DNA-hydrolyzing IgG antibodies from the blood of patients with acquired immune deficiency syndrome

DNAase activity of 110 samples of IgG from the blood of AIDS patients was analyzed. It was shown that the relative activity of preparations varies very much from patient to patient, but 96% preparations show detectable level of DNAase activity. Several rigid criteria were applied and it was shown that DNAase activity is an intrinsic property of antibodies from AIDS patients. It was shown that catalytic activity could posses not only intact IgG, but also separated light chains of polyclonal antibodies. The abzymes catalyze DNA hydrolysis effectively in a wild range of pH (5.0-9.5). K(M) and V(MaKC) values of antibody-dependent hydrolysis of DNA was estimated.     

Limited plasticity of the rabbit visual cortex and hippocampal neurons in the oddball test

The activity of 41 visual cortex and 20 hippocampal neurons from field CA1 was registered in experiments using oddball-stimulation with different color stimuli varied in intensity. 34% cortical and 37% hippocampal neurons demonstrated plasticity reactions. The significant increase of latest phases of neuronal activity (200-500 and 200-1000 ms after stimulation for cortical neurons and 300-550 ms for hippocampal neurons) was shown in responses to rare deviant stimuli, which had a less intensity than frequently standards. The quantity of the earliest neuronal phase of activity (40-120 ms after stimulation) was stabilized in responses to deviants and standards during the experiment. We propose that such increase of the latest phases of neuronal activity (the limited plasticity) may reflect the mechanisms of orienting reaction.     

Role of lipolytic enzymes in the development of hyperlipidemia induced by administration of heterologous albumin in rabbits

Experiments on rabbits have shown hyperlipidemia to develop within the first 48 h after a single intravenous injection of bovine serum albumin (BSA, fraction V). The mean concentration of blood plasma triglycerides (TG) was considerably higher than normal (by 262% after 24 and by 625% after 48 h). The cholesterol content was also elevated (by 80 and 270%, respectively). Following 7 and 14 days the lipid concentration returned to normal. The plasma post-heparin lipoprotein lipase activity (PHP-LPL) was lower 24 h and 7 days after BSA injection and the hypotriglyceridemic effect of heparin was less pronounced. The data obtained support the hypothesis that hyperlipidemia provoked by a single intravenous injection of BSA to rabbits results from low PHP-LPL activity and possible changes in TG-rich lipoprotein substrate affinity for the enzyme.     

Monaural phasic sensitivity of auditory system neurons in the cat

During monaural stimulation, studies have been made of impulse reactions in neurons from the cochlear nucleus and inferior colluculus to changes in the phase value (psi) of one of the harmonics in two-tone signals. It was shown that about 70% of the neurons from the cochleaur nucleus with a tonic discharge pattern may sharply change their activity (from maximum one up to a complete inhibition) due to changes in psi value. Unlike the cochlear nucleus units, neurons from the inferior colliculus change their tonic or burst-like activity rather seldomly and to a slight extent. At the same time, their phasic on- and especially off-responses exhibit high sensitivity to changes in psi value.     

Neuronal activity in the amygdaloid structure during the sleep-wake cycle of the rat

The pattern of neuronal spike activity in the amygdaloid structure was studied in the sleep-wake cycle during experiments on unrestrained rats. It was shown that most neurons of the dorsomedial portion of the amygdala display greater spike activity during active wakefulness (80%) and paradoxical sleep (66.7%) than during slow-wave sleep. Most neurons of the basolateral amygdaloid region discharged at high frequency during active wakefulness (84.6%) and during paradoxial sleep (38.4%) compared with the frequency of firing during slow-wave sleep. Some neurons were found whose rate of discharge rose during slow-wave sleep in comparison with a similar period of paradoxical sleep (38.4%) and of active wakefulness (7.7%). Our findings show how the pattern of neuronal activity in the dosromedial and basolateral regions of the amygdaloid structure differs at various stages of the sleep-wake cycle. It is postulated that this structure serves mainly to regulate emotionally motivated processes rather than helping to govern the basic mechanisms of the sleep-wake cycle.Beritashvili Institute of Physiology, Academy of Sciences of the Georgian SSR, Tbilisi. Translated from Neirofiziologiya, Vol. 17, No. 6, pp. 747–756, November–December, 1985.     

