrate concentrations has revealed the effect of substrate activation. Kinetic constants Ks and beta for high substrate concentrations are calculated. It is suggested that the similarity of alpha- and beta-thrombin action on arginine esters and sharp differences in their effect on fibrinogen may be a result of a disturbance of substrate-binding region of beta-thrombin active site.


[Determination of the kinetic parameters of individual stages of M(alpha)-arylsulfonyl-L-arginine ester hydrolysis by thrombin and trypsin]. The N(alpha)-arylsulfonyl-L-arginine ethyl- and propyl esters were synthesized and the kinetics of their hydrolysis by thrombin was studied. The values of kcat and Km were shown to depend on the structure of the leaving group and to decrease in the line: OCH3 greater than OC2H5 greater than OC3H7. Using methanol as an additional nucleophile, the kinetic parameters - k2, k3 and Ks - were measured for both thrombin- and trypsin-catalysed reactions. A similarity of two enzymes at the stage of Michaelis complex formation was revealed: the Ks values for both enzymes were practically identical (18.10(-5)M). The differences between thrombin and trypsin were observed at the stages of chemical conversion of substrates and were especially well-pronounced at the stage of acylation. It was shown that the k2 values for thrombin were lower than that for trypsin and the k2/k3 ratio of TAME hydrolysis by trypsin was equal to 21, while that for thrombin was 4.5. This finding is indicative of an essential role of the acylation step in thrombin-catalysed hydrolysis of the esters under study.


[Disintegration and fractionation of barley chloroplast membranes at different concentrations of digitonin and chloroplasts]. Chloroplasts pretreated with digitonin were fractionated by electrophoresis in order to study the arrangement of chloroplast membranes. Different types of particles resulting from the electrophoresis are shown to be due to fractionation of the chloroplast membranes of the same type rather than to the isolation of particular types from the chloroplasts of different ages. The stopped-flow measurement of quickly changing light scattering demonstrated that the membrane solubilization increased with the increase in the digitonin concentration. Meanwhile the fractionation result (the amount of particles in the electrophoretic zones and their spectral properties) remain very similar over a wide range of the detergent and chloroplast concentrations. This is explained by the fact that the increasing detergent concentrations do not cause particle destruction. Thus the method used revealed the particles which different from one another before they were isolated from the membrane. They are fractionated following the natural boundaries between them and the containing membrane. In other words, chloroplast membranes have a discrete set of submembrane particles of different types.


[Conformation of gamma-aminobutyric acid and its receptors]. The preferable conformations of the inhibitory transmitter gamma-aminobutyric acid (GABA) and its specific inhibitor bicuculline are due to the occupation of the GABAreceptor by a part of the bicuculline molecule that is isosteric with the biologically active conformation of GABA. In the present review are described characteristics of sodium-independence receptor sites and evaluated the regional distribution of postsynaptic receptor binding for GABA in the central nervous system. The postsynaptic GABA receptor has been labeled by direct binding of 3H-GABA. The structural analogues and antagonists of GABA were investigated for inhibition of GABA binding. The developmental changes in the activity of glutamic acid decarboxylase, the GABA uptake mechanism and GABA receptor binding in embryo brain were examined. Different models for the GABA-receptor interaction are considered. Data of the stoichiometry of GABA-receptor interactions indicate that the interaction is probabilistic and that 3 molecules of GABA are needed to activate the receptor probabilistically. It was found that 1 molecule of picrotoxin was capable of blocking the interaction with 3 GABA molecules. The hypothetical model of cooperative action of GABA-recepto-inophore complexes involving membrane mobility is also discussed. The transitive complex GABA with ligand binding of Na+, Cl- and the conformational changes of GABA-receptor is proposed.


[Purification and properties of 3-hexulosephosphate synthase from facultative methylotroph Pseudomonas oleovorans]. 3-Hexulosephosphate synthase (HPS), the key enzyme of the hexulosephosphate cycle of formaldehyde fixation, was isolated from facultative methylotroph Pseudomonas oleovorans. Enzyme was purified 100-fold. The purification procedure involved fractionation with ammonium sulfate, gel-filtration on Sephadex G-150 and chromatography on DEAE-Sephadex A-50. The purified enzyme gave single band on analytical polyacrylamide gel electrophoresis. Optimal conditions for activity of HPS are: pH 7,0, temperature 50 degrees C. The molecular weight was calculated to be 45 000 from gel-filtration experiments. HPS is active only in the presence of Mg2+ or Mn2+. Ribulose-5-phosphate is the sole acceptor of formaldehyde. Activity of the enzyme is inhibited by NADH and NADPH.


[Chemical modification of epsilon-NH2 groups of lysine residues in horseradish peroxidase. Accessibility of these groups to different modifying agents]. Modification of epsilon-NH2 groups of lysine residues in horseradish peroxidase by acetic, propionic, butyric, valeric and succinic anhydrides and trinitrobenzenesulfonic acid (TNBS) is carried out. All the anhydrides modify at 0 degrees C four epsilon-NH2 lysine groups, and TNBS--three groups. TNBS modifies all 6 peroxidase epsilon-NH2 groups at 40 degrees C. Possible spacing of lysine epsilon-NH2 groups in the enzyme molecule is discussed.


[Model for auxin receptor].

[Model for auxin receptor]. Literary data on correlation between the structure and auxin activity of aryl-, arylalkan- and aryloxyalkancarboxylic acids and those of 3-indolylacetic acid are reviewed. The structure of auxins is compared to that of their inactive structural analogs in terms of a literary hypothesis on the bifunctional binding of auxins to the receptor, which allowed to construct a model for the structure of the auxin receptor (site responsible for intrinsic binding of auxins). The geometric parameters of this receptor site and specific arrangement of specific auxin binding sites have been proposed. It is demonstrated that during auxin absorption on the receptor the conformation of the latter is changed, which may be accounted for as a primary functional effect of auxins on the receptor.


[Study of evolutionary changes in subfraction composition of histone H1 in birds]. Analysis of electrophoretic mobility of histone H1 subfractions from liver, brain and erythrocytes of 41 bird species was carried out. Subfractions of erythrocyte H1 histones from each species were compared with those of thrush Turdus musicus. The majority of species proved to possess a set of electrophoretically similar subfractions. Interspecific differences in histone H1 were mainly due to the differences in the ratio of those subfractions. The identity of the electrophoretic mobility and similar contribution to H1 histone from the respective tissues in different species permits to consider certain subfractions as homologous ones. The conservation of electrophoretic mobility  for homologous subfractions of the birds of different orders shows a high evolutionary conservatism of the corresponding genes. It seems that in the course of evolution only a change in the expression of some of those genes occurs.


[Stabilization of polyribosomal mRNA in rat liver cells under protein synthesis inhibition by cycloheximide]. It was found that in the course of centrifugation of the postmitochondrial fraction from rat liver in the sucrose concentration gradient, mRNA located in the EDTA-resistant structures sedimented together with polyribosomes. The same structures were detected in the polyribosomal fraction of liver cells after injection of ethionine to the animals. Centrifugation of polyribosomes through a layer of 2.0 M sucrose at 105 000 g for 4 hrs and subsequent centrifugation through a layer of 1.0 M sucrose resulted in the disappearance of the complexes from the polyribosomal fraction. Evidence for the absence of destruction of labelled polyribosomal mRNA in liver cells following the injection of cycloheximide were obtained. Actinomycin D also stabilized polyribosomal mRNA in liver cells, however, in a much lesser degree as compared to cycloheximide. Possible mechanisms of the stabilizing effects of cycloheximide and actinomycin D on polyribosomal mRNA in liver cells are discussed.


[Proteolytic enzymes bound to Bac. thuringiensis crystals]. It was demonstrated that crystals of entomopathogenic protein from Bac. thuringiensis contain admixture of proteinase either adhered to their surface on inconponated into crystal lattice defects. A proteolytic action, particularly when enhanced by crystal dissolution, causes progressive degradation of crystal proteins with molecular weights of 140 000--129 000 down to the components with smaller molecular weights. This may, at least, partially account for the contradictions in the literature data on crystal composition. Using synthetic peptide substrates and specific inhibitors, it was shown that the enzymes incorporated into crystals belong to serine and metalloproteases. The presence of leucine aminopeptidase was also noted. A method for enzyme separation from crystal has been developed.


[Isolation and some properties of restriction endonuclease from Bacillus amyloliquefaciens]. A partially purified preparation of restriction endonuclease Bam I was isolated from the cells of Bacillus amyloliquefaciens. The purification procedure was a modification of a method described by Wilson and Young. Isolation and purification of the enzyme involved disruption of the cells by ultrasonication, treatment with streptomycin sulfate, fractionation by ammonium sulfate, chromatography on DEAE-cellulose and hydroxylapatite and rechromatography on DEAE-cellulose. Restrictase Bam I splits the linear double-chain DNA molecule of phage gamma into six fragments. The enzyme retained its stability under storage on the ice for 1,5--2 months in a Na- or K-phosphate buffer with beta-mercaptoethanol (10 mM).


