Pierre Champy

Annonacin, a Natural Mitochondrial Complex I Inhibitor, Causes Tau Pathology in Cultured Neurons

A neurodegenerative tauopathy endemic to the Caribbean island of Guadeloupe has been associated with the consumption of anonaceous plants that contain acetogenins, potent lipophilic inhibitors of complex I of the mitochondrial respiratory chain. To test the hypothesis that annonacin, a prototypical acetogenin, contributes to the etiology of the disease, we investigated whether annonacin affects the cellular distribution of the protein tau. In primary cultures of rat striatal neurons treated for 48 h with annonacin, there was a concentration-dependent decrease in ATP levels, a redistribution of tau from the axons to the cell body, and cell death. Annonacin induced the retrograde transport of mitochondria, some of which had tau attached to their outer membrane. Taxol, a drug that displaces tau from microtubules, prevented the somatic redistribution of both mitochondria and tau but not cell death. Antioxidants, which scavenged the reactive oxygen species produced by complex I inhibition, did not affect either the redistribution of tau or cell death. Both were prevented, however, by forced expression of the NDI1 nicotinamide adenine dinucleotide (NADH)-quinone-oxidoreductase of Saccharomyces cerevisiae, which can restore NADH oxidation in complex I-deficient mammalian cells and stimulation of energy production via anaerobic glycolysis. Consistently, other ATP-depleting neurotoxins (1-methyl-4-phenylpyridinium, 3-nitropropionic, and carbonyl cyanide m-chlorophenylhydrazone) reproduced the somatic redistribution of tau, whereas toxins that did not decrease ATP levels did not cause the redistribution of tau. Therefore, the annonacin-induced ATP depletion causes the retrograde transport of mitochondria to the cell soma and induces changes in the intracellular distribution of tau in a way that shares characteristics with some neurodegenerative diseases.

Quantification of Acetogenins in Annona muricata Linked to Atypical Parkinsonism in Guadeloupe

Atypical parkinsonism in Guadeloupe has been associated with the consumption of fruit and infusions or decoctions prepared from leaves of Annona muricata L. (Annonaceae), which contains annonaceous acetogenins, lipophilic inhibitors of complex I of the mitochondrial respiratory chain. We have determined the concentrations of annonacin, the major acetogenin in A. muricata, in extracts of fruit and leaves by matrix‐assisted laser desorption‐ionization mass spectrometry. An average fruit is estimated to contain about 15 mg of annonacin, a can of commercial nectar 36 mg, and a cup of infusion or decoction 140 μg. As an indication of its potential toxicity, an adult who consumes one fruit or can of nectar a day is estimated to ingest over 1 year the amount of annonacin that induced brain lesions in rats receiving purified annonacin by intravenous infusion.

Natural lipophilic inhibitors of mitochondrial complex I are candidate toxins for sporadic neurodegenerative tau pathologies

Annonacin, a natural lipophilic inhibitor of mitochondrial complex I has been implicated in the etiology of a sporadic neurodegenerative tauopathy in Guadeloupe. We therefore studied further compounds representing the broad biochemical spectrum of complex I inhibitors to which humans are potentially exposed. We determined their lipophilicity, their effect on complex I activity in submitochondrial particles, and their effect on cellular ATP levels, neuronal cell death and somatodendritic redistribution of phosphorylated tau protein (AD2 antibody against pS396/pS404-tau) in primary cultures of fetal rat striatum. The 24 compounds tested were lipophilic (logP range 0.9-8.5; exception: MPP(+) logP=-1.35) and potent complex I inhibitors (IC(50) range 0.9 nM-2.6 mM). They all decreased ATP levels (EC(50) range 1.9 nM-54.2 microM), induced neuronal cell death (EC(50) range 1.1 nM-54.5 microM) and caused the redistribution of AD2(+) tau from axons to the cell body (EC(5) range 0.6 nM-33.3 microM). The potency of the compounds to inhibit complex I correlated with their potency to induce tau redistribution (r=0.80, p<0.001). In conclusion, we propose that the widely distributed lipophilic complex I inhibitors studied here might be implicated in the induction of tauopathies with global prevalence.

