Contents of aldosterone and electrolytes in blood plasma and the myocardium of rats after single physical exertion during acute alcoholic intoxication]

It was proved in experiments with male Wistar rats that acute alcohol intoxication caused significant changes in electrolytes balance in blood plasma and myocardium remaining for a long time after ethanol injection. SPL test makes it possible to reveal inadequate reaction of blood plasma aldosterone in alcohol injected rats. This fact may be considered as the fact of involving extracardiac component of compensation of myocardial functional insufficiency conditioned mainly by electrolyte unbalance. The tolerance to physical loading is significantly higher in alcohol injected rats than in intact animals.     

Principles of reciprocal changes in the cardiomyocyte and hepatocyte lysosomal enzyme activities in experimental diphtheria intoxication in rabbits]

The dynamics of two lysosomal enzymes' free activity was studied in cardiomyocytes, hepatocytes and blood serum during experimental diphtheria intoxication in rabbits. The principle of reciprocal changes in interrelations of the same and different enzymes in cardiomyocytes and hepatocytes was revealed.     

Method of determinating the protective capacity of C1. perfringens toxoid]

The authors describe a model of experimental gas gangrene in guinea pigs; it was produced by the administration of the vegetative form of C. perfringens; the cells were completely washed of the lethal toxin and no toxic or necrotizing agents were added. A possibility of development of gangrenous process without any preliminary depression of the resistance of body tissues in the area of injection of the causative agent was revealed. Apart from the local process and general intoxication gas gangrene, caused by intramuscular injection of C1. perfringens to guinea pigs, was accompanied by bacteriemia and microbial contamination of the internal organs. A method of the animal infection was ascertained and the causes of their death was assessed. The method is recommended for determination of the immunological efficacy of C1. perfringens toxoids.     

Lipid interaction of diphtheria toxin and mutants with altered fragment B. 2. Hydrophobic photolabelling and cell intoxication

The membrane insertion of diphtheria toxin and of its B chain mutants crm 45, crm 228 and crm 1001 has been followed by hydrophobic photolabelling with photoactivatable phosphatidylcholine analogues. It was found that diphtheria toxin binds to the lipid bilayer surface at neutral pH while at low pH both its A and B chains also interact with the hydrocarbon chains of phospholipids. The pH dependence of photolabelling of the two protomers is different: the pKa of fragment B is around 5.9 while that of fragment A is around 5.2. The latter value correlates with the pH of half-maximal intoxication of cells incubated with the toxin in acidic mediums. These results suggest that fragment B penetrates into the bilayer first and assists the insertion of fragment A and that the lipid insertion of fragment B is not the rate-controlling step in the process of membrane translocation of diphtheria toxin. crm 45 behaves as diphtheria toxin in the photolabelling assay but, nonetheless, it is found to be three orders of magnitude less toxic than diphtheria toxin on acid-treated cells, suggesting that the 12-kDa COOH-terminal segment of diphtheria toxin is important not only for its binding to the cell receptor but also for the membrane translocation of the toxin. It is suggested that crm 1001 is non-toxic because of a defect in its membrane translocation which occurs at a lower extent and at a lower pH than that of the native toxin; as a consequence crm 1001 may be unable to escape from the endosome lumen into the cytoplasm before the fusion of the endosome with lysosomes.     

Modulation of the intracellular stability and toxicity of diphtheria toxin through degradation by the N-end rule pathway.

The enzymatically active A-fragment of diphtheria toxin enters the cytosol of sensitive cells where it inhibits protein synthesis by inactivating elongation factor 2 (EF-2). We have constructed a number of diphtheria toxin mutants that are degraded by the N-end rule pathway in Vero cells, and that display a wide range of intracellular stabilities. The degradation could be inhibited by the proteasome inhibitor lactacystin, indicating that the proteasome is responsible for N-end rule-mediated degradation in mammalian cells. Previously, the N-end rule has been investigated by studying the co-translational degradation of intracellularly expressed beta-galactosidase. Our work shows that a mature protein entering the cytosol from the exterior can also be degraded by the N-end rule pathway with a similar, but not identical specificity to that previously found. We found a correlation between the intracellular stability of the mutants and their toxic effect on cells, thus demonstrating a novel manner of modulating the toxicity of a protein toxin. The data also indicate that the inactivation of EF-2 is the rate-limiting step in the intoxication process.     

