Hepatic glutathione concentrations and the release of pyrrolic metabolites of the pyrrolizidine alkaloid, monocrotaline, from the isolated perfused liver. We have examined the relationship between the metabolism of the pyrrolizidine alkaloid, monocrotaline, and glutathione concentration in the isolated, perfused rat liver. On perfusion of monocrotaline (300 microM) through the isolated liver, high concentrations (1.1 mM) of its metabolite glutathionyldehydroretronecine are released into bile, while much lower amounts (4.86 microM; 0.05 mumol/g liver) accumulate in the perfusate over a 1 hr perfusion period. Metabolite concentration in both the bile and perfusate increase when the level of monocrotaline perfused is increased to 900 microM. Metabolite release is also elevated in livers pretreated with phenobarbital. Monocrotaline perfusion lowered glutathione concentrations in the liver from 30 min onwards. Livers from animals treated with buthionine sulfoximine or chloroethanol showed much lower glutathione levels after 60 min perfusion. Livers from chloroethanol-treated (but not buthionine sulfoximine-treated) animals showed significantly lower release of pyrroles into the bile on perfusion with monocrotaline, but there is no effect on the rate of build-up of pyrrolic metabolites in the perfusate. We conclude that hepatic glutathione concentrations and the release of pyrrolic metabolites of monocrotaline mutually interact. Exposure of the liver to monocrotaline reduces glutathione concentrations, while marked depletion of liver glutathione concentration leads to a decrease in the release of monocrotaline metabolites.


Effects induced by bothropstoxin, a component from Bothrops jararacussu snake venom, on mouse and chick muscle preparations. Bothropstoxin, a 13,700 mol. wt myotoxic phospholipase homologue isolated from the venom of Bothrops jararacussu and devoid of PLA2, proteolytic or hemolytic activities, inhibited muscle twitch tension, evoked either directly or indirectly through stimulation of the motor nerve in the mouse phrenic-diaphragm preparations. The compound action potential of the muscle was also abolished with a similar time course. In addition, the toxin (0.7 mM) evoked membrane depolarization which was inhibited in the presence of 10 mM Ca2+. In chick biventer cervicis muscle, the toxin (2 mM) induced a contracture that reached its maximum amplitude in 44.8 +/- 15.6 min (n = 6) and was not blocked by either d-tubocurarine or tetrodotoxin. The time to maximum amplitude was reduced to 5.5 +/- 1.0 min (n = 4) in nominally Ca(2+)-free Krebs solution and was completely abolished in Ca(2+)-free Krebs solution containing 1 mM EGTA.


Antivenom and biological effects of ar-turmerone isolated from Curcuma longa (Zingiberaceae). A potent antivenom against snakebite was isolated from Curcuma longa, a plant commonly used in traditional Brazilian medicine. The fraction consisting of ar-turmerone neutralized both the hemorrhagic activity present in Bothrops jararaca venom, and the lethal effect of Crotalus durissus terrificus venom in mice. Immunological studies demonstrated that this fraction also inhibited the proliferation and the natural killer activity of human lymphocytes.


Comparison of the potency of three Brazilian Bothrops antivenoms using in vivo rodent and in vitro assays. BIASG (Butantan Institute Antivenom Study Group). Three Brazilian polyspecific Bothrops antivenoms were compared using standard W.H.O. rodent in vivo and in vitro assays of their ability to neutralize the principal venom activities of pooled whole Bothrops jararaca venom. On a volume basis, the antivenoms were equally effective in neutralizing lethal activity in mice, and there were only minor differences in their ability to neutralize venom-induced haemorrhage, necrosis and procoagulant activity. Antivenom efficacy in neutralizing defibrinogenation varied. However, when equal amounts of antivenom IgG were compared, it was found that the FUNED antivenom best neutralized lethality, haemorrhage, necrosis and fibrinogen clotting activity. Vital Brazil and FUNED antivenoms were equally effective in neutralizing plasma coagulant activity but Vital Brazil antivenom was the more effective in neutralizing defibrinogenation.


Citrate is an endogenous inhibitor of snake venom enzymes by metal-ion chelation. Citrate levels in selected snake venoms were determined by an enzymatic assay coupled to NADP+ reduction. Citrate concentrations in different viper venoms (n = 5) varied from 95 to 150 mM, in crotalids (n = 3) from 63 to 142 mM, and in elapids (n = 4) from 17 to 163 mM. In Bothrops asper venom Ca(2+)-ion concentrations varied from 2.5 to 3.6 mM, suggesting that the high relative citrate levels may serve to chelate endogenous divalent metal cations, thereby inactivating divalent cation requiring enzymes. Control experiments with B. asper phospholipase A2 MIII in the presence of 2.5 mM Ca2+, showed that the enzyme is completely inhibited by 20 mM citrate. Crotalus adamanteus 5'-nucleotidase and phosphodiesterase are also inhibited 100 and 75%, respectively, by 100 mM citrate. By forming complexes with divalent metal ions, citrate markedly reduces the activities of selected enzymes in snake venoms. Secretion of high concentrations of citrate may represent an important mechanism by which snakes protect themselves against the toxic effects of their own venoms.


Effects of sand viper (Cerastes cerastes) venom on isolated smooth muscle and heart and on haematological and cardiovascular parameters in the guinea-pig. The effects of the venom of the sand viper (Cerastes cerastes) on haematological and cardiovascular parameters and on isolated ileum, trachea, pulmonary artery and atrium from the guinea-pig were studied. In concentrations from 0.2 micrograms/ml to 0.6 mg/ml, snake venom caused concentration-dependent relaxation of the longitudinal ileal segments and the epinephrine-precontracted pulmonary artery rings, increased the tone of the tracheal rings and increased the rate of the spontaneously beating atrium but depressed the amplitude of its contraction. Low doses (0.04-0.4 mg) of the venom caused transient small depression of both heart rate and contractility of the isolated perfused heart. A higher dose (1.0 mg) significantly inhibited both parameters and delayed their recovery. An i.v. injection of 0.12 mg/kg of the snake venom to anaesthetized guinea-pigs had no effect on red blood cell count, haemoglobin concentration or the haematocrit value but significantly reduced the circulating white blood cells, the plasma clotting time and erythrocyte deformability. Also, i.v. injections of 0.02-0.2 mg/kg of the venom caused initial depression of the systolic and the diastolic blood pressure followed by complete or partial recovery and subsequent hypotension. These observations indicate that C. cerastes venom has multiple sites of action which may help in better understanding the pathology of bites by this snake.


Cardiovascular effects of Acanthaster planci venom in the rat: possible involvement of PAF in its hypotensive effect. Cardiovascular effects of the crowns-of-thorns starfish (Acanthaster planci) venom were examined in rats. The crude venom extracted from the spines of A. planci caused systemic hypotension associated with an increase in heart rate and a decrease in renal cortical blood flow when given i.v. The hypotensive effect of the venom was not inhibited by pretreatment with atropine, indomethacin or aprotinin, but was significantly inhibited by SRI 63-441, a platelet activating factor (PAF) antagonist. The venom caused dose-dependent vasorelaxation of the isolated rat aortic ring preparation precontracted by noradrenaline, an effect which was significantly attenuated by pretreatment with SRI 63-441, methylene blue or parabromophenacyl bromide. Denudation of the endothelium also diminished the vasorelaxing effect of the venom. Both the vasorelaxing and the hypotensive effects showed tachyphylaxis. These results suggest the release of PAF or a PAF-like substance from the endothelium by the venom.


The toxicity of macrocyclic trichothecenes administered directly into the rat brain. The tested macrocyclic trichothecenes are produced by Myrothecium fungi and by the plant Baccharis megapotamica. The toxicity of five macrocyclic trichothecenes has been measured by intracerebral and subcutaneous injection into rats. It is assumed that the toxic effects are based on inhibition of protein synthesis. Intoxication of rats by these compounds finds expression in slowly progressing respiratory depression and paralysis of skeletal muscles. The macrocyclics are derived from verrucarol, which lacks ring D and exhibits only low toxicity. The high toxicity of the macrocyclics, established by intracerebral and subcutaneous applications, may thus be attributed to the presence of the large ring D.


Occurrence of the mycotoxin chlamydosporol in Fusarium species. The mycotoxin chlamydosporol (C11H14O5) has been independently isolated from three different Fusarium species (F. chlamydosporum, F. acuminatum and F. culmorum), suggesting that its distribution may be widespread. We have developed a sensitive, specific fluorometric assay for chlamydosporol that allows quantitation in crude culture extracts. Using this assay we have observed production of chlamydosporol on rice medium by 23 of 40 isolates (58%) of Fusarium sp. examined in amounts as high as 0.8% (wt/wt) dried culture medium.


Experimental myonecrosis induced by the venoms of South American Micrurus (coral snakes). Venoms from 11 taxa of Micrurus (coral snakes) from Brazil and Colombia were tested for myotoxic activity in mice. All venoms, except that of M. surinamensis, induced myotoxicity as judged by the increase in plasma creatine kinase levels and by histological analysis. Qualitatively, these venoms induced a similar necrotic pattern, although there were conspicuous quantitative differences between them.


European viper venoms: haemorrhagic and myotoxic activities. Thirty-one venom samples from European vipers (genera Vipera and Daboia) were tested for haemorrhagic and myotoxic activity by intramuscular injection into mice. Most venoms exhibited haemorrhagic activity and fewer had myotoxic activity, both of which are not strictly related.


Microcystin-like toxins in different freshwater species of Oscillatoria. In January and September of 1989 and March 1990 blooms of Oscillatoria rubescens, Oscillatoria tenuis and Oscillatoria mougetii were found in Lake Simbirizzi and Lake Flumendosa in Sardinia, and in Lake San Puoto in the Lazio region of Italy. By using different extraction methods and HPLC analysis, two microcystin-like toxins (RR-like and YR-like), similar to some of the toxic compounds produced by the Cyanophycea Microcystis aeruginosa, were detected in these blooms.


Comparison of the purity and efficacy of affinity purified avian antivenoms with commercial equine crotalid antivenoms. Antivenoms were raised in laying hens by repeated immunizations with detoxified crotalid snake venoms and purified from egg yolks by affinity chromatography. While the affinity purified avian antivenoms were essentially pure IgG, commercial equine (Wyeth) and W.H.O. international reference antivenoms (Trimeresurus flavoviridis) contained several non-immunoglobulin contaminants. In standard mouse protection assays, the purified avian Crotalus atrox and T. flavoviridis antivenoms were 6.3 and 2.0 times as potent, respectively, as these equine antivenoms in neutralizing venom lethality. The purity, efficacy, and ease of manufacture of avian antivenoms, and their inability to fix mammalian complement, make them an attractive alternative to equine and other mammalian antivenoms.