Ginger prevents Th2-mediated immune responses in a mouse model of airway inflammation

It is well documented that compounds from rhizomes of Zingiber officinale, commonly called ginger, have anti-inflammatory properties. Here, we show that ginger can exert such functions in vivo, namely in a mouse model of Th2-mediated pulmonary inflammation. The preparation of ginger aqueous extract (Zo.Aq) was characterized by mass spectrometry as an enriched fraction of n-gingerols. Intraperitoneal injections of this extract before airway challenge of ovalbumin (OVA)-sensitized mice resulted in a marked decrease in the recruitment of eosinophils to the lungs as attested by cell counts in bronchoalveolar lavage (BAL) fluids and histological examination. Resolution of airway inflammation induced by Zo.Aq was accompanied by a suppression of the Th2 cell-driven response to allergen in vivo. Thus, IL-4, IL-5 and eotaxin levels in the lungs as well as specific IgE titres in serum were clearly diminished in ginger-treated mice relative to their controls after allergen sensitization and challenge. Finally, we found that [6]-gingerol, a major constituent of ginger, was sufficient to suppress eosinophilia in our model of inflammation. This is the first evidence that ginger can suppress Th2-mediated immune responses and might thus provide a possible therapeutic application in allergic asthma.

Annonacin in Asimina triloba fruit: Implication for neurotoxicity

INTRODUCTION The acetogenin, annonacin, from the tropical annonaceous plant Annona muricata, is a lipophilic, mitochondrial complex I inhibitor reported to be more toxic than rotenone to mesencephalic neurons. The temperate annonaceous plant Asimina triloba (pawpaw) is native to the Eastern United States and products are available online. This study determined whether annonacin is in the pawpaw fruit pulp and whether it or the crude ethyl acetate extract is toxic to cortical neurons. METHODS Pawpaw extract was prepared by pulp extraction with methanol and liquid-liquid partitioning with ethyl acetate (EtOAc). Annonacin was isolated from the crude EtOAc extract via column chromatography using a gradient solvent system of increasing polarity. Mass spectroscopy, nuclear magnetic resonance and infrared spectroscopy were used to compare isolated material with synthetic annonacin data and a natural annonacin sample. Toxicity of isolated annonacin and the total EtOAc extract was determined in primary rat cortical neurons using the MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay. RESULTS The average concentration of annonacin in the fruit pulp was 0.0701±0.0305mg/g. Purified annonacin (30.07μg/ml) and crude EtOAc extract (47.96μg/ml) induced 50% death of cortical neurons 48h post treatment. Annonacin toxicity was enhanced in the presence of crude extract. DISCUSSION Pawpaw fruit contains a high concentration of annonacin, which is toxic to cortical neurons. Crude fruit extract also induced neurotoxicity, highlighting the need for additional studies to determine the potential risks of neurodegeneration associated with chronic exposure to pawpaw products.

MALDI-TOF MS Profiling of Annonaceous Acetogenins in Annona muricata Products for Human Consumption

Annonaceous acetogenins are proposed as environmental neurotoxicants consumed through medicinal and alimentary habits and responsible for atypical parkinsonian syndromes observed in tropical areas. Potential sources of exposure still have to be determined, as, to date, only a few batches of products for human consumption were searched for these compounds. To assess the presence of acetogenins, we propose a fast, sensitive and accurate method of screening, using MALDI-TOF MS, with minimal sample preparation. Development of the technique is discussed. Its application to leaves of herbal tea, pulp and bottled nectar of Annona muricata is presented.

Annonacin, a natural lipophilic mitochondrial complex I inhibitor, increases phosphorylation of tau in the brain of FTDP-17 transgenic mice.

Both genetic and environmental factors likely contribute to the neuropathology of tauopathies, but it remains unclear how specific genetic backgrounds affect the susceptibility towards environmental toxins. Mutations in the tau gene have been associated with familial tauopathies, while annonacin, a plant-derived mitochondrial inhibitor, has been implicated in an environmental form of tauopathy. We therefore determined whether there was a pathogenic synergy between annonacin exposure and the expression of the R406W-tau mutation in transgenic mice. We found that annonacin exposure caused an increase in the number of neurons with phosphorylated tau in the somatodendritic compartment in several brain areas in R406W(+/+) mice as opposed to mice that had only the endogenous mouse tau (R406W(-/-)). Western blot analysis demonstrated a concomitant increase in total tau protein without increase in tau mRNA, but reduced proteasomal proteolytic activity in R406W(+/+), but not R406W(-/-) mice, upon annonacin-treatment. Phosphorylated tau levels exceeded the increase in total tau protein, along with increased levels of different tau kinases, foremost a striking increase in the p25/p35 ratio, known to activate the tau kinase Cdk5. In summary, we observed a synergistic interaction between annonacin exposure and the presence of the R406W-tau mutation, which resulted in reduced degradation, increased phosphorylation and redistribution of neuronal tau.

