Daniéli Gerhardt

Protective effect of resveratrol against oxygen-glucose deprivation in organotypic hippocampal slice cultures : Involvement of PI3-K pathway

Here we investigated the neuroprotective effect of resveratrol in an in vitro model of ischemia. We used organotypic hippocampal cultures exposed to oxygen-glucose deprivation (OGD). In OGD-vehicle exposed cultures, about 46% of the hippocampus was labeled with PI, indicating a robust percentage of cell death. When cultures were treated with resveratrol 10, 25 and 50 microM, the cell death was reduced to 22, 20 and 13% respectively. To elucidate a possible mechanism by which resveratrol exerts its neuroprotective effect, we investigated the phosphoinositide3-kinase (PI3-k) pathway using LY294002 (5 microM) and mitogen-activated protein kinase (MAPK) using PD98059 (20 microM). The resveratrol (50 microM) neuroprotection was prevented by LY294002 but was not by PD98059. Immunoblotting revealed that resveratrol 50 microM induced the phosphorylation/activation of Akt and extracellular signal-regulated kinase-1 and -2 (ERK1/2) and the phosphorylation/inactivation of glycogen synthase kinase-3beta (GSK-3beta). Our results suggest that PI3-k/Akt pathway are involved in the neuroprotective effect of resveratrol.

Conditioned medium from mesenchymal stem cells induces cell death in organotypic cultures of rat hippocampus and aggravates lesion in a model of oxygen and glucose deprivation.

Cell therapy using bone marrow-derived mesenchymal stem cells (MSC) seems to be a new alternative for the treatment of neurological diseases, including stroke. In order to investigate the response of hippocampal tissue to factors secreted by MSC and if these factors are neuroprotective in a model of oxygen and glucose deprivation (OGD), we used organotypic hippocampal cultures exposed to conditioned medium from bone marrow-derived MSC. Our results suggest that the conditioned medium obtained from these cells aggravates lesion caused by OGD. In addition, the presence of the conditioned medium alone was toxic mainly to cells in the CA1, CA2 and CA3 areas of the hippocampal organotypic culture even in basal conditions. GABA stimulation and NMDA and AMPA receptors antagonists were able to reduce propidium iodide staining, suggesting that the cell death induced by the toxic factors secreted by MSC could involve these receptors.

Aβ1-42 and Aβ25-35 peptides induce similar toxicity in organotypic hippocampal slice cultures and this effect can be prevented by resveratrol

Background: Alzheimer’s disease (AD) is the prevalent neurodegenerative disorder affecting over 15 million individuals worldwide. Abnormal biosynthesis and accumulation of the neurotoxic b-amyloı̈d (Ab) peptide in senile plaques is suggested to play a central role in the pathogenesis of AD and investigating mechanisms involved in these processes remains essential to elucidate origin and evolution of this disease. Brain is isolated from the whole body by the blood-brain barrier (BBB) which restricts the exchanges of solutes such as Ab peptide between brain and blood. The cellular mechanisms involved in these exchanges remain poorly understood. We focused our interest on RAGE (Receptor for advanced glycation end-products) and LRPs (Low density lipoprotein receptor-related proteins), suspected to be implicated respectively in influx and in efflux of Ab peptides Methods: In our laboratory, we have developed an in vitro BBB model consisting of a co-culture of bovine brain capillary endothelial cells (BCECs) and rat glial cells that closely mimics the in vivo situation. Using this model, Ab1-40 and Ab1-42 peptide transports through the BCECs were evaluated. Competitive inhibitions were performed using RAP (Receptor-associated Protein) and CML (Carboxymethyl-lysine), specific ligands of LRP family members and RAGE respectively. Results: We have demonstrated that Ab1-40 and Ab1-42 peptides were not toxic for the integrity of the BBB. Moreover, we observed an asymmetric transport across the BBB suggesting the involvement of transporters and/or receptors. RAGE and LRP1 were both expressed by BCECs. Our results showed that RAP inhibited by 28% the Ab peptide efflux while CML decreased Ab peptide influx into the brain by 23%. Conclusions: These results reinforce the hypothesis of involvement of RAGE and LRPs receptors in Ab peptide transport across the BBB and provide interesting perspectives on the use of our BBB model in the study of these processes.

Boldine Attenuates Cancer Cell Growth in an Experimental Model of Glioma In vivo

Plants are important sources of biologically active natural products which differ in terms of structure and biological properties. One of the biological activities of plant compounds that attracts great interest is the ability to exert anticancer activity in different cancer types. Boldine is a natural alkaloid that occurs abundantly in the leaves and bark of Peumus boldus. It has been demonstrated to present anticancer properties in bladder carcinoma and in glioma cancer cells in vitro. In the present study, we evaluated the effect of boldine in an in vivo experimental model of glioma and investigated some parameters of toxicity in healthy animals. Our results show that boldine did not cause any toxicity in the animals treated. In rats with implanted glioma and treated with boldine a significant reduction in tumor size was observed. Pathological analysis of implanted gliomas showed a reduction in the mitotic index as well as in other histological characteristics in boldine treated animals, indicating a less invasive and proliferative tumor. No change was observed in the VEGF expression in treated animals. Our results suggest that boldine could be a promise adjuvant therapy in the treatment of malignant gliomas

Involvement of PI3-K pathway and β-catenin signaling in curcumin protecting against Aβ1-42-induced neurotoxicity

Alzheimer’s disease (AD) is the most prevalent form of dementia. New treatments to manage this complex illness will require full understanding of the pathophysiological mechanisms involved, which include amyloid β (Aβ)-induced toxicity. Substantial evidence indicates that curcumin has neuroprotective properties in AD; however, the molecular mechanisms involved in this process remain poorly understood. The aim of this study was investigate whether curcumin possesses a neuroprotective effect against Aβ-induced toxicity organotypic hippocampal cultures. Slices were exposed to Aβ, curcumin and/or LY294002, an inhibitor of phosphoinositide-3-kinase (PI3-K) pathway. Cell death was measured by propidium iodide uptake and some cell signaling pathways were investigated by Western blot assay. Moreover, we measured the synaptophysin expression, involved in the regulation of synaptic plasticity. Our results show that Aβ caused about 30% of cell damage in hippocampal slices, a significant increase when compared to controls cultures. The treatments with 5 and 10 μM of curcumin decreased the cell death significantly. The curcumin treatment prevented the decreased in synaptophysin expression after exposure to Aβ peptide. Aβ treatment increased the phosphorylated (Ser45) β-catenin and decreased βcatenin immunocontent, and the curcumin treatment prevented this β-catenin destabilization. Additionally, the curcumin neuroprotection was prevented by LY294002 and curcumin induced the phosphorylation/activation of Akt and the phosphorylation/inactivation of glycogen synthase kinase-3β (GSK-3β). These results reinforce the neuroprotective effect of curcumin and add some evidence that its mechanism may involve the PI3-K pathway and β-catenin signaling, a key transducer of the Wnt signaling pathway.