Non-steroidal anti-inflammatory drugs (NSAIDs) inhibit cyclooxygenase-1 (COX-1) and COX-2 enzymes. The NLRP3 inflammasome is a multi-protein complex responsible for the processing of the ...proinflammatory cytokine interleukin-1β and is implicated in many inflammatory diseases. Here we show that several clinically approved and widely used NSAIDs of the fenamate class are effective and selective inhibitors of the NLRP3 inflammasome via inhibition of the volume-regulated anion channel in macrophages, independently of COX enzymes. Flufenamic acid and mefenamic acid are efficacious in NLRP3-dependent rodent models of inflammation in air pouch and peritoneum. We also show therapeutic effects of fenamates using a model of amyloid beta induced memory loss and a transgenic mouse model of Alzheimer's disease. These data suggest that fenamate NSAIDs could be repurposed as NLRP3 inflammasome inhibitors and Alzheimer's disease therapeutics.
Recent data have revealed that soluble oligomeric amyloid-beta peptide (Abeta) may be the proximate effectors of neuronal injuries and death in Alzheimer's disease (AD) by unknown mechanisms. ...Consistently, we recently demonstrated the critical role of a redox-sensitive cytosolic calcium-dependent phospholipase A2 (cPLA2)-arachidonic acid (AA) pathway in Abeta oligomer-induced cell death. According to the involvement of oxidative stress and polyunsaturated fatty acids like AA in the regulation of sphingomyelinase (SMase) activity, the present study underlines the role of SMases in soluble Abeta-induced apoptosis. Soluble Abeta oligomers induced the activation of both neutral and acidic SMases, as demonstrated by the direct measurement of their enzymatic activities, by the inhibitory effects of both specific neutral and acidic SMase inhibitors, and by gene knockdown using antisense oligonucleotides. Furthermore, soluble Abeta-mediated activation of SMases and subsequent cell death were found to be inhibited by antioxidant molecules and a cPLA2-specific inhibitor or antisense oligonucleotide. We also demonstrate that sphingosine-1-phosphate is a potent neuroprotective factor against soluble Abeta oligomer-induced cell death and apoptosis by inhibiting soluble Abeta-induced activation of acidic sphingomyelinase. These results suggest that Abeta oligomers induce neuronal death by activating neutral and acidic SMases in a redox-sensitive cPLA2-AA pathway.
Despite the key role in neuronal development of a deficit in the methyl donor folate, little is known on the underlying mechanisms. We therefore studied the consequences of folate deficiency on ...proliferation, differentiation, and plasticity of the rat H19‐7 hippocampal cell line. Folate deficit reduced proliferation (17%) and sensitized cells to differentiation‐associated apoptosis (+16%). Decreased production (–58%) of S‐adenosylmethionine (the universal substrate for transmethylation reactions) and increased expression of histone deacetylases (HDAC4,6,7) would lead to epigenomic changes that may impair the differentiation process. Cell polarity, vesicular transport, and synaptic plasticity were dramatically affected, with poor neurite outgrowth (–57%). Cell treatment by an HDAC inhibitor (SAHA) led to a noticeable improvement of cell polarity and morphology, with longer processes. Increased homocysteine levels (+55%) consecutive to folate shortage produced homocysteinylation, evidenced by coimmunoprecipitations and mass spectrometry, and aggregation of motor proteins dynein and kinesin, along with functional alterations, as reflected by reduced interactions with partner proteins. Prominent homocysteinylation of key neuronal proteins and subsequent aggregation certainly constitute major adverse effects of folate deficiency, affecting normal development with possible long‐lasting consequences.—Akchiche, N., Bossenmeyer‐Pourié, C., Kerek, R., Martin, N., Pourié, G., Koziel, V., Helle, D., Alberto, J.‐M., Ortiou, S., Camadro, J.‐M., Léger, T., Guéant, J.‐L., Daval, J.‐L. Homocysteinylation of neuronal proteins contributes to folate deficiency‐associated alterations of differentiation, vesicular transport, and plasticity in hippocampal neuronal cells. FASEB J. 26, 3980–3992 (2012). www.fasebj.org
