Fumonisin- and AAL-Toxin-Induced Disruption of Sphingolipid Metabolism with Accumulation of Free Sphingoid Bases.

Fumonisins (FB) and AAL-toxin are sphingoid-like compounds produced by several species of fungi associated with plant diseases. In animal cells, both fumonisins produced by Fusarium moniliforme and AAL-toxin produced by Alternaria alternata f. sp. lycopersici inhibit ceramide synthesis, an early biochemical event in the animal diseases associated with consumption of F. moniliforme-contaminated corn. In duckweed (Lemna pausicostata Heglem. 6746), tomato plants (Lycopersicon esculentum Mill), and tobacco callus (Nicotiana tabacum cv Wisconsin), pure FB1 or AAL-toxin caused a marked elevation of phytosphingosine and sphinganine, sphingoid bases normally present in low concentrations. The relative increases were quite different in the three plant systems. Nonetheless, disruption of sphingolipid metabolism was clearly a common feature in plants exposed to FB1 or AAL-toxin. Resistant varieties of tomato (Asc/Asc) were much less sensitive to toxin-induced increases in free sphinganine. Because free sphingoid bases are precursors to plant "ceramides," their accumulation suggests that the primary biochemical lesion is inhibition of de novo ceramide synthesis and reacylation of free sphingoid bases. Thus, in plants the disease symptoms associated with A. alternata and F. moniliforme infection may be due to disruption of sphingolipid metabolism.

Antagonist activities of mecamylamine and nicotine show reciprocal dependence on beta subunit sequence in the second transmembrane domain.

We show that a portion of the TM2 domain regulates the sensitivity of beta subunit-containing rat neuronal nicotinic AChR to the ganglionic blocker mecamylamine, such that the substitution of 4 amino acids of the muscle beta subunit sequence into the neuronal beta4 sequence decreases the potency of mecamylamine by a factor of 200 and eliminates any long-term effects of this drug on receptor function. The same exchange of sequence that decreases inhibition by mecamylamine produces a comparable potentiation of long-term inhibition by nicotine. Inhibition by mecamylamine is voltage-dependent, suggesting a direct interaction of mecamylamine with sequence elements within the membrane field. We have previously shown that sensitivity to TMP (tetramethylpiperidine) inhibitors is controlled by the same sequence elements that determine mecamylamine sensitivity. However, inhibition by bis-TMP compounds is independent of voltage. Our experiments did not show any influence of voltage on the inhibition of chimeric receptors by nicotine, suggesting that the inhibitory effects of nicotine are mediated by binding to a site outside the membrane's electric field. An analysis of point mutations indicates that the residues at the 6' position within the beta subunit TM2 domain may be important for determining the effects of both mecamylamine and nicotine in a reciprocal manner. Single mutations at the 10' position are not sufficient to produce effects, but 6' 10' double mutants show more effect than do the 6' single mutants.

An overview of rodent toxicities: liver and kidney effects of fumonisins and Fusarium moniliforme.

Fumonisins are produced by Fusarium moniliforme F. verticillioides) and other Fusarium that grow on corn worldwide. They cause fatal toxicoses of horses and swine. Their effects in humans are unclear, but epidemiologic evidence suggests that consumption of fumonisin-contaminated corn contributes to human esophageal cancer in southern Africa and China. Much has been learned from rodent studies about fumonisin B1(FB1), the most common homologue. FB1 is poorly absorbed and rapidly eliminated in feces. Minor amounts are retained in liver and kidneys. Unlike other mycotoxins, fumonisins cause the same liver cancer promotion and subchronic (studies (3/4) 90 days) liver and kidney effects as (italic)F. moniliforme. FB 1 induces apoptosis of hepatocytes and of proximal tubule epithelial cells. More advanced lesions in both organs are characterized by simultaneous cell loss (apoptosis and necrosis) and proliferation (mitosis). Microscopic and other findings suggest that an imbalance between cell loss and replacement develops, a condition favorable for carcinogenesis. On the molecular level, fumonisins inhibit ceramide synthase, and disrupt sphingolipid metabolism and, theoretically, sphingolipid-mediated regulatory processes that influence apoptosis and mitosis. Liver sphingolipid effects and toxicity are correlated, and ceramide synthase inhibition occurs in liver and kidney at doses below their respective no-observed-effect levels. FB1 does not cross the placenta and is not teratogenic in vivoin rats, mice, or rabbits, but is embryotoxic at high, maternally toxic doses. These data have contributed to preliminary risk evaluation and to protocol development for carcinogenicity and chronic toxicity studies of FB1 in rats and mice.

Open window thoracostomy and plastic surgery with muscle flaps in the treatment of chronic empyema.