Requirement of specific receptors for efficient translocation of diphtheria toxin A fragment across the plasma membrane

The role of specific receptors in the translocation of diphtheria toxin A fragment to the cytosol and for the insertion of the B fragment into the cell membrane was studied. To induce nonspecific binding to cells, toxin was either added at low pH, or biotinylated toxin was added at neutral pH to cells that had been treated with avidin. In both cases large amounts of diphtheria toxin became associated with the cells, but there was no increase in the toxic effect. There was also no increase in the amount of A fragment that was translocated to the cytosol, as estimated from protection against externally added Pronase E. In cells where specific binding was abolished by treatment with 12-O-tetradecanoyl-phorbol 13-acetate, trypsin, or 4,4'-diisothiocyano-2,2'-stilbenedisulfonic acid, unspecific binding did not induce intoxication or protection against protease. This was also the case in untreated L cells, which showed no specific binding of the toxin. When Vero cells with diphtheria toxin bound to specific receptors were exposed to low pH, the cells were permeabilized to K+, whereas this was not the case when the toxin was bound nonspecifically at low pH or via avidin-biotin. The data indicate that the cell-surface receptor for diphtheria toxin facilitates both insertion of the B fragment into the cell membrane and translocation of the A fragment to the cytosol.     

The diagnostic effectiveness of an immunoenzyme test system for determining diphtheria toxin antigens in the blood serum in a clinical laboratory trial

The data on the approbation of the diagnostic value of the enzyme immunoassay (EIA) system for the determination of diphtheria toxin in the blood sera of diphtheria patients and persons suspected for diphtheria are presented. The EIA system was prepared on the basis of F(ab)2 fractions of purified antidiphtheria antibodies. 240 serum samples from diphtheria and tonsillitis patients and from healthy persons were studied. Diphtheria toxin was determined in all patients with the toxic form of diphtheria and in 41.3% of patients with its localized forms. Blood was taken mainly of the first week of the disease. In healthy persons the results of EIA were negative. Thus, the trial of the assay system in a clinical laboratory showed its good diagnostic effectiveness. The use of this EIA system in medical practice is believed to be quite promising.     

Evidence that diphtheria toxin and modeccin enter the cytosol from different vesicular compartments

Inhibition of protein synthesis in Vero cells was measured at different periods of time after treatment with diphtheria toxin and the related plant toxin modeccin. Diphtheria toxin acted much more rapidly than modeccin. Cells were protected against both toxins with antiserum as well as with agents like NH4Cl, procaine, and the ionophores monensin, FCCP, and CCCP, which increase the pH of intracellular vesicles. Antiserum, which is supposed to inactivate toxin only at the cell surface, protected only when it was added within a short period of time after modeccin. Compounds that increase the pH of intracellular vesicles, protected even when added after 2 h, indicating that modeccin remains inside vesicles for a considerable period of time before it enters the cytosol. After addition of diphtheria toxin to the cells, compounds that increase the pH of intracellular vesicles protected only approximately to the same extent as antitoxin. This indicates that after endocytosis diphtheria toxin rapidly enters the cytosol. At 20 degrees C, the cells were more strongly protected against modeccin than against diphtheria toxin. The residual toxic effect of diphtheria toxin at 20 degrees C could be blocked with NH4Cl whereas this was not the case with modeccin. This indicates that at 20 degrees C the uptake of diphtheria toxin occurs by the normal route, whereas the uptake of modeccin occurs by a less efficient route than that dominating at 37 degrees C. The results indicate that after endocytosis diphtheria toxin rapidly enters the cytosol from early endosomes with low pH (receptosomes). Modeccin enters the cytosol much more slowly, possibly after fusion of the endocytic vesicles with another compartment.     

Different response of the knockout mice lacking b-series gangliosides against botulinum and tetanus toxins

We assessed the response in knockout mice lacking the b-series (G(D2), G(D1b), G(T1b) and G(Q1b)) gangliosides against Clostridium botulinum (types A, B and E) and tetani toxins. We found that botulinum toxins were fully toxic, while tetanus toxin was much less toxic in the knockout mice. Combining the present results with our previous finding that tetanus toxin and botulinum types A and B toxins showed essentially no toxic activity in the knockout mice lacking both the a-series and b-series gangliosides (complex gangliosides), we concluded that the b-series gangliosides is the major essential substance for tetanus toxin, while b-series gangliosides may be not the essential substance for botulinum toxins, at the initial step during the intoxication process in mouse.     

Action of cytoplasmic factors from the regenerating liver on cholesterol esterification in the blood serum of rats with tetrachloromethane liver damage

The effect of proliferation stimulating factors (PSF) obtained from regenerating liver on the cholesterol esterification in blood serum under deep tetrachloromethane intoxication has been studied in experiments with white rats. Three phases in dynamics of cholesterol content in blood serum have been picked out after a single tetrachloromethane injection. The content of total cholesterol and cholesterol esters reduces during the first phase (24 h), during the second phase (3-7th day) it increases and during the third phase (15th day) these values become normal. PSF injection does not produce any influence on the nature of blood cholesterol under deep tetrachloromethane intoxication, but stimulates the processes of cholesterol ester formation in blood serum.     