Effects of Rodiola extract on the cytotoxic activity of natural killer cells of the liver, spleen, lungs and small intestine in rats after partial hepatectomy

In experiments on Wistar rats the cytotoxic activity of NK on the 1, 2, 10 days after partial hepatectomy (PH) and the application of Rodiola extract (RE) was studied. After 5 injections of RE the NK activity in gut increased by 112%, and after 12 ones (towards the end of experiment) by 222% in the spleen. The decreasing of this index in a first day after PH in lung, liver and gut was shown to restore in these tissues to the end of experiment. The absence of NK cytotoxicity diminishing just after PH in all the tissues was shown in operated animals, receiving RE and the decreasing of this index was found only in the lungs (by 335%).     

Genetic heterogeneity at the glucose-6-phosphate dehydrogenase locus in southern Italy: a study on the population of Naples

Summary Glucose-6-phosphate dehydrogenase (G6PD) electrophoretic phenotype was determined in red cells from 979 male subjects born in Naples (Southern Italy). In 0.7% of the cases no activity could be detected in haemolysates, while in 1.3% of the cases G6PD activity was approximately 20% of normal and electrophoretic mobility was altered. Moveover in two subjects a G6PD with altered mobility and normal activity was shown. G6PD was characterized in 10 subjects with variant phenotype. We conclude that the G6PD(-) phenotype in the population of Naples consists of at least six different G6PD variants associated with mild deficiency and at least one, G6PD Mediterranean, associated with severe deficiency.     

Identification of the cell wall receptor for bacteriophage E79 in Pseudomonas aeruginosa strain PAO.

Bacteriophage E79 was shown to interact with the lipopolysaccharide (LPS) of Pseudomonas aeruginosa strain PAO. LPS isolated from an E79-sensitive, smooth strain inactivated the phage, exhibiting a Phl50 value (concentration of LPS that caused a 50% decrease in the titer of phage during 1 h of incubation at 37 degrees C) of 0.04 microgram/ml, whereas the LPS isolated from a rough mutant derived from the wild type showed no neutralizing activity towards E79. EDTA and sodium deoxycholate were demonstrated to abolish the neutralizing capacity of the smooth LPS. One E79 receptor site was shown to be equivalent to 10(-16) g of LPS.     

On the mechanism of action of lead in the testis: in vitro suppression of FSH receptors, cyclic AMP and steroidogenesis

Previous evidence has shown that prenatal and neonatal exposure to low levels of Pb result in decreased FSH binding and steroidogenesis in the testes at the onset of puberty. The purpose of the present study was to determine by in vitro methods, if Pb acts by interfering directly with hormone binding, cyclic AMP production and steroidogenic enzyme activity. Sertoli cells were isolated from testes of prepubertal rats and cultured in the presence of 2.64 x 10(-4)M of either NaAc (control) or PbAc for 1, 4, 24, 48, 96 or 144 hr. There was no reduction in FSH binding and in FSH-induced cyclic AMP after a 1-4 hr exposure to Pb. After a 24-hr exposure to Pb, the cells exhibited a 10-20% decrease in FSH binding and cyclic AMP production and after 96 hr there was a 75% decrease in these 2 parameters. The inhibition was greater in cells from 16 day old than from 20 day old rats, so that in the former, after a 144 hr exposure the FSH-induced cyclic AMP of the Pb exposed cells was only 3% of the amount produced by the NaAc exposed cells (i.e. a 97% inhibition). After in vitro exposure to Pb for 48 hr, the steroidogenic activity (progesterone conversion to steroid metabolites) of Sertoli cells was significantly reduced and their steroidogenesis was no longer stimulated by FSH. A crude testicular enzyme preparation containing 3 beta-hydroxysteroid dehydrogenase (3 beta-HSD) exhibited approximately 25% reduction in activity if the assay buffer contained PbCl2 instead of the equivalent in NaCl. Prolonged in vivo exposure to Pb resulted in approximately 50% reduction in 3 beta-HSD activity. This is the first indication that in the testis Pb may act directly (immediate effect) by suppressing enzyme activities, and indirectly (long term effect) by reducing gonadotropin-receptor binding and the resultant cyclic AMP production.     

