No description available.

Harmful algae studies, in particular toxic dinoflagellates, and their response to global warming in the Red Sea are still limited. This study was aimed to be the first to characterize the identity, thermal responses and toxin production of four Prorocentrum strains isolated from the Central Red Sea, Saudi Arabia. Morphological and molecular phylogenetic analysis identified the strains as P. elegans, P. rhathymum and P. emarginatum. However, the identity of strain P. sp.6 is currently unresolved, albeit sharing close affinity with P. leve. Growth experiments showed that all species could grow at 24-32°C, but only P. sp.6 survived the 34°C treatment. The optimum temperatures (Topt) estimated from the Gaussian model corresponded to 27.17, 29.33, 26.87, and 27.64°C for P. sp.6, P. elegans, P. rhathymum and P. emarginatum, respectively. However, some discrepancy with the Topt derived from the growth performance were observed for P. elegans and P. emarginatum, as thermal responses differed from the typical Gaussian fit. The Prorocentrum species examined showed a sharp decrease after the optimum temperature resulting in very high activation energies for the fall slope, especially for P. elegans and P. emarginatum. The minimum critical temperature limit for growth was not detected within the range of temperatures examined. Subsequently, high performance liquid chromatography coupled with mass spectrometry (HPLC-MS) analysis revealed all species as non okadaic acid (OA, common toxin of the Prorocentrum genus) producers at any temperature treatment. However, other forms of toxin (i.e. fast acting toxins) not examined here could be produced. Therefore, further investigations are required. The results of this study provided significant contribution to our knowledge regarding the presence, thermal response and toxin production of four Prorocentrum species from the Central Red Sea, Saudi Arabia.

Toxins produced by harmful algal blooms (HABs) are known to pose contamination risks to seafood products (e.g. fish and shellfish) consumed by human. In order to control contamination risks, monitoring regimes have to be performed rigorously. The effort to monitor the amount of toxins in consumable products has to rely on continuous supply to analytical standards. The current work presents the strategy in optimising the production of major diarrhetic shellfish poisoning (DSP) toxins, OA and DTX1, from Prorocentrum lima. The organism is also known to produce peridinin, a carotenoid pigment that has been found to have pharmaceutical potential. Results from this study showed that cultivation of P. lima CCAP 1136/11 was still, although not completely, reliant on supply of natural seawater. Characterisation of compounds produced by P. lima CCAP 1136/11 in batch culture identified three major bioactive compounds (OA, DTX1 and peridinin) and two minor OA-related compounds. Recovery of these major compounds was further optimised with two-stage extraction procedure. Several important considerations for the cultivation process include standardisation of inoculum age and initial cell density. These and several other growth parameters such as temperature, light and CO2 supplementation have been shown to affect the growth and production of DSP toxins and peridinin in the culture. One of the main highlights in this study revealed that providing culture with light and dark cycle at frequency of 0.5 hour benefit in the enhancement of OA, DTX1 and peridinin yield from P. lima CCAP 1136/11. As the last part of this study, a simple and scalable design of reactor has been proposed. Contrary to common observations for dinoflagellate culture, P. lima CCAP 1136/11 was found to be able to withstand increased sparging within the culture system, resulting in concomitant increased of growth and production of OA, DTX1 and peridinin. Future works have been suggested to focus on: (1) exploitation of different cultivation system, such as continuous or semicontinuous systems, and (2) exploration on genetic modification to enable commercial scale production of DSP toxins and peridinin.

No description available.