[Soluble high molecular weight derivatives of pancreatic trypsin inhibitor. Isolation and properties of the inhibitor bound to carboxymethyl cellulose and diethylaminoethyl dextran]. Soluble preparations of pancreatic trypsin inhibitor, which is bound to carboxymethyl cellulose- and diethylaminoethyl dextran activated by chlorine derivatives of s-triazine were isolated. The activity of the pancreatic inhibitor bound to carboxymethyl cellulose treated by 2-amino-4,6-dichloro-s-triazine was completely retained. Simultaneously, the reaction cyanuric chloride activation of water-soluble polysaccharide carriers in a non-aqueous solvent was studied by assaying the amount of chlorine atoms on the treated matrix. The gel-filtration experiments show that the cross-linkages between polysaccharide molecules appear during the activation reaction. A structural model of a carboxymethyl cellulose-bound pancreatic inhibitor is proposed.


[Effect of phloridzin on photosynthetic electron transport]. Phloridzin (2',4',6',4-tetraoxyhydrochalcon-2'-glucoside) was used to study the localization of synthesis of ATP in the electrontransporting chain of photosynthesis. It was shown that phloridzin inhibits the rate of photoreduction of NADP+ by isolated pea chloroplasts by 40%, electron transport via cytochrome f by 100% and via plastocyanin--by 50%. The "crossover" experiments demonstrated that phloridzin inhibits ADP-induced photoreduction of cytochrome f, having no effect on plastocyanin under identical conditions. It is assumed that the site of ATP synthesis is localized on the reduced site of cytochrome f, while the carrier itself is located in the electron transporting chain coupled to phosphorylation. It is possible that only part of the plastocyanin molecules are located in the phosphorylating pathway of electron transport.


[Electrophoretic and chromatographic studies of zein]. Total zein is isolated from fat-free maize flour and studied using electrophoresis in acrylamide gel gradient (7-30%) and gel filtration through Sephacryl S-200 (Superfine) and Sepharose CL-6B. Molecular weights of electrophoretic and chromatographic fractions are determined. 12 electrophoretic and 5-7 chromatographic fractions are found in zein, their molecular weight being 40000-280000 daltons.


[Effect of hydrocortisone on the properties of rat liver polyribosomes, metabolism, and template activity of polysomal poly-A-containing RNA]. The effect of hydrocortisone on the amount of newly synthesized polyribosomal poly-A+-RNA and its translation activity and the distribution of polyribosomes in the induction dynamics according to their size were studied. It was shown that 3-5 hours after intraperitoneal injection of hydrocortisone the incorporation of labelled precursors into polyribosomal poly-A+-mRNA is increased, which is accompanied by rapid accumulation of mRNA in the polyribosomes. Under prolonged induction those parameters come down to the initial level. 4-7 hours after the injection of the hormone the relative amount of heavy polyribosomes (350-412S) in liver cells is increased. It was found that hydrocortisone significantly changes the specific translation activity of polysomal poly-A+-mRNA: it shows an increase 2-4 hours after the hormone injection and returns to the initial level 12 hours after the injection.


[The absence of coupling between the formation of membrane potential and electron transfer via the alternative pathway of cyanide-resistant mitochondria]. The estimation of membrane potential of cyanide-resistant mitochondria of Candida lipolytica yeast was carried out using positively charged dye phenosafranine. The electron transfer via alternative pathway of cynide resistant mitochondria was shown not to be coupled with the formation of potential on membrane mitochondria.


Sephadex-gel filtration (SGF) in infant and adult Gunn rats. SGF was compared in infant and adult homozygous Gunn rats. Without any drug application, the test was negative in the adult animals, whereas a remarkable percentage of positive SGF was obtained in 5-7-day-old rats, especially in those undernourished and/or intensely icteric. Unexpectedly, in infant rats the rate of positive SGF after sulfadimethoxine injection was lower than in the untreated control group. Moreover, in 9-10-day-old animals who had positive SGF before the injection, the test became negative 15-20 min after sulfadimethoxine application in vivo. No convincing explanation could be given, but drug interference with the binding capacities of the Sephadex column could be excluded by appropriate in vitro tests.


Undersulfated urinary glycosaminoglycans in preterm newborns. The glycosaminoglycans (GAG) isolated from pooled urine of preterm newborns were fractionated by stepwise elution from a Dowex 1-X2 column. Analytical reactions, cellulose acetate electrophoresis and enzymatic digestions with chondroitinases and testicular hyaluronidase were performed on each fraction. Chondroitin-4-sulfate and chondroitin-6-sulfate constitute about 60% of urinary GAG; heparan sulfate amounts to about 20%, while chondroitin represents 12% of total GAG; dermatan sulfate, hyaluronic acid and keratan sulfate are present in small traces. Approximately 30% of the total is constituted by nonsulfated GAG.


Fetal and placental growth in the rat following differential reduction of litter size. The effects on fetal and placental growth of differential litter size reduction on day 10 (LR10) and day 13 (LR13) were determined in the rat. LR10 resulted in significant increments in placental weight, fetal body and brain weight, and brain protein content in the survivors at day 21. A marginal effect on brain cellularity (DNA) was observed. Values for these parameters generally followed the extent of reduction. Placenta weight was also related to original litter size. In contrast, LR13 which was associated with survival of supernumerary placentae (those without a fetus) produced no growth enhancement. Prenatal development was discussed in terms of general systemic influences and hormonal regulatory mechanisms.


Materno-fetal transport of creatine in the rat. The transfer of 14C-creatine to the rat fetus was studied following continuous i.v. infusion into the mother. In the presence of a relatively constant maternal plasma 14C-creatine concentration, creatine was accumulated by the chorioallantoic placenta and visceral yolk sac to concentrations higher than that found in maternal or fetal plasma. The ability of the extraembryonic membranes to accumulate creatine changed during gestation; nevertheless, these membranes concentrated creatine against a gradient throughout the period studied (14-22 days of gestation). Neither 14C-creatine nor 14C-urea were concentrated in the placentae or fetal plasma when compared to maternal plasma. Simultaneous infusion of beta-guanidinopropionic acid with 14C-creatine reduced both movement and accumulation of creatine into the fetoplacental unit. It is concluded that the accumulation of creatine by the chorioallantoic placenta and by the visceral yolk sac is an active process with creatine diffusing down its concentration gradient into the fetal circulation.


Orotic acid: its influence on nuclear RNA and microsomal protein synthesis in brain cells of undernourished rat pups. Malnourished rat pups were supplemented with orotic acid. RNA-synthesizing activity of isolated brain cell nuclei and microsomal protein synthesis was measured in vitro. A marked increase was found in the activity of nuclear RNA synthesis but not in microsomal protein synthesis after orotic acid treatment for 7 days.


The effects of glucose and alanine infusion on urea production and gluconeogenesis in the starved newborn guinea pig. We studied the effects of glucose, alanine, and saline infusion in newborn guinea pigs. After acute (24-30 h after delivery) or prolonged (96-100 h after delivery) starvation, alanine or glucose infusion significantly increased liver glycogen concentration. Alanine infusion increased plasma glucose concentration and urea production rate (UrP). After acute starvation, a continuous 12-hour glucose infusion did not affect either plasma alanine concentration or UrP; however, after prolonged starvation, glucose infusion significantly reduced both variables. After prolonged starvation, the newborn guinea pig mobilizes alanine for gluconeogenesis. Increased protein catabolism is a consequence of this process. Glucose infusion negates the need for gluconeogenesis and spares body protein.


Cord and maternal serum meperidine concentrations and clinical status of the infant. The possible relationship between the use of meperidine during labor and fetal or neonatal distress was investigated by correlating clinical parameters with maternal and cord serum concentrations of meperidine. The serum concentration of meperidine was determined by a radioimmunoassay which is 100-fold more sensitive than previously employed techniques and can detect as little as 0.02 nmol meperidine/ml. The incidence of fetal or neonatal depression following meperidine administration was higher (6 of 17 infants) than in a control group but no correlation between suppression and the route of meperidine administration or the serum meperidine concentrations at the time of delivery was found.


Longitudinal studies of infants with the Wilson-Mikity syndrome. Clinical, radiological and mechanical correlations. Simple clinical measurements were performed on a group of 23 very low birth weight babies (less than 1500 g and less than 32 weeks gestation), and combined with pulmonary function studies in 13 cases. The studied included a special single-breath analysis, or Tidal Resistance Profile, which enabled us to detect air trapping. 2 infants developed classical Wilson-Mikity syndrome and 2 further infants exhibited a milder, shorter form of the same disorder. All 4 survived. Results suggest that all infants in this maturity group are at risk of developing Wilson-Mikity syndrome, and that it comprises a wide spectrum of respiratory dysfunction.


An in vivo method to evaluate the effect of materials upon arterial thrombosis. An in vivo method to evaluate the effect of materials upon arterial thrombosis was developed that minimized the effect of surgical artifacts and provided a test which was critically sensitive to the surface properties of the materials. The procedure has general applicability to elastomers, either solvent cast or mold polymerized, and involves the implantation of segments of the test materials within canine femoral and carotid arteries. The sensitivity of this technique was demonstrated by comparing two surface preparations of a segmented polyurethane: as-cast and ion sputtered. There was a striking difference in the rate of thrombus development on as-cast polyurethane implants compared with sputtered polyurethane implants. After 1 hr. of implantation the surface of the as-cast polyurethane was covered with a monolayer of platelets and leukocytes, whereas thrombus development progressed more rapidly on the sputtered polyurethane surface and at 1 hr. it was covered with pillars of platelets and leukocytes with fibrin accumulation between pillars. The method was also useful for long-term studies, which showed that a thin layer of thrombus developed on both polyurethane surfaces and by 1 wk. after implantation the surfaces of the thrombi became endothelialized.