Open window thoracostomy (OWT) and plastic surgery using myocutaneous flaps of extrathoracic muscles, was done in nine patients with chronic empyema, after conventional therapy had failed. In all these patients, the pleural cavity and bronchopleural fistulas, when present, closed within eight to 61 days (mean 30), after the second stage of surgery comprising the myoplasty; this in turn was performed between three to seven (mean 3.0) months after the OWT. All patients were considered cured with a minimal deformity of the chest wall. The final scar had an acceptable cosmetic appearance.

Inhibition of protein synthesis in rat primary hepatocytes by extracts of Fusarium moniliforme-contaminated corn.

Mycological screening of two separate lots of corn samples that caused field cases of equine leukoencephalomalacia (ELEM) revealed heavy contamination with the fungus Fusarium moniliforme. Neutral and acidic fractions of a chloroform-methanol (1:1, v/v) extract of the corn were evaluated for toxicity using rat primary hepatocytes. The extracts had little effect on the release of lactate dehydrogenase from the hepatocytes, and were without effect on unscheduled DNA synthesis, indicating low cell lethality and lack of genotoxicity. However, neutral extracts of the corn were found to contain potent inhibitor(s) of protein synthesis as measured by incorporation of [3H]valine into the hepatocytes. When an isolate of F. moniliforme obtained from the corn samples or an isolate of F. moniliforme from South Africa that had previously been shown to cause ELEM (MRC 826) were grown on autoclaved seed corn, neutral extracts of the culture materials similarly inhibited protein synthesis. Whether the compound(s) responsible for inhibition of protein synthesis is associated with any of the toxic syndromes associated with F. moniliforme remains to be determined. The use of primary hepatocytes may be a useful bioassay for elucidating biologically active secondary metabolites of fungi.

Fusaric acid and modification of the subchronic toxicity to rats of fumonisins in F. moniliforme culture material.

Fumonisins and fusaric acid (FA) are mycotoxins produced by Fusarium moniliforme and other Fusarium which grow on corn. Fumonisins cause animal toxicities associated with F. moniliforme and, like F. monliforme, they are suspected human oesophageal carcinogens. Toxic synergism was obtained by simultaneous administration of FA and fumonisin B1 to chicks in ovo. To determine the effect of FA on in vivo toxicity of F. moniliforme culture material (CM), male rats (12 groups, n = 5/ group) were fed diets containing 0.025, 0.10 or 2.5% CM (providing dietary levels of 3.4, 18.4 or 437 ppm fumonisins, respectively) to which, at each CM level, 0, 20, 100 or 400 ppm FA were added. Additionally, an FA control group was fed 400 ppm FA only and an untreated control group was given neither FA nor culture material. Apoptosis and other effects consistent with those caused by fumonisins were present in the kidneys of animals fed 0.025% or more CM and in the livers of animals fed 2.5% CM. FA was without effect. No differences between the untreated and FA control groups were noted and no differences among the four groups (0-400 ppm FA) fed 0.025% CM, the four groups fed 0.10% CM or the four groups fed 2.5% CM were apparent. Thus, FA exerted no synergistic, additive or antagonistic effects on the subchronic in vivo toxicity of fumonisin-producing F. moniliforme.

Effects of Fusarium mycotoxins on levels of serotonin, melatonin, and 5-hydroxytryptophan in pineal cell cultures.

Analysis of melatonin (MEL) in pineal cell cultures by enzyme linked immunosorbent assay showed its concentration was increased by fusaric acid (FA), a mycotoxin produced by Fusarium species and associated with toxic duck and ostrich feeds. Subsequent cell culture studies demonstrated the precursors of MEL, 5-hydroxytryptophan (5HTP) and serotonin (5HT), were also affected by FA as well as other Fusarium mycotoxins. Herein we describe a technique for the analysis of 5HTP and 5HT in pineal cell cultures using HPLC with electrochemical detection (EC), and report on the effects of FA alone and in combination with fumonisin B1 (FB1) and deoxynivalenol (DON) on the levels of these MEL precursors.

Use of drugs during cardiopulmonary resuscitation: a review of the literature.

The use of drugs during cardiopulmonary resuscitation is often a crucial factor in saving the lives of patients suffering cardiac arrest. However, inappropriate use of these agents may produce deleterious effects that may increase morbidity and adversely affect the outcome of resuscitative measures. It is imperative to realize that cardiac arrest is a frank disease entity with very specific signs and symptoms deserving specific and precise treatment.

Analysis of endophyte toxins: fescue and other grasses toxic to livestock.

