Effect of a short-term in vitro exposure to the marine toxin domoic acid on viability, tumor necrosis factor-alpha, matrix metalloproteinase-9 and superoxide anion release by rat neonatal microglia.

BACKGROUND: The excitatory amino acid domoic acid, a glutamate and kainic acid analog, is the causative agent of amnesic shellfish poisoning in humans. No studies to our knowledge have investigated the potential contribution to short-term neurotoxicity of the brain microglia, a cell type that constitutes circa 10% of the total glial population in the brain. We tested the hypothesis that a short-term in vitro exposure to domoic acid, might lead to the activation of rat neonatal microglia and the concomitant release of the putative neurotoxic mediators tumor necrosis factor-alpha (TNF-alpha), matrix metalloproteinases-2 and-9 (MMP-2 and -9) and superoxide anion (O2-). RESULTS: In vitro, domoic acid [10 microM-1 mM] was significantly neurotoxic to primary cerebellar granule neurons. Although neonatal rat microglia expressed ionotropic glutamate GluR4 receptors, exposure during 6 hours to domoic acid [10 microM-1 mM] had no significant effect on viability. By four hours, LPS (10 ng/mL) stimulated an increase in TNF-alpha mRNA and a 2,233 % increase in TNF-alpha protein In contrast, domoic acid (1 mM) induced a slight rise in TNF-alpha expression and a 53 % increase (p < 0.01) of immunoreactive TNF-alpha protein. Furthermore, though less potent than LPS, a 4-hour treatment with domoic acid (1 mM) yielded a 757% (p < 0.01) increase in MMP-9 release, but had no effect on MMP-2. Finally, while PMA (phorbol 12-myristate 13-acetate) stimulated O2- generation was elevated in 6 hour LPS-primed microglia, a similar pretreatment with domoic acid (1 mM) did not prime O2- release. CONCLUSIONS: To our knowledge this is the first experimental evidence that domoic acid, at in vitro concentrations that are toxic to neuronal cells, can trigger a release of statistically significant amounts of TNF-alpha and MMP-9 by brain microglia. These observations are of considerable pathophysiological significance because domoic acid activates rat microglia several days after in vivo administration.

Marine pharmacology in 1999: antitumor and cytotoxic compounds.

During 1999 marine antitumor pharmacology research involved researchers in Austria, Australia, England, France, Germany, Greece, Holland, Italy, Japan, Spain, Taiwan and the United States. Thirty six papers were published in peer-reviewed journals describing the antitumor and cytotoxic properties of 30 marine natural products belonging to four structural types, namely polyketides, terpenes, nitrogen-containing compounds and polysaccharides. The organisms yielding these bioactive marine compounds comprised a diverse group of marine animals, algae, fungi and bacteria. A variety of antitumor pharmacological studies were conducted with 17 marine natural products with established mechanisms of action in a number of experimental and clinical models. Didemnin B, a tunicate-derived depsipeptide with potent antitumor effects, completed a Phase II anticancer clinical trial which resulted indeterminate in respect to activity against human melanoma due to anaphylactoid reactions. In vitro cytotoxicity data with murine and human cell lines were reported for 14 novel marine chemicals with as yet undetermined mechanisms of action. This 1999 literature overview thus highlights the fact that the multinational effort aimed at the discovery of novel marine antitumor agents remained at the same level of research activity as during 1998.

Isolation and characterization of microcystins from a river nile strain of Oscillatoria tenuis Agardh ex Gomont.

The River Nile is the major source of drinking water in Egypt, however, increased eutrophication due to agricultural, municipal and industrial runoff has contributed to the growth of toxin producing cyanobacteria. This study describes the isolation and characterization of microcystins (MCYSTs), cyclic heptapeptide hepatotoxins, from a rare strain of Oscillatoria tenuis, isolated from the River Nile at Sohag province in July 1995. The MCYST concentration of laboratory-cultured O. tenuis strain E6 was found to be 0.3 mg/g freeze-dried weight determined by enzyme-linked immunosorbent assay (ELISA). Two microcystins, 1 and 2, were isolated from lyophilized cells using solid phase extraction and reversed-phase high performance liquid chromatography (HPLC). Structures were assigned based upon their amino acid analyses, electrospray ionization mass spectrometry (ESIMS, ESIMS-CID-MS), high resolution fast atom bombardment mass spectrometry, and nuclear magnetic resonance data ((1)H and (1)H COSY NMR). Toxin 1 was identified as MCYST-LR, and toxin 2, a new MCYST, as MCYST-LHArg ([L-homoarginine(4)]). Previous studies indicate that Oscillatoria agardhii strains produce demethylated MCYSTs (containing D-Asp and/or dehydroalanine). This is the first report of a toxic O. tenuis, strain E6, one which produces a fully methylated MCYST, MCYST-LR and a new L-homoarginine containing MCYST, MCYST-LHArg.