Microglial glutamate release evoked by α-synuclein aggregates is prevented by dopamine.

When activated, microglial cells have the potential not only to secrete typical proinflammatory mediators but also to release the neurotransmitter glutamate in amounts that may promote excitotoxicity. Here, we wished to determine the potential of the Parkinson's disease (PD) protein α-Synuclein (αS) to stimulate glutamate release using cultures of purified microglial cells. We established that glutamate release was robustly increased when microglial cultures were treated with fibrillary aggregates of αS but not with the native monomeric protein. Promotion of microglial glutamate release by αS aggregates (αSa) required concomitant engagement of TLR2 and P2X7 receptors. Downstream to cell surface receptors, the release process was mediated by activation of a signaling cascade sequentially involving phosphoinositide 3-kinase (PI3K) and NADPH oxidase, a superoxide-producing enzyme. Inhibition of the Xc- antiporter, a plasma membrane exchange system that imports extracellular l-cystine and exports intracellular glutamate, prevented the release of glutamate induced by αSa, indicating that system Xc- was the final effector element in the release process downstream to NADPH oxidase activation. Of interest, the stimulation of glutamate release by αSa was abrogated by dopamine through an antioxidant effect requiring D1 dopamine receptor activation and PI3K inhibition. Altogether, present data suggest that the activation of microglial cells by αSa may possibly result in a toxic build-up of extracellular glutamate contributing to excitotoxic stress in PD. The deficit in dopamine that characterizes this disorder may further aggravate this process in a vicious circle mechanism.

Contributive Role of TNF-α to L-DOPA-Induced Dyskinesia in a Unilateral 6-OHDA Lesion Model of Parkinson's Disease.

Our present objective was to better characterize the mechanisms that regulate striatal neuroinflammation in mice developing L-DOPA-induced dyskinesia (LID). For that, we used 6-hydroxydopamine (6-OHDA)-lesioned mice rendered dyskinetic by repeated intraperitoneal injections of 3,4-dihydroxyphenyl-L-alanine (L-DOPA) and quantified ensuing neuroinflammatory changes in the dopamine-denervated dorsal striatum. LID development was associated with a prominent astrocytic response, and a more moderate microglial cell reaction restricted to this striatal area. The glial response was associated with elevations in two pro-inflammatory cytokines, tumor necrosis factor-α (TNF-α) and interleukin-1ß. Treatment with the phytocannabinoid cannabidiol and the transient receptor potential vanilloid-1 (TRPV-1) channel antagonist capsazepine diminished LID intensity and decreased TNF-α levels without impacting other inflammation markers. To possibly reproduce the neuroinflammatory component of LID, we exposed astrocyte and microglial cells in culture to candidate molecules that might operate as inflammatory cues during LID development, i.e., L-DOPA, dopamine, or glutamate. Neither L-DOPA nor dopamine produced an inflammatory response in glial cell cultures. However, glutamate enhanced TNF-α secretion and GFAP expression in astrocyte cultures and promoted Iba-1 expression in microglial cultures. Of interest, the antidyskinetic treatment with cannabidiol + capsazepine reduced TNF-α release in glutamate-activated astrocytes. TNF-α, on its own, promoted the synaptic release of glutamate in cortical neuronal cultures, whereas cannabidiol + capsazepine prevented this effect. Therefore, we may assume that the release of TNF-α by glutamate-activated astrocytes may contribute to LID by exacerbating corticostriatal glutamatergic inputs excitability and maintaining astrocytes in an activated state through a self-reinforcing mechanism.

Rifampicin and Its Derivative Rifampicin Quinone Reduce Microglial Inflammatory Responses and Neurodegeneration Induced In Vitro by α-Synuclein Fibrillary Aggregates.

: Aggregated forms of the synaptic protein α-synuclein (αS) have been proposed to operate as a molecular trigger for microglial inflammatory processes and neurodegeneration in Parkinson´s disease. Here, we used brain microglial cell cultures activated by fibrillary forms of recombinant human αS to assess the anti-inflammatory and neuroprotective activities of the antibiotic rifampicin (Rif) and its autoxidation product rifampicin quinone (RifQ). Pretreatments with Rif and RifQ reduced the secretion of prototypical inflammatory cytokines (TNF-, IL-6) and the burst of oxidative stress in microglial cells activated with αS fibrillary aggregates. Note, however, that RifQ was constantly more efficacious than its parent compound in reducing microglial activation. We also established that the suppressive effects of Rif and RifQ on cytokine release was probably due to inhibition of both PI3K- and non-PI3K-dependent signaling events. The control of oxidative stress appeared, however, essentially dependent on PI3K inhibition. Of interest, we also showed that RifQ was more efficient than Rif in protecting neuronal cells from toxic factors secreted by microglia activated by αS fibrils. Overall, data with RifQ are promising enough to justify further studies to confirm the potential of this compound as an anti-parkinsionian drug.

New use for an old drug: COX-independent anti-inflammatory effects of sulindac in models of cystic fibrosis.

BACKGROUND AND PURPOSE: Pulmonary disease is the main cause of morbidity and mortality in cystic fibrosis (CF) patients due to exacerbated inflammation. To date, the only anti-inflammatory drug available to CF patients is high-dose ibuprofen, which can slow pulmonary disease progression, but whose cyclooxygenase-dependent digestive adverse effects limit its clinical use. Here we have tested sulindac, another non-steroidal anti-inflammatory drug with an undefined anti-inflammatory effect in CF airway epithelial cells. EXPERIMENTAL APPROACH: Using in vitro and in vivo models, we NF-κB activity and IL-8 secretion. In HeLa-F508del cells, we performed luciferase reporter gene assays in order to measure i) IL-8 promoter activity, and ii) the activity of synthetic promoter containing NF-κB responsive elements. We quantified IL-8 secretion in airway epithelial CFBE cells cultured at an air-liquid interface and in a mouse model of CF. KEY RESULTS: Sulindac inhibited the transcriptional activity of NF-κB and decreased IL-8 transcription and secretion in TNF-α stimulated CF cells via a cyclooxygenase-independent mechanism. This effect was confirmed in vivo in a mouse model of CF induced by intra-tracheal instillation of LPS, with a significant decrease of the induction of mRNA for MIP-2, following treatment with sulindac. CONCLUSION AND IMPLICATIONS: Overall, sulindac decrease lung inflammation by a mechanism independent of cycolooxygenase. This drug could be beneficially employed in CF.

COMMD1 modulates noxious inflammation in cystic fibrosis.

Cystic fibrosis (CF) is an autosomal recessive disease caused by mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene, which encodes an epithelial anion channel. Morbidity is mainly due to lung disease, which is characterized by chronic neutrophilic inflammation. Deregulation of inflammatory pathways is observed in the airways of CF patients, as evidenced by exaggerated NF-κB activity, causing an increase in the local release of pro-inflammatory cytokines such as IL-8. COMMD1, a pleiotropic protein, was recently shown to interact with CFTR and to promote CFTR cell surface expression. The effect of COMMD1 on the NF-κB pathway was assessed in CF and non-CF bronchial epithelial cells by knockdown and overexpression experiments. Results showed that (i) COMMD1 knockdown induced NF-κB-dependent transcription, (ii) COMMD1 overexpression inhibited NF-κB activity and was associated with a decrease in IL-8 transcript level and protein secretion. These data demonstrate the anti-inflammatory properties of COMMD1 in bronchial epithelial cells and open new therapeutic avenues in CF.