Spinal fixation using acrylic bone cement: mechanical property measurements. Following dorsal laminectomies (L2-L3), the resultant spinal instabilities were stabilized by placing four Steinmann pins and embedding them in acrylic bone cement. The cement was molded about the instability and incorporated the pins and articular process. Mechanical testing was performed to evaluate the strength of the resultant union between the two vertebrae. Three types of samples, in vivo, in vitro and normal were tested using a specially built apparatus. The results showed average maximum loads of 150, 180 and 180 Newtons respectively which represent 29, 42 and 37 MPa in calculated shear stresses. The results indicate that the use of bone cement with pins can stabilize the injured vertebral discs easily iwth the maximum load or shear strength equivalent to that of normal vertebral discs.


A numerical method to model excitable cells. We have extended a fast, stable, and accurate method for the numerical solution of cable equations to include changes in geometry and membrane properties in order to model a single excitable cell realistically. In addition, by including the provision that the radius may be a function of distance along an axis, we have achieved a general and powerful method for simulating a cell with any number of branched processes, any or all of which may be nonuniform in diameter, and with no restriction on the branching pattern.


High gradient magnetic separation of erythrocytes. The high gradient magnetic separation technique has been applied to separate paramagnetic erythrocytes from a cell suspension that also contained diamagnetic cells. Paramagnetism was induced in the red blood cells by oxidizing the iron atoms in the cell hemoglobin to the ferric state (methemoglobin). Diamagnetic cells were either untreated erythrocytes, containing oxyferrohemoglobin, or leukocytes in a suspension of mouse spleen cells. Cell suspensions were passed through a column containing 40 micron diameter stainless steel wire in a high magnetic field (33 kG). The paramagnetic cells were retained on the surface of the wire while the diamagnetic cells passed through. Elution of the paramagnetic cells was accomplished by removing the column from the magnet, in effect turning off the field.


Semiempirical calculations of model deoxyheme. Variation of calculated electromagnetic properties with electronic configuration and distance of iron from the plane. The electronic structure of deoxyheme units and the electric field gradient at the iron nucleus are calculated as a function of the displacement of the iron from the heme plane. The high spin state has a calculated minimum energy with the iron out of plane at a distance similar to that observed for metMb. This could correspond to a relaxed form of deoxyHb, while the greater displacement observed in the intact protein could be strained. The calculated electric field gradient is relatively insensitive to iron displacement from the heme plane. This insensitivity could account for the very similar values of quadrupole splittings observed in low-affinity deoxyhemoglobin A and its separate high-affinity alpha and beta subunits, and still allow small differences in iron displacement consistent with the stereochemical trigger mechanism of cooperativity.


Electrostatic calculations for an ion channel. I. Energy and potential profiles and interactions between ions. The electrostatic energy profile of one, two, or three ions in an aqueous channel through a lipid membrane is calculated. It is shown that the previous solution to this problem (based on the assumption that the channel is infinitely long) significantly overestimates the electrostatic energy barrier. For example, for a 3-A radius pore, the energy is 16 kT for the infinite channel and 6.7 kT for an ion in the center of a channel 25 A long. The energy as a function of the position of the ion is also determined. With this energy profile, the rate of crossing the membrane (using the Nernst-Planck equation) was estimated and found to be compatible with the maximum conductance observed for the gramicidin A channel. The total electrostatic energy (as a function of position) required to place two or three ions in the channel is also calculated. The electrostatic interaction is small for two ions at opposite ends of the channel and large for any positioning of the three ions. Finally, the gradient through the channel of an applied potential is calculated. The solution to these problems is based on solving an equivalent problem in which an appropriate surface charge is placed on the boundary between the lipid and aqueous regions. The magnitude of the surface charge is obtained from the numerical solution for a system of coupled integral equations.


The kinetics of thymine dimer excision in ultraviolet-irradiated human cells. We have investigated the kinetics of the loss of thymine dimers from the acid-insoluble fraction of several ultraviolet (UV)-irradiated cultured human cell lines. Our results show that UV fluences between 10 and 40 J/m2 produce an average of 21-85 x 10(5) thymine dimers per cell and an eventual maximal loss per cell of 12-20 x 10(5) thymine dimers. The time for half-maximal loss of dimers ranged from 12-22 h after UV irradiation. In contrast, the time for half-maximal repair synthesis of DNA measured by autoradiography was 4.5 h. This figure agrees well with reported half-maximal repair synthesis times, which range from 0.5 to 3.6 h based on our analysis. The discrepancy in the kinetics of the loss of thymine dimers from DNA and repair synthesis is discussed in terms of possible molecular mechanisms of thymine dimer excision in vivo and in terms of possible experimental artifacts.


Excision repair of ultraviolet damage in mammalian cells. Evidence for two steps in the excision of pyrimidine dimers. The incidence of pyrimidine dimer formation and the kinetics of DNA repair in African green monkey kidney CV-1 cells after ultraviolet (UV) irradiation were studied by measuring survival, T4 endonuclease V-sensitive sites, the fraction of pyrimidine dimers in acid-insoluble DNA as determined by thin layer chromatography (TLC), and repair replication. CV-1 cells exhibit a survival curve with extrapolation number n = 7.8 and Do = 2.5 J/m2. Pyrimidine dimers were lost from acid-insoluble DNA more slowly than endonuclease-sensitive sites were lost from or new bases were incorporated into high molecular weight DNA during the course of repair. Growth of CV-1 cultures in [3H]thymidine or X-irradiation (2 or 10 krads) 24 h before UV irradiation had no effect on repair replication induced by 25 J/m2 of UV. These results suggest that pyrimidine dimer excision measurements by TLC are probably unaffected by radiation from high levels of incorporated radionuclides. The endonuclease-sensitive site and TLC measurements can be reconciled by the assumption that pyrimidine dimers are excised from high molecular weight DNA in acid-insoluble oligonucleotides that are slowly degraded to acid-soluble fragments.


Ionic channels in excitable membranes. Current problems and biophysical approaches. Ionic channels are gated aqueous pores whose conformational changes are driven by the electric field in the membrane. Gating may be studied by three electrical methods: ionic current transients, ionic current fluctuations, and "gating current," and probably occurs through a series of conformational changes in the channel leading to an all-or-nothing opening of the pore. When the potential is held constant, the gating steps come to equilibrium rather than reaching an energy-dissipating, cyclic steady state. The kinetic models now in use eventually need to be changed to correct disagreements with several recent studies. Diffusion of ions through open channels is very fast but involves many interactions of ions, pore, and solvent that lead to ionic selectivity, saturation, block, and flux coupling. Our description of the ionic fluxes can be improved by abandoning continuum models in favor of more structured ones. Problems to be solved include determining how many ions occupy a channel at once and what to be solved include determining how many ions occupy a channel at once and what kind of energy barriers they must cross in traversing the membrane. Ultimately we will need to know the chemical structure of the whole system to understand how it functions.


Interactions between intrinsic membrane protein and electric field. An approach to studying nerve excitability. We have approached the problem of nerve excitability through three questions: (a) What is the diagram for a channel? That is, what conformational states can the protein assume, and what transitions between these conformations are permitted? (b) What is the channel conductance associated with each conformation the channel can assume? (c) How do the rates for conformational transition depend upon membrane potential? These three questions arise from a standard statistical mechanical treatment of a nerve membrane containing several classes of identical, independent channels. Gating of channels, in this view, is associated with conformational changes of the channel protein, and it is assumed these conformations are distinct. The precise formulation of these questions is presented in terms of the theoretical treatment, and the approaches we have taken to answer the questions are indicated. Our present results indicate: Transition rates should depend exponentially on membrane potential over a limited voltage range, but probably will show a more complex dependence for extremes of the range; channels probably can take on only two conductances, open and shut, but more complicated situations are not entirely excluded; the diagram for a channel cannot be determined from standard voltage clamp data alone, but by studying gating currents and conductance fluctuations, it should be possible to select between alternative plausible physical mechanisms.


The effects of calcium2+ and magnesium2+ on the electrophoretic mobility of chromaffin granules measured by electrophoretic light scattering. Electrophoretic light scattering was used to determine the electrophoretic mobility distributions of isolated bovine adrenal chromaffin granules as a function of divalent metal ion concentrations. Changes in the electrophoretic mobility reflected changes in the surface charge density of the granules. Ca2+ and Mg2+ (0.10--2.0 mM) were equally effective in reducing the electrophoretic mobilities. These findings are consistent with recent studies of the binding of Ca2+ and Mg2+ to the surface of chromaffin granules and are further evidence that the specific role of Ca2+ in exocytosis is due to effects other than the ability of Ca2+ to decrease the electrostatic repulsion between negatively charged membranes.


Profiles in altered metabolism. II.--Accumulation of homogentisic acid in serum and urine following acetylsalicylic acid ingestion. Homogentisic acid elevations have been found in urine and serum of patients on long term heavy salicylate therapy and those acutely intoxicated with acetylsalicylic acid. Loading of two normal volunteers with single oral acetylsalicylate doses produced transient elevations in urinary homogentisic acid excretion. These findings suggest that heavy salicylate use results in the partial inhibition of homogentisic acid oxidase in vivo.


Competitive reactions of protonated polyamines in the gas phase. The chemical ionization mass spectra of polyamines have been used to determine the importance of competing intramolecular reactions of protonated molecules in the gas phase. Multifunctional molecules react if possible in the order of intramolecular displacement-cyclization reactions, stabilization of the protonated molecular ion through collision stabilized intramolecular hydrogen bond formation and fragmentation of an isolated protonated function. Intramolecular nucleophilic displacement reactions closely parallel the relative rates of cyclization reactions in solution. These reactions may occur from protonated functions which are neither the initial nor the most stable sites of protonation.