COMPLICATIONS OF GOLD THERAPY AND THEIR MANAGEMENT

Early recognition of manifestations of gold intoxication is important to the treatment of such complications. Proper dosage schedules should be followed and blood and urine frequently examined.Most toxic manifestations subside, but those which become worse or which do not subside on withdrawal of the gold should be treated with BAL (2, 3-Dimercaptopropanol).BAL has a toxicity of its own and is painful on injection. Since BAL combines with gold, the therapeutic effect of the metal may be lost after such treatment.The beneficial effects of methionine and methionine plus BAL in treatment of experimentally induced gold intoxication of animals suggests such combined therapy in the treatment of clinical complications of gold poisoning. A schedule of combined antidotes is outlined.     

Diphtheria toxin entry into cells is facilitated by low pH

At neutral pH, NH4Cl and chloroquine protected cells against diphtheria toxin. A brief exposure of the cells to low pH (4.5-5.5) at 37 degrees completely abolished this protection. When, to cells preincubated with diphtheria toxin and NH4Cl, neutralizing amounts of anti-diphtheria toxin were added before the pH was lowered, the toxic effect was considerably reduced, but it was not completely abolished. A much stronger toxic effect was seen when antibodies were added immediately after incubation at low pH. Upon a short incubation with diphtheria toxin at low pH, the rate of protein synthesis in the cells decreased much faster than when the normal pH was maintained. The data suggest that, at low pH, diphtheria toxin (or its A fragment) penetrates directly through the surface membrane of the cell. The possibility is discussed that, when the medium has a neutral pH, the entry of diphtheria toxin involves adsorptive endocytosis and reduction of the pH in the vesicles possibly by fusion with lysosomes. Low pH did not facilitate the entry of the closely related toxins abrin, ricin, and modeccin.     

New concepts in characterization of ischemically injured myocardium by MRI

New concepts regarding the assessment of ischemic myocardial injuries have been addressed in this Minireview using magnetic resonance imaging (MRI). MRI, with its different techniques, brings not only anatomic, but also physiologic, information on ischemic heart disease. It has the ability to measure identical parameters in preclinical and clinical studies. MRI techniques provide the ideal package for repeated and noninvasive assessment of myocardial anatomy, viability, perfusion, and function. MR contrast agents can be applied in a variety of ways to improve MRI sensitivity for detecting and assessing ischemically injured myocardium. With MR contrast agents protocol, it becomes possible to identify ischemic, acutely infarcted, and peri-infarcted myocardium in occlusive and reperfused infarctions. Necrosis specific and nonspecific extracellular contrast-enhanced MRI has been used to assess myocardial viability. Contrast-enhanced perfusion MRI can explore the disturbances in large (angiography) and small coronary arteries (myocardial perfusion) as the underlying cause of myocardial dysfunction. Perfusion MRI has been used to measure myocardial perfusion (ml/min/g) and to demonstrate the difference in transmural myocardial blood flow. Information on no-reflow phenomenon is derived from dynamic changes in regional signal intensity after bolus injection of MR contrast agents. Another development is the near future availability of blood pool MR contrast agents. These agents are able to assess microvascular permeability and integrity and are advantageous in MR angiography (MRA) due to their persistence in the blood. Noncontrast-enhanced MRI such as cine MRI at rest/stress, sodium MRI, and MR spectroscopy also have the potential to noninvasively assess myocardial viability in patients. Futuristic applications for MRI in the heart will focus on identifying coronary artery disease at an early stage and the beneficial effects of new therapeutic agents such as intra-arterial gene therapy. MR techniques will have great future in the drug discovery process and in testing the effects of drugs on myocardial biochemistry, physiology, and morphology. Molecular imaging is going to bloom in this decade.     

A decrease in the acute toxicity of ethanol produced by zinc sulfate

It was shown that zinc sulphate injection during the acute alcohol intoxication resulted in the decrease of anaesthetic and toxic effects of ethanol. The most effective dose was 15 mkg/kg i.p. The possible mechanisms are discussed.     

The cellular immune response in different forms of diphtheria in adults

In 109 adult diphtheria patients with different forms of the disease (toxic, subtoxic, localized) and carriers, the latter including 6 persons having had the localized form of diphtheria, 12 different subpopulations of T and B lymphocytes and neutrophils were studied by the methods of rosette formation with sheep, mouse and bovine red blood cells. The study showed that, starting from week 2 of the disease, an increase in the content of the subpopulations under study was registered in all forms of diphtheria; the disease, as well as in patients with the localized form of the disease accompanied by complications and in long-term carriers. In all forms of diphtheria a decrease in the content of neutrophil rosette-forming cells was shown. The marker tests for prognostication of the complicated course of the disease, viz. the elevated content of M-rosette-forming cell and T gamma-rosette-forming cells during the first week of then disease, were established.     

