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T Yasumoto

Publications and source records attributed to T Yasumoto.

At least 19 recordsLinked to original sources
PubMed2006

Changes in membrane potential: an early signal triggered by neurologically active phycotoxins.

Most common phycotoxin poisoning syndromes have important neurological symptoms. However, little is known of the cellular and molecular targets of many of the phycotoxins that produce those human intoxications. We explore the effect of representative toxins on the membrane potential in human neuroblastoma cells by using a fluorimetric assay. Results presented in this study demonstrate that maitotoxin, palytoxins, brevetoxins, and ciguatoxins triggered a dose-dependent membrane depolarization. Mechanisms responsible for the toxins-induced changes in membrane potential are always related to a direct action of the compounds on membrane ion fluxes. This initial screening of the phycotoxins effect is the starting point to lately develop functional methods of detection.

Cell Line, Tumor↗
PubMed2005

Actin cytoskeleton of rabbit intestinal cells is a target for potent marine phycotoxins.

Biotoxins produced by harmful marine microalgae (phycotoxins) can be accumulated into seafood, representing a great risk for public health. Some of these phycotoxins are responsible for a variety of gastrointestinal disturbances; however, the relationship between their mechanism of action and toxicity in intestinal cells is still unknown. The actin cytoskeleton is an important and highly complicated structure in intestinal cells, and on that basis our aim has been to investigate the effect of representative phycotoxins on the enterocyte cytoskeleton. We have quantified for the first time the loss of enterocyte microfilament network induced by each toxin and recorded fluorescence images using a laser-scanning cytometer and confocal microscopy. Our data show that pectenotoxin-6, maitotoxin, palytoxin and ostreocin-D cause a significant reduction in the actin cytoskeleton. In addition, we found that the potency of maitotoxin, palytoxin and ostreocin-D to damage filamentous actin is related to Ca(2+) influx in enterocytes. Those results identify the cytoskeleton as an early target for the toxic effect of those toxins.

Acrylamides↗
PubMed2004

Short-term oral toxicity of homoyessotoxins, yessotoxin and okadaic acid in mice.

A short-term toxicity study after 7 days oral daily administration of yessotoxin (YTX; 2 mg/kg/day), homoYTX (1 mg/kg/day), 45-hydroxy-homoYTX (1 mg/kg/day) and of the main diarrhoetic shellfish toxin okadaic acid (OA; 1 mg/kg/day) was carried out in mice. Symptoms, lethality, food consumption, body and organ weights, gross pathology and histopathology of the main organs and tissues, leukocytes formula as well as plasmatic levels of transaminases, lactate dehydrogenase and creatinine phosphokinase were evaluated. Heart tissue was studied also hystochemically for the presence of apoptotic nuclei and by transmission electron microscopy. No mortality, signs of toxicity or cumulative effects were induced by the repeated oral exposure to YTXs. Only ultrastructural changes in the cardiac muscle cells near the capillaries, such as package of rounded mitochondria and alteration of the cells boundary were observed, without any increase of lactate dehydrogenase, an index of cardiac damage. OA induced diarrhoea, body weight loss, reduced food consumption, and the death of 2/5 mice after 5 days. Necroscopy and/or light microscopy analysis revealed toxic effects mainly at forestomach (ulceration and hyperplasia), liver and, indirectly to body weight loss of mice, atrophic signs in the lymphoid organs and exocrine pancreas. Electron microscopy of heart tissue showed alterations of mitochondria and fibers in myocardiocytes, although no apoptotic change was recorded.

Administration, Oral↗
PubMed2004

Detection of sodium channel activators by a rapid fluorimetric microplate assay.

Marine toxins such as brevetoxins and ciguatoxins are produced by dinoflagellates and can accumulate in seafood. These toxins affect humans through seafood consumption. Intoxication is mainly characterized by gastrointestinal and neurological disorders and, in most severe cases, by cardiovascular problems. To prevent the consumption of food contaminated with these toxins, shellfish have been tested by mouse bioassay. However, this method is expensive, time-consuming, and ethically questionable. The objective of this study was to use a recently developed fluorimetric microplate assay to rapidly detect brevetoxins and ciguatoxins. The method is based on the pharmacological effect of brevetoxins and ciguatoxins known to activate sodium channels and involves (i). the incubation of excitable cells in 96 well microtiter plates with the fluorescent dye bis-oxonol, whose distribution across the membrane is potential-dependent, and (ii). dose-dependent cell depolarization by the toxins. Our findings demonstrate that measuring changes in membrane potential induced by brevetoxins and ciguatoxins allowed their quantitation. Active toxins could be reliably detected at concentrations in the nanomolar range. The simplicity, sensitivity, and possibility of being automated provide the basis for development of a practical alternative to conventional testing for brevetoxins and ciguatoxins.