Molecular properties and structure-function relationships of lethal peptides from venom of Wagler's pit viper, Trimeresurus wagleri. Two new lethal peptides (waglerins) were purified from the venom of Trimeresurus wagleri, and sequenced. We found them to be analogs of lethal peptides (waglerins) I and II reported previously (Weinstein et al., Toxicon 29, 227-236, 1991), with an additional Ser-Leu on the amino terminus. Three of the four waglerins were synthesized and the products were chemically and biologically equivalent to the naturally occurring counterparts in venom. Murine i.p. LD50 for synthetic waglerins I, SL-I and II were 0.33, 0.22, and 0.51 mg/kg, respectively. The single, intramolecular disulfide bond in each synthetic peptide formed rapidly in high yield. The reduced (cysteine-containing) forms of the peptides appeared to have significant toxicities, even without prior disulfide bond formation, but synthetic analogs with serine substituted for cysteine were not toxic. The synthetic dimer of waglerin I, formed by two intermolecular disulfide bonds, was not toxic, but rapidly rearranged to lethal, monomeric waglerin I at alkaline pH upon the addition of 5 mM beta-mercaptoethanol. Waglerin I was inactivated by cleavage at Tyr-15 with chymotrypsin.


Separation of insecticidal components from an extract of the roots of male Piper guineense (west African black pepper) by gas chromatography. A petroleum ether extract of Piper guineense male roots showed insecticidal activity when tested against Musca domestica. Gas chromatography of the extract yielded four active fractions, one of which was a pure component which was identified by spectral and chemical methods as pellitorine (N-isobutyl-2E,4E-decadienamide). The root extract lost about half of its insecticidal potency during passage down the GLC column, and much of the residual activity was due to the presence of pellitorine. It was concluded that GLC facilitates the isolation of active components in a pure form, but causes some insecticidal components of the root extract to be lost.


Identification of insecticidal peptides from venom of the trap-door spider, Aptostichus schlingeri (Ctenizidae). Nine insecticidal peptides were isolated from the venom of the trap-door spider, Aptostichus schlingeri. Seven of these toxins cause flaccid paralysis of insect larvae within 10 min of injection and all were lethal within 24 hr. The complete amino acid sequences (32-76 residues) of six peptides are presented. The identified peptides contain three or four disulfide bonds and the larger peptides (74-76 residues) are quite similar in sequence with carboxyl termini as free acids, not amidated.


Neuromuscular effects of four phospholipases A2 from the venom of Pseudechis australis, the Australian king brown snake. Four homologous single chain phospholipases A2 (Pa-1G, Pa-5, Pa-12C and Pa-15) were tested for neuromuscular effects on chick biventer cervicis and mouse hemidiaphragm nerve-muscle preparations. The four isozymes blocked directly elicited (mouse hemidiaphragm) and indirectly elicited (mouse and chick nerve-muscle preparations) twitch responses in concentrations of 1-30 micrograms/ml. The order of potency seen in both types of preparations was Pa-1G = Pa-5 greater than Pa-12C much greater than Pa-15. All four isozymes caused slow-onset, sustained contractures and reduction of muscle membrane potentials. In the chick preparation, responses to acetylcholine, carbachol and KCl were reduced by exposure to the toxins. It is concluded that the toxins act primarily postsynaptically to depress muscle contractility, perhaps by directly damaging muscle fibres. The order of potency agrees with their phospholipase A2 activity. Pa-1G is unusual because it is an acidic molecule, most toxic phospholipases being basic.


Potential-dependent alpha-latrotoxin interaction with black lipid membranes. The influence of membrane potential on alpha-latrotoxin insertion into bilayer lipid membranes (BLM) has been investigated. It was found that positive potentials cis to toxin application stimulated the formation of channels in the bilayer. A two-step model of latrotoxin/membrane interaction is put forward to explain these data. In the first step, latrotoxin irreversibly binds to the bilayer without forming conductive structures. The second step of the process represents rapid insertion of the protein molecule into the bilayer with the formation of the conducting channel. We imagine the driving force for this process to be the interaction of charged groups in the toxin molecule with the electric field applied across the BLM. Our results are compared with known data on the interaction of LTX with synaptosomal membranes.


Activation of bovine factor V by an activator purified from the venom of Naja naja oxiana. The crude venom of many elapid snakes appeared to contain proteins that activated blood coagulation factor V. The factor V activator present in the venom of Naja naja oxiana was purified to homogeneity by chromatography on a mono-S column. The activator was a single chain protein with an apparent mol. wt of 48,000, as judged by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate and by gel permeation chromatography on Sephacryl S200. Activation of bovine factor V by the purified venom activator was accompanied by proteolytic cleavage of factor V and resulted in the formation of two major polypeptide chains with mol. wts of about 90,000 and 77,000. The final product obtained was compared with thrombin-activated factor V for its ability to function as cofactor in factor Xa-catalysed prothrombin activation in the presence of negatively charged phospholipid vesicles (5 mole% phosphatidylserine/95 mole% phosphatidylcholine). The Km for prothrombin obtained at a saturating amount of venom-activated factor Va was nine-fold higher than with thrombin-activated factor V (0.83 microM vs 0.09 microM, respectively) whereas both factor Va molecules stimulated the Vmax of thrombin formation some 6000-fold. Both forms of factor Va promoted the binding factor Xa to negatively charged phospholipid vesicles. However, the apparent Kd for factor Xa was less favorable in the presence of venom-activated factor V (0.67 x 10(-9) M) than in the presence of thrombin-activated factor V (0.043 x 10(-9) M). Thrombin cleaved a peptide bond in the 77,000 mol. wt polypeptide chain of venom-activated factor V, which resulted in the formation of a normal factor Va light chain. This peptide bond cleavage was, however, not associated with a change of cofactor activity. Venom treatment of thrombin-activated factor V, on the other hand, did remove a small fragment (mol. wt approximately 4000) from the heavy chain of factor Va (94,000), yielding a molecule with reduced cofactor activity. The diminished cofactor activity of venom-activated factor V is, therefore, likely due to the fact that a small peptide fragment, involved in the interaction with prothrombin and factor Xa, is missing from the heavy chain of venom-activated factor V.


Pharmacological studies of jumper ant (Myrmecia pilosula) venom: evidence for the presence of histamine, and haemolytic and eicosanoid-releasing factors. Jumper ant venom was prepared by extraction of venom sacs in distilled water and centrifugation to remove insoluble material. Jumper ant venom (2 micrograms/ml) produced a biphasic response on isolated guinea-pig ileum, i.e. an initial rapid contraction followed by a slower prolonged contraction. The histamine antagonist mepyramine (0.1 microM) inhibited the first phase of this response by greater than 90%. In the isolated rat stomach fundus strip (which is insensitive to histamine), jumper ant venom (6 micrograms/ml) produced only a single contraction. No tachyphylaxis was observed to repeated doses of jumper ant venom in guinea-pig ileum or rat fundus strip. Responses to jumper ant venom of the egg-albumin-sensitised guinea-pig ileum were not significantly different before and after an in vitro anaphylactic response induced by egg albumin (0.5 mg/ml). Fluorometric assay revealed a mean value of 0.9 +/- 0.2% of the dry weight as histamine in jumper ant venom. Both the lipoxygenase/cyclo-oxygenase inhibitor BW755C and the cyclooxygenase inhibitor indomethacin significantly inhibited the second phase response to jumper ant venom of the guinea-pig ileum, and the response of the rat fundus strip. The muscarinic receptor antagonist atropine (0.1 microM), the bradykinin antagonist [Thi5,8,D-Phe7]-bradykinin (10 microM) and the angiotensin converting enzyme inhibitor captopril (20 microM) did not affect either phase of the venom response in guinea-pig ileum. Jumper ant venom caused haemolysis of guinea-pig blood. The degree of haemolysis was significantly reduced when boiled venom was used. These results suggest that jumper ant venom contains histamine and may cause the release of cyclo-oxygenase products. It also contains a heat-sensitive haemolytic factor.


Structure determination and toxicity of a new microcystin from Microcystis aeruginosa strain 205. A new hepatotoxic microcystin was isolated from the cyanobacterium Microcystis aeruginosa strain 205. Its structure was found to be [Dha7]microcystin-RR as determined by amino acid analysis, mass spectrometry and 1H NMR spectroscopy. LD50 value (i.p. mouse) of this toxin was 180 micrograms/kg. The 48 hr lethal concentration (48-hr-LC50) of the toxin for larvae of the yellow fever mosquito, Aedes aegypti, was 14.9 micrograms/ml.


Pathological and biochemical changes induced in mice after intramuscular injection of venom from newborn specimens of the snake Bothrops asper (Terciopelo). Venom from newborn Bothrops asper snakes has higher lethal, hemorrhagic, edema-forming, proteolytic and defibrinating activities than venom from adult B. asper specimens. Electrophoretic analysis confirmed the variation between these venoms. Intramuscular injection of 100 micrograms of venom from newborn specimens in mice induced defibrination, together with moderate increments of serum levels of lactate dehydrogenase, creatine kinase, hemoglobin and total proteins. A conspicuous hemorrhage developed in injected muscle rapidly after envenomation, probably due to a drastic alteration in capillaries and larger blood vessels. Other histological alterations included moderate myonecrosis, lung collapse and prominent renal damage, characterized by tubular necrosis and hyalinization. Polyvalent antivenom effectively neutralized lethal, hemorrhagic and indirect hemolytic activities of newborn B. asper venom, although requiring higher antivenom doses than neutralization of venom from adult B. asper.


Precursor structure of omega-conotoxin GVIA determined from a cDNA clone. The Ca2+ channel blocking neurotoxin, omega-conotoxin GVIA, is a 27-amino acid peptide with three disulfide bonds. We have determined the precursor structure of this peptide by analyzing a cDNA clone obtained from a Conus geographus venom duct library. The omega-conotoxin GVIA prepropeptide is 73 amino acids in length comprising a 22 amino acid signal sequence, an intervening region of 23 amino acids, and finally, a 27 amino acid toxin region. A C-terminal glycine residue is later processed to a C-terminal amide moiety. The omega-conotoxin GVIA precursor exhibits regions of strong homology to the previously characterized King-Kong peptide precursor, but shows surprising divergence as well. The possible significance of the precursor organization is discussed.