A Doença de Alzheimer (DA) é uma doença cerebral progressiva que resulta em prejuízos na memória e disfunção cognitiva global. Entre as principais características neuropatológicas associadas a DA estão à presença de placas senis, emaranhados neurofibrilares e a hiperfosforilação da proteína Tau. A hiperativação do sistema glutamatérgico tem sido implicada na fisiopatologia da DA. O excesso de glutamato na fenda sináptica causa hiperativação do seu receptor ionótropico N-metill-D-aspartato (NMDA) o que favorece o aumento do influxo de cálcio e morte neuronal. A administração intracerebral de ácido ocadáico (AO) causa alterações morfológicas e funcionais similares à DA. O AO promove a inibição da proteína fosfatase 2A (PP2A) favorecendo as atividades cinásicas de proteínas como a cinase dependente de ciclina 5 (Cdk5). A memantina (MN) é uma das principais drogas utilizadas no tratamento da DA e o seu mecanismo de ação envolve um antagonismo não competitivo de baixa afinidade pela subunidade NR2B do receptor NMDA. Neste trabalho, foram avaliados efeitos do pré-tratamento com MN em um modelo semelhante a DA induzido pela administração intrahipocampal de AO em ratos. O pré-tratamento com MN preveniu o déficit na memória especial causado pela infusão intrahipocampal de AO. Os mecanismos envolvidos nestes efeitos neuroprotetores envolvem a prevenção do aumento de glutamato no liquido cefalorraquidiano, juntamente com a regulação da expressão de Cdk5 e em conseqüência a prevenção do aumento da fosforilação de Tau. Desta maneira, a MN pode ser um alvo terapêutico para prevenir as alterações comportamentais e neuroquímicas em um modelo similar a DA induzido pelo AO. / Alzheimer's disease (AD) is a progressive brain disease that causes memory loss and global cognitive dysfunction. The neuropathological alterations associated with AD include senile plaques, neurofibrillary tangles and Tau protein hyperphosphorylation. The glutamatergic system is implicated in the pathophysiology of AD. Indeed, the excessive glutamate levels in the synaptic cleft may cause hyperactivation of glutamate ionotropic N-metill-Daspartate (NMDA), which favors increase calcium influx and neuronal death. The intracerebral administration of okadaic acid (OA) causes morphological and functional alterations similar to AD. The OA inhibits the protein phosphatase 2A (PP2A) thus overstimulating the kinases activities. Memantine (MN) is a drug currently used in the treatment of AD, which mechanism involves a noncompetitive low affinity antagonism for NR2B subunit of NMDA receptors. In this work we evaluate the effects of pretreatment with MN in an AD-like model in rats induced by intrahippocampal administration of OA. The pretreatment with MN could prevent the spatial memory deficits caused by OA intrahipocampal administration in rats. The mechanisms underlying this neuroprotective effects involves the prevention of the increase in brain glutamate levels along with regulation of Cdk5 and, in consequence, downstream phosphorylation of Tau (ser199/202) protein. To conclude, MN has potential therapeutic role in preventing behavioral and neurochemical alterations caused by an AD like model induced by OA.

A Doença de Alzheimer (DA) é uma doença cerebral progressiva que resulta em prejuízos na memória e disfunção cognitiva global. Entre as principais características neuropatológicas associadas a DA estão à presença de placas senis, emaranhados neurofibrilares e a hiperfosforilação da proteína Tau. A hiperativação do sistema glutamatérgico tem sido implicada na fisiopatologia da DA. O excesso de glutamato na fenda sináptica causa hiperativação do seu receptor ionótropico N-metill-D-aspartato (NMDA) o que favorece o aumento do influxo de cálcio e morte neuronal. A administração intracerebral de ácido ocadáico (AO) causa alterações morfológicas e funcionais similares à DA. O AO promove a inibição da proteína fosfatase 2A (PP2A) favorecendo as atividades cinásicas de proteínas como a cinase dependente de ciclina 5 (Cdk5). A memantina (MN) é uma das principais drogas utilizadas no tratamento da DA e o seu mecanismo de ação envolve um antagonismo não competitivo de baixa afinidade pela subunidade NR2B do receptor NMDA. Neste trabalho, foram avaliados efeitos do pré-tratamento com MN em um modelo semelhante a DA induzido pela administração intrahipocampal de AO em ratos. O pré-tratamento com MN preveniu o déficit na memória especial causado pela infusão intrahipocampal de AO. Os mecanismos envolvidos nestes efeitos neuroprotetores envolvem a prevenção do aumento de glutamato no liquido cefalorraquidiano, juntamente com a regulação da expressão de Cdk5 e em conseqüência a prevenção do aumento da fosforilação de Tau. Desta maneira, a MN pode ser um alvo terapêutico para prevenir as alterações comportamentais e neuroquímicas em um modelo similar a DA induzido pelo AO. / Alzheimer's disease (AD) is a progressive brain disease that causes memory loss and global cognitive dysfunction. The neuropathological alterations associated with AD include senile plaques, neurofibrillary tangles and Tau protein hyperphosphorylation. The glutamatergic system is implicated in the pathophysiology of AD. Indeed, the excessive glutamate levels in the synaptic cleft may cause hyperactivation of glutamate ionotropic N-metill-Daspartate (NMDA), which favors increase calcium influx and neuronal death. The intracerebral administration of okadaic acid (OA) causes morphological and functional alterations similar to AD. The OA inhibits the protein phosphatase 2A (PP2A) thus overstimulating the kinases activities. Memantine (MN) is a drug currently used in the treatment of AD, which mechanism involves a noncompetitive low affinity antagonism for NR2B subunit of NMDA receptors. In this work we evaluate the effects of pretreatment with MN in an AD-like model in rats induced by intrahippocampal administration of OA. The pretreatment with MN could prevent the spatial memory deficits caused by OA intrahipocampal administration in rats. The mechanisms underlying this neuroprotective effects involves the prevention of the increase in brain glutamate levels along with regulation of Cdk5 and, in consequence, downstream phosphorylation of Tau (ser199/202) protein. To conclude, MN has potential therapeutic role in preventing behavioral and neurochemical alterations caused by an AD like model induced by OA.