Research on livestock toxicoses caused by Acremonium (endophyte)-infected grasses strongly implicate the ergopeptine alkaloids with A. coenophialum-infected fescue and paxilline and the lolitrem alkaloids with A. lolii-infected perennial ryegrass as the causative agents. Isolation, identification, and detection of these toxins involves extraction with appropriate solvents, clean-up procedures, and chromatographic methods with known standards. Thin-layer, high-performance liquid and gas chromatography along with ultraviolet and mass spectrometric (i.e., electron impact, chemical ionization, tandem mass) characterizations have been reported. These methods have varying degrees of success depending on the matrix from which the alkaloids have been extracted. Ergovaline is the primary ergopeptine alkaloid isolated from cultures of A. coenophialum and also from infected fescue grass and seeds toxic to livestock. Other compounds isolated from the endophyte-infected fescue include: lysergic acid amide (ergine), the clavine class of ergot alkaloids (chanoclavine I, agroclavine, elymoclavine, penniclavine), the pyrrolizidine alkaloids (N-formylloline, N-acetylloline, N-methyloline, N-acetylnorloline), and the unique pyrrolopyrazine alkaloid peramine. The loline alkaloids and peramine have been more associated with the insect-deterrent properties of the endophyte-infected fescue than with livestock toxicoses. Also, both peramine and the ergopeptine alkaloids (ergovaline, ergotamine) have been isolated from A. lolii-infected perennial ryegrass. More recently, paxilline and lolitrem B have been detected in laboratory cultures of A. coenophialum isolated from tall fescue. The ergot alkaloids in endophyte-infected perennial ryegrass may be more related to decreased animal productivity (weight gains, reproduction problems), whereas the lolitrems cause the staggers syndrome. The detection, isolation, identification, and analyses of these compounds from Acremonium-infected grasses is presented.

Effects of fescue toxicosis on reproduction in livestock.

Fescue toxicosis in livestock is due to ingestion of endophyte (Acremonium coenophialum) -infected tall fescue. Understanding mechanisms responsible for decreased calving and growth rates, delayed onset of puberty, and impaired function of corpora lutea in heifers at puberty consuming endophyte-infected fescue is an emerging field in reproductive toxicology. The condition decreases overall productivity through a reduction in reproductive efficiency, reduced weight gains, and lowered milk production. Reproduction in cattle may be further compromised by winter coat retention, increased susceptibility to high environmental temperatures, and light intolerance. Endocrine effects in steers associated with infected tall fescue include reduced prolactin and melatonin secretions and altered neurotransmitter metabolism in the hypothalamus, the pituitary, and pineal glands. Ewes have decreased prolactin and lengthened intervals from introduction of the ram until conception. The endophyte induces prolonged gestation, thickened placentas, large, weak foals, dystocia, and agalactia in pregnant mares. Ergot peptide alkaloids, produced by the endophyte, are suggested as the primary cause of fescue toxicosis. These compounds reduce prolactin, increase body temperatures, and have powerful vasoconstrictive effects. Neurohormonal imbalances of prolactin and melatonin, with restricted blood flow to internal organs, may be the principal causes of aberrant reproduction, growth, and maturation in livestock consuming endophyte-infected tall fescue.

Neuroendocrine measurements in steers grazed on endophyte-infected fescue.

Dopamine (DA), serotonin (5HT) and selected precursors and metabolites were measured in the anterior pituitary gland, hypothalamus and pineal gland, along with serum prolactin (PRL) and average daily gains (ADG), in steers (n = 6/group) grazing endophyte (Acremonium coenophialum)-infected and noninfected fescue (Festuca arundinacea Schreb). Paddocks (two/treatment) were designated 100F and 0F (100 and 0% infection, respectively). After 6 wk, three animals from one of the 100F paddocks were exchanged with three animals from one of the 0F paddocks, yielding 0F, 100F/0F, 0F/100F and 100F groups (n = 3). Compared to 0F steers, 100F steers had reduced serum PRL (9.23 vs 32.55 ng/ml, P less than or equal to .0001) and trial ADG (-.07 vs .28 kg, P less than or equal to .0002) but increased pituitary dihydroxyphenylacetic acid (DOPAC, a major metabolite of DA; 108 vs 59 ng/g, P less than or equal to .02) and 5-hydroxyindoleacetic acid (5HIAA, a major metabolite of 5HT; 265 vs 148 ng/g, P less than or equal to .04). Pituitary 5HIAA was greater in the steers rotated from the 0F to 100F paddocks than in steers maintained on the 0F paddocks (296 vs 148 ng/g, P less than or equal to .04). In addition pineal 5-hydroxytryptophan (5HTP, a precursor of 5HT) was increased (502 vs 280 ng/ml; P less than .08), whereas 5-methoxyindoleacetic acid (MIAA, a major metabolite of 5HIAA) and the 5HT/5HTP ratio were reduced (P less than .07) in 100F vs 0F steers. No differences among the treatment groups were observed in hypothalamic neurotransmitter and metabolite concentrations.(ABSTRACT TRUNCATED AT 250 WORDS)

Melatonin and pineal neurochemicals in steers grazed on endophyte-infected tall fescue: effects of metoclopramide.