Alteration of the Amount of the Chloroplast Phosphate Translocator in Transgenic Tobacco Affects the Distribution of Assimilate between Starch and Sugar

Tobacco plants (Nicotiana tabacum L.) transformed with sense and antisense constructs of a cDNA encoding the tobacco phosphate-triose phosphate-3-phosphoglycerate translocator (phosphate translocator) were shown to contain altered amounts of phosphate translocator mRNA and protein. Phosphate translocator activity in intact chloroplasts isolated from transformed plants showed a 15-fold variation, from 20% of the wild-type activity in antisense transformants to 300% of the wild-type activity in sense transformants. However, the maximal rates of photosynthesis and the rates of photosynthetic carbon assimilation in ambient CO2 showed no consistent differences between transformants. Starch content was decreased by 20% and total soluble sugars were increased by 20% in leaves of antisense transformants compared to sense transformants. The 40% decrease in the ratio of starch to total soluble sugars in antisense transformants relative to sense transformants indicates that distribution of assimilate between starch and sugar had been altered. However, the amount of sucrose in the leaves was unchanged. The changes in total soluble sugars were accounted for completely by changes in glucose and fructose, suggesting the existence of a homeostatic mechanism for maintaining sucrose concentrations in the leaves at the expense of glucose and fructose.     

Effects of chick brain extract on the proliferation of chick neuroblasts cultured in media supplemented with low and high serum concentrations

The proliferative activity of chick neuroblasts cultured in a medium containing a low (5%) or a high (20%) concentration of fetal calf serum (FCS) was analyzed and the influence of a chick brain extract was investigated. Morphological observations and tritiated thymidine incorporation measurements have shown that neuroblasts from 6 day-old chick embryo cerebral hemispheres proliferate more actively in the medium with 5% FCS compared to the medium with 20% FCS. The medium containing 5% FCS favoured the maintenance of neuronal cells in a neuroblast stage as shown by electron microscopy. The stimulatory effect of brain extract on the proliferation of neuroblasts is stronger in the low serum culture condition. These findings indicate that a low serum-containing medium is an adequate condition to study neuronal proliferation and effects of growth factors on these cells.     

Effects of environmental factors on circadian activity in the flesh fly, Sarcophaga crassipalpis

Abstract.The diel locomotor activity patterns of wandering larvae in the flesh fly, Sarcophaga crassipalpis Macquart (Diptera: Sarcophagidae), were examined using a novel apparatus and shown to be primarily diurnal, but with a minority (37%) showing nocturnal activity. In response to the environmental stress of heat shock, a significantly larger proportion (72%) of the larvae became nocturnal. In comparison, adult circadian activity also was predominantly diurnal, but not correlated with the larval activity patterns. In addition, adult patterns showed age-related changes in entrainment and free running period. Finally, the phase of circadian-gated adult eclosion was shown to be entrained by a 3-day exposure to light–dark cycles delivered prior to pupariation, with the phase maintained throughout pupal–adult metamorphosis under constant dark conditions. These results demonstrate that environmental changes may have profound effects on the expression of 24-h activity patterns and circadian rhythms during different life stages throughout development.     

The effects of hypoxia and pH on phenoloxidase activity in the Atlantic blue crab, Callinectes sapidus

In its natural coastal and estuarine environments, the blue crab, Callinectes sapidus, often encounters hypoxia, accompanied by hypercapnia (increased CO2) and an associated decrease in water pH. Previous studies have shown that exposure to hypercapnic hypoxia (HH) impairs the crab's ability to remove culturable bacteria from its hemolymph. In the present study we demonstrate that the activity of phenoloxidase (PO), an enzyme critical to antibacterial immune defense in crustaceans, is decreased at the low levels of hemolymph O2 and pH that occur in the tissues of blue crabs exposed to HH. Hemocyte PO activity was measured at tissue O2 levels that occur in normoxic (5% and 15% O2, approximate venous and arterial hemolymph, respectively) and hypoxic (1% O2) crabs and compared to PO activity in air-saturated conditions (21% O2). PO activity decreased by 33%, 49% and 70% of activity in air at 15%, 5% and 1% O2, respectively. When O2 was held at 21% and pH lowered within physiological limits, PO activity decreased with pH, showing a 16% reduction at pH 7.0 as compared with a normoxic pH of 7.8. These results suggest that decreased PO activity at low tissue O2 and pH compromises the ability of crustaceans in HH to defend themselves against microbial pathogens.     