IgG antibodies to Loxosceles sp. spider venom in human envenoming. The presence and specificity of IgG antibodies produced by patients with loxoscelism were studied. The loxoscelism diagnosis was supported mainly by clinical parameters. A search for IgG antibodies anti-Loxosceles gaucho venom in patients with loxoscelism submitted to serumtherapy showed antibodies in four out of 20 patients. The IgG antibodies were detected as early as 9 days and as late as 120 days after bite. The highest IgG antibody titer was 1:640 and the lowest was 1:80. Immunoblotting tests showed that human anti-L. gaucho IgG antibodies recognize preferentially the components responsible for the dermonecrotic and lethal activities of the venom. A comparison of the clinical picture, the level of serum IgG antibodies and the dose of antivenom administered suggest that there is no relationship between these parameters.


Histopathological changes in WEHI-3B leukemia cells following intoxication by cytotoxin P4 from Naja nigricollis nigricollis venom. The histopathological changes in WEHI-3B leukemia cells were followed by light and electronmicroscopy at different time intervals following exposure to cytotoxin P4 from Naja nigricollis nigricollis venom. At 1 hr after exposure to the toxin (2.5 x 10(-7) M) deformation was detected, primarily in the mitochondria, followed by vacuolization in the cytoplasm and an increase in lysosome number at 2 hr post-intoxication. Thereafter, the endoplasmic reticulum assumed a microsomal-like appearance, the plasma membrane was disrupted and, finally, the cells released their content to the culture medium. It is postulated that cytotoxin P4 may affect mitochondria either indirectly, by stimulating intracellular processes after binding to the cell membrane, or directly by interaction with the mitochondria after penetrating into the cell.


Characterization of snake venom components acting on blood coagulation and platelet function. Snake venoms can affect blood coagulation and platelet function in various ways. The physicochemical properties and the mechanisms of actions of the snake venom components affecting blood coagulation and platelet function are discussed.


Cross-reacting antigens in the butter clam (Saxidoma giganteus) and their relationship to total paralytic shellfish poison toxicity. A specific protein with an apparent mol. wt of 23,000 was identified in foot homogenate derived from paralytic shellfish poisoning (PSP) contaminated butter clams and was found to cross-react with crab-saxitoxin-induced protein (SIP) antiserum. Antiserum, once cross-absorbed against non-toxic shellfish material, was incubated with tissue homogenate derived from 52 butter clams with varying total PSP toxicities in a prototype ELISA. A significant (r = 0.83; P less than 0.001) correlation existed between soluble clam antigen content in foot homogenate and total PSP toxicity; the latter measured by the mouse lethality bioassay. From the ELISA results, a soluble antigen threshold of 0.1% total protein was successfully used to distinguish between PSP toxic and non-toxic butter clams. It is proposed that this type of screening assay could be used in conjunction with the standard mouse bioassay to increase PSP monitoring and potentially reduce unnecessary animal testing.


Ricin-induced hepatic lipid peroxidation, glutathione depletion, and DNA single-strand breaks in mice. The ability of ricin, the glycoprotein toxin from the castor bean (Ricinus communis), to stimulate oxidative stress was investigated. Following the i.p. administration of 25 micrograms ricin/kg or the vehicle to female CF-1 mice, the effects of ricin on hepatic lipid peroxidation, nonprotein sulfhydryl (reduced glutathione) content and DNA single-strand breaks were determined at 0, 12, 24, 36, 48 and 72 hr post-treatment. Hepatic lipid peroxidation significantly increased 3.4-, 3.8-, and 3.0-fold relative to control values at 24, 36, and 48 hr post-treatment, respectively. Hepatic nonprotein sulfhydryl concentrations decreased significantly to approximately 51%, 61% and 65% of control values, at 24, 36, and 48 hr, respectively. The incidence of hepatic DNA single-strand breaks increased by 2.9-, 2.8-, and 2.4-fold relative to the zero time values at 24, 36, and 48 hr after treatment with ricin, respectively. No significant differences were observed in either lipid peroxidation or nonprotein sulfhydryl concentrations at 12 or 27 hr post-treatment. Decreases in liver and intestinal weight to body weight ratios were observed in ricin-treated animals, while no changes were observed in spleen and kidney weight to body weight ratios. These results indicate that in the liver of mice, ricin induces an oxidative stress which is maximal at approximately 36 hr post-treatment.


Purification and characterization of an antithrombin III inactivating enzyme from the venom of the African night adder (Causus rhombeatus). A serine proteinase was isolated from the venom of the night adder (Causus rhombeatus) by fast protein liquid chromatography (anion-exchange, gel filtration and hydrophobic interaction). The protein (termed CR-serpinase) had an estimated mol. wt of 45,500 as determined by SDS-PAGE, pI of 4.7 and a carbohydrate content of 18.9%. Incubation of CR-serpinase with purified human antithrombin III at a molar ratio of 1:66 resulted in a loss of more than 90% of the initial AT III activity within 10 min. The reaction was dependent on heparin. In SDS-PAGE inactivation of human antithrombin III was correlated with the occurrence of two cleavage products. The cleavage site in the antithrombin III molecule was determined to be Arg 393-Ser 394 by amino-terminal sequencing. CR-Serpinase had no thrombin-like activity since no fibrinogen conversion was induced and had no procoagulant activity. CR-Serpinase activity was not inhibited by antithrombin III-heparin and was not decreased by a 10-min preincubation in normal human plasma. Inactivation of antithrombin III by CR-serpinase appeared to be very specific.


Influence of unilateral ovarian torsion to the contralateral ovary. An experimental study was planned to show the suppression of contralateral ovary in unilateral ovarian torsions. In the study, 55 New Zealand female rabbits were used. Basal and stimulation 17 beta estradiol values were measured and unilateral torsion in 20 rabbits, bilateral torsion in ten rabbits and bilateral oophorectomy in five rabbits were performed. Eight, 16, 24 hours after the procedure, 17 beta estradiol levels were measured again and the second stimulation values maintained. The control group consisted of 20 rabbits. Values were analyzed statistically and the results were significant.


Effects of endurance training under hyperoxia on serum and tissue lipid levels in rats. Effects of 6 weeks endurance training (5 days per week) under hyperoxia (60% O2 plus 40% N2) on serum and tissue lipid levels were investigated in male rats. Rats were divided into 4 groups: normoxia-control (NC), hyperoxia-control (HC), normoxia-training (NT), and hyperoxia-training (HT). NT and HT groups were run on a treadmill in a chamber at 20 m per min with a 6 degree gradient for 30 min. The chamber was perfused with hyperoxic gas. After training under hyperoxia, high density lipoprotein cholesterol (HDL-C) was higher (p less than 0.01) than in the normoxic condition, but there were no changes in serum lipids and glucose, or liver and aorta cholesterol. Skeletal muscle TG and liver glycogen in hyperoxic condition tended to show the higher values. Basal lipolysis of adipose tissue was increased (p less than 0.05) by hyperoxia, and decreased (p less than 0.01) by training, but there was no change of norepinephrine induced lipolysis in any group. The results suggest that endurance training or resting under hyperoxia induces increased HDL-C or lipolytic activity of adipose tissue. These effects might be caused by greater fat oxidation during exercising or resting under hyperoxia.


Progressive facial hemiatrophy with localized scleroderma. A 46-year-old woman who presented with progressive facial hemiatrophy (PFH) following localized scleroderma is described. The patient had a markedly deformed and depressed plaque surrounded by erythema on the right cheek. She also showed linear scleroderma with hair loss on the occipital area and morphea lesions on the neck and shoulder. Histological findings of the facial and other atrophic lesions were consistent with localized scleroderma. Therefore, the PFH and localized scleroderma may have had the same pathological background in this case.


Cardiac tamponade in the blastic crisis of chronic myelogenous leukemia. A 36-year-old man with Philadelphia chromosome-positive chronic myelogenous leukemia (CML) developed hemorrhagic pericarditis with tamponade as a terminal manifestation of the blastic crisis. Cardiac tamponade should be kept in mind as an uncommon cause of death of CML patients. Based on a literature review, symptomatic pericarditis in patients with CML blast crisis suggests imminent death. This is in contrast to long-term survival for patients in the chronic phase.


Plasmapheresis in the Tokai University Medical Center, 1983-1989. There were 634 instances of plasmapheresis performed on 152 patients in the Kidney Center, Tokai University Hospital, in the seven year period from Jan. 1983 to Dec. 1989. The average number of plasmapheresis treatments per patient was highest for those with neurological and dermatological disease (6.7), followed by liver disease (5.6), and drug intoxication (1.2). The major treatment modalities for the liver, collagen, drug intoxication, hematologic, neurologic and hyperlipidemic disorders were plasma exchange (PE) and double filter plasmapheresis (DFPP). Recently, however, the use of plasma perfusion (PP) and immunoadsorbent plasma perfusion (IPP) has been increasing because of their ability to selectively adsorb various pathologic macromolecules.


[The morphofunctional characteristics of the dystrophic and reparative processes in experimental autodermoplasty using a CO2 laser]. The aim of this work was research of the morphofunctional features of rabbit free skin flaps after CO2 laser and scalpel interventions. pO2, histologic and histochemical data were analyzed. The results evidence an insignificant delay of the pO2 growth in laser-treated flaps early after surgery. This fact hints at a low speed of laser-treated flaps vascularization because of wound coagulation necrosis. However, this does not influence the course of the reparative process. Our conclusions are confirmed by histologic and histochemical findings.


[A temperature assessment of the treatment of exacerbated chronic periodontitis with the immobilized proteinase preparation profezim]. Presents the results of studies of the gingival thermal response in patients with chronic periodontitis exacerbations. Profezyme, an immobilized proteinase enzyme, was used in the treatment of 51 patients (test group) and chymotrypsin was administered to 43 controls. Anti-inflammatory action of profezyme was demonstrated. Reduction of the gingival temperature and pain alleviation evidence a high efficacy of the drug.


[The results of the clinical use of the immobilized-protease enzyme profezim in treating exacerbated chronic periodontitis]. The authors characterize the action of profezyme, an immobilized proteinase enzyme, used in the treatment of patients with exacerbations of chronic periodontitis. Time course of symptomatology was followed up in 65 patients in various periods of periodontitis therapy. Prolonged enzymic therapy helped reduce the length of treatment by 2-3 times.


[The efficacy of clinical indices in determining periodontal status]. The accuracy and reproducibility of a number of clinical indices used in assessment of the status of periodontal tissues was evaluated in clinical examinations of 240 students. The Green-Vermillion index and the CDL periodontal index were found the most accurate and reproducible, their efficacies ranging within 86-97%.


[The microbial status of the periodontal pocket]. Analysis of components helped reveal the true relationships between the counts of individual representatives of the bacterial association in the contents of 18 periodontal pouches. Though the authors did their best to select a group of patients with the periodontitis course as similar as possible, the multidimensional grouping method has demonstrated the heterogeneity of this group. Four types of periodontal pouches were distinguished, differing by their bacterial status; this probably requires a differentiated approach to its correction.