Medicinal plants and finished marketed herbal products used in the treatment of malaria in the Ashanti region, Ghana

ETHNOPHARMACOLOGICAL RELEVANCE Ethnobotanical survey was performed to document medicinal plants employed in the management of malaria in the Bosomtwe and Sekyere East Districts of the Ashanti Region (Ghana), in comparison with the plant ingredients in herbal antimalarial remedies registered by the Ghana Food and Drug Administration. MATERIALS AND METHODS Two hundred and three (203) herbalists from 33 communities within the two districts were interviewed on medicinal plants they use to manage malaria. A literature search was made to determine already documented plants. In addition, 23 finished marketed herbal products indicated for the management of malaria were identified and their labels examined to find out which of the plants mentioned in our survey were listed as ingredients and whether these products are in anyway regulated. RESULTS Ninety-eight (98) species of plants were cited for the management of malaria. In comparison with literature citations, 12 (12.2%) species were reported for the management of malaria for the first time and 20 (20.4%) others for the first time in Ghana. Twenty-three (23) finished marketed herbal antimalarial products examined contained aerial or underground parts of 29 of the plants cited in our survey as ingredients. Twenty-two (22) of these products have been registered by the Ghana Food and Drugs Authority, four (4) of which were included in the recommended herbal medicine list for treating malaria in Ghana. CONCLUSION This study provides new additions to the inventory of medicinal plants used for the management of malaria and reports the commercial availability and regulation of finished marketed labelled herbal products intended for the treatment of malaria in Ghana.

Mutagenic effect, antioxidant and anticancer activities of six medicinal plants from Burkina Faso

The antiproliferative activities of six medicinal plant extracts from Burkina Faso were evaluated in order to justify their traditional use for the treatment of cancer. The SOS chromotest method was used in vitro on Escherichia coli PQ37 to evaluate the mutagenic effect of the plant extracts. The DPPH method was used to evaluate the antioxidant activity of each plant. The antiproliferative activity was evaluated by MTS method on normal cells (Vero and MCR5) and cancer cells (KB) in contact with the extracts for 72 h. The results showed that the studied plants are not genotoxic. Lantana ukambensis and Acacia macrostachya induced a very significant antiproliferative effect against cancer cells with 94% and 95%, respectively. They also developed a strong antioxidant activity. The IC50 values were 5.96 ± 0.40 µg mL−1 for L. ukambensis and 4.30 ± 0.26 µg mL−1 for A. macrostachya. These two plants are therefore potential sources for isolating new antioxidant and anticancer molecules.

Structural study of acetogenins by tandem mass spectrometry under high and low collision energy

Collision-induced dissociation experiments of seven annonaceous acetogenins were carried out under high and low collision energy conditions. Each compound was studied as protonated or deprotonated and lithium- or sodium- cationized molecules, using ElectroSpray Ionisation (ESI) with a hybrid linear trap/orbitrap mass spectrometer (LTQ-Orbitrap®). The same ion species were studied with a Matrix-Assisted Laser Desorption Ionisation (MALDI) tandem mass spectrometer in a high collision energy regime (1 or 2 keV). Although each of the techniques showed some limitations in the detection of functional groups, unambiguous structural identification of the acetogenins was obtained. MALDI ToF-ToF has the advantage over ESI-based methods to provide mass spectra rich in informative fragments which allows the confirmation of some functional groups position. By contrast, ESI-LTQ-Orbitrap® analysis has the advantage over MALDI that the mass spectra are relatively simple with only fragments close to the functional groups. However, this technique needs to carry out experiments both in negative and positive ionization modes. Copyri