A Doença de Alzheimer (DA) é uma doença cerebral progressiva que resulta em prejuízos na memória e disfunção cognitiva global. Entre as principais características neuropatológicas associadas a DA estão à presença de placas senis, emaranhados neurofibrilares e a hiperfosforilação da proteína Tau. A hiperativação do sistema glutamatérgico tem sido implicada na fisiopatologia da DA. O excesso de glutamato na fenda sináptica causa hiperativação do seu receptor ionótropico N-metill-D-aspartato (NMDA) o que favorece o aumento do influxo de cálcio e morte neuronal. A administração intracerebral de ácido ocadáico (AO) causa alterações morfológicas e funcionais similares à DA. O AO promove a inibição da proteína fosfatase 2A (PP2A) favorecendo as atividades cinásicas de proteínas como a cinase dependente de ciclina 5 (Cdk5). A memantina (MN) é uma das principais drogas utilizadas no tratamento da DA e o seu mecanismo de ação envolve um antagonismo não competitivo de baixa afinidade pela subunidade NR2B do receptor NMDA. Neste trabalho, foram avaliados efeitos do pré-tratamento com MN em um modelo semelhante a DA induzido pela administração intrahipocampal de AO em ratos. O pré-tratamento com MN preveniu o déficit na memória especial causado pela infusão intrahipocampal de AO. Os mecanismos envolvidos nestes efeitos neuroprotetores envolvem a prevenção do aumento de glutamato no liquido cefalorraquidiano, juntamente com a regulação da expressão de Cdk5 e em conseqüência a prevenção do aumento da fosforilação de Tau. Desta maneira, a MN pode ser um alvo terapêutico para prevenir as alterações comportamentais e neuroquímicas em um modelo similar a DA induzido pelo AO. / Alzheimer's disease (AD) is a progressive brain disease that causes memory loss and global cognitive dysfunction. The neuropathological alterations associated with AD include senile plaques, neurofibrillary tangles and Tau protein hyperphosphorylation. The glutamatergic system is implicated in the pathophysiology of AD. Indeed, the excessive glutamate levels in the synaptic cleft may cause hyperactivation of glutamate ionotropic N-metill-Daspartate (NMDA), which favors increase calcium influx and neuronal death. The intracerebral administration of okadaic acid (OA) causes morphological and functional alterations similar to AD. The OA inhibits the protein phosphatase 2A (PP2A) thus overstimulating the kinases activities. Memantine (MN) is a drug currently used in the treatment of AD, which mechanism involves a noncompetitive low affinity antagonism for NR2B subunit of NMDA receptors. In this work we evaluate the effects of pretreatment with MN in an AD-like model in rats induced by intrahippocampal administration of OA. The pretreatment with MN could prevent the spatial memory deficits caused by OA intrahipocampal administration in rats. The mechanisms underlying this neuroprotective effects involves the prevention of the increase in brain glutamate levels along with regulation of Cdk5 and, in consequence, downstream phosphorylation of Tau (ser199/202) protein. To conclude, MN has potential therapeutic role in preventing behavioral and neurochemical alterations caused by an AD like model induced by OA.

