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Title: Toxicogenomic evaluation of microcystin-LR treated with ultrasonic irradiation

Abstract

Microcystins are a family of toxins produced by cyanobacteria found throughout the world in marine and freshwater environments. The most commonly encountered form of microcystin is microcystin-LR (MC-LR). Humans are exposed to MC-LR by drinking contaminated water. The toxin accumulates rapidly in the liver where it exerts most of its damage. Treatment of water containing MC-LR by ultrasonic irradiation leads to the breakdown of the toxin. Both the parent toxin and the treated toxin reaction products (TTRP) were evaluated for toxic effects in mice. Animals were exposed to purified MC-LR or an equivalent dose of the TTRP and sacrificed after 4 h or 24 h. Serum was collected and assayed for lactate dehydrogenase (LDH) activity as an indicator of hepatotoxicity. LDH activity was detected in the serum of MC-LR exposed mice indicative of liver damage, but not in control mice. Only a fraction of that activity was detectable in mice exposed to TTRP. Liver RNA was used for microarray analysis and real-time PCR. Individual animals varied in their overall genomic response to the toxin; however, only 20 genes showed consistent changes in expression. These include chaperones which may be part of a generalized stress response; cytochrome P450 which may bemore » involved in metabolizing the toxin; and lipid dystrophy genes such as lipin-2, uridine phosphorylase and a homolog to tribbles, a stress-inducible gene involved in cell death. Of the genes that responded to the MC-LR, none showed significant changes in expression profile in response to TTRP. Taken together, the data indicate that ultrasonic irradiation of MC-LR effectively reduces hepatotoxicity in mice and therefore may be a useful method for detoxification of drinking water.« less

Authors:
 [1];  [2];  [2];  [3]
  1. Division of Marine Biology and Fisheries, Rosenstiel School of Marine and Atmospheric Science, University of Miami, Virginia Key, FL 33149 (United States). E-mail: Alice.Hudder@wayne.edu
  2. Department of Chemistry and Biochemistry, Florida International University, University Park, FL 33199 (United States)
  3. Centre for Advanced Research in Environmental Genomics (CAREG), University of Ottawa, 30 Marie Curie, Ottawa, ON, K1N 6N5 (Canada)
Publication Date:
OSTI Identifier:
20976928
Resource Type:
Journal Article
Resource Relation:
Journal Name: Toxicology and Applied Pharmacology; Journal Volume: 220; Journal Issue: 3; Other Information: DOI: 10.1016/j.taap.2007.02.004; PII: S0041-008X(07)00075-0; Copyright (c) 2007 Elsevier Science B.V., Amsterdam, The Netherlands, All rights reserved; Country of input: International Atomic Energy Agency (IAEA)
Country of Publication:
United States
Language:
English
Subject:
63 RADIATION, THERMAL, AND OTHER ENVIRONMENTAL POLLUTANT EFFECTS ON LIVING ORGANISMS AND BIOLOGICAL MATERIALS; APOPTOSIS; BIOLOGICAL STRESS; CYANOBACTERIA; DAMAGE; DETOXIFICATION; DOSE EQUIVALENTS; DRINKING WATER; GENES; IRRADIATION; LACTATE DEHYDROGENASE; LIVER; MICE; POLYMERASE CHAIN REACTION; RNA; TOXICITY; TOXINS; URIDINE; XENOBIOTICS