[A comparative study of the efficacy of the action of a number of local antimicrobial preparations on the species and quantitative composition of the microbial flora in periodontal pockets]. The effects of a number of antibiotics were studied with the use of phase contrast microscopy. The time course of the qualitative and quantitative changes of the periodontal pouch microflora evidences that a mixture of trichopol with chlorohexidine is the most effective therapy for periodontal pouch sanitization.


[The morphofunctional characteristics of the free cells and microvessels of the gingiva in inflammatory periodontal diseases]. A comprehensive assessment of the gingival histohematic barrier components in patients with gingivitis and periodontitis was carried out. Analysis of the intratissular correlations has led the authors to a hypothesis on the presence of a peculiar 'break-off' of the interstructural relationships determined by the severity of the periodontal injury. Gingivitis is characterized by a change, though minimal, of correlations, as against the intact periodontium. Involvement of the entire tissue complex of the periodontium in the process is associated with elevation of the counts of both positive and negative relationships between free stromal cells. Accumulation of free stromal cells and disorder of their cooperation is parallel with lowering of alkaline phosphatase activity and isolation of polymorphonuclear leukocytes, this forming the structural basis for the progress of the inflammatory dystrophic process in the periodontium.


[A morphological assessment of the efficacy of the deaggregation therapy of chronic catarrhal gingivitis]. Light optics, electron microscopy, morphometry were used to study the structural changes in the epithelial cells and endotheliocytes of gingival papillary capillaries of patients with chronic catarrhal gingivitis after papaverin therapy. The studies have revealed signs of disappearance of intercellular edema and a higher degree of cellular differentiation; signs of normalization of transcellular fluid transport were seen in the endotheliocytes, and aggregations of formed elements of the blood disappeared.


[Indices characterizing the expression of hyperesthesia of the hard dental tissues in patients with periodontitis]. Clinical examinations of 226 patients with periodontitis have revealed in 61% of them hyperesthesia of the hard dental tissues, that was mostly generalized, characterized by a marked reaction to thermal, chemical, and tactile stimulants. Study of the mineral metabolism has demonstrated that growing bone tissue loss in dental biopsy specimens and gingival fluid is associated with reducing levels of P, Ca, and Mg. Combination of clinical manifestations of periodontitis and hyperesthesia necessitate search for combined therapy aimed at simultaneous elimination of hyperesthesia and inflammation.


[Ways to improve the treatment of patients with phlegmons of the maxillofacial area]. The examination has involved a total of 216 patients (111 in the study group and 105 in the reference one) with maxillofacial phlegmons. Patients of the main group were administered multiple-modality treatment with surgical treatment of the purulent focus, involving a primary suture on the wound and intraarterial local drug therapy. Partial necrectomy was found possible in the majority of cases with the surgical treatment of the purulent focus; therefore to prevent dissemination of the inflammatory process and liquidate it as soon as possible surgery had to be performed against the background of the maximal permissible antibiotic concentration in the adjacent tissues. Primary suturing of the purulent wound had to be combined with draining and continuous dialysis of the postoperative wound. The postoperative wound healed by primary tension in 91.89% of patients in the main group; the incidence of complications was lower in them than in the reference group; the length of hospital stay and invalidity periods were reduced almost twofold.


[Lipid peroxidation in suppurative-inflammatory diseases of the maxillofacial area]. The levels of lipid peroxidation products were measured in patients with various pyoinflammatory conditions of the maxillofacial area: abscess, phlegmons, chronic mandibular osteomyelitis). Blood plasma and red cell levels of lipoperoxides were found increased and alpha-tocopherol levels lowered in all the examinees.


[The clinico-immunological aspects of dental implantology]. Analyzes the clinical and immunologic data evidencing that implantation, as a rule, induces immunologic shifts in patient's body. Traces the direct relationship between the frequency and severity of complications and the type of the patient's immune response to implantation, the intensity of this response being directly related to the implantation material and volume. Basing on the relationship between the clinical and immunologic data, recommends examination of the patients' immune status for the prediction of the results of the surgical stage of dental implantation and for the assessment of new implantation material intended for introduction into practical dentistry.


[The bone tissue of the jaws in congenital and acquired deformities]. Biophysical parameters of the jam bones were examined in patients with congenital and acquired deformations. Ash content, density, mineral content, and microhardness were estimated in 58 patients aged 17 to 28. The mean values of these characteristics were found to be unrelated to the individual features and the type of the changes in the bone tissue, associated with deformations of the jaws. Frequency ranging of the individual values helped obtain a more objective picture of the bone tissue status. Bone tissue density was found increased in congenital maxillary and mandibular deformations. These data evidence a lowered plasticity of the bone tissue in the patients with congenital deformations of the jaw bones.


[The regional blood circulation of the parotid gland and the correction of its disorders in chronic interstitial parotitis]. Elevated vascular tone, lengthened time of intraglandular arterial bed filling, fibrinogenemia, elevated plasma tolerance to heparin were detected in 32 patients with chronic interstitial parotitis (CIP) during exacerbation and in a number of CIP patients with the remission of the disease. Correction of microcirculation disturbances is advisable during CIP exacerbation; in remission it may be effective in patients with the late stage of parotitis, in those with parotitis coursing along with diabetes mellitus, essential hypertension, or against the background of prolonged oral drug therapy (e. g. clofelin). Drugs correcting the microcirculatory disorders are conducive to a sooner alleviation of the inflammation and better and longer preservation of the function of the parotid glands.


[The regional blood circulation in the parotid gland area in Sjögren's disease (syndrome)]. Rheographic investigations of the parotid gland area in 103 patients with Sjogren's disease (syndrome) that developed in the presence of chronic nonspecific sialadenitis during its various stages and inflammation phases have demonstrated that the blood stream volumic rate directly depended on the inflammation phase and intensity of this phase. During remission the circulation in the salivary glands was close to normal.


[The late results of observing and treating patients with histiocytosis X and involvement of the jaw bones]. Eighty-one patients with histiocytosis X involving the jaw bones were followed up for 30 years; the patients presented with involvement of the lungs, skin, mucosa, other bones of the skeleton, and diabetes insipidus signs. In 58 of the 81 patients eosinophilic granuloma eventuated in a generalized process: chronic xanthomatosis or Hand-Schüller-Christian's disease. This fact permits a conclusion that eosinophilic granuloma is an initial stage of the same systemic disease, histiocytosis X, and that subdivision of histiocytosis X into 3 forms, accepted in literature, is undesirable, for it erroneously presents the essence of the process. Therapeutic recommendations are given.


[Body immunological reactivity in patients with chronic nontumorous diseases of the parotid salivary glands]. Examinations of 81 patients and 30 healthy donors have revealed a variety of immunologic disturbances in different forms of chronic nontumorous conditions of the parotid glands. Lymphocyte sensitization to salivary gland antigens was observed in all the patients, but was the most marked in those with Sjogren's disease. The studies permit considering the detected shifts in the immunity system an important component of the pathogenesis of chronic nontumorous diseases of the parotid glands. These findings may be useful in the differential diagnosis, prognosis, and adequate therapy of salivary gland diseases.


[The dynamics of fracture healing in the bones of the zygoma area]. Investigation of the reparative histogenesis in patients with zygomatic bone fractures suffered in different periods in the past included histologic study of the tissue specimens taken from fracture sites during surgical intervention, studies of the osteogenesis, topographic-anatomic position of bone fragments, their mobility and configuration, and clinical and pathomorphologic changes in the soft tissues adjacent to the fracture site. Specific features of the reparative osteogenesis of the facial skull flat bones and changes in the soft tissues adjacent to the fracture site were detected.


[The importance of the surgical treatment method in cancer of the oral mucosa with the primary tumor 4 cm in size and larger]. The authors analyze the results of surgical treatment of patients with local disseminated squamous-cell carcinoma of the buccal mucosa. The data on the survival rate of patients treated with different surgical methods are presented and variants of combined operations described.


[The modelling of mandibular osteomyelitis]. The method suggested by the authors permits a reliable reproduction of mandibular osteomyelitis in white rats. The primary localization and pattern of the infectious process in this model well correspond to odontogenic osteomyelitis, this permitting a preclinical study of new drugs and physiotherapeutic methods intended for patients with odontogenic diseases.


[The experience of using the 1-day hospital in maxillofacial surgery]. The authors describe for the first time the experience gained by a one-day hospital for oral surgery. The efficacy of this hospital is analyzed and economic effect estimated.


[A method for performing stimulation electromyography of the masticatory muscles in man]. A principal block scheme of an apparatus complex is suggested and a method for examination of masticatory muscle function is described. The method is based on electrostimulation of masticatory nerve followed by registration and analysis of the masticatory muscle action evoked potentials. The block scheme of the apparatus complex includes a universal electrostimulator ESU-2. Examinations of 30 healthy volunteers and 30 patients with unilateral fractures of the mandible helped define the major parameters of stimulation electromyography. The patients were treated by splints applied on the teeth with fixation on both jaws. By day 28 of the investigation the M-response amplitude dropped more than twofold and its length by 1.5 times as against the mean parameters in health.


[The characteristics of treating odontogenic osteomyelitis against a background of malaria in the population of the Republic of Burundi]. All the patients were administered preoperative antimalarial therapy, antibiotics and sulfanilamides, and agents maintaining the defense factors, as well as detoxifying and desensitizing therapy. Rational nutrition was prescribed. All these measures helped liquidate the inflammation foci not only in the bones, but in the soft tissues of the paramaxillary area as well.


[An x-ray method for determining the position of the tongue in the oral cavity]. The suggested modified technique for assessing the tongue position in the oral cavity is based on teleroentgenography in the lateral projection and estimation of cephalometric parameters. Theoretical rationale is given for the use of skeletal check points in the mandibular area and cervical portion of the spine and of the mandibular and spinal reference lines. Indications for the use of this method are defined and its advantages described.


[The comparative characteristics of mastication tests]. Changes in the values of Manly's [correction of Manley's], Rubinov's, and the author's mastication tests were studied in subjects with intact dentition, orthognathic occlusion; the tests were carried out 3 times each with intervals of 2-3 weeks. Correlations of the data of Dahlberg's [correction of Dalberg's], Manley's, Rubinov's and the author's tests were studied and the pattern of changes in the data of the tests in minor dentition defects analyzed. Repeated tests yielded better results; the data of various tests were found incompatible; the suggested test was found the most sensitive and accurate. The basic requirements to a mastication test are formulated.


