Toxicity and kinetics of [3H]microcystin-LR in isolated perfused rat livers

Pace, J.; Robinson, Nancy; Miura, George A.; Matson, Charles F.; Geisbert, Thomas W.; White, James R.

doi:10.1016/0041-008x(91)90303-v

Cited by 31 publications

(18 citation statements)

References 20 publications

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“…3-6). These findings are basically consistent with earlier studies showing the functional and morphological mitochondrial alterations caused by microcystins (13)(14)(15)(16).…”

Section: Discussionsupporting

confidence: 82%

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Microcystic Cyanobacteria Causes Mitochondrial Membrane Potential Alteration and Reactive Oxygen Species Formation in Primary Cultured Rat Hepatocytes

Ding¹,

Shen²,

Shen³

et al. 1998

Environmental Health Perspectives

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Cyanobacteria contamination ofwater has become a growing public health problem worldwide.Microcystis aerginosa is one of the most common toxic cyanobacteria. It Contamination of water by toxic blooms of cyanobacteria (blue-green algae) has occurred widely in many regions of the world and poses a serious public health problem (1,4. Among the huge family of cyanobacteria, Microcystis aeruginosa is the most common toxic species. Microcystis is able to produce microcystins, a group of cyclic heptapeptide compounds with potent hepatotoxicity and tumor promotion activity (3-5).At present, the exact mechanisms by which microcystins induce hepatotoxicity and tumor promotion have not been fully elucidated. One of the well-studied mechanisms is that microcystins are potent inhibitors of protein phosphatase 1 and 2A, leading to increased protein phosphorylation, which is directly related to their cytotoxic effects and tumor-promoting activity (1,6). There is also some preliminary evidence indicating that oxidative damage plays an important role in the hepatotoxicity of microcystins. An earlier study in our laboratory demonstrated that oxidative stress is implicated in the hepatotoxic effects of cyanobacteria extract in cultured rat hepatocytes (7), which is consistent with some other reports showing the inhibitory effects of antioxidants on the toxicity of microcystins (8-10).Mitochondria are among the most important subcellular organelles in maintaining cellular structure and function by providing more than 80% of energy requirements through ATP production. Moreover, mitochondria are the main source of intracellular reactive oxygen species (ROS) formation and integrally involved in oxidative stress and cellular injury (11,12 (20) with modifications. As more than 90% of algae were Microcystis aeruginosa, it is believed that microcystins were the main toxins extracted. Briefly, lyophilized algae cells (25 mg) were first dissolved in 2.5 ml nbutanol:methanol:water (1:4:15, v/vlv) with high-speed stirring at room temperature for 1 hr, followed by centrifugation at 16,000g for 30 min. The precipitant was reextracted two more times as described above. The supernatant from three extractions was pooled and evaporated to dryness at 560C, and then dissolved in 5 ml 20% methanol. The extracted

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“…3-6). These findings are basically consistent with earlier studies showing the functional and morphological mitochondrial alterations caused by microcystins (13)(14)(15)(16).…”

Section: Discussionsupporting

confidence: 82%

“…Some earlier studies indicated that ATP depletion might be involved in this process (14,24). For instance, Pace et al (14) found that after the rat liver was perfused with microcystin-LR, one of the major toxic compounds produced by cyanobacteria, the isolated mitochondria displayed more than 50% inhibition ofcellular respiration.…”

Section: Discussionmentioning

confidence: 99%

Microcystic Cyanobacteria Causes Mitochondrial Membrane Potential Alteration and Reactive Oxygen Species Formation in Primary Cultured Rat Hepatocytes

Ding¹,

Shen²,

Shen³

et al. 1998

Environmental Health Perspectives

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“…These observations led MacKintosh et al (16) to describe the binding as "extremely strong," "tight binding," and "essentially stoichiometric," which indicate that the binding could have been covalent. Furthermore, hepatic glycogen depletion 60 min after injection of MCYST-LR (100 &kg, IP) in nonfasted rats (32), release of glucose into the medium during perfusion of isolated liver with MCYST-LR (15), and dose-dependent activation of phosphorylase a after exposure of isolated hepatocytes to MCYST-YM (33), are all consistent with these toxins causing inhibition of PP1 and PP2A.…”