Plasma and pineal melatonin (MEL) and selected pineal neurochemicals (5-hydroxytryptophan, serotonin, N-acetylserotonin, dopamine, norepinephrine) associated with MEL synthesis were determined in steers grazing Acremonium coenophialum (endophyte)-infected 'Kentucky-31' tall fescue paddocks. Paddock treatments included low (LE, 33%) or high (HE, 74%) endophyte at either low (134 kg.ha-1 x yr-1, LN) or high (335 kg.ha-1 x yr-1, HN) nitrogen fertilization. Twelve pairs of yearling Angus steers were randomly assigned to three replications of the paddock treatments (LEHN, HEHN, LELN, and HELN). One steer in each of the 12 paddocks received per os either a dopamine antagonist, metoclopramide (MC; 15 mg/kg), or sucrose (S; 15 mg/kg) three times weekly for 10 wk. Blood was collected via jugular cannulas during the day and night for plasma MEL analysis and pineal glands were collected at termination. Day and night plasma MEL in the S/HEHN steers was reduced by 31.7 and 35.4% (P < .05), respectively, compared with that in S/LEHN steers. Mean night plasma MEL in the S/HELN steers was reduced by 26.7% (P < .05) compared with that in S/LELN steers. Metoclopramide reduced mean day and night plasma MEL by 22.9 and 38.3% (P < .05), respectively, in the LEHN steers and increased night MEL in the HELN animals by 35.1% (P < .05). During the day and night, there was a MC x pasture treatment interaction (P < .05). No differences were observed in either pineal MEL or the pineal neurochemicals. Acremonium coenophialum-infected fescue reduced plasma concentrations of MEL in steers, whereas treatment with MC altered plasma MEL biphasically.(ABSTRACT TRUNCATED AT 250 WORDS)

Evaluation of sorghum ergot toxicity in broilers.

Three experiments evaluated the performance of broilers fed sorghum ergot consisting of sphacelia/sclerotia of Claviceps africana present in tailings removed by conditioning of seed from grain sorghum hybrid seed production fields near Uvalde (Experiments 1 and 2) and Dumas (Experiment 3), Texas. Percentage sphacelia/sclerotia and total alkaloid content, respectively, in sorghum ergot tailings were 8% and 11.3 ppm for Uvalde and 75% and 235 ppm for Dumas. Sorghum ergot and control sorghum diets were based on the NRC (1994) requirements for starting broilers. In Experiment 1, neither growth nor feed efficiency were significantly reduced in male broilers fed sorghum ergot from hatch to 3 wk of age, but liver weights were significantly greater than those in the control. In Experiment 2, straight-run broilers were raised to 6 wk of age in floor pens using a three-phase feeding program. Sorghum ergot significantly reduced gain in 4-wk-old broilers and cumulative body weight at 5 wk. Feed conversion was significantly reduced during all three phases of feeding. In Experiment 3, control sorghum and the 75% ergot tailings were added to corn-soy basal diets at rates of 2.5, 5, and 10% by weight and fed to male broilers from hatch to 3 wk of age. Sorghum ergot did not significantly reduce growth, but, during Weeks 2 and 3, feed-to-gain ratios were higher. Neither type nor concentration of sorghum ergot significantly affected relative liver weights. We did not observe significant mortality or obvious symptoms of ergot toxicity, such as necrotic lesions of the feet or vesicular dermatitis of the comb, in any of the three experiments.

[Ergot alkaloids and alkaloids from other endophytes, responsible for causing toxic syndrome in cattle after eating contaminated grass]. / Alakloidy sporyn'i i drugikh éndofitov, vyzyvaiushchie toksichnyi sindrom u skota pri pitanii zarazhennymi travami.

Different signs of ergotism in livestock associated with animals consuming feed grains and pasture grasses infected by Clavicipitaceae (e.g. Balansia spp. and Acremonium spp.) have been considered. Endophyte-infected tall fescue causes the damage to livestock, and its toxicity and ryegrass in livestock are associated with ergopeptide alkaloid ergovaline and indole-isoprenoid lolitrems produced by A. coenophialum and A. lolii. The role of another compound attendant to endophyte-infected fescue such as peramine and loline alkaloid is discussed. Concentration of alkaloids depends upon the season and the period of growing plant-host. The advantages and the problems of endophyte-grass mutualistic symbiosis in pasture conditions are considered. The perspective of genetic manipulation of grass and fungus to produce endophyte-infected grasses with stress and insect deterences without affecting ruminant health.