[The clinical evaluation of the use of metal ceramic dentures with a Simet glass-ceramic coating]. Good results of clinical follow-up of patients with cermet dentures coated with cimet (glass ceramic) permit recommending this type of coating for wide practical use. Reconstructive prosthetics with glass ceramic and metal and cermet dentures should involve denture modeling in the articulator and a prophylactic check-up at least once a year with oral hygiene and functional occlusion control.


[An automated system for the collection and processing of clinico-diagnostic information in developing methods for treating dental defects by using dental implants]. A dialogue medical information system "Dental Implant" is described, intended for the collection and analysis of clinico-diagnostic information with the aim of developing methods for the treatment of patients with dentition defects, making use of dental implants. This automated system permits a rapid and effective analysis of surgical and orthodontic complications and is intended for operation by a non-programmer.


[Kalipsol-based balanced intravenous anesthesia in pediatric outpatient dental practice]. Basing on clinical studies of anesthesia in 568 children, central hemodynamic studies, analysis of the blood acid-base balance and gases, glycemia level, and assessment of the psychoneurologic status, the authors validate the usefulness of calypsol-based anesthesia in practical outpatient pedodontics. Such anesthesia did not influence the stability of central hemodynamics parameters, nor did it change the acid-base balance and blood sugar level; it provided an adequate protection of the patient from surgical stress with the adaptation mechanisms remaining intact. The child may leave the clinic 2.5-3 h after the intervention and a test for adequate adaptation to the environment. Calypsol-based balanced anesthesia improves outpatient care of the children, introduction of this method into practice will bring about a good economic effect.


Minor and trace sterols in marine invertebrates 65. 23-Epidihydrocalysterol: a new cyclopropane-containing sponge sterol. A new cyclopropane-containing sterol was isolated from the marine sponge Cribrochalina vasculum. The new sterol was characterized by nuclear magnetic resonance and mass spectrometry and the structure was shown to be (23R,24S,28R)-dihydrocalysterol. Implications concerning the biosynthesis of cyclopropane and cyclopropene sterols in sponges are discussed.


[A comparative study of the role of atresia and stenosis of the outflow duct of a salivary gland in the pathogenesis of dystrophic and inflammatory changes]. Experiments were carried out with submandibular salivary glands of 20 dogs. Complete obturation of the submandibular duct (in 10 animals) was achieved by binding this duct. Partial impairment of the saliva flow from the gland was induced in 10 animals by fixing a metal ring on the duct. The experiment has demonstrated that both complete and partial obturation of the salivary duct result in the development of chronic sialadenitis. Partial obturation of salivary outflow from the gland did not result in an acute inflammation but in a flaccid process, slowly developing dystrophic changes in acinar cells, gradually developing sclerosis of the glandular parenchyma and stroma. These data indicate a direct relationship between the intensity of dystrophic changes in the salivary gland and the severity of impairment of salivary discharge.


Persistent estrogen responsiveness of ras oncogene-transformed mouse mammary epithelial cells. Ovarian steroids are associated with the proliferation of normal as well as tumorigenically transformed mammary epithelial cells. The experiments performed in this study were designed to establish that (1) tumorigenic transformation induced by the ras oncogene is associated with alterations in estradiol biotransformation, (2) altered endocrine responsiveness persists in the fully transformed tumor cell phenotype and (3) specific perturbations induced by the ras oncogene can be experimentally downregulated. The ras transfectant pH06T and the tumor-derived T1/Pr1 cells exhibited 3- and 43-fold increases, respectively, in C-16 alpha hydroxylation of estradiol relative to the parental mouse mammary epithelial cells (P less than 0.0001). At the cellular level, this alteration corresponded with approximately 90-fold increase in the anchorage-independent growth of T1/Pr1 cells (P less than 0.0001). Estrogen responsiveness of T1/Pr1 cells was demonstrated by their suppression of growth in phenol red-free and/or tamoxifen-supplemented medium and by the reversal of antiproliferative effect of tamoxifen by phenol red and estradiol. Indole-3-carbinol, a naturally occurring tumor suppressive agent, was able to upregulate C-2 hydroxylation at the expense of C-16 alpha hydroxylation of estradiol. Treatment of T1/Pr1 cells with indole-3-carbinol resulted in a substantial decrease in anchorage-independent growth.


[The characteristics of the occlusal surface relief of the molars and its role in caries susceptibility in Nenets children]. Analysis of the principal odontological and odontogliphic parameters of 1088 molars of caries-susceptible and caries-resistant Russian and Nenets children has demonstrated that molar caries-resistance of the Nenets children is explained by the prevalence of the basic and supplementary grooves and elements that create a more intricate fissure pattern, the tooth size, groove length, number of tubers and fossae at the site of fissure fusion.


[Changes in the oral cavity of children as possible diagnostic signs of leukemia]. In 15 of the 65 children the initial signs of leukemia coincided with oral diseases running an uncommon course: acute or chronic recurrent herpetic stomatitis, periostitis, parotitis. An infectious process showing no tendency to resolution, stubborn infiltration of tissues in inflamed foci, hemorrhages, and changed total status of a child may be regarded as probable diagnostic signs of leukemia, requiring a profound examination, carried out by a dentist and hematologist.


The effect of dihydrotestosterone and culture conditions on proliferation of the human prostatic cancer cell line LNCaP. Cell density, nutritional state, and serum factors modify the growth response of LNCaP human prostatic cancer cells to dihydrotestosterone. Evaluation of growth response to dihydrotestosterone requires logarithmic transformation of cell count or thymidine incorporation data. Under conditions of dose response, growth increases with cell density but no significant interaction of dihydrotestosterone with cell density was found under optimal culture conditions. The frequency of media change was a significant factor in modulating dose response. When cells from cultures maintained at different feeding periods were plated at different cell densities of (trypan blue) viable cells, significant effects of plating density on dihydrotestosterone response were found. Dihydrotestosterone protects cells under the adverse effects of media deprivation. Under the extreme adverse effects of serum deprivation, cells respond to dihydrotestosterone even under conditions of increasing cell loss. The effects of dihydrotestosterone on final cell density were significant. In the absence of serum, the elongated cells of LNCaP assume a round shape, but many remain adherent to the culture dish and can be restored to normal morphology by serum. A number of growth factors fail to restore normal morphology that was completely restored by a combination of fibronectin and dihydrotestosterone. We have not developed a practicable serum-free system for LNCaP.


Steroid metabolism in teleost gonads: purification and identification of metabolites by high-performance liquid chromatography. A simple, efficient, and comprehensive technique for the purification, identification, and quantitation of the common steroid metabolites synthesized by the gonads of teleosts involving five systems of high-performance liquid chromatography (HPLC) was developed. Steroid standards were identified in HPLC by UV absorption at 254 nm or 280 nm, by differential refractive index, or by using radioactive standards. Metabolites that do not absorb UV light and are not resolved in the isocratic HPLC systems were identified in thin-layer chromatography following purification by HPLC. By using this technique, most of the steroid metabolites, including some polar metabolites, synthesized by the gonadal tissues of the teleosts can be purified within three steps of chromatography. The HPLC systems reported here are also useful in identifying the chromium trioxide oxidized products of metabolites, such as triols and tetrols, which considerably narrows down the number of probable metabolites.


[The morphological parameters of the dental arches in children with unilateral complete cleft lip and palate]. The size and proportions of dentitions and relationships between longitudinal, transverse, and vertical parameters of the upper and lower dentitions and palatine processes were examined in 42 children (22 girls and 20 boys), aged 5-6 years, with unilateral complete labiopalatine clefts before uranoplasty. Analysis of correlations in the findings was carried out and 12 reliable coefficients of correlations obtained.


A highly specific heterologous enzyme-linked immunosorbent assay for measuring testosterone in plasma using antibody-coated immunoassay plates or polypropylene tubes. A highly specific and sensitive enzyme-linked immunosorbent assay, using a heterologous combination of antiserum raised against testosterone-3-(O-carboxymethyl) oxime-bovine serum albumin and penicillinase-labeled testosterone-11 beta-carboxymethyl ether, was developed for measuring testosterone in human plasma. Immunoassay plates (96 wells) provided a sensitivity of 2.5 pg/well. This was achieved by maintaining the molar ratios of steroid to enzyme between 10 and 20. The assay was very specific for testosterone and did not show any cross-reaction with the related C19 steroids tested. Replacement of immunoassay plates with the locally available polypropylene tubes raised the detection limits to 25 pg/tube, but improved the range of doses of testosterone that could be measured up to 10,000 pg. The antiserum to testosterone derivative was linked to both immunoassay plates and polypropylene tubes through immunochemical bridges. Comparison of testosterone values of 52 plasma specimens obtained by both solid phase methods with those of radioimmunoassay showed good correlation.


Comparison of estrogen sulfotransferase and pregnenolone sulfotransferase of guinea pig. Guinea pig adrenal estrogen sulfotransferase from either sex was eluted as a single peak, irrespective of buffer salt concentration, when subjected to fast protein liquid chromatography on gel filtration columns. The same enzyme was consistently eluted in two distinct peaks during chromatofocusing. Adrenal pregnenolone sulfotransferase was eluted during gel filtration in a heterogeneous pattern, dependent on salt concentration. These properties have made possible almost complete separation of the two sulfotransferases in one step, although adrenal estrogen sulfotransferase may possess a minute intrinsic ability to catalyze sulfation of pregnenolone. Pregnenolone sulfotransferase had no measurable activity toward estrone. Pregnenolone sulfotransferase from both sexes yielded variable elution patterns during chromatofocusing. Estrogen sulfotransferase from the adrenal, as well as that of guinea pig chorion, was strongly inhibited by N-ethylmaleimide and to a lesser degree by iodoacetamide and iodoacetate. Adrenal and chorion estrogen sulfotransferases were thermolabile and were activated, although not protected from the effect of heat, by binding to 3'-phosphoadenosine 5'-phosphosulfate. Adrenal pregnenolone sulfotransferase was inhibited only by high concentrations of N-ethylmaleimide and not at all by iodoacetamide or iodoacetate. It was more thermostable than the estrogen sulfotransferase and was not activated by binding to 3'-phosphoadenosine 5'-phosphosulfate.


Secular trends in stroke incidence and mortality. The Framingham Study. The reduction in US stroke mortality has been attributed to declining stroke incidence. However, evidence is accumulating of a trend in declining stroke severity. We examined secular trends in stroke incidence, prevalence, and fatality in Framingham Study subjects aged 55-64 years in three successive decades beginning in 1953, 1963, and 1973. No significant decline in overall stroke and transient ischemic attack incidence or prevalence occurred. In women, but not men, incidence of completed ischemic stroke declined significantly. Stroke severity, however, decreased significantly over time. Stroke with severe neurological deficit decreased in later decades, with a fall in rates of severe stroke cases in which patients were unconscious on admission to the hospital. There was no substantial change in the case mix of infarcts and hemorrhages and no decline in hemorrhage to account for the decline in severity. The proportion of isolated transient ischemic attacks increased significantly over the 30 years studied, yielding an apparent and significant decline in case-fatality rates in men only. Secular trends in stroke incidence and fatality did not follow a clear or definite pattern of decline. While a significant decline in stroke severity occurred over three decades, incidence of infarction fell only in women. The decline in total case fatality rates occurred only in men and resulted largely from an increased incidence of isolated transient ischemic attacks. The severity of completed stroke was significantly lower in the later decades under study.