Mass spectrometry and gas chromatography of trimethylsilyl derivatives of catecholamine related molecules. The trimethylsilyl derivatives of approximately 50 compounds related in structure to biogenic catecholamines have been studied in terms of their gas chromatographic and mass spectrometric behavior. The electron impact mass spectra of the trimethylsilyl derivatives of 3,4-dihydroxyphenylethylamine, 3,4-dihydroxyphenylacetic acid and 3,4-dihydroxyphenylethylamine and deuterated isotopic variants were compared to determine fragmentation characteristics of the amines, acids, alcohols and amino acids within this class of compounds. Analysis of shifts in the masses of major diagnostic ions in the spectra of structural analogs of these compounds has shown that structural modification of the structure can be identified and localized within these molecules. The gas chromatographic characteristics of these derivatives are reported, in terms of methylene unit values.


A comparison of t-butyldimethylsilyl and trimethylsilyl ether derivatives for the characterization of urinary metabolites of prostaglandin F2alpha by gas chromatography mass spectrometry. t-Butyldimethylsilyl and trimethylsilyl ether derivatives of a series of methylated prostaglandin F2alpha metabolites have been compared with respect to their gas chromatographic and mass spectrometric properties. The t-butyldimethylsilyl derivatives had considerably higher retention indices (approximately 2.2 C units per silyl group) than their trimethylsilyl ether counterparts when analysed on the (non-polar) OV-1 stationary phase. Electron impact induced fragmentation patterns were strongly dependent upon the type of silyl ether employed and on the nature of the prostaglandin omega sidechain; the mass spectra of pairs of t-butyldimethylsilyl and trimethylsilyl ether derivatives were found to differ appreciably in several respects and to afford complementatary structural information.


Determination of the amino acid sequence of the C-terminal cyanogen bromide fragment of myoglobin from bottle nosed dolphin by mass spectrometric peptide mixture analysis. The C-terminal cyanogen bromide fragment of myoglobin from bottle nosed dolphin was digested with trypsin and two different derivatives of the digest were prepared, one being deuteroacetylated and deuteropermethylated, the other being submitted to one step of Edman degradation and then acetylated and permethylated. Comparison of the results obtained by computerized interpretation of the mass spectra of these two derivatives, combined with previous information about the sample, facilitated the sequence determination of all the peptides in the sample. Alignment of the peptides could then be carried out by homology with myoglobin from other cetaceans.


Mass spectrometry of some permethylated apiosyl nucleosides. The mass spectra of the permethyl derivatives of a group of synthetic D-apio-L-furanosyl nucleosides were collected and compared with those of their ribosyl analogs. While most of the m/e values are the same between analogous spectra, some of the relative intensities differ markedly. Each spectrum contains characteristics ions which are probably due to fragmentation of the dissimilar sugar moieties.


Taxonomy and phylogeny of chytrids.

Taxonomy and phylogeny of chytrids. Thallus development, zoospore size, zoospore ultrastructural complexity and organization, and flagellum length are cited as important in phylogeny of the Chytridiales (chytrids) and should be the bases for this classification. Discharge of zoospores is also cited as important although emphasis is not placed on operculation. It is proposed that simple, eucarpic, monocentric chytrids which discharge zoospores following dissolution of the sporangium wall evolved into multipapilliate species of Rhizophydium and 2 lines of evolution from these species are documented with examples. In one line they evolved into single-pored species such as R. pollinis-pini, then to Chytridium and tendochytrium-like chytrids and to polycentric chytrids such as Nowakowskiella and Cladochytrium. The second line evolved from multipapilliate Rhizophydium species to Phlyctochytrium and Entophlyctis, and to the order Blastocladiales. Rhizophlyctis rosea and some olpidiaceous chytrids, because of a fibrillar rhizoplast in their zoospores do not appear close to Rhizophydium-related chytrids and may require separate subordinal rank in classification. It is proposed that zoospores of the advanced members of the class Chytridiomycetes have reached limits for further zoospore development because of size; however, the zoospore is no longer essential for them because they have developed a filamentous habit of growth.


Phylogenetic implications of the microbody-lipid globule complex in zoosporic fungi. Chytridiomycetous fungal zoospores contain a unique and intricate association of organelles, the 'microbody-lipid globule complex' (MLC). The spatial arrangement of organelles in the MLC appears important in the utilization of lipid globules for energy, but in addition, the structural association of organelles in the MLC reveals phylogenetic trends within this diverse group of organisms. Variations in the structure of the MLC correlate well with current phylogenetic concepts of aquatic fungi, yet suggest new relationships among these posteriorly uniflagellate zoospores. Based upon the organization of organelles in the MLC, 4 basic patterns of MLCs can be recognized, and these correspond to the 4 orders of Chytridiomycetes. The MLC in its simplest form consists of a microbody appressed to the edge of a lipid globule. In more highly organized MLCs, mitochondria subtend the microbody and a cisterna surmounts one side of the lipid globule. The organization and structure is still more complex in other MLCs where ER is elaborated into a tubular network of membranes or where small microbodies or mitochondria fuse into 'giant' organelles. The structural organization of the MLC provides an additional criterion by which the phylogeny of awuatic fungi can be evaluated.


Microbial toxins, their functional role and phylogenetic validity. Microbially produced toxins, which appear to lack a role in microbial survival, may be antimicrobial compounds of significance to the producers. These toxin/antibiotics may act against cell metabolism shared by man or animals and other microorganisms. Protein toxin/antibiotics are produced by single species of bacteria. Those from fungi and algae are nonprotein secondary metabolites and several microorganisms may make the same or similar toxin/antibiotics.


Phylogenetic affinities between eukaryotic cells and a thermophilic mycoplasma. Thermoplasma acidophilum, a thermophilic mycoplasma, has several unusual features suggesting a possible relationship to eukaryotic cells. One feature is a histone-like protein that is associated with the DNA, condensing it into subunits similar to those in eukaryotic chromatin. A second feature is an association of cytoplasmic proteins that resembles eukaryotic actin and myosin. These two components are widely distributed in different groups of eukaryotic cells, but are typically lacking in prokaryotic cells. Furthermore, T. acidophilum lacks cytochromes and respires by enzymes that apparently are not coupled to oxidative phosphorylation. This primitive type of respiration resembles that of microbodies, another feature which is represented in the cytoplasm of all groups of eukaryotic cells. Furthermore, since T. acidophilum lacks a cell wall and appears to have a primitive correlate of endocytosis, it would appear to be mechanically capable of acquiring a symbiotic mitochondrion. Thus, our observations are consistent with the symbiotic hypothesis for the origin of eukaryotic cells. We suggest that an organism similar to T. acidophilum was the host cell for the original symbiosis, becoming the nucleus and cytoplasm of modern eukaryotic cells.


Some advantages and limitations of mitosis as a phylogenetic criterion. The advantages and disadvantages of mitosis as a phylogenetic character are discussed. Mitosis is suggested to be a good character on the grounds that it is universally applicable and that it reflects features of the basic organism since it is unlikely to have arisen through multiple endosymbiotic events. Since mitosis is an absolutely essential feature of the cell cycle it is subject to a great deal of evolutionary pressure and one would not expect a priori that relatively inefficient mitotic apparatuses would persist. It is suggested that supposedly primitive mitotic apparatuses are not inefficient and that the evidence that they are functionally different from those of higher organisms is largely negative electron microscopic evidence. This type of evidence is not very satisfactory since it can result from poor preservation and consequently it should be continually retested. Another weakness of mitosis as a phylogenetic criterion (shared with other essentially morphological criteria) is that there is no easy way to relate structural change with genetic change. Thus, while mitosis is a useful phylogenetic character, its usefulness is limited.


[Effect of electrical stimulation of the inferior central raphe nucleus on spontaneous and induced thalamic neuron activity in awake cats]. Using microelectrode technique in chronic experiments it was shown that electrical stimulation of the inferior raphe nucleus inhibited spontaneous activity of the thalamic neurons and that evoked by stimulation of radial nerves. Changes in the activity of the same neurons were absent in case of stimulation of the bulbar reticular formation.


[Spread of excitation from the ventromedial hypothalamus to the limbic-reticular structure of the brain]. The sequence of origination of the evoked potentials in different regions of the septum amygdale and the reticular formation in response to the gradually increasing stimulation of the hypothalamic ventromedial nucleus was studied. As demonstrated, excitation that initially occurs in the hypothalamic ventromedial nuclei embraced first the structures of the septum and rostral reticular formation and then more caudal region of the reticular formation and amygdala.


[Effect of stimulating the medial and lateral zones of the respiratory center on the electrical activity of the diaphragm, intercostal muscles and phrenic neurons]. Experiments on cats showed that the nucleus of the solitary tract displayed zones whose stimulation provoked separately stimulation or inhibition of the electrical activity of the phrenic neurons and the diaphragm. Stimulation in the nucleus ambiguus of such zones caused stimulation and inhibition of electrical activity of the intercostal inspiratory muscles. In stimulation of the corresponding zone in the giant cell nucleus the electrical activity of both groups of the inspiratory muscles proved to change. It is suggested that the action of stimulation of the giant cell nucleus zones on both groups of inspiratory muscles is mediated through the neurons of the solitary tract and the nucleus ambiguus.