Identification of host cell factors required for intoxication through use of modified cholera toxin

We describe a novel labeling strategy to site-specifically attach fluorophores, biotin, and proteins to the C terminus of the A1 subunit (CTA1) of cholera toxin (CTx) in an otherwise correctly assembled and active CTx complex. Using a biotinylated N-linked glycosylation reporter peptide attached to CTA1, we provide direct evidence that ~12% of the internalized CTA1 pool reaches the ER. We also explored the sortase labeling method to attach the catalytic subunit of diphtheria toxin as a toxic warhead to CTA1, thus converting CTx into a cytolethal toxin. This new toxin conjugate enabled us to conduct a genetic screen in human cells, which identified ST3GAL5, SLC35A2, B3GALT4, UGCG, and ELF4 as genes essential for CTx intoxication. The first four encode proteins involved in the synthesis of gangliosides, which are known receptors for CTx. Identification and isolation of the ST3GAL5 and SLC35A2 mutant clonal cells uncover a previously unappreciated differential contribution of gangliosides to intoxication by CTx.     

Endogenous intoxication of the body in acute myocardial infarct during lymph stimulation]

It was stated experimentally in dogs that the elevation of the lymph toxicity was more expressed than that of the blood in acute myocardial infarction. The shortening of the half-life period of paramecia evidenced the above mentioned fact. The injection of the lymphogogue preparations (obsidan, heparin, rheogluman) after coronary artery occlusion resulted in distinct rise of blood and lymph toxicity in early periods because of the "washing out" of toxic products from the ischemic myocardium, followed by normalization that had been more quicker than in controls.     

Selenium metabolism in rats after administration of toxic doses of selenite

We studied organ concentration, excretion and excreted forms of selenium in young and adult rats after a single s.c. injection of a sublethal dose of 75Se-selenite. In the young about a 10-fold higher concentration of 75Se in blood, liver, kidneys and In the young, about a 10-fold higher concentration of 75Se in blood, liver, kidneys and heart was found at all the experimental intervals studied (1-7 days). The highest 75Se concentration in the young was in the liver while in the adults it was found in the kidneys. The spectrum of radioselenium metabolites in the urine was the same in both groups. However, the main product excreted by young rats was 75Se-glutathione selenotrisulphide and an unidentified neutral substance while it was the trimethylselenonium ion in the adults. Ontogenetic differences in selenium metabolism could be one of the factors underlying the differences in the response of the young and the adult rats to toxic doses of selenite.     

Cultured human fibroblasts as a model for evaluation of potential in vivo toxicity of membrane damaging antibiotics

The toxic side effects of certain antimicrobial agents are probably related to their membrane damaging properties. Thus it should be possible to use measurement of membrane damage in vitro for evaluation of the potential toxicity in vivo of such antibiotics. We estimated the membrane damage induced in cultured human fibroblasts by anti-microbial agents, such as polyene antibiotics, sodium fusidate and polymyxin B as well as derivatives of some of these. Degree and character of membrane damage was determined on basis of leakage of three defined cytoplasmic markers from prelabelled cells after treatment with test substance.By comparing the minimal inhibitory concentrations against the target microbial cells (MIC) with the amounts needed to cause membrane damage of human cells (ED₅₀) a ‘therapeutic dose range’ was obtained (ED₅₀/MIC). The therapeutic dose range and the character of induced membrane damage were compared with the relative toxicities in vivo of each test substance. Highly toxic agents caused large functional ‘holes’ and/or showed a narrow therapeutic dose range, whereas less toxic substances induced smaller functional holes and/or had a larger therapeutic dose range. These parameters, evaluated in the presented model system, should be useful for an indication of potential toxicity in vivo.     

Efficacy of cow's kumiss in the treatment of large intestine dysbacteriosis and its impact on tolerability of antituberculosis agents in patients with respiratory tract tuberculosis]

Dysbacteriosis of the large intestine is one of severe complications of long-term use of antituberculosis agents in the treatment of respiratory tract tuberculosis that results in a significant decrease of tolerability of antituberculosis agents, persistence of tuberculosis intoxication and slower involution of the tuberculosis process in the lungs. When the complex treatment with antituberculosis agents was accompanied by the use of cow's kumiss for correction of the large intestine dysbacteriosis, the intoxication signs disappeared in 12% of the patients in the main group, while in the patients of the control group the level of the intoxication syndrome increased twice. The rate of the tuberculosis lesions regression evident from the lung roentgenograms was 2.7-fold higher in the main group vs. the control (62 and 23% respectively). The indices of the lung functional capacity recovery in the patients of the main group vs. the control were also higher (41 and 33% respectively). Hepatic toxic reactions in the patients not given cow's kumiss for correction of dysbacteriosis were 8 times more frequent vs. the control. The results of the study made it possible to develop recommendations for phthisiologists in the use of cow's kumiss as one of the methods of pathogenetic therapy in complex treatment of patients with respiratory tract tuberculosis in sanatoria.