Animals↗
PubMed2003

Oral and intraperitoneal acute toxicity studies of yessotoxin and homoyessotoxins in mice.

The acute toxicity of yessotoxin (YTX), homoyessotoxin (homoYTX) and 45-hydroxy-homoyessotoxin (45-OH-homoYTX) has been studied in comparison to that of okadaic acid (OA), the main diarrhogenic toxin, both after intraperitoneal (i.p.) and oral administration. After i.p. administration, homoYTX and YTX showed similar lethality (LD(50)=444 microg/kg and 512 microg/kg), higher than that of OA (LD(50)=225 microg/kg), while 750 microg/kg of 45-OH-homoYTX did not cause death. OA induced the already known toxic signs: before death, mice were motionless and cyanotic; small intestine and liver damage were shown at post-mortem. Mice treated with YTX and homoYTX were restless and jumped before death; necroscopy did not show major changes. After oral treatment, 2 mg/kg of OA induced diarrhoea and body weight loss, causing 4/5 deaths; necroscopy and/or histology revealed degenerative lesions to small intestine, forestomach and liver (confirmed by increased plasma transaminase), but no myocardium alterations. On the contrary, the oral treatment with YTX (1 and 2 mg/kg) and its derivatives (1 mg/kg) did not cause any death or signs of toxicity, except some ultrastructural myocardiocyte alterations, adjacent to capillaries, such as cytoplasmic protrusions (YTX, 1 and 2 mg/kg), fibrillar alteration (YTX, 1 mg/kg) or mitochondria assemblage (45-OH-homoYTX). Altogether, our data show that YTX and its derivatives are less toxic than OA after acute oral and i.p. treatments, at doses which may represent up to 100 times of the possible human daily intake.

Administration, Oral↗
PubMed2003

The G1 domain of aggrecan released from porcine articular cartilage forms stable complexes with hyaluronan/link protein.

OBJECTIVE: To raise peptide antibodies recognizing the C-terminal amino acid sequence in the G1 domain of porcine aggrecan, generated by the action of either aggrecanase or neutral metalloproteinase(s), in rabbits and to use them to investigate the release of aggrecan from porcine articular cartilage. METHOD: An explant culture system was used to investigate the release of the G1 domain of aggrecan from porcine articular cartilage treated with retinoic acid or interleukin 1beta and to study how the activity of these agents is modified by the proteinase inhibitor, batimastat (BB94). RESULTS: Retinoic acid and interleukin 1beta induced both enzyme activities and the release of the G1 domain into the culture medium. Proteinase activity was significantly reduced when the tissue was incubated in the presence of BB94. The functional properties of the enzyme-generated G1 domain were studied using large-pore, agarose/polyacrylamide gel electrophoresis, and it was shown to interact with hyaluronan and link protein. CONCLUSIONS: The results show that there must be a mechanism for removing a functional G1 domain from aggrecan during tissue turnover using this culture system.

Aggrecans↗
PubMed2002

Comparison of oral and intraperitoneal toxicity of yessotoxin towards mice.

Currently, yessotoxin is regulated among the toxins in the diarrhetic shellfish poisoning (DSP) complex. Yessotoxin is equally acutely toxic towards mice upon intraperitoneal injections as those algal toxins giving diarrhea, but is not diarrheagenic. Its presence in mussels may therefore lead to overestimation of risk of DSP in consumers when the standard mouse bioassay is used. Arguments are presented for the use of analytical methods instead of the mouse bioassay for the diarrheagenic DSP toxins and yessotoxin. Yessotoxin was found to be more than ten times less toxic to mice via the oral route, compared with intraperitoneal injections. Even at 10mg/kg body weight, the highest dose ever tested orally, yessotoxin did not kill the mice. By means of light microscopy of several organs, moderate changes were only observed in the heart. Ultrastructural studies revealed swelling of heart muscle cells leading to separation of the organelles. Effects were most pronounced close to the capillaries. The pathological changes were clearly dose dependent, and the lowest oral dose where any effects were seen was 2.5mg yessotoxin per kg.