Increase of a calcium independent transglutaminase activity in the erythrocyte during the infection with Plasmodium falciparum

We have studied the activity of a calcium dependent transglutaminase (EC 2.3.2.13) during the growth of the parasite Plasmodium falciparum inside the infected human erythrocyte. There is only one detectable transglutaminase in the two-cell-system, and its origin is erythrocytic. No activity was detected in preparations of the parasite devoid of erythrocyte cytoplasm. The Michaelis Menten constants (Km) of the enzyme for the substrates N'N' dimethylcaseine and putrescine were undistinguishable whether the cell extracts used in their determination were obtained from normal or from infected red cells. The total activity of transglutaminase in stringently synchronized cultures, measured at 0.5 mM Ca2+, decreased with the maturation of the parasite. However, a fraction which became irreversibly activated and independent of calcium concentration was detected. The proportion of this fraction grew with maturation; it represented only 20% of the activity in 20 hr-old-trophozoites while in 48-hr-schizonts it was more than 85% of the total activity. The activation of this fraction of transglutaminase did not depend on an increase in the erythrocyte cytoplasmic calcium, since most of the calcium was shown to be located in the parasite.     

Evidence for the presence of non-lipoprotein factors in diabetic serum capable of stimulating rat hepatic cholesterol-7 alpha-hydroxylase in vitro

Experimental diabetes in the rat has been shown to result in marked increases in bile acid pool and synthesis. In this study, mechanisms responsible for the increased bile acid synthesis was examined in rats made diabetic with streptozotocin. Our results indicate that a) in diabetic rats, hepatic cholesterol-7 alpha-hydroxylase activity is increased by 100%, b) this increased activity is not due to a higher stimulating activity of cell supernatant factors, but c) may be due to a non-lipoprotein factor(s) in diabetic serum capable of stimulating (by 100%) cholesterol-7 alpha-hydroxylase activity in control livers to the level noted in diabetic animals.     

Geographic distribution and genetics of killer phenotypes for the yeast Pichia kluyveri across the United States.

Representative strains (n = 61) of the yeast Pichia kluyveri from across the United States were studied for their ability to kill 71 other strains (representing 25 species) of yeast. This survey showed killing activity in 69% of the P. kluyveri strains tested. More extensive analysis of killer activity of 197 P. kluyveri strains against strains of five tester species showed comparable activity (67% of strains tested). This activity was shown to be equally variable within localities, within regions, and across the continent. The genetic basis of the variability was ascertained by tetrad analysis and is most likely due to alleles segregating at three epistatic loci. Evidence for the idea that killer toxins have a role in excluding other yeasts from particular habitats is discussed.     

Biosynthesis of ascaridole: iodide peroxidase-catalyzed synthesis of a monoterpene endoperoxide in soluble extracts of Chenopodium ambrosioides fruit

Ascaridole, an asymmetric monoterpene endoperoxide with anthelmintic properties, occurs as a major constituent (60-80%) in the volatile oil of American wormseed fruit (Chenopodium ambrosioides: Chenopodiaceae), and as a lesser component in the leaf pocket oil of the boldo tree (Peumus boldus: Monimiaceae). Determination of optical activity and chromatographic resolution of naturally occurring ascaridole, and several synthetic derivatives, showed that both wormseed and boldo produce ascaridole in racemic form. The biosynthesis of ascaridole from the conjugated, symmetrical diene alpha-terpinene (a major component of the oil from wormseed) was shown to be catalyzed by a soluble iodide peroxidase isolated from homogenates of C. ambrosioides fruit and leaves. The enzymatic synthesis of ascaridole was confirmed by capillary gas-liquid chromatography and mass spectrometry of the product, which was also shown to be racemic. Optimal enzymatic activity occurred at pH 4.0 in the presence of 2.5 mM H2O2 and 1 mM NaI. Soluble enzyme extracts were fractionated by gel filtration on both Sephacryl S-300 and Sephadex G-100, and were shown to consist of a high-molecular-weight peroxidase component (Mr greater than 1,000,000, 30% of total activity) and two other peroxidase species having apparent molecular weights of 62,000 and 45,000 (major component). Peroxidase activity was susceptible to proteolytic destruction only after periodate treatment, suggesting an association of the enzyme(s) with polysaccharide material. Ascaridole biosynthesis from alpha-terpinene was inhibited by cyanide, catalase, and reducing agents, but not by compounds that trap superoxide or quench singlet oxygen. A peroxide transfer reaction initiated by peroxidase-generated I+ is proposed for the conversion of alpha-terpinene to ascaridole.     