L’apparition de l’obésité et du cancer du sein est influencée par l’exposition aux polluants environnementaux. Ces polluants interfèrent avec le fonctionnement normal de l’organisme causant des modifications dans le comportement cellulaire. Ils peuvent agir chacun seul ou sous forme de cocktail. Dans ce contexte, nous avons choisi trois polluants dans notre projet : le bisphénol A (perturbateur endocrinien), le glyphosate (désherbant) et l’acide okadaïque (toxine marine). Nous avons étudié leur effet seul ou en cocktail sur les cellules de cancer du sein MCF7 et MDA-MB231, les cellules épithéliale mammaire humaine HME1 et les préadipocytes humain HWP. Dans une première partie, nous avons étudié leur effet sur la viabilité cellulaire. L’électrophorèse 2D couplée à MS/MALDI-TOF, le dosage de l’activité des protéines phosphatases ont été menés pour comprendre le mécanisme d’action des polluants sur la viabilité cellulaire. La partie suivante a porté sur la détection et la quantification des polluants dans le milieu de culture afin de déterminer leur voie d’action : extra ou intra cellulaire. Finalement, nous avons étudié l’effet des polluants sur le cycle de vie des adipocytes et la lipogenèse. Également, l’effet de sécrétome des HWP (+/-polluants) sur la viabilité cellulaire des cellules cancéreuses a été étudié. / The development of obesity and breast cancer is influenced by exposure to environmental pollutants. These pollutants interfere with the normal body function causing changes in cells behavior. They can act each alone or as a cocktail. In this context, we have chosen three pollutants in our project : bisphenol A (endocrin disruptor), glyphosate (herbicide), okadaic acid (marin toxine). We have studied their effect of alone or in combination in cellular models : human breast cancer cells MCF7 and MDA-MB 231, human mammary epithelial cells HME1 and human preadipocytes HWP. In the first part, we have studied the effect of pollutants on cells viability. 2D electrophoresis coupled to MS/MALDI-TOF, measurement of proteins phosphatase activity were conducted to understand pollutants action mechanism on cells viability. In the following part we detected and quantified pollutants in culture medium to determine their pathway of action : extra or intra cellular. Finally, we studied the effect of pollutants on adipocytes life cycle and lipogenesis. Also, effect of secretome from HWP (+/- pollutants) on cancer cells viability was studied.

The purposes of this study were to characterize the complexation of toxic metals with algal toxins and to determine the effects of arsenic and copper on the growth of Karenia brevis under specific experimental conditions. Microcystins, pahayokolides, brevetoxins and okadaic acid were used as representatives of algal toxins while arsenic, copper, cadmium, cobalt, iron, manganese and mercury were selected as typical toxic metals (including metalloids here) in the aquatic environment. The stabilities of the toxin-metal complexes were determined using equilibrium dialysis and/or centrifugal ultrafiltration technique. A direct exposure of arsenic and copper to the K. brevis was carried out to determine the effects of these metals to the growth of the algal cell. The results indicated that Cu2+, Hg2+, Co2+, Cd2+ and Fe2+ were capable of complexing with the algal toxins. Moreover, the exposure experiments demonstrated that the high concentration of arsenic and copper could affect the growth of the K. brevis.

Humans are exposed to a variety of chemicals including environmental pollutants, cosmetics, food preservatives and drugs. Some of these substances might be harmful to the human body. Traditional toxicological and behavioural investigations performed in animal models are not suitable for the screening of a large number of compounds for potential toxic effects. There is a need for simple and robust in vitro cellular models that allow high-throughput toxicity testing of chemicals, as well as investigation of specific mechanisms of cytotoxicity. The overall aim of the thesis has been to evaluate neuronally differentiated mouse embryonal carcinoma P19 cells (P19 neurons) as a model for such testing. The model has been compared to other cellular models used for neurotoxicity assessment: retinoic acid-differentiated human neuroblastoma SH-SY5Y cells and nerve growth factor-treated rat pheochromocytoma PC12 cells. The chemicals assessed in the studies included the neurotoxicants methylmercury, okadaic acid and acrylamide, the drug of abuse MDMA (“ecstasy”) and a group of piperazine derivatives known as “party pills”. Effects of the chemicals on cell survival, neurite outgrowth and mitochondrial function have been assessed. In Paper I, we describe a fluorescence-based microplate method to detect chemical-induced effects on neurite outgrowth in P19 neurons immunostained against the neuron-specific cytoskeletal protein βIII-tubulin. In Paper II, we show that P19 neurons are more sensitive than differentiated SH-SY5Y and PC12 cells for detection of cytotoxic effects of methylmercury, okadaic acid and acrylamide. Additionally, in P19 neurons and differentiated SH-SY5Y cells, we could demonstrate that toxicity of methylmercury was attenuated by the antioxidant glutathione. In Paper III, we show a time- and temperature-dependent toxicity produced by MDMA in P19 neurons. The mechanisms of MDMA toxicity did not involve inhibition of the serotonin re-uptake transporter or monoamine oxidase, stimulation of 5-HT2A receptors, oxidative stress or loss of mitochondrial membrane potential. In Paper IV, the piperazine derivatives are evaluated for cytotoxicity in P19 neurons and differentiated SH-SY5Y cells. The most toxic compound in both cell models was TFMPP. In P19 neurons, the mechanism of action of TFMPP included loss of mitochondrial membrane potential. In conclusion, P19 neurons are a robust cellular model that may be useful in conjunction with other models for the assessment of chemical-induced neurotoxicity.