Lipoprotein and apolipoprotein profile in men with ischemic stroke. Role of lipoprotein(a), triglyceride-rich lipoproteins, and apolipoprotein E polymorphism. The role of lipoprotein abnormalities in the development of ischemic cerebrovascular disease has not been sufficiently clarified. The aim of this study was to identify the lipoprotein profile in ischemic cerebrovascular disease and the possible role of apolipoprotein E polymorphism. The relation between the concentrations of lipoprotein(a), intermediate density lipoproteins, apolipoprotein A-I, apolipoprotein B, apolipoprotein E, and other lipoproteins was studied in 100 men with ischemic cerebrovascular disease (48 atherothrombotic, 28 lacunar, and 24 of unknown type) and in 100 healthy age-matched men as a control group. Patients with ischemic cerebrovascular disease had significantly higher levels of lipoprotein(a), lipids carried by intermediate density lipoproteins, and low density lipoprotein cholesterol and lower levels of high density lipoproteins than control subjects. Patients with atherothrombotic infarction had higher total serum cholesterol and low density lipoprotein cholesterol concentrations than patients with lacunar infarction. To assess lipoprotein abnormalities in normolipidemic subjects, a subgroup of 38 patients with ischemic cerebrovascular disease and 53 control subjects, both with serum cholesterol levels < 5.2 mmol/l (200 mg/dl) and triglycerides < 2.3 mmol/l (200 mg/dl), was analyzed. Serum lipoprotein(a), lipids carried by very low density lipoproteins and intermediate density lipoproteins, and low density lipoprotein triglycerides were significantly higher in normolipidemic patients compared with normolipidemic control subjects, whereas high density lipoprotein cholesterol levels were lower. Apolipoprotein E polymorphism in our ischemic cerebrovascular patients differed from that of the control group, with the epsilon 4 allele being more prevalent. Increased serum lipoprotein(a) levels and intermediate density lipoprotein abnormalities together with decreased high density lipoprotein levels are major risk factors for ischemic cerebrovascular disease, even in normocholesterolemic and normotriglyceridemic subjects. Finally, the epsilon 4 allele could probably be a predisposing genetic marker for ischemic cerebrovascular disease.


Cerebral cysticercosis as a risk factor for stroke in young and middle-aged people. A probable association between cerebral cysticercosis and susceptibility to stroke, especially among young and middle-aged patients, has been reported. We examined the association between cerebral cysticercosis and stroke and the possible factors causing this association. In 169 stroke patients (75 males and 94 females) under 65 years of age admitted to our neurology department, we evaluated the following possible risk factors: arterial hypertension, diabetes, cardiac disease, hyperlipidemia, smoking, alcohol abuse, and cerebral cysticercosis. In 169 control patients under 65 years of age matched by sex and age, we evaluated the same possible risk factors for stroke. In the univariate matched analyses, the frequencies of cerebral cysticercosis (p < 0.001), arterial hypertension (p < 0.001), cardiac disease (p < 0.001), hyperlipidemia (p < 0.05), and alcohol abuse (p = 0.05) were higher in the stroke patients than in the control patients. After controlling for possible confounding factors, we found that arterial hypertension (p < 0.001), cardiac disease (p < 0.001), and cerebral cysticercosis (p < 0.001) were independent risk factors for stroke. Cerebral cysticercosis should be considered a risk factor for stroke in young and middle-aged individuals.


Early time course of N-acetylaspartate, creatine and phosphocreatine, and compounds containing choline in the brain after acute stroke. A proton magnetic resonance spectroscopy study. The early time course after acute stroke of cerebral N-acetylaspartate, creatine and phosphocreatine, and compounds containing choline was studied in vivo by means of localized water-suppressed proton magnetic resonance spectroscopy. Eight patients with acute stroke were studied serially in the acute phase, 1 week after, and 2-4 weeks after the onset of clinical symptoms. Ten healthy volunteers served as controls. A stimulated echo (STEAM) sequence was used for measurement of the brain metabolites in a volume of interest located within the infarcted area as visualized by magnetic resonance imaging. For quantification, the unsaturated water signal was used as the internal standard. Regional cerebral blood flow in the infarcted area was measured relative to a symmetrically located unaffected area by means of single-photon emission computed tomographic scanning, using 99mTc-labeled d,l-hexamethylenepropyleneamine oxime as the flow tracer. Relative regional cerebral blood flow was considerably reduced in the infarcted area in the acute phase. After 1 week, hyperemia was seen in all but one patient. The N-acetylaspartate content was significantly reduced, with the loss appearing to occur between 6 and 24 hours after the stroke incident. The reduction in N-acetylaspartate content was greater in the central part than in the peripheral part of the infarcted area. Creatine and phosphocreatine were also reduced in the infarcted area, whereas no significant change was seen in the choline content. Assuming that N-acetylaspartate content reflects neuronal survival or loss, our results may suggest that treatment procedures with restoration of blood flow to severely ischemic areas should be initiated within the first 6 hours after stroke onset.


Imaging of the intracranial vertebrobasilar system using color-coded ultrasound. Anatomic variety and difficult accessibility of the vertebrobasilar arteries pose considerable problems to conventional ultrasound. We evaluated the diagnostic potential of transcranial color-coded sonography in the distal part of this system. We insonated the intracranial section of the vertebrobasilar arteries through the foramen magnum window in 24 healthy individuals using a Doppler color flow imaging system in connection with a 2.5-MHz sector transducer. Magnetic resonance images in special inclination planes were performed and compared with the color-coded duplex images in five cases. The B-mode image of the craniocervical junction and the intracranial parenchymal structures in addition to the color-coded blood flow allowed an unambiguous identification of the vertebrobasilar arteries (vertebral artery, 96%; basilar artery, 79%; and posterior inferior cerebellar artery, 50%). Blood flow velocities were measured considering the insonation angles: vertebral arteries, 50/24 cm/sec (30 degrees); basilar artery, 59/28 cm/sec (4 degrees); and posterior inferior cerebellar artery, 56/30 cm/sec (20 degrees) [peak systolic/end diastolic blood flow velocity (mean angle correction)]. Transcranial color-coded sonography enables accurate identification and differentiation of the intracranial vertebrobasilar arteries and improves accuracy of Doppler measurements. It may prove useful for evaluation of tortuosity and for hemodynamic studies in this vascular territory.


Risks and benefits of shunting in carotid endarterectomy. The International Transcranial Doppler Collaborators. Controversy continues about the pathogenesis of perioperative stroke in carotid endarterectomy and the use of shunting. The purpose of this study was to determine, using transcranial Doppler ultrasonography, the severity of ischemia during clamping of the carotid artery as a basis for analysis of complications in patients operated on with and without shunting. In a retrospective study, 11 centers contributed 1,495 carotid endarterectomies monitored with transcranial Doppler. The cases were divided into groups with severe, mild, and no ischemia, and each group was subdivided according to shunt use. The perioperative rate of severe stroke attributable to intraoperative ischemia, in addition to total perioperative stroke, was determined for each subgroup. Severe ischemia occurred in 7.2% of our cases but cleared spontaneously in about half of these. In those with persisting ischemia the rate of severe stroke was very high, while shunting protected against stroke in such cases. If ischemia did not occur, the stroke rate was higher with shunting, although not so high as in unshunted cases with severe ischemia. Slightly more than one third of the severe strokes were due to postoperative cerebral hemorrhage or carotid thrombosis, unrelated to clamp-induced ischemia or shunting. Carotid endarterectomy complications might be reduced by selectively shunting only for severe persisting ischemia. Monitoring of cerebral ischemia would be essential to selective shunting.


Prediction of cerebral ischemia by ophthalmoscopy after carotid occlusion in gerbils. The Mongolian gerbil provides a unique model of unilateral focal cerebral ischemia because of the lack of posterior communicating arteries in all gerbils as well as an absence of an anterior communicating artery in approximately 20% of the gerbil population. It is unclear how to identify unequivocably the subpopulation of animals that would suffer a severe focal cerebral ischemia after unilateral carotid occlusion. Ninety-three male gerbils were exposed to unilateral occlusion of the right common carotid artery. The severity of neuronal loss was evaluated histologically in gerbils selected as having significant focal ischemia based on either behavioral criteria (i.e., the demonstration of stereotypical behavior within 1 hour after occlusion) or ophthalmoscopic criteria (i.e., interruption of the retinal arterial perfusion within 10 minutes of carotid ligation as assessed with an ophthalmoscope). After 3 hours of unilateral carotid occlusion, cerebral blood flow was reinstated for 24 hours before fixation for histological analysis. The viability of the CA1 region of the hippocampus, lateral cortex, and medial cortex was scored on a scale of 0-4 based on the percentage of apparent neuronal loss (e.g., 0, no damage; 4, > 75% damage (the Viability Index). Twenty-eight percent of the gerbils met the behavioral selection criteria, and 17% met the ophthalmoscopic criteria. In the specimens selected by behavioral criteria (n = 7), 30% demonstrated no evidence of postischemic neuronal loss; the mean +/- SEM Viability Index scores for CA1, lateral cortex, and medial cortex were 1.6 +/- 0.6, 1.0 +/- 0.3, and 0.3 +/- 0.2, respectively. Of the animals selected by ophthalmoscopic criteria (n = 12), 100% had severe ischemic tissue damage to the ipsilateral hemisphere; the Viability Index scores for CA1, lateral cortex, and medial cortex were 3.5 +/- 0.1, 3.1 +/- 0.2, and 1.2 +/- 0.2, respectively; all scores were significantly larger than those observed in the behaviorally selected group. Selection of animals by ophthalmoscopic criteria provides a reliable, consistent method to predict animals with severe focal cerebral ischemia.