Administration, Oral↗
PubMed2002

Isolation of Prorocentrum lima (Syn. Exuviaella lima) and diarrhetic shellfish poisoning (DSP) risk assessment in the Gulf of California, Mexico.

A benthic toxic dinoflagellate identified as Prorocentrum lima (Syn. Exuviaella lima), and designated as strain PRL-1, was isolated from the coast of El Pardito (Coyote) Island in Baja California Sur, Mexico, after a fisherman poisoning incident involving consumption of liver from Lutjanus colorado, and Mycteroperca prionura fish. Purification and culturing was done in ES-Si medium, under 12:12 light/dark cycle (4 x 20 W cool-white fluorescent lamps), at 22 degrees C and constant stirring during 28 days. Whole cells were toxic to Artemia franciscana and its methanolic extract to mouse and to the marine yeast Debaryomyces hansenii. Chromatographic analysis (TLC and HPLC-MS) of such extract indicated an unusual proportion (1:2) okadaic acid (OA) and dinophysistoxin-1 (DTX-1). Estimated total toxin content by mouse bioassay (based on OA toxicity) was 19 pg/cell, a value significantly higher than that found by HPLC-MS (about 5.2 pg/cell, taking into account OA and DTX-1 only), suggesting that additional toxic components of unidentified nature are detected with the bioassay. This is the first report of a successful isolation and culturing of a toxic dinoflagellate from the Gulf of California, Mexico.

Animals↗
PubMed2002

Azaspiracid shellfish poisoning: unusual toxin dynamics in shellfish and the increased risk of acute human intoxications.

A number of recent acute human intoxications in Europe from the consumption of Irish mussels have been attributed to the presence of a new class of toxins named azaspiracids. The study demonstrates that azaspiracids behave differently from other polyether toxins, and this accounts for most false-negative results in the mouse bioassay employed by regulatory agencies to detect azaspiracids. Typically, polyether toxins are concentrated in the digestive glands of shellfish, but this is not always the situation with azaspiracids. Liquid chromatography-mass spectrometry (LC-MS), especially multiple tandem MS methods, have been applied to demonstrate that azaspiracid (AZA1) and its methyl- and demethyl- analogues, AZA2 and AZA3 respectively, are distributed throughout shellfish tissues. Using conventional mouse bioassay protocols, only 0-40% of the total azaspiracid content of shellfish was used in the assay, which could directly account for false-negative results. It was also observed that the toxin profiles differed significantly in various mussel tissues with AZA1 as the predominant toxin in the digestive glands and AZA3 predominant in the remaining tissues.

Acute Disease↗
PubMed2001

Electrospray ionization mass spectrometry of tetrodotoxin and its analogs: liquid chromatography/mass spectrometry, tandem mass spectrometry, and liquid chromatography/tandem mass spectrometry.

Tetrodotoxin (TTX), a powerful sodium channel blocker, usually exists as a mixture of its analogs (TTXs) in natural sources. Due to the structural variation, some analogs are difficult to detect using the postcolumn liquid chromatography-fluorescent detection (LC-FLD) system. Liquid chromatography/electrospray ionization mass spectrometry (LC/ESI-MS) analysis of TTXs can be achieved by a combination of chromatography on a reversed-phase column with long carbon chains (C30) and the mobile phase containing an ion pair reagent (ammonium heptafluorobutyrate). The relationship between the amount of applied standard TTX and its peak area on the mass chromatogram (m/z 320) showed good linearity over a range of 50-1000 pmol. The detection limit for TTX in the selected ion monitoring (SIM) mode was estimated to be 0.7 pmol (signal to noise ratio: 2). The tandem mass spectrometry (MS/MS) scan for the fragment ions of eight TTXs arising from the molecular ions provided characteristic spectra, and the structures of the origins of the prominent fragment ions were proposed. The intense fragment ions of TTX and 11-deoxyTTX were applicable to LC/MS/MS operated in the selected reaction monitoring mode. This method might be useful for further identification of TTXs.

Animals↗
PubMed2001

Quantitative determination of marine toxins associated with diarrhetic shellfish poisoning by liquid chromatography coupled with mass spectrometry.