Geographic distribution and genetics of killer phenotypes for the yeast Pichia kluyveri across the United States.

Representative strains (n = 61) of the yeast Pichia kluyveri from across the United States were studied for their ability to kill 71 other strains (representing 25 species) of yeast. This survey showed killing activity in 69% of the P. kluyveri strains tested. More extensive analysis of killer activity of 197 P. kluyveri strains against strains of five tester species showed comparable activity (67% of strains tested). This activity was shown to be equally variable within localities, within regions, and across the continent. The genetic basis of the variability was ascertained by tetrad analysis and is most likely due to alleles segregating at three epistatic loci. Evidence for the idea that killer toxins have a role in excluding other yeasts from particular habitats is discussed.     

Partial purification and characterization of phosphatidic acid-specific phospholipase A(1) in porcine platelet membranes

We have shown previously that the phospholipase A (PLA) activity specific for phosphatidic acid (PA) in porcine platelet membranes is of the A(1) type (PA-PLA(1)) [J. Biol. Chem. 259 (1984) 5083]. In the present study, the PA-PLA(1) was solubilized in Triton X-100 from membranes pre-treated with 1 M NaCl, and purified 280-fold from platelet homogenates by sequential chromatography on blue-Toyopearl, red-Toyopearl, DEAE-Toyopearl, green-agarose, brown-agarose, polylysine-agarose, palmitoyl-CoA-agarose and blue-5PW columns. In the presence of 0.1% Triton X-100 in the assay mixture, the partially purified enzyme hydrolyzed the acyl group from the sn-1 position of PA independently of Ca(2+) and was highly specific for PA; phosphatidylcholine (PC), phosphatidylethanolamine (PE), phosphatidylserine (PS), and phosphatidylinositol (PI) were poor substrates. The enzyme exhibited lysophospholipase activity for l-acyl-lysoPA at 7% of the activity for PA hydrolysis but no lipase activity was observed for triacylglycerol (TG) and diacylglycerol (DG). At 0.025% Triton X-100, the enzyme exhibited the highest activity, and PA was the best substrate, but PE was also hydrolyzed substantially. The partially purified PA-PLA(1) in porcine platelet membranes was shown to be different from previously purified and cloned phospholipases and lipases by comparing the sensitivities to a reducing agent, a serine-esterase inhibitor, a PLA(2) inhibitor, a Ca(2+)-independent phospholipase A(2) inhibitor, and a DG lipase inhibitor.     

Peroxisomal participation in the cellular response to the oxidative stress of endotoxin

Exposure to a sublethal dose of endotoxin offers protection against subsequent oxidative stresses. The cellular mechanisms involved in generating this effect are not well understood. We evaluated the effect of endotoxin on antioxidant enzymes in liver peroxisomes. Peroxisomes have recently been shown to contain superoxide dismutase (SOD) and glutathione peroxidase (GPX) in addition to catalase. Peroxisomes were isolated from liver homogenates by differential and density gradient centrifugations. Endotoxin treatment increased the specific activity of SOD and GPX in peroxisomes to 208% and 175% of control activity, respectively. These findings correlated with increases in peroxisomal SOD and GPX proteins observed by immunoblot. Although the quantity of catalase protein was increased when assessed by immunoblot analysis, the specific activity of catalase was decreased to 68% of control activity. Activation of catalase with ethanol only restored catalase activity to control levels suggesting that catalase had undergone irreversible inactivation. The observed increase in GPX activity may represent a compensatory mechanism triggered by accumulating H₂O₂. The data presented here suggest for the first time that mammalian peroxisomal antioxidant enzymes are altered during the oxidative injury of endotoxin treatment.