Diffusion-weighted imaging studies of cerebral ischemia in gerbils. Potential relevance to energy failure. Diffusion-weighted magnetic resonance imaging has been shown to be particularly suited to the study of the acute phase of cerebral ischemia in animal models. The studies reported in this paper were undertaken to determine whether this technique is sensitive to the known ischemic thresholds for cerebral tissue energy failure and disturbance of membrane ion gradients. Diffusion-weighted images of the gerbil brain were acquired under two sets of experimental conditions: as a function of cerebral blood flow after controlled graded occlusion of the common carotid arteries (partial ischemia), as a function of time following complete bilateral carotid artery occlusion (severe global ischemia), and on deocclusion after 60 minutes of ischemia. During partial cerebral ischemia, the diffusion-weighted images remained unchanged until the cerebral blood flow was reduced to 15-20 ml.100 g-1.min-1 and below, when image intensity increased as the cerebral blood flow was lowered further. This is similar to the critical flow threshold for maintenance of tissue high-energy metabolites and ion homeostasis. After the onset of severe global cerebral ischemia, diffusion-weighted image intensity increased gradually after a delay of approximately 2.5 minutes, consistent with complete loss of tissue adenosine triphosphate and with the time course of increase in extracellular potassium. This hyperintensity decreased on deocclusion following 60 minutes of ischemia. The data suggest that diffusion-weighted imaging is sensitive to the disruption of tissue energy metabolism or a consequence of this disruption. This raises the possibility of imaging energy failure noninvasively. In humans, this could have potential in visualizing brain regions where energy metabolism is impaired, particularly during the acute phase following stroke.


Cerebral vasoconstriction in response to hypocapnia is maintained after ischemia/reperfusion injury in newborn pigs. Hypocapnic cerebral vasoconstriction is used therapeutically to reduce elevated intracranial pressure caused by cerebral edema. Because cerebral ischemia/reperfusion injury causes a selective loss of prostanoid-dependent responses, including vasodilation to hypercapnia, we designed these experiments to examine the effect of ischemia/reperfusion on hypocapnic cerebral vasoconstriction. Microvascular responses were studied in 10 newborn pigs (closed cranial window) in response to hyperventilation-induced hypocapnia (PaCO2, 22 +/- 2 mm Hg) both before and 45 minutes after 20 minutes of global cerebral ischemia. Responses to hypercapnia (PaCO2, 63 +/- 3 mm Hg), topical isoproterenol (10(-7) M), and norepinephrine (10(-4) M) were also studied before and after ischemia in the same animals for comparison. Before ischemia/reperfusion, pial arterioles vasoconstricted to hypocapnia (-17 +/- 2%) and norepinephrine (-35 +/- 4%) and vasodilated to CO2 (37 +/- 7%) and isoproterenol (25 +/- 2%). After ischemia/reperfusion, the constriction of pial arterioles to hypocapnia (-19 +/- 2%) was similar to that before ischemia. This is in contrast to the loss of dilation to hypercapnia. Dilation to isoproterenol and constriction to norepinephrine were not affected by ischemia. Hypocapnic cerebral vasoconstriction is maintained after ischemia/reperfusion. Since prostanoid-dependent responses, such as hypercapnic dilation, are lost following cerebral ischemia, these data suggest that hypocapnic constriction is not dependent on an intact prostanoid system and that cerebral vascular responses to CO2 involve multiple mechanisms, depending on whether CO2 is increasing or decreasing from baseline.


Fructose-1,6-bisphosphate stabilizes brain intracellular calcium during hypoxia in rats. Exogenously administered fructose-1,6-bisphosphate reduces neuronal injury from hypoxic or ischemic brain insults. To test the hypothesis that fructose-1,6-bisphosphate prevents changes in intracellular calcium ([Ca2+]i) and high-energy phosphate levels, we measured [Ca2+]i, intracellular pH (pHi), and adenosine triphosphate in cultured rat cortical astrocytes and cortical brain slices during hypoxia. The fluorescent indicators fura-2 and bis-carboxyethyl-carboxyfluorescein were used to simultaneously measure [Ca2+]i and pHi with a fluorometer. Exposure to hypoxia (95% N2, 5% CO2) or 100 microM sodium cyanide produced transient increases in [Ca2+]i in astrocytes and sustained increases in [Ca2+]i in brain slices. Adenosine triphosphate levels fell in slices exposed to hypoxia or cyanide. Fructose-1,6-bisphosphate (3.5 mM) blocked increases in [Ca2+]i and prevented depletion of adenosine triphosphate. Fructose-1,6-bisphosphate also partially prevented adenosine triphosphate depletion in brain slices incubated in glucose-free medium. Iodoacetate (a specific inhibitor of glycolysis) elevated [Ca2+]i and partially prevented these actions of fructose-1,6-bisphosphate. Changes in pHi during hypoxia were not affected by fructose-1,6-bisphosphate. Fructose-1,6-bisphosphate supports adenosine triphosphate production via stimulation of glycolysis and results in the maintenance of normal [Ca2+]i during hypoxia or hypoglycemia.


Effects in cats of inhibition of nitric oxide synthesis on cerebral vasodilation and endothelium-derived relaxing factor from acetylcholine. We investigated the chemical identity of the endothelium-derived relaxing factor generated by acetylcholine in cerebral microvessels by studying the effects and mechanism of action of inhibitors of nitric oxide synthesis from arginine on the vasodilation and endothelium-derived relaxing factor production induced by topical application of acetylcholine in cerebral arterioles. We determined cerebral arteriolar dilation and endothelium-derived relaxing factor production by bioassay in anesthetized cats equipped with cranial windows during superfusion of 10(-7) M acetylcholine before and after administration of either NG-monomethyl L-arginine or NG-nitro-L-arginine, two inhibitors of nitric oxide synthesis. NG-Nitro-L-arginine abolished the vasodilation from acetylcholine and eliminated the production of endothelium-derived relaxing factor in the bioassay experiments. NG-Monomethyl L-arginine had no effect on the response to acetylcholine in the absence of pretreatment. However, after pretreatment with the detergent sodium dodecyl sulfate to increase cell membrane permeability, the inhibitor had effects identical to those of NG-nitro-L-arginine. L-Arginine reversed the effects of the inhibitors of nitric oxide synthesis. Neither inhibitor affected baseline vascular caliber, nor did they generate a vasoconstrictor agent in the bioassay experiments. The two inhibitors of nitric oxide synthesis did not affect the response to nitroprusside or adenosine, showing that the effect on responses to acetylcholine was specific. Also, the blockade of the response to acetylcholine induced by the inhibitors of nitric oxide synthesis was unaffected by treatment with superoxide dismutase and catalase, showing that the effect was not mediated by oxygen radicals. The endothelium-derived relaxing factor generated by acetylcholine in cerebral arterioles of cats is either nitric oxide or a nitric oxide-containing substance. The effect of these inhibitors on the response to acetylcholine is mediated by inhibition of the synthesis of nitric oxide. There is no involvement of radicals, and no vasoconstrictor agent is generated.


Neuroprotective effects of carvedilol, a new antihypertensive agent, in cultured rat cerebellar neurons and in gerbil global brain ischemia. Free radical generation mediates part of the ischemic neuronal damage caused by the excitatory amino acid glutamate. Carvedilol, a novel multiple-action antihypertensive agent, has been shown to scavenge free radicals and inhibit lipid peroxidation in swine heart and rat brain homogenates. Therefore, we studied the neuroprotective effect of carvedilol on cultured cerebellar neurons and on CA1 hippocampal neurons of gerbils exposed to brain ischemia. Neuroprotective mechanisms were studied using an in vitro ischemia model of cultured rat cerebellar granule cell neurons exposed to either glutamate or oxygen free radical-generating systems. Prevention of lipid peroxidation by carvedilol was studied by measuring the formation of thiobarbituric acid-reactive substance. Gerbil CA1 neuron survival was examined by direct neuronal count 7 days after 6 minutes of global ischemia with reperfusion. Carvedilol protected cultured neurons in a dose-dependent manner against glutamate-mediated excitotoxicity (inhibitory concentration [IC50] = 1.1 microM) as well as against a 20-minute oxidative challenge (IC50 = 5 microM). The IC50 against the oxidative challenge was lowered to 1.3 microM by growing neurons for 24 hours in the presence of carvedilol. At 10 microM carvedilol inhibited lipid peroxidation 50% and 73% (n = 4, p < 0.001) in neurons exposed to two different free radical-generating systems. Neuroprotection of 52% (n = 22, p = 0.009 versus vehicle) of gerbil CA1 hippocampal neurons was achieved by pretreatment and posttreatment with subcutaneous injection of 3 mg/kg carvedilol twice a day for 4 and 3 days, respectively. Carvedilol provided neuroprotection in both in vitro and in vivo models of neuroinjury, where oxygen radicals are likely to play an important role. Therefore, carvedilol may reduce the risk of cerebral ischemia and stroke by virtue of both its antihypertensive action and its antioxidative properties.


Release of proinflammatory and prothrombotic mediators in the brain and peripheral circulation in spontaneously hypertensive and normotensive Wistar-Kyoto rats. We reported previously that stroke risk factors prepared the brain stem for the development of ischemia and hemorrhage and induced the production of tumor necrosis factor following an intrathecal injection of lipopolysaccharide, a prototypic monocyte-activating stimulus. This study evaluates whether blood or brain cells of hypertensive rats produce more proinflammatory and prothrombotic mediators than do blood or brain cells of normotensive rats. Levels of tumor necrosis factor, platelet-activating factor, 6-ketoprostaglandin F1 alpha, and thromboxane B2 in the cerebrospinal fluid and blood of spontaneously hypertensive and normotensive Wistar-Kyoto rats were monitored before and after a challenge with lipopolysaccharide. Little or no activity from these mediators was found in the cerebrospinal fluid or blood of saline-injected control animals. Intravenous administration of lipopolysaccharide (0.001, 0.1, and 1.8 mg/kg) produced dose-dependent increases in blood levels of all mediators in hypertensive rats. In normotensive rats the levels were less than in hypertensive rats and were not clearly dose-related. When lipopolysaccharide was injected intracerebroventricularly, more tumor necrosis factor was measured in the cerebrospinal fluid than in the blood, suggesting local synthesis of this cytokine. Levels of tumor necrosis factor and platelet-activating factor in the cerebrospinal fluid were higher in hypertensive than in normotensive rats. The thromboxane A2/prostacyclin ratio was not altered significantly between the two rat strains. It is suggested that the higher incidence of brain stem ischemia and hemorrhage after the intrathecal injection of lipopolysaccharide in hypertensive rats than in normotensive rats might be related to the higher levels of the two cytotoxic factors tumor necrosis factor and platelet-activating factor produced in response to such challenge.