Whereas brief acute or intermittent episodes of hypoxia have been shown to exert a protective role in the central nervous system and to stimulate neurogenesis, other studies suggest that early ...hypoxia may constitute a risk factor that influences the future development of mental disorders. We therefore investigated the effects of a neonatal "conditioning-like" hypoxia (100% N₂, 5 min) on the brain and the cognitive outcomes of rats until 720 days of age (physiologic senescence). We confirmed that such a short hypoxia led to brain neurogenesis within the ensuing weeks, along with reduced apoptosis in the hippocampus involving activation of Erk1/2 and repression of p38 and death-associated protein (DAP) kinase. At 21 days of age, increased thicknesses and cell densities were recorded in various subregions, with strong synapsin activation. During aging, previous exposure to neonatal hypoxia was associated with enhanced memory retrieval scores specifically in males, better preservation of their brain integrity than controls, reduced age-related apoptosis, larger hippocampal cell layers, and higher expression of glutamatergic and GABAergic markers. These changes were accompanied with a marked expression of synapsin proteins, mainly of their phosphorylated active forms which constitute major players of synapse function and plasticity, and with increases of their key regulators, i.e. Erk1/2, the transcription factor EGR-1/Zif-268 and Src kinase. Moreover, the significantly higher interactions between PSD-95 scaffolding protein and NMDA receptors measured in the hippocampus of 720-day-old male animals strengthen the conclusion of increased synaptic functional activity and plasticity associated with neonatal hypoxia. Thus, early non-injurious hypoxia may trigger beneficial long term effects conferring higher resistance to senescence in aged male rats, with a better preservation of cognitive functions.
As a hepatic receptor for triglyceride-rich lipoproteins, the lipolysis-stimulated lipoprotein receptor (LSR) may be involved in the dynamics of lipid distribution between the liver and peripheral ...tissues. Here, we explore the potential role of leptin in regulating LSR. At physiological concentrations (1-10 ng/ml), leptin increased LSR protein and mRNA levels in Hepa1-6 cells through an ERK1/2-dependent and α-amanitin-sensitive pathway. In vivo, leptin treatment of C57BL6/Rj mice (1 μg 2x/d, 8 d) led to a significant increase in hepatic LSR mRNA and protein, decreased liver triglycerides and increased VLDL secretion as compared to controls. LSR⁺/⁻ mice with elevated postprandial lipemia placed on a high-fat (60% kcal) diet exhibited accelerated weight gain and increased fat mass as compared to controls. While plasma leptin levels were increased 3-fold, hepatic leptin receptor protein levels and phosphorylation of ERK1/2 were significantly reduced. Therefore, leptin is an important regulator of LSR protein levels providing the means for the control of hepatic uptake of lipids during the postprandial phase. However, this may no longer be functional in LSR⁺/⁻ mice placed under a chronic dietary fat load, suggesting that this animal model could be useful for the study of molecular mechanisms involved in peripheral leptin resistance.--Stenger, C., Hanse, M., Pratte, D., Mbala, M.-L., Akbar, S., Koziel, V., Escanyé, M.-C., Kriem, B., Malaplate-Armand, C., Olivier, J.-L., Oster, T., Pillot, T., Yen, F. T. Up-regulation of hepatic lipolysis stimulated lipoprotein receptor by leptin: a potential lever for controlling lipid clearance during the postprandial phase.