Quantitative determination by liquid chromatography (LC) coupled with mass spectrometry (MS) was achieved for the following 10 toxins found in association with diarrhetic shellfish poisoning: okadaic acid (OA), dinophysistoxin-1 (DTX1), 7-O-palmitoylokadaic acid (palOA), 7-O-palmitoyldinophysistoxin-1 (pa1DTX1), pectenotoxin-1 (PTX1), pectenotoxin-2 (PTX2), pectenotoxin-2 seco acid (PTX2SA), pectenotoxin-6 (PTX6), yessotoxin (YTX), and 45-hydroxyyessotoxin (YTXOH). Toxins in 2 g of the adductor muscle or the digestive glands of scallops, Patinopecten yessoensis, were extracted with 18 ml of methanol-water (9:1, v/v), freed of polar contaminants by partition between chloroform and water, and treated by solid-phase extraction on a silica cartridge column. Samples containing YTXOH were purified separately on a buffered reversed-phase column. Chromatographic separation was achieved by the following combinations of columns and mobile phases: a Symmetry C18 column with acetonitrile-0.05% acetic acid (7:3, v/v) for OA, DTX1, PTX6 and PTX2SA; a Develosil ODS column with the same mobile phase for PTX1 and PTX2; a Capcellpak column with methanol-2.5% acetic acid (98:2, v/v) for palOA and palDTX1; and an Inertsil ODS column with methanol-0.2 M ammonium acetate (8:2, v/v) for YTX and YTXOH. Carboxylic acid toxins were selectively monitored on [M-H]- ions, sulfated toxins on [M-Na]-ions, and neutral toxins on [M+NH4]+ ions. Average recoveries of the toxins spiked to tissue homogenates ranged from 70 to 134%. Detection limits in the muscle ranged from 5 to 40 ng/g and those in the digestive glands from 10 to 80 ng/g.

Chromatography, Liquid↗
PubMed2001

Absolute configuration at C14 and C85 in prymnesin-2, a potent hemolytic and ichthyotoxic glycoside isolated from the red tide alga Prymnesium parvum.

Prymnesin-2 is a potent red tide toxin characterized by a highly oxidized C(90) carbon chain and multiple functional groups. Succeeding the assignment of the relative stereochemistry in the polycyclic-ether segment, the absolute configurations of two chiral centers in linear chain parts were elucidated. The configuration at C14 bearing an amino group was determined to be S by using a chiral anisotropic reagent and that at chlorinated C85 to be S by fluorimetric chiral HPLC comparison between a degradation product and synthetic references.

Animals↗
PubMed2001

The chemistry and biological function of natural marine toxins.

Studies on ciguatera fish poisoning led to clarification of the absolute stereochemistry of ciguatoxin, gambierol, gambieric acids, and maitotoxin. Anisotropic NMR reagents and fluorometric chiral HPLC reagents were effectively used together with synthesis of partial structures. Structures of 16 ciguatoxin congeners were successfully elucidated by FAB/MS/MS using samples of 5 microg or less. Stereochemical assignments were also achieved on dinophysistoxin-1, pectenotoxins, yessotoxins, polycavernoside-A, azaspiracid, and prymnesins. The toxins possessed poly-cyclic-ether structures and originated from unicellular algae. Biological functions are briefly described.

Animals↗
PubMed2001

Monthly variations in diarrhetic toxins and yessotoxin in shellfish from coast to the inner part of the Sognefjord, Norway.

Monthly concentrations of diarrhetic shellfish poisoning (DSP) toxins and yessotoxin (YTX) in mussels from the coast to the inner part of the Sognefjord were determined. Mussels from nine locations were sampled from March to November 1997. The DSP toxins and YTX were analysed by a colorimetric protein phosphatase 2A (PP2A) inhibition assay or fluorometric HPLC, respectively. The mouse bioassay for DSP toxins was performed including either chloroform or diethyl ether in the final step of extraction. Using ether in the final step normally facilitated extraction of the DSP toxins, okadaic acid (OA) and dinophysis toxin-1 (DTX-1), while chloroform extraction included a wider spectrum of toxins, including YTX and a fast acting toxin(s) with neurotoxic effects. The concentrations of DSP toxins and YTX in mussels increased with distance from the coast. The highest concentrations of YTX (574 microg YTX/100 g mussel meat) and diarrhetic toxins (349 microg OA equivalents/100 g mussel meat) were measured in May and August, respectively, at locations in the inner part of the fjord. Since concentrations of DSP toxins and YTX in mussels increased with distance from the coast, the locations for mussel farming in the Sognefjord close to the coast, seem to be preferable.

Animals↗
PubMed2001

Maitotoxin-induced calcium entry in human lymphocytes: modulation by yessotoxin, Ca(2+) channel blockers and kinases.