Section: Discussionsupporting

confidence: 61%

Association of microcystin‐LR and its biotransformation product with a hepatic‐cytosolic protein

Robinson

Matson

Pace

1991

J. Biochem. Toxicol.

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Microcystin-LR (MCYST-LR), a cyclic peptide hepatotoxin, associates with high-molecular-weight, liver cytosolic components. Repetitive cycles of heat denaturation and pronase digestion released 80 +/- 6% of the bound radiolabel from these components, parent toxin (22%), and two biotransformation products, with high-performance liquid chromatography (HPLC) retention times of 6.7 (52%) and 5.6 (13%) min. Both parent and the biotransformed (6.7 min) toxin appeared to be covalently bound to a monomeric protein of molecular weight 40,000 (protein plus radiolabeled toxin). Binding and biotransformation reactions were time- and temperature-dependent and did not require endogenous molecules less than 6,000 daltons. The binding appeared to be saturable with a maximum of 20 pmol MCYST-LR bound per mg protein. The binding protein(s) and biotransformation activity were present in rat liver, brain, kidney, heart, lung, small intestine, large intestine, testes, skeletal muscle, and to a lesser extent, in fat. Okadaic acid, a specific protein phosphatase inhibitor, showed a concentration-dependent inhibition of [3H]MCYST-LR binding to hepatic cytosol. The molecular weight and organ distribution of the binding protein(s), and inhibition of binding by okadaic acid were consistent with one of the binding sites being the catalytic subunit of protein phosphatase type 2A.

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“…Microcystins are highly liver specific and their hepatotoxicity has been well studied (9)(10)(11). However, the exact mechanisms by which microcystins cause hepatotoxicity have not been fully elucidated.…”

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confidence: 99%

Microcystic cyanobacteria extract induces cytoskeletal disruption and intracellular glutathione alteration in hepatocytes.

Ding

Shen

Ong

2000

Environ Health Perspect

130

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Microcystins are a group of highly liver-specific toxins, although their exact mechanisms of action remain unclear. We examined the effects of microcystic cyanobacteria extract (MCE) collected from a contaminated water source on the organization of cellular microtubules (MTs) and microfilaments (MFs) in hepatocytes. We also investigated the effects on lactate dehydrogenase (LDH) leakage and intracellular glutathione (GSH). Primary cultured rat hepatocytes exposed to MCE (equivalent to 125 /mL lyophilized algae cells) showed a characteristic disruption of MTs and MFs in a time-dependent manner. Under these conditions, MCE caused aggtion of MTs and MFs and a severe loss of MTs in some cells. Moreover, MCE-induced cytoskeletal alterations preceded the LDH leakage. On the other hand, the treatment ofcells with MCE led to a dose-dependent increase of intracellular GSH. However, time-course study showed a biphasic change of intracellular GSH levels with a significant increase in the initial stage followed by a decrease after prolonged treatment. Furthermore, pretreatnent with N-acetylcystein (NAC), a GSH precursor, significanty hanced the intracellular GSH level and decreased the MCEinduced cytotoxicity as well as cytoskeleton changes. In contrast, buthionine-(G51-sulfoximine, a specific GSH synthesis inhibitor, increased the cell susceptibility to MCEinduced cytotoxicity by depleting the intracellular GSH level. These findings suggest that intracellular GSH plays an important role in MCEinduced cytotoxicity and cytoskeleton changes in primary cultured rat hepatocytes. Increasing intracellular GSH levels protect cdls from MCE-induced cytotoxicity and cytoskeleton changes.

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Toxicity and kinetics of [3H]microcystin-LR in isolated perfused rat livers

Cited by 31 publications

References 20 publications

Microcystic Cyanobacteria Causes Mitochondrial Membrane Potential Alteration and Reactive Oxygen Species Formation in Primary Cultured Rat Hepatocytes

Microcystic Cyanobacteria Causes Mitochondrial Membrane Potential Alteration and Reactive Oxygen Species Formation in Primary Cultured Rat Hepatocytes

Association of microcystin‐LR and its biotransformation product with a hepatic‐cytosolic protein

Microcystic cyanobacteria extract induces cytoskeletal disruption and intracellular glutathione alteration in hepatocytes.

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