Citation Formats

Hudder, Alice, Song Weihua, O'Shea, Kevin E., and Walsh, Patrick J.. Toxicogenomic evaluation of microcystin-LR treated with ultrasonic irradiation. United States: N. p., 2007. Web. doi:10.1016/j.taap.2007.02.004.
Hudder, Alice, Song Weihua, O'Shea, Kevin E., & Walsh, Patrick J.. Toxicogenomic evaluation of microcystin-LR treated with ultrasonic irradiation. United States. doi:10.1016/j.taap.2007.02.004.
Hudder, Alice, Song Weihua, O'Shea, Kevin E., and Walsh, Patrick J.. Tue . "Toxicogenomic evaluation of microcystin-LR treated with ultrasonic irradiation". United States. doi:10.1016/j.taap.2007.02.004.
@article{osti_20976928,
title = {Toxicogenomic evaluation of microcystin-LR treated with ultrasonic irradiation},
author = {Hudder, Alice and Song Weihua and O'Shea, Kevin E. and Walsh, Patrick J.},
abstractNote = {Microcystins are a family of toxins produced by cyanobacteria found throughout the world in marine and freshwater environments. The most commonly encountered form of microcystin is microcystin-LR (MC-LR). Humans are exposed to MC-LR by drinking contaminated water. The toxin accumulates rapidly in the liver where it exerts most of its damage. Treatment of water containing MC-LR by ultrasonic irradiation leads to the breakdown of the toxin. Both the parent toxin and the treated toxin reaction products (TTRP) were evaluated for toxic effects in mice. Animals were exposed to purified MC-LR or an equivalent dose of the TTRP and sacrificed after 4 h or 24 h. Serum was collected and assayed for lactate dehydrogenase (LDH) activity as an indicator of hepatotoxicity. LDH activity was detected in the serum of MC-LR exposed mice indicative of liver damage, but not in control mice. Only a fraction of that activity was detectable in mice exposed to TTRP. Liver RNA was used for microarray analysis and real-time PCR. Individual animals varied in their overall genomic response to the toxin; however, only 20 genes showed consistent changes in expression. These include chaperones which may be part of a generalized stress response; cytochrome P450 which may be involved in metabolizing the toxin; and lipid dystrophy genes such as lipin-2, uridine phosphorylase and a homolog to tribbles, a stress-inducible gene involved in cell death. Of the genes that responded to the MC-LR, none showed significant changes in expression profile in response to TTRP. Taken together, the data indicate that ultrasonic irradiation of MC-LR effectively reduces hepatotoxicity in mice and therefore may be a useful method for detoxification of drinking water.},
doi = {10.1016/j.taap.2007.02.004},
journal = {Toxicology and Applied Pharmacology},
number = 3,
volume = 220,
place = {United States},
year = {Tue May 01 00:00:00 EDT 2007},
month = {Tue May 01 00:00:00 EDT 2007}
}
  • Isolated rat livers were perfused for 60 min with either 0.3 or 0.5 microgram/ml (initial volume, 119 ml) of (3H)microcystin-LR at a constant flow of 10 ml/min in a recirculating system. During the 60-min exposure, toxin caused stimulation of glycogenolysis, liver engorgement, and cessation of bile flow. Electron micrographs of liver showed dilation of bile canaliculi and the space of Disse, loss of sinusoidal lining architecture, and decreased hepatocyte intercellular contacts. Although hepatocytes did not exhibit overt necrosis, mitochondria were hydropic, occasionally encircled by whorls of rough endoplasmic reticulum, and desmosomal tonofilaments were decreased on the plasma membrane lateral surface.more » Isolated mitochondria displayed inhibition of state 3 respiration and a 50-60% decrease in the respiratory control index, characteristic of hydropism. Distribution of radiolabel was 1.7% to bile, 79% to perfusate, and 16% to liver. Two to four percent was recovered in perfusate that leaked from the surface of the liver. Of the radiolabel found in bile and perfusate, 78 and 100% were associated with parent toxin, respectively. The radiolabel in liver, associated with the cytosolic fraction (S-100), corresponded to parent toxin (15%) and to a more-polar component(s) (85%). The elimination half-life from perfusate was 130 +/- 10 min (0.5 microgram/ml) and the hepatic extraction ratio 0.07 +/- 0.01. Although the calculated hepatic extraction ratio was low, there was a significant accumulation of microcystin in the liver. Many toxic effects of microcystin in the perfused liver mimicked those observed in the whole animal, suggesting that this model can be used as an alternative to whole animals for screening of potential therapeutic agents.« less