[Extravagal respiratory reflex from pulmonary nociceptors]. A restricted damage of the lung parenchyma before vagotomy evokes an increase of inspiratory discharges of the diaphragm and an increase of the rate of respiration. Similar damage in vagotomized animals evokes only an increase of inspiratory discharges. The rate of respiration does not alter at all or changes insignificantly. An increase of the depth of respiration evoked by the lung nociceptors innervated by the extravagal afferents is of great importance because the lung ventilation is best provided by deep respiration. Probably not only the afferent fibers of the vagus nerves, but also the extravagal lung afferents must be taken into consideration in analysis of the relationship between the rate and depth of lung ventilation, particularly under pathological conditions.


[Optimal parameters for electrical auricular defibrillation]. In acute experiments on dogs the chest was opened and auricular flutter and fibrillation were induced. Current, energy and charge thresholds which eliminated this arrhythmia by the monopolar direct impulse of 1--15 ms duration were determined. The current and energy values that evoked the auricular defibrillation during direct application of the electrodes to them were minimal when the impulses of 8 ms were used: 113 +/- 13.7 mA and 10.4 +/- 2.6 mWs in elimination of the flutter, and 275 +/- 18.2 mA and 62.3 +/- +/- 9.0 mWsec in elimination of the fibrillation. The current that restored the nomotopic rhythm during direct auricular defibrillation was 50 times less than that determined during the transthoracic auricular defibrillation in dogs. The efficacy of direct auricular defibrillation indicated the necessity of the elaboration of an adequate method for its application in clinical practice.


[Properdin in the lymph in traumatic stress]. The changes of properdin level in different kinds of lymph and blood of dogs in case of burn of hind paws (the 1st group), and of craniocerebral injury (the 2nd group) were investigated. The properdin level proved to increase in the lymph of the cervical duct and the afferent lymph of the popliteal node in both the groups of dogs. The properdin level in the afferent lymph of the popliteal lymph node, thoracic duct and in blood was reduced in case of the burn and rose in cranicerebral injury. The total protein level in the afferent and efferent lymph of the popliteal lymph node, the cervical and thoracic ducts rose, whereas the total blood protein level fell.


[Effect of estrogens on reconstructed nucleohistones]. The action of estron, 17beta-estradiol and estriol on the extent of histone binding with DNA in the reconstituted nucleohistones was studied; also the influence of these hormones on the thermal denaturation of nucleoproteins was investigated. As revealed, estrogens in a concentration of 10(-5) M failed to alter the protein/DNA weight ratio in the patially reconstituted nucleohistones F1, F2a, F2b, F3 and in the sum total nucleohistone reconstituted from the DNA and the equimolar complex of histones F2a, F2b, F3. Treatment with estrogens in the mentioned concentration did not alter the melting point of all the nucleohistones under study. The influence of estrogens on the thermal denaturation of the sum total nucleohistone and of the partially reconstituted nucleohistones F2a, F2b, and F3 consisted of the enhancement of hyperchromic effect of the first melting phase and a corresponding reduction of hyperchromicity of the rest of the phases. Estrogens failed to alter the heat denaturation profile of the partially reconstructed nucleohistone F1.


[Incorporation of 14-c-1-glycine into tissue proteins under the influence of insulin and its derivatives]. Effect of parenterally injected insulin and its composite sulpho-A- and B-chains on the rate of protein synthesis by the organs of the rat was investigated. It has been shown that insulin accelerates the incorporation of I-C14-glycine into proteins of the liver, kidneys, pancreas, spleen, sceletal muscules, thyroid gland, thymus and adrenal glands but does not influence this process in the cardiac muscle and diaphragm. A- and B-chains also activated protein metabolism in some organs, but despite some specificity of their action its extent was more limited as compared to that of insulin.


[Effect of the cytotoxin of Central Asia cobra venom on the functional state of rat liver mitochondria]. A study was made of the action of various concentrations of cytotoxin (C), free of phospholipase A admixtures, on the respiration, oxidative phosphorylation of the rat liver mitochondria and their "ghosts", and also on the permeability of their internal membranes for various ions. Low C concentrations (10--25 micrometer) failed to influence the functional parameters of the intact mitochondria, but sharply increased the permeability of the internal membranes for monovalent ions. Apparently the uncoupling action of C on the mitochondrial "ghosts" was caused by the formation in the membrane of nonspecific conductivity "canals" with poor selectivity to the ion size and charge.


[Intensity of 14C-2-acetate incorporation into guinea pig brain and spinal cord phospholipids and cholesterol under normal conditions and following triorthocresylphosphate poisoning]. A severe form of chronic intoxication was caused in guinea pigs by a single intracutaneous inoculation of triorthocresyl phosphate (TOCP); the intensity of acetate-2(14)C incorporation into the brain stem and spinal cord phospholipids and cholesterol was investigated in vivo. In paralytic stage of the disease incorporation of 14C into phospholipids and cholesterol was found to be markedly depressed; the inhibition of the synthesis of these lipids was observed not only in the most vulnerable lumbo-sacral region of the spinal cord, but also in the brain stem. This finding is in favour of the systemic character of lipid metabolism disorder in the central nervous system and indicates alteration of oligodendrocyte metabolism caused by TOCP.


[Effect of neuroleptics on acetylcholine metabolism in the basal ganglia of the brain]. Neuroleptics (haloperidol) closapine, pimozid, chlorpromazine) diminished the level of free (functionally active) form of acetylcholine (ACh), and, to some extent, the bound form of ACh; they changed the content of the labile-bound (vesicular) form of ACh and weakly influenced the choline-acetyltranspherase activity in the basal ganglia of the rat brain 5 to 30 min after the injection. In contrast to the inhibitory action on the acetyl-cholinesterase (AChE) activity in vitro, most of the neuroleptics, except closapine, increased the AChE activity in vivo. These results indicate that the neuroleptics activate ACh-metabolism and probably stimulate the cholinergic structure in the basal ganglia of the brain; the AChE activity may serve as a criterion of such stimulating action of neuroleptics.


[Effect of the central cholinolytic cyclosil on several functional characteristics of rat liver mitochondria]. The effectiveness of macroergic compounds formation was found to increase in the mitochondria of the rat liver incubated in the presence of cyclosil (0.04 mg/ml). The rate of O2 consumption was unchanged. A tendency to increase the K+ transport into mitochondria in the presence of cyclosil was noted. The data obtained testify to an increase of the mitochondrial energization in the presence of cyclosil.


[Effect of verapamil on cerebral circulation]. Acute experiments were conducted on anesthetized cats; verapamil in a dose of 0.05, 0.1, 0.5, and 1 mg/kg, injected intravenously, caused a reduction of the tone of the cerebral vessels and of the general arterial pressure. Changes in the volume velocity of the cerebral circulation were phasic in character and depended on the tone of the cerebral vessels and the arterial pressure level. The circulation increased more in the cerebral cortex than in the reticular formation; PO2 correlated with the circulation in the cerebral cortex. The diameter of the pial arteries became greater both in case of the intravascular verapramil administration and its local application. The block of beta-adrenoceptors, M-cholinoreceptors, histamine receptors and preliminary administration of the sympatholytic failed to eliminate the vasodilatory action of verapamil. It is supposed that vasodilation was myotropic in character, and was possible connected with the electrolyte metabolism.


[Characteristics of the estrogen receptor system of the guinea pig uterus]. Physico-chemical parameters of estradiol-receptor interaction in guinea pig uterine cytosol (velocity constants of association and dissociation, half-life time of estradiol-receptor complex, and the value of free energy change) were studied. The number of receptor sites per cell was calculated. It was shown that under conditions of different degree of approximation to the equilibrium, the affinity percentage in the system under study remained unchanged for the majority of steroids, this pointing to the achievement of equilibrium in these cases. The steroid affinity to the analyzed R-system depended on presence of intact 3- and 17beta-hydroxils with somewhat greater significance of 3-(phenolic) hydroxil of steroid molecule.


[Vital microscopy and histochemical study of the autonomic nervous system of the small intestine in animals with thermal burns]. By means of vital microscopy a possibility of histochemical detection of redox enzymes and of study of changes in their activity in intramural nerve plexuses of the small intestine on a live animal was demonstrated. Severe thermal trauma induced a disturbance of permeability of mitochondrial membranes of the Auerbach's plexus nerve cells; as a result, there was a change in the activity of redox enzymes localized within the mitochondria. Lumbar novocain block performed soon after the burn trauma normalized the state of the mitochondrial membranes of the nerve cells.


[Activation of rat heart atrial and auricular cells after left ventricular infarct]. The number of the DNA-synthesizing nuclei in the connective tissue cells of the auricle and the auricula atril rose from the 1st to the 15th day after the experimental induction of left ventricular infarction in rats. In the myocytes of the auricula atril the amount of the labeled nuclei peaked on the 5th day, and that of mitoses--on the 7th day. In myocytes of the auricle the number of the nuclei incorporating 3H-thymidine was insignificant and only occasional abnormal mitoses were observed. In the rat hearts with healed infarctions the amount of polyploid nuclei was increased both in the myocytes of the auricle and of the auricula atrii.