We have studied the effect of the ciguatera-related toxin maitotoxin (MTX) on the cytosolic free calcium concentration ([Ca(2+)]i) of human peripheral blood lymphocytes loaded with the fluorescent probe Fura2 and the regulation of MTX action by different drugs known to interfere in cellular Ca(2+) signalling mechanisms and by the marine phycotoxin yessotoxin (YTX). MTX produced a concentration-dependent elevation of [Ca(2+)]i in a Ca(2+)-containing medium. This effect was stimulated by pretreatment with YTX 1 microM and NiCl(2) 15 microM. The voltage-independent Ca(2+) channel antagonist 1-[beta-[3-(4-methoxyphenyl)propoxyl]-4-methoxyphenyl]-1H-imidazole hydrochloride (SKF96365) blocked the MTX-induced [Ca(2+)]i elevation, while the L-type channel blocker nifedipine had no effect. Pretreatment with NiCl(2) or nifedipine did not modify YTX-induced potentiation of MTX effect, and SKF96365-induced inhibition was reduced in the presence of YTX, which suggest different pathways to act on [Ca(2+)]i. Preincubation with N-[2-(p-bromocinnamylamino)ethyl]-5-isoquinolinesulfonamide.2HCl (H-89) or genistein (10 microM) also had no effect on the MTX-induced [Ca(2+)]i increment. In contrast, the PKC inhibitor bisindolilmaleimide I (GF109203X 1 microM) potentiated the MTX effect, whereas phosphatidylinositol (PI) 3-kinase inhibition with wortmannin (10 nM) reduced the MTX-elicited Ca(2+) entry. In summary, MTX produced Ca(2+) influx into human lymphocytes through a SKF96365-sensitive, nifedipine-insensitive pathway. The MTX-induced [Ca(2+)]i elevation was stimulated by the marine toxin YTX through a mechanism insensitive to SKF96365, nifedipine or NiCl(2). It was also stimulated by the divalent cation Ni(2+) and PKC inhibition and was partially inhibited by PI 3-kinase inhibition.

Calcium↗
PubMed2001

Purification, characterization, and cDNA cloning of a novel soluble saxitoxin and tetrodotoxin binding protein from plasma of the puffer fish, Fugu pardalis.

Some species of puffer fish have been reported to possess both of tetrodotoxin and saxitoxin, which share one binding site on sodium channels. We purified a novel soluble glycoprotein that binds to these toxins from plasma of the puffer fish, Fugu pardalis, and named puffer fish saxitoxin and tetrodotoxin binding protein (PSTBP). PSTBP possessed a binding capacity of 10.6 +/- 0.97 nmol x mg(-1) protein and a K(d) of 14.6 +/- 0.33 nm for [(3)H]saxitoxin in equilibrium binding assays. [(3)H]Saxitoxin (10 nm) binding to PSTBPs was half-inhibited by the presence of tetrodotoxin and saxitoxin at 12 microm and 8.5 nm, respectively. From the results of gel filtration chromatography (200 kDa) and SDS/PAGE (104 kDa), PSTBP was suggested to consist of noncovalently linked dimers of a single subunit. PSTBP was completely deglycosylated by glycopeptidase F, producing a single band at 42 kDa. Two highly homologous cDNAs to each other coding PSTBP (PSTBP1 and PSTBP2, the predicted amino-acid identity 93%), were obtained from a cDNA library of F. pardalis liver. These proteins consisted to two tandemly repeated homologous domains. The predicted amino-acid sequences of PSTBP1 and 2 were not homologous to that of saxiphilin, a reported saxitoxin binding protein, or sodium channels, but their N-terminus sequences were homologous to that of the reported tetrodotoxin binding protein from plasma of Fugu niphobles, which has not been fully characterized. The partially homologous cDNA sequences to PSTBP1 and 2 were also found in expressed sequence tag clones of nontoxic flounders liver. Presumably, PSTBP is involved in accumulation and/or excretion of toxins in puffer fish.

Amino Acid Sequence↗
PubMed2001

Structure elucidation of ostreocin D, a palytoxin analog isolated from the dinoflagellate Ostreopsis siamensis.

The structure of ostreocin D, a palytoxin analog isolated from the marine dinoflagellate Ostreopsis siamensis, was found to be 42-hydroxy-3,26-didemethyl-19,44-dideoxypalytoxin by detailed 2D NMR analyses of intact ostreocin D and its ozonolysis products. Partial stereochemical assignments were done. This result indicates that the dinoflagellate O. siamensis is one of the biogenetic origins of palytoxin.

Acrylamides↗