  • The distribution, excretion and hepatic metabolism of (3H)microcystin-LR (sublethal i.v.) were measured in mice. Plasma elimination was biexponential with alpha- and beta-phase half-lives of 0.8 and 6.9 min, respectively. At 60 min, liver contained 67 +/- 4% of dose. Through the 6-day study the amount of hepatic radioactivity did not change whereas 23.7 +/- 1.7% of the dose was excreted; 9.2 +/- 1.0% in urine and 14.5 +/- 1.1% in feces. Approximately 60% of the urine and fecal radiolabel 6 and 12 hr postinjection was the parent toxin. Hepatic cytosol, which contained 70 +/- 2% of the hepatic radiolabel (1more » hr through 6 days), was prepared for high-performance liquid chromatography analysis by heat denaturation, pronase digestion and C18 Sep Pak extraction. At 1 hr, 35 +/- 2% of the radiolabel was insoluble or C18 Sep Pak-bound; 43 +/- 3% was associated with a peak of retention time (rt) 6.6 min, and 16 +/- 3% with the parent toxin (rt 9.4 min). After 6 days, 8 +/- 1% was C18 Sep Pak-bound or insoluble; 5 +/- 0% occurred at rt 6.6 min, 17 +/- 1% with parent and 60 +/- 2% was associated with rt 8.1 min. Two other peaks, rt 4.9 and 5.6 min, appeared transiently. Analysis of hepatic cytosol by desalting chromatography under nondenaturing and denaturing conditions revealed that all of the radiolabel was associated with cytosolic components, and 83 +/- 5% was bound covalently through 1 day. By day 6 the amount of covalently bound isotope decreased to 42 +/- 11%. This is the first study to describe the long-term hepatic retention of microcystin toxin and documents putative detoxication products.« less
  • Microcystins (MCs), a cyclic heptapeptide hepatotoxins, are mainly produced by the bloom-forming cyanobacerium Microcystis, which has become an environmental hazard worldwide. Long term consumption of MC-contaminated water may induce liver damage, liver cancer, and even human death. Therefore, in addition to removal of MCs in drinking water, novel strategies that prevent health damages are urgently needed. Sulforaphane (SFN), a natural-occurring isothiocyanate from cruciferous vegetables, has been reported to reduce and eliminate toxicities from xenobiotics and carcinogens. The purpose of the present study was to provide mechanistic insights into the SFN-induced antioxidative defense system against MC-LR-induced cytotoxicity. We performed cell viabilitymore » assays, including MTS assay, colony formation assay and apoptotic cell sorting, to study MC-LR-induced cellular damage and the protective effects by SFN. The results showed that SFN protected MC-LR-induced damages at a nontoxic and physiological relevant dose in HepG2, BRL-3A and NIH 3 T3 cells. The protection was Nrf2-mediated as evident by transactivation of Nrf2 and activation of its downstream genes, including NQO1 and HO-1, and elevated intracellular GSH level. Results of our studies indicate that pretreatment of cells with 10 {mu}M SFN for 12 h significantly protected cells from MC-LR-induced damage. SFN-induced protective response was mediated through Nrf2 pathway.« less
  • Microcystins (MCs), the products of blooming algae Microcystis, are waterborne environmental toxins that have been implicated in the development of liver cancer, necrosis, and even fatal intrahepatic bleeding. Alternative protective approaches in addition to complete removal of MCs in drinking water are urgently needed. In our previous work, we found that sulforaphane (SFN) protects against microcystin-LR (MC-LR)-induced cytotoxicity by activating the NF-E2-related factor 2 (Nrf2)-mediated defensive response in human hepatoma (HepG2) and NIH 3T3 cells. The purpose of this study was to investigate and confirm efficacy the SFN-induced multi-mechanistic defense system against MC-induced hepatotoxicity in an animal model. We reportmore » that SFN protected against MC-LR-induced liver damage and animal death at a nontoxic and physiologically relevant dose in BALB/c mice. The protection by SFN included activities of anti-cytochrome P450 induction, anti-oxidation, anti-inflammation, and anti-apoptosis. Our results suggest that SFN may protect mice against MC-induced hepatotoxicity. This raises the possibility of a similar protective effect in human populations, particularly in developing countries where freshwaters are polluted by blooming algae. - Graphical abstract: Display Omitted Research Highlights: > SFN protected against MC-LR-induced liver damage and animal death in BALB/c mice. > The dose of SFN is at a nontoxic and physiologically relevant dose. > The protection included activities of anti-oxidation, anti-inflammation, and anti-apoptosis. > SFN may protect mice against MC-induced hepatotoxicity.« less
  • Microcystin (MC)-LR, a cyclic heptapeptide, is a potent reproductive system toxin. To understand the molecular mechanisms of MC-induced reproductive system cytotoxicity, we evaluated global changes of miRNA and mRNA expression in mouse Sertoli cells following MC-LR treatment. Our results revealed that the exposure to MC-LR resulted in an altered miRNA expression profile that might be responsible for the modulation of mRNA expression. Bio-functional analysis indicated that the altered genes were involved in specific cellular processes, including cell death and proliferation. Target gene analysis suggested that junction injury in Sertoli cells exposed to MC-LR might be mediated by miRNAs through themore » regulation of the Sertoli cell-Sertoli cell pathway. Collectively, these findings may enhance our understanding on the modes of action of MC-LR on mouse Sertoli cells as well as the molecular mechanisms underlying the toxicity of MC-LR on the male reproductive system. - Highlights: • miRNAs were altered in Sertoli cells exposed to MC-LR. • Alerted genes were involved in different cell functions including the cell morphology. • MC-LR adversely affected Sertoli cell junction formation through the regulating miRNAs.« less