Detection of intracranial emboli in patients with symptomatic extracranial carotid artery disease. Cerebral embolism from extracranial sources is an important cause of ischemic stroke. The purpose of this limited study using long-term transcranial Doppler ultrasonographic monitoring was to estimate the frequency of clinically silent intracranial embolisms in patients with symptomatic extracranial carotid artery disease. By means of a 2-MHz pulsed-wave transcranial Doppler instrumentation, three consecutive patients with extracranial internal carotid artery stenosis (n = 2) or occlusion (n = 1) and recurrent ipsilateral ischemic events were monitored (19 hours total recording time). In addition, 10 control subjects without cerebrovascular disease were studied (25 hours total recording time). Formed-element emboli were defined as distinct signals within the fast Fourier-transform Doppler spectrum that were < 70 msec in duration and > 9dB greater in intensity than the background signal. Clinically silent formed-element embolism of ophthalmic or cerebral arteries was demonstrated in all three patients. Embolic events occurred only in the territory of the symptomatic internal carotid artery. The average rate of cerebral embolization at transcranial Doppler ultrasonography was 4.1/hr, with a mean signal duration of 47 msec. No emboli were found in control subjects. The observed high frequency of silent embolism of the intracranial arteries detected by transcranial Doppler monitoring in patients with recurrently symptomatic extracranial carotid artery disease should encourage studies of the prognostic and therapeutic implications of this method.


Irish brogue after stroke.

Irish brogue after stroke. We describe a patient who presented a unique variation of the previously described acquired foreign accent syndrome. A 65-year-old women developed an Irish brogue immediately after a deep left hemisphere stroke. The newly accented speech possibly represented a previously learned speech pattern. Suppressed prosodic speech patterns may reemerge in the setting of brain injury.


Bilateral medial medullary infarction with oculomotor disorders. We describe the first case of bilateral medial medullary infarction demonstrated by magnetic resonance imaging. We discuss the relation between this lesion and the oculomotor signs that were observed clinically. A 71-year-old man initially presented with pure motor hemiparesis, which progressed to complete quadriplegia. He also developed nearly complete vertical and horizontal ophthalmoplegia. Magnetic resonance imaging revealed upper medial medullary infarctions bilaterally that extended to the pontomedullary junction. We propose that the vertical oculomotor disorders resulted from involvement of the oculomotor system in the caudal brain stem, especially the caudal paramedian pontine reticular formations on both sides.


The role of transesophageal echocardiography in the acute onset of paraplegia. Acute paraplegia must be investigated promptly to exclude reversible causes. In this report we illustrate the usefulness of transesophageal echocardiography in identifying the vascular etiologies of acute paraplegia. Two patients presented with acute paraplegia, one spontaneously and the other after removal of an intra-aortic balloon pump catheter. Through the use of transesophageal echocardiography, we excluded aortic dissection and identified protruding atherosclerotic plaques in the descending thoracic aorta of each patient. Embolization of atheromatous material from the thoracic aorta was considered the most likely etiology of paraplegia in both cases. Embolization from atherosclerotic plaques in the thoracic aorta may be an underestimated cause of acute paraplegia. Transesophageal echocardiography provides a safe, rapid, and reliable tool for investigating a vascular etiology of acute paraplegia.


Is cerebral angiography indicated in infective endocarditis? Patients with infective endocarditis may develop intracranial mycotic aneurysms. Whether these patients should undergo cerebral angiography followed by prophylactic surgery if an aneurysm is detected is an unresolved question. We estimated the probability of survival 12 weeks after the diagnosis of infective endocarditis on the basis of data available in the literature. For a 40-year-old female patient with right-sided hemiplegia, the 12-week survival is estimated to be 83.75% without angiography and 83.65% with angiography; the specific mortality of intracranial mycotic aneurysms is relatively small but increases by 40% (from 0.25% to 0.35%) if angiography is performed. The risk of aneurysm rupture in infective endocarditis and the mortality from rupture appear to be the most important factors that affect the analysis. Cerebral angiography should not be performed routinely in patients with infective endocarditis. Specific subgroups in whom such a policy might be beneficial have not yet been identified.


Experimental evidence of ischemic thresholds and functional recovery. Impaired blood flow below certain critical levels and an insufficient supply of energy-rich substrates cause failure of neuronal function, triggering biochemical disturbances that eventually lead to ischemic cell damage. This article reviews experimental studies that attempted to define those ischemic thresholds using functional and histological markers and, more recently, chemical markers of ischemic damage. The duration of ischemia causing irreversible cell damage still is ill defined. Reestablishment of sufficient perfusion must be induced very early after an ischemic attack to ameliorate the potentially harmful biochemical sequelae of transient ischemia.


Leukoaraiosis and dementia in hypertensive patients. Although our previous study demonstrated that dementia of the Binswanger type may be a disconnection dementia caused by leukoaraiosis, some hypertensive patients with marked leukoaraiosis do not develop dementia. The goal of the present study is to elucidate the pathophysiology of nondemented hypertensive patients with leukoaraiosis. We performed clinical and neuroradiological studies, including positron emission tomography, in eight hypertensive patients with leukoaraiosis. Four patients were demented, and two among the other four who were not demented at the first examination developed dementia during the follow-up period. Digital subtraction angiography of the cervical and intracranial arteries demonstrated stenotic lesions in only one patient. Cerebral blood flow and oxygen metabolism in patients with dementia were markedly reduced in the white matter (59-67% of control values). In contrast, cerebral blood flow in the white matter of patients without dementia was reduced less markedly (74% of control), oxygen extraction fraction in the white matter was significantly increased (130% of control), and oxygen metabolism remained at almost-normal levels not only in the white matter but also in the cortical area. Hypertension-caused arteriosclerotic changes of the long penetrating medullary arteries may cause misery perfusion and later ischemic damage in the periventricular white matter. Preserved oxygen metabolism in hypertensive patients with leukoaraiosis may represent the early stage of vascular dementia of the Binswanger type.


[The nature of the deposit of substances in gunshot injuries from a smooth-bore hunting rifle]. Direct microscopic examinations of plain-barrel hunting gun-shot wounds may reveal on the victim's skin or clothes starch or paraffin particles or impressions thereof (a secondary shell trace). These data may evidence a specific charge of the cartridge and be used to specify the distance from which the shot was made, with an obligatory experimental shot made in each case.


[The forensic medical expertise of the victims of large-scale fires]. The difficulties of present-day forensic medical expert evaluation of poisonings in fires are explained by exposure of man to highly toxic compounds that form during burning of polymers. Analyzing forensic medical expert evaluations of a large number of victims dead in a large-scale fire, the authors emphasize the necessity of measuring not only dead people's blood levels of HbCO, but of some other toxic combustion products that may exert combined effects on human body. The authors think it desirable that criteria of forensic medical evaluation of combined poisoning in fire be defined.


[The combination of mixed agglutination and immunofluorescence reactions for the simultaneous detection of 2 ABO-system antigens in isolated cells]. A combination of two immunologic tests, mixed agglutination and immunofluorescence, is suggested, that permits the detection of two AB0 system antigens in the same cell within one process. Such mode better validates the conclusion on the group appurtenance of cells, for instance, in analyses of mixed stains originating from different subjects with different blood groups.


[The use of immunoenzyme analysis for determining ABH antigens in traces of saliva and sperm]. The authors suggest a simple sensitive technique for enzyme immunoassay (EIA) of ABH antigens in saliva and semen. A two-staged dot blot solid-phase EIA on nitrocellulose membranes was employed with anti-ABH monoclonal antibodies obtained in immunization of mice with human red cells. 4-chloro-1-naphthol substrate solution was used to visualize the peroxidase label. The results of analysis of salivary and spermatic samples obtained from donors of various groups evidence that this EIA variant may be useful in forensic medicine.


[The use of trace deposits for establishing the weapon of an injury]. The author analyzes the value of traces detectable on stabbing and cutting instruments (traces of blood, isolated cells of the injured organs and tissues of human body, trace particles of textile fabrics) for the identification of the instrument of injury. A comprehensive assessment of the fact of detection of material traces and of the specific features of injuries of victim's body and clothes helps in some cases identify the instrument of injury.


[The determination of phoxim in cadaveric material by gas-liquid chromatography]. The developed gas-liquid chromatographic procedure for forensic chemical detection and measurement of phoxim in cadaveric material permits the detection in 25 g of an organ of 44 to 52% of added phoxim. The threshold of phoxim measurement in 100 g of the liver is 0.04 mg, the least volume of phoxim detectable being 0.02 g. The developed gas-liquid chromatographic technique should be used as an additional method together with thin-layer chromatography, for a more objective evaluation of the results of forensic chemical analysis of cadaveric material for phoxim.


Presuicide attempt communications between parasuicides and consulted caregivers. A high percentage of parasuicides visit professional caregivers prior to the attempted suicide. The content or outcome of these consultations is unknown. We interviewed hospitalized attempters and the professional caregivers they identified as having been consulted prior to their attempts. About half of these patients directly disclosed suicidal symptoms or intentions, especially to mental health professionals. These professionals more often inquired about suicidal ideations than did nonpsychiatric physicians. However, few caregivers noted suicidal thinking or probed suicidal symptoms. The data suggest that professional caregivers and especially nonpsychiatric physicians should be more sensitive and responsive to the signs and symptoms of suicidality.


Intensive follow-up does not decrease the risk of repeat suicide attempts. We carried out a randomized controlled trial to determine whether an intensive intervention after a suicide attempt could decrease by half the risk of a repeat attempt in the following two years. After initial assessment and randomization, experimental subjects attended 18 therapy appointments over one year, including one home visit, and measures to improve attendance. Control subjects received the usual care. Of 63 experimental subjects, 35% made a repeat attempt, and so did 30% of 63 control subjects. The study had a 99% power to detect the desired decrease of risk (30% to 15%). Clearly, the intervention did not achieve its objective.


Adolescents' experience with and response to suicidal peers. The present study investigated a sample of 325 suburban high school students' knowledge of suicidal peers, whether they had ever talked to a suicidal peer, and, if so, what they actually did in that situation. Sixty-eight percent of the females and 42.5% of the males reported knowing a teen who had committed or attempted suicide. Ninety-seven students reported having talked to a peer who was definitely considering suicide; of these, 63% talked to their peer about his or her concerns, 24.7% told an adult, and 12% did nothing in response to the encounter. Ninth graders were significantly more likely to do nothing as compared to eleventh graders. A mixed pattern of results was found as to the relationship of the response of youth to suicidal peers and their general experience with suicidal peers. The results confirm the importance of adolescents themselves for the prevention of youth suicide, and the need to convince adolescents to report at-risk peers to an adult.


[The cost price of expertise as an objective index of the work-share coefficient of the forensic medical expert]. The author suggests a method for estimating the net cost of a forensic medical expert investigation, consisting in calculation of an objective real coefficient of the ratio of various expert investigations to the reference value. The ratio between forensic medical investigation budget and the total number of these investigations forms the basis for the estimation of the net cost of an expert evaluation.