[Morphologic and biochemical correlations in the muscle tissue of the atria and ventricles]. Electron microscope study of the heart muscle tissue demonstrated a difference between the cardiomyocytes of the auricles and ventricles. Three varieties of cells distinguished by the character of relations between the organoids and the number of specific secretory granules in them were revealed in the muscle tissue of the auricles. In studying the activity distribution of the lactate dehydrogenase (LDH) and its isoenzymatic pattern by disc electrophoresis and isoelectric focusing, the enzymatic spectra of LDH from the auricles and ventricles were found to differ considerably. It is suggested that the sources of energy and the mechanisms of energy production in the muscle cells of the auricles and ventricles are different.


[Ultrastructure of epithelial cells and cell contacts in the urinary bladder of frogs treated with aminazine]. Chlorpromazine induced swelling and vacuolization of cells of the frog urinary bladder mucosa and eliminated their capacity to react to antidiuretic hormone by increased water permeability. The structure of the tight junction and desmosoma is unchanged, this possibly determining the water-tightness of the epithelium treated with chlorpromazine.


[Ultrastructural changes in the posterior lobe of the pituitary in tetanus poisoning]. Ultrastructural studies (electron microscopy) of the caudal part of the hypothalamo-hypophysial neurosecretory system were conducted in rats during experimental tetanus. Its activation was found to occur as soon as the initial period after the injection of the toxin, this being apparently connected with the administration of the heterogenous protein, because inactivated toxin proved to induce the same effect. Changes in the vascular permeability, lipid metabolism and the blood coagulation system were also observed.


[Ultrastructural and functional features of the reversibility of sclerotic liver changes in rats treated with exogenous organ specific RNA]. Ultrastructural peculiarities of hepatocytes and Kupffer's cells in rats during the spontaneous reversibility of the experimental cirrhosis of the liver under the usual conditions and also under the influence of exogenous RNA, administered by different methods, were studied. The changes in cell ultrastructure proved to correlate with those of the quantitative indices of the protein synthesizing the liver function during the hepatic cirrhosis reversibility. A positive role of the RNA influence of short duration on the intracellular reparative hepatocytes regeneration, and a negative one in case of RNA influence of long duration was revealed. Participation in the resorption of collagen both of the parenchymal cells and of the stroma was shown.


[Characteristics of the blood system of inbred and mongrel mice]. Peripheral blood and bone marrow indices were studied in inbred (101/H, CC57W, A2G, CBA/Lac, B6WV, C57BL/6, ACR) and mongrel mice. Normal values of the morphological composition of the blood and bone marrow were established. Comparative analysis demonstrated the existence of differences in the mentioned indices not only between the inbred and mongrel mice, but also within the individual animal strains.


[Effect of ligation of the pancreatic ducts on endogenous serotonin distribution in the organs of the gastrointestinal tract]. The content of endogenous serotonin in the mucosa of various parts of the stomach and the small intestine and also in the homogenates of the pancreatic tissue and the peripheral blood was traced in the course of 12 months after the ligation of the pancreatic ducts in dogs. Marked and prolonged disturbances were revealed in the deposition of the endogenous serotonin characterized by definite regularities in all the tissues under study, connected with the changes of the relative constancy of the enteral medium due to the absence of pancreatic secretion in the lumen of the digestive tube.


[External respiration and gas metabolism in orthostatic syncopes]. Cardiorespiratory reactions to tilt tests were compared in 80 healthy male subjects with an adequate orthostatic tolerance and in 19 subjects who fainted during tilting. They showed significant differences in the gas exchange, hemodynamics, and external respiration. Variations in the heart rate, pulmonary ventilation and the alveolar CO2 tension were most demonstrative. The findings, particularly the lack of the expected decrease o= oxygen consumption in the presyncopal state contribute to the concepts of the pathogenesis of the orthostatic collapse.


[Effect of different leukocyte pyrogen fractions on hemopoiesis]. Fractionation of leukocyte pyrogen on a column of Sephadex G-75 made it possible to obtain separately the fraction stimulating the hemopoiesis and the fraction possessing the pyrogenic activity and inhibiting the hemopoiesis. Judging by the elution profile of Sephadex column G-75, substances of high molecular weight produced a stimulating action, and of low molecular weight--pyrogenic and inhibitory action. Possibly pyrogenic and inhibitory activities are connected with different substances. The nature of the inhibitory factor requires further investigation. It may be supposed that it is a substance of chalone type.


[Effect of cholesterol administration on the state of the adrenal cortex in severe stress]. The two weeks' effect of cholesterol on the rat adrenal cortex response under conditions of a severe stress was studied. In comparison with control, the degree of the gland activation and its alteration was found to decrease. It is suggested that the effect of cholesterol depends upon the action on the hypothalamus, and upon the adrenocortical tissue as well. The action of cholesterol on the adrenocortical tissue is bound with the cholesterol capacity to retard peroxydation reactions.


[Bioelectrical activity of the uterus of white rats with intrauterine streptococcal infection and exposed to the use of pulsing, local negative pressure with oxygen]. Intrauterine streptococcal infection of albino rats hematogenically during pregnancy (implantation, placentation) leads to different disorders of the bioelectrical activity of the uterus, which become normalized after the institution of pulsating local negative pressure with periodic oxygen supply.


[Effect of adrenalectomy on the gastric juice secretion evoked by food or histamine in dogs]. Comparison was made of the effect of total adrenalectomy on the gastric secretion in chronic experiments on dogs with the Pavlov's stomach and Basov's fistula. A decrease of the maximal secretion level of gastric juice was associated with the alteration of the organ hemodynamics. A tendency to reduction of the acid production in the stomach was revealed. Essential differences were noted in the character of proteolytic enzymes secretion with different agents stimulating the secretion. Specific nature of the gastric secretory system for each stimulant, and different effects of adrenalectomy on the secretion induced by these stimulants was shown.


[Ribonuclease activity of the "myofibrillar" fraction of normal and autolyzed muscle tissue]. The separation in a sucrose gradient of the "myofibrillar" fraction of normal and autolysed muscle tissue gave 4 components. During the post-mortem destruction of the tissue there was observed a slight decrease of the "myofibrillar" fraction yield and also certain changes in the distribution of protein between different components. Under the selected conditions the RNA-ase activity was found in all 4 components. During the course of autolysis the enzymatic activity increased in the whole "myofibrillar" fraction, as well as in the lysosomal-mitochondrial components of "myofibrillae".


[Optical properties of tissues and organs in relation to laser rays with a wave length of 632.8 and 488 nm]. The authors measure the coefficient of attenuation of the equivalent depth of penetration in tissues and organs of rats of helium-neon and argon lasers with the generation wave length of 632.8 and 488 nm, respectively. Blood and organs with numerous capillaries proved to have the greatest capacity to absorb laser ray energy. With the reduction of the generation wave length there occurred a sharp partial elevation of the absorptive capacity of the blood. This permits to expect a greater blood photoreactivity to laser rays generating in the short wave part of the visible wave length range.


[Effect of thyroxine on radioactive iodine accumulation in the thyroid glands of hemithyroidectomized rats of different ages]. Experiments were carried out on hemithyroidectomized male rats aged from 2 to 20 months. Thyroxin administration in a dose of 1 microgram per 100 g of body weight on the 5th day of the experiment led to significantly greater suppression of radioiodine accumulation in the lobe of the thyroid gland remaining after the operation in 16--20-month-old rats in comparison with 2--2 1/2-month-old animals. The data obtained confirmed a conclusion drawn earlier on the reduction with ageing of the hypothalamo-hypophysial threshold to the homeostatic thyroxin inhibition.


[Study of the chemical receptor for diphtheria toxin in cell membranes]. Adsorption of diphtheria toxin on immobilized cell membrane preparations in the presence of gangliosides was studied. A mixture of gangliosides completely inhibited the adsorption of diphtheria toxin on the cell membranes, both sensitive (HeLa cells, macrophages) and resistant (L-cells) to its action. Neuraminidase-treated gangliosides were less effective. Immobilized gangliosides adsorbed diphtheria toxin effectively in the presence of a protective colloid and detergent triton X-100. The results obtained suggest gangliosides to be diphtheria toxin receptors.


[Immunogenicity of H-antigen polymers obtained by different methods]. The authors studied the properties of two polymeres prepared from flagellin by various methods: precipitation with ammonium sulfate with the subsequent centrifugation or treatment with glutaric aldehyde in solution and subsequent gel-chromatography. Molecules of the former (P-I) and of the latter (P-II) were similar by shape, but the mol wt of P-I was greater. The preparations contained common H-antigen and were similar by serological activity. P-I had a very high "priming" activity for mice. P-II was highly immunogenic only within the narrow range of doses, was less immunogenic in comparison with P-II, and, in difference from P-I, induced formation of some amount of 7S-antibodies.


[Stimulation and suppression of contact dermatitis in mice with a low-molecular thymic humoral factor]. The authors studied the influence of the low molecular lymphocyte-stimulating substance (LSS) of the thymus on the development of contact sensitivity to picrylchloride in mice. As revealed, low LSS doses enhanced, and high doses suppressed this type of delayed hypersensitivity. Contact sensitivity was passively transfered by means of the lymph node and spleen cells isolated on the 6th day after the immunization. Experiments showed mice given high LSS doses to contain cells suppressing passive transfer of contact sensitivity by immune cells. This suppression was annulled after the treatment of cells with theta-antiserum and the complement. A conclusion was drawn that suppressor cells influenced the effector phase of contact sensitivity.


[Death of adhering spleen cells (macrophages) in vitro in delayed hypersensitivity to microbial antigens]. The cytotoxic action of immune lymphocytes on the adhering cells of the spleen obtained from guinea pigs sensitized and nonsensitized with BCG was studied in autologous and allogenous systems. A low cytotoxic effect revealed in cultivation with tuberculin of a suspension of spleen cells obtained from sensitized guinea pigs increased considerably after addition of lymph node cells from the same animal. Determination of death of the adhering cells of the spleen, as well as of the adhering cells of the lymph nodes could be used as a sensitive method for detection of hypersensitivity of a delayed type. Use of the splenic cells in the capacity of target cells is more convenient, since there are much more adhering cells in the spleen than in the suspension of lymph node cells.


[Mechanism of mammary cancer regression in lactating rats]. Dependence of the incidence of regression of the transplantable carcinoma of the mammary gland RMK-1 in albino rats on lactation conditions was studied. The tumour was subject to regression in 47% of the animals nursing 8 +/- 1 ratlings. Following ovariectomy and cortisone and oxytocine administration which produced mediated inhibitory effect of FSH secretion of the hypophysis the frequency of the tumour regression in rats nursing the same number of ratlings rose to 71--81%. In rats with prolonged lactation nursing 8 +/- 1 ratlings for 2--2.5 months and in rats with intensive lactation nursing 13 +/- 2 ratlings the tumour regression practically failed to occur. The data obtained confirmed the suggestion that along with high LTH of the hypophysis secretion of importance for regression of mammary carcinoma in the course of lactation was reduction of the secretion of FSH of the hypophysis.


[Release of the nucleoli and karyoplasm into the cytoplasm of reticular cells in lymph node cultures]. Successive stages of the process characterizing the release of the nucleoli and part of the nuclei contents into the reticular cell cytoplasm were recorded in the primary cultures of the lymph nodes of albino mice with transplantable leukemia. Comparison of the results obtained with the autoradiographic data pointing to the high proliferative and metabolic activity of the reticular cells permits a suggestion that the recorded phenomenon reflected the secretory function of the stroma cells of the hemopoietic organs at the morphological level.


[Plasmatic membrane protein synthesis in cells of the regenerating liver]. Protein synthesis in the cells of the regenerating rat liver was studied. The rate of 3H-glycine incorporation into the total proteins of the liver, those of microsomal fraction, proteins of hyaloplasm, and plasmatic membrane proteins, soluble and non-soluble in 0.05 M K2CO3, was determined. The rate of 3H-glycine incorporation into soluble proteines of plasma membranes became maximal one hour after partial hepatectomy. The peak of the rate of synthesis of proteins of other fractions fell on the end of the G1-period. A sharp increase of the synthesis rate of plasma membrane proteins seems to be one of the earliest biochemical events in the regenerating liver hepatocytes ready for division.


[Morphological and functional interrelationships of the thymus and adrenals in inbred mice]. The authors carried out morphological examination of the thymus, adrenal glands and the spleen in CBA, C57BL/6, AKR mice and studied the immunological reactivity of mice to sex antigen of the skin transplant with the determination of the blood plasma corticosterone level. In comparison with C57BL/6 and AKR mice, CBA mice displayed a narrow cortical layer in the thymus, wide thymus-dependent zones in the spleen, a high corticosterone level, and the absence of any reaction to the sex antigen of the skin transplant. Adrenalectomy in CBA mice led to a sharp dilatation of the thymus cortex, the appearance of numerous lymph follicles in the spleen and development of an immunological reaction to sex antigen of the skin transplant. Atrophy of the cortical layer of the adrenal glands witha sharp reduction of subanophilic lipids was noted in CBA B-mice.


[Origin of Kupffer macrophages in the regenerating liver]. Male Wistar rats were injected intraperitoneally with H3-thymidine (5 muC/g). Two thirds of the liver was removed one hour later. Sham operated animals served as control. A moderate elevation of the labeled nuclei indices of hepatocytes (LNI) was seen by the third hour, increased significantly by the ninth, and especially by the 24th hour after the operation. The indices decreased by the 48th hour after partial hepatectomy. In 3 hours the LNI was 10 times higher than in the control; in 9 hours it reached the maximum, decreasing by the 24th and 48th postoperative hours. The origin of Kupffer's macrophages in the regenerating liver is discussed.


[Characteristics of mitosis pathology in a Chinese hamster cell culture with disorders of the protein-synthesizing system]. Disturbance of protein synthesis with puromycin and transcription of chromosomal and ribosomal RNA with actinomycin D was followed by marked changes in the normal course of mitosis. There was noted an increase in number of colchicin-like mitoses (C-mitoses), and particularly, sometimes, fragmentation of their cytoplasm with the formation of cluster-like structures. It is suggested that development of C-mitosis was connected not only with destruction in the mitotic apparatus formation system, but also with the block of synthesis of one of the chromosomal proteins, stabilizing DNA strands spiralization. Another form of pathological mitosis occurring under conditions of suppression of the metabolic processes ("hollow metaphase plate") was linked not only with the chromosomal alterations. Selective suppression of the ribosomal RNA transcription led to an evident anaphase delay and to coupling of telomere regions of chromosomes. This phenomenon is apparently associated with destruction of the "protective cover" of chromosomes, formed by RNA of the disintegrating nucleoli and RNA of perichromatin granules.


[Electron microscopic autoradiography of the liver after corticosterone-3H administration]. Electron microscope autoradiography was applied to the study of the distribution of autographs corticosterone-3H in the cells of the liver against the background of adrenalectomy. Deficiency of endogenous glucocorticoids aided more intensive incorporation of corticosterone-3H into the cells of the liver, this being evidenced by the appearance of tracks over the hepatocyte nuclei and an increase of their number over various cytoplasmic formations.


[Microspectrophotometric study of the sialomucin content of the glomerular filter in the nephrotic syndrome]. A microspectrophotometric study of sialomucins in human glomerular filter of the kidney in some diseases was carried out. Sialomucins were detected in paraffine sections stained after Haim, and examined on scanning microspectrophotometer. Lipid nephrosis, secondary amyloidosis, membranous-proliferative glomerulonephritis with the nephrotic syndrome were accompanied by a significant decrease in the amount of sialic acid. No reduction of sialomucins occurred in subacute glomerulonephritis with the nephrotic syndrome. A connection between the selective proteinuria and a decrease of the amount of sialomucins in the glomerulus is suggested.


[Immunocytochemical detection of the nonfibrillar stage of amyloid formation in the mouse myocardium]. The indirect electron microscope immunoperoxidase method utilizing pure rabbit antibodies to protein of mouse amyloid fibrils was used for studying casein-induced amyloidosis in mice. At the early stages of amyloidosis there appeared deposits of fine granular material in the mouse myocardium. These deposits contained an antigen similar to the amyloid fibrils antigen, but the deposits had no fibrillar ultrastructure. The results testify to the presence of early nonfibrillar stage of amyloid formation.


[Cytospectrophotometric study of GABA-transaminase in the rat cerebellar cortex]. Modification of the histochemical method for detection of GABA-transaminase activity is suggested. Optimal concentrations of the substrates and cofactors are chosen on the basis of kinetic study of the enzymatic reaction in the cryostat sections of the rat cerebellar cortex by the quantitative microspectrophotometric method. The method is intended for the quantitative histochemical analysis of GABA-transaminase activity in the brain sections.


[Device for the quantitative analysis of sum total pulse activity]. The authors present a circuit and the work principle of a device analysing the sum total electrical activity of the nerves. The device works by the principle of a frequency meter, it is made of integral elements, has a simple scheme, and is linear.


[Electronic device for determining motor activity]. The authors present a short description and the principal scheme of an electron apparatus for the determination of the motor activity of laboratory animals. This apparatus is based on the principle of recording the changes in volume occurring during the movement of animals.


Inhibition of platelet aggregation by protease inhibitors. Possible involvement of proteases in platelet aggregation. The possible participation of proteases in human platelet aggregation was explored using various protease inhibitors and substrates. Protease inhibitors used included naturally occurring inhibitors of serine proteases and synthetic inhibitors that modify the active site of protease. Substrates used were synthetic substrates for the trypsin type as well as for the chymotrypsin type of protease. All these inhibitors and substrates inhibited platelet aggregation and serotonin release induced by ADP, collagen, epinephrine, or thrombin. In ADP- and epinephrine-induced platelet aggregation the second phase of aggregation was most efficiently inhibited. The inhibitors suppressed the formation of malondialdehyde during platelet aggregation. Release by aggregating agents of arachidonate and its metabolites from indomethacin-treated platelets as well as nontreated platelets was also inhibited. The inhibitors apperar to interact with stimulated platelets but not with unstimulated platelets. These observations suggest that the interaction of an aggregating agent with its platelet receptor activates a unique precursor serine protease that in turn activates platelet phospholipase to liberate arachidonic acid (the precursor of the potent platelet aggregating agent thromboxane A2) from platelet phospholipids.


Intermediate syndrome of platelet dysfunction. We studied eight patients with intermittent bleeding episodes usually following trauma and associated with the ingestion of medicine known to interfere with platelet function. All patients had a normal or minimally prolonged baseline bleeding time. All had a normal platelet count, glass bead retention test, and platelet serotonin content and a variable pattern of abnormalities in prothrombin consumption and platelet factor 3 availability. However, all showed abnormal platelet aggregation reactions using epinephrine, adenosine diphosphate, and collagen. Following the administration of 975 mg aspirin, our patients' bleeding times became prolonged to a greater extent than the bleeding times of normal controls (range 13 to greater than 20 min). Review of the literature showed approximately 5% of "normal" controls had findings similar to those we report. We believe we are describing a group of individuals with an intermediate form of platelet dysfunction. Although their bleeding diathesis is not as severe as that of patients with platelet dysfunction syndromes previoulsy described, they do bleed significantly when subjected to trauma following the ingestion of drugs such as aspirin. We propose that this defect is common and should be screened for. The aspirin tolerance test is a simple test for detecting these patients.


Production of eosinophil-rich guinea pig peritoneal exudates. A variety of methods for production of eosinophil-rich guinea pig peritoneal exudates were tested in order to procure large numbers of purified eosinophils. Repeated peritoneal lavage with saline did not produce peritoneal eosinophilia, nor did intraperitoneal injection of dextran beads. Mast cell degranulating agents polymyxin B and, to a lesser degree, compound 48/80 produced significant peritoneal eosinophilia after 6-8 wk of weekly intraperitoneal injections (p less than or equal to 0.05). After more than 8 wk of treatment a mean of 52.1 x 10(6) eosinophils comprising an average of 62% of the peritoneal exudate cells was recovered from polymyxin B-treated guinea pigs. However, polymyxin B-induced peritoneal eosinophilia was abolished by pretreatment with the antihistamine diphenhydramine. Metiamide, an H2 receptor antagonist, did not block the production of peritoneal eosinophils. This is a highly satisfactory method for preparing peritoneal eosinophils; however, the mechanism of action remains to be clearly elucidated.


Physiologic features of hemolysis axxociated with altered cation and 2,3-diphosphoglycerate content. A hemolytic disorder characterized by altered RBC cation composition (increases Na, decreases K), reduced monovalent cation content (decreased Na + K/liter RBC), and decreased levels of 2,3-diphosphoglycerate (2,3-DPG) is described. The etiology of these RBC abnormalities was not elucidated following extensive laboratory evaluation, although two important physiologic principles were manifested by this case: (1) Hemolysis was relatively well compensated (41% hematocrit) despite a significantly decreased RBC survival (51 Cr t 1/2 = 10.5 days). This effect presumably was due to reduced 2,3-DPG content (1.9 mumol/ml RBC) and the associated increase in whole blood oxygen affinity (P50 = 19.6 mm hg). (2) RBC size and water content were normal in spite of marked cation depletion. This anomaly was thought to reflect the osmotic effects of reduced polyvalent anion (2,3-DPG) content.


Regulation of proliferation of murine megakaryocyte progenitor cells by cell cycle. The extent to which mouse megakaryocyte progenitor cells (colony-forming unit-megakaryocyte, CFU-M) can proliferate in semisolid cultures prior to endomitosis, and conditions that may regulate that differentiation step, were investigated. The proliferative capacity of CFU-M was determined by estimating the number of megakaryocytes per colony. A bimodal distribution was observed (modal values, 10-15 and 25-30 cells/colony), indicating that separate megakaryocyte progenitor cells may be biased in their capacity for proliferation versus endomitosis. Differences were observed in the cell cycle characteristics of CFU-M as determined in vivo and in vitro that suggest that maturation of CFU-M into megakaryocytes may be regulated within the marrow by control of the cell cycle of the megakaryocyte precursor cell.


Terminal stages of erythroid differentiation: persistent production of globin messenger RNA is necessary to sustain globin synthesis. The kinetic relationship between the globin mRNA accumulation and the rate of synthesis of globin chains was studied during the terminal stages of differentiation in erythroid cells derived from the yolk sac of mouse fetuses. RNA derived from the whole cells and from different cell compartments were hybridized to DNA complementary to embryonic globin mRNA. The relative proportion of embryonic globin RNA molecules and their absolute number per cell were estimated on the 11th, 12th, and 13th days of mouse fetal development. During erythroid terminal differentiation globin mRNA became progressively predominant on polyribosomes along with the progressive specialization of cell functions. The number of embryonic globin RNA molecules per cell remained constant while yolk sac erythroid cells underwent two rounds of cell division. These data indicate that the transcription of globin genes is operative throughout the last stages of terminal differentiation and that there is no detectable storage of globin RNA sequences in these cells. The rates of accumulation of mRNA molecules and of globin synthesis both seem correlated to the length of the cell cycle.


Red cell 2,3-diphosphoglycerate and oxygen affinity of hemoglobin in patients with thyroid disorders. We measured red blood cell 2,3-diphosphoglycerate (2,3-DPG), adenosine triphosphate (ATP), and the P50 value in vitro of the oxyhemoglobin dissociation curve, which is the oxygen tension at half saturation of hemoglobin, in order to quantitate red blood cell oxygen transport function in individuals who were diagnosed as hypothyroid, euthyroid, or hyperthyroid based on measurements of thyroxine (T4), triiodothyronine (T3), thyrotropin (TSH), and their clinical status. Hypothyroid (mean T4 2.8 microgram/dl, T3 49 ng/dl, TSH 37 microU/ml) and hyperthyroid (mean T4 14 microgram/dl, T3 271 ng/dl, TSH less than 0.7 microU/ml) patients had normal red cell 2,3-DPG and ATP levels and normal P50 values in vitro. The known changes in oxygen consumption produced by alterations in thyroid hormone levels in patients with hypothyroidism or hyperthyroidism did not affect red blood cell oxygen transport function.


Abnormalities of iron metabolism and erythropoiesis in vitamin E-deficient rabbits. Rabbits fed a vitamin E-deficient diet developed severe muscular dystrophy in 3-4 wk, but they did not become anemic. Nevertheless, reticulocyte counts increased in deficient rabbits (3.2%) compared to control rabbits (0.9%), and erythroid hyperplasia was evident in the bone marrow. Comparing deficient rabbits to controls, the plasma iron concentration was lower (134.4 versus 206.6 microgram/dl); the TIBC was higher (335.9 versus 228.3 microgram/dl); the whole blood protoporphyrin concentration was higher (131.6 versus 81.7 microgram/dl); and the total iron content was lower in spleen (71 versus 153 microgram), higher in skeletal muscle (4956 versus 3054 microgram), and unchanged in bone marrow, liver, and heart. Studies of iron absorption and excretion using 59Fe showed no abnormalities in deficient rabbits. There were abnormalities of ferrokinetics, however. The half-time of disappearance of 59Fe was shorter (100.6 versus 169.4 min), the plasma iron turnover was greater (1.25 versus 0.95 mg/dl blood/day), and the reappearance of 59Fe  in circulating erythrocytes at day 9 was greater (77.2% versus 57.2%) in deficient rabbits. Anemia induced by phlebotomy accentuated the abnormal iron metabolism of deficient rabbits, and the animals were unable to correct the anemia. These findings show that vitamin E deficiency in rabbits causes abnormal erythropoiesis associated with abnormal iron metabolism and sequestration of iron in skeletal muscle.


Erythropoiesis-stimulating factor production by rabbit kidney cultures from "programmed" rabbits. Rabbit kidney tissue either from normal rabbits or from rabbits "programmed" by phlebotomy or varying periods of hypoxia was cultured and shown to release into the culture medium an erythropoietically active substance. The cultures established from rabbits programmed by 6-12 hr of hypoxia yielded the highest levels of erythropoiesis-stimulating activity.


[Effect of heparin and ASA on changes of haemostasis induced by venous occlusion (author's transl)]. The influence of venous occlusion on plasmatic coagulation, on platelets, and on fibrinolysis was examined. After occlusion, activated factors XI and X could be demonstrated. Simultaneously, platelet aggregation induced by both collagen and epinephrine was increased. Fibrinolysis was found to be moderately enhanced. In patients taking acetylsalicylic acid (ASA), platelet functions were not altered by occlusion but the activation of plasmatic clotting factors was not influenced. Low dose heparin inhibited plasmatic activation but had no influence on the increase of platelet activities. By simultaneous administration of both substances, an additive effect was observed resulting in inhibition of plasmatic and platelet activation due to venous occlusion.


[Inhibition of the carrageenen-induced inflammatory rat paw oedema by systemic application of rat fibrinogen and fibrinopeptides (author's transl)]. The acute inflammatory rat paw oedema induced by subplantar injection of carrageenan was significantly inhibited by intraperitoneal injection of rat fibrinogen, intraperitoneal and intracardial injection of fibrinoepetides A and B. In contrast the fibrin split products derived from plasmin digestion proved to be ineffective.


Prognostic aspects of aplastic anemia in pregnancy. Experience on six cases and review of the literature. Our resent experience on six cases of aplastic anemia complicated with pregnancy is described. In addition, 43 similar cases were collected from the literature and reviewed to analyze some prognostic aspects of this relatively rare but potentially serious complication. Clinical and hematological data were treated to extract some clinically meaningful factors in relation to the success and failure of pregnancy. Among initial hematological parameters, no significant difference was found between successful and unsuccessful cases with an exception of hemoglobin concentration. The patients diagnosed as aplastic anemia prior to conception demonstrated an better outcome of pregnancy as well as survival rate of mother when compared with those diagnosed during pregnancy. Mortality has apparently improved after the late 1950's. Success rate of pregnancy before 1958 was 21%, while it was 67% and 71% in the era of 1959-1969 and after 1970, respectively. However, hemorrhage and infection remained to be two major causes of maternal death in both eras. Based on these observations, the currently recommendable attitude to this complication is discussed.