Methyl donor deficiency (MDD) during pregnancy influences intrauterine development. Ghrelin is expressed in the stomach of fetuses and influences fetal growth, but MDD influence on gastric ghrelin is ...unknown. We examined the gastric ghrelin system in MDD-induced intrauterine growth retardation. By using specific markers and approaches (such as periodic acid–Schiff, bromodeoxyuridine, homocysteine, terminal deoxynucleotidyl transferase biotin-dUTP nick end labeling, immunostaining, reverse transcription-polymerase chain reaction), we studied the gastric oxyntic mucosa cellular organization and ghrelin gene expression in the mucosa in 20-day-old fetuses and weanling pups, and plasma ghrelin concentration in weanling rat pups of dams either normally fed or deprived of choline, folate, vitamin B6, and vitamin B12 during gestation and suckling periods. MDD fetuses weighed less than controls; the weight deficit reached 57% at weaning ( P < 0.001). Both at the end of gestation and at weaning, they presented with an aberrant gastric oxyntic mucosa formation with loss of cell polarity, anarchic cell migration, abnormal progenitor differentiation, apoptosis, and signs of surface layer erosion. Ghrelin cells were abnormally located in the pit region of oxyntic glands. At weaning, plasma ghrelin levels were decreased (−28%; P < 0.001) despite unchanged mRNA expression in the stomach. This decrease was associated with lower body weight. Taken together, these data indicate that one mechanism through which MDD influences fetal programming is the remodeling of gastric cellular organization, leading to dysfunction of the ghrelin system and dramatic effects on growth.
Increased amounts of bilirubin, the end product of heme degradation, are known to be detrimental to the central nervous system, especially in preterm newborns. In an attempt to delineate the cellular ...mechanisms by which unconjugated bilirubin exerts its toxic effects on neuronal cells in the developing brain, bilirubin (0.25–5 μM) was added to the extracellular medium of 6-day-old primary cultured neurons from the embryonic rat forebrain, and cell alterations were studied over the ensuing 96 h. Bilirubin decreased cell viability dose dependently with an ED50 around 1 μM. At the dose of 0.5 μM, it triggered delayed cell death that affected 24% of the neurons. Nuclear incorporation of the fluorescent dye DAPI (4,6-diamidino-2-phenylindole) depicted the presence of apoptosis (16%). Apoptosis features were confirmed by DNA fragmentation reflected by a progressive loss of 3Hthymidine and sequential changes in macromolecular synthesis, as shown by the time course of 3Hleucine incorporation, as well as by the beneficial effects of cycloheximide and caspase inhibitors. In parallel, treatments with glutamate receptor antagonists showed that MK-801, but not NBQX, protected neurons against bilirubin neurotoxicity, suggesting a role for NMDA receptors in bilirubin effects. Coupled with previous work about glutamate toxicity in the same culture model, these data support the hypothesis that low levels of free bilirubin may promote programmed neuronal death corresponding to an apoptotic process which involves caspase activation and requires the participation of NMDA receptors, along with bilirubin-induced inhibition of protein kinase C activity.
Abstract Alzheimer's disease (AD) is a major public health concern in all countries. Although the precise cause of AD is still unknown, a growing body of evidence supports the notion that soluble ...amyloid β-peptide (Aβ) may be the proximate cause of synaptic injuries and neuronal death early in the disease. AD patients display lower levels of docosahexaenoic acid (DHA, C22:6 ; n-3) in plasma and brain tissues as compared to age-matched controls. Furthermore, epidemiological studies suggest that high DHA intake might have protective properties against neurodegenerative diseases. These observations are supported by in vivo studies showing that DHA-rich diets limits the synaptic loss and cognitive defects induced by Aβ peptide. Although the molecular basis of these neuroprotective effects remains unknown, several mechanisms have been proposed such as (i) regulation of the expression of potentially protective genes, (ii) activation of anti-inflammatory pathways, (iii) modulation of functional properties of the synaptic membranes along with changes in their physicochemical and structural features.
To assess temporal brain deficits consecutive to severe birth hypoxia, newborn rats were exposed for 20 min to 100% N
2. This treatment induced a long-term growth retardation and a delayed, but only ...transient, neuronal loss (∼25%) in the CA
1 hippocampus and parietal cortex, starting from 3 days and peaking at 6 days post-hypoxia. The expression profiles of various apoptosis-regulating proteins (including Bcl-2, Bax, p53 and caspase-3) were well correlated to the alterations of nuclear morphology depicted by 4,6-diamidino-2-phenylindole (DAPI). Whereas they confirmed a gradual histological recovery, specific DNA fragmentation patterns suggested that birth hypoxia may transiently reactivate the developmental programme of neuronal elimination. Although they successfully achieved various behavioral tests such as the righting reflex, negative geotaxis, locomotor coordination, and the eight-arm maze tasks, both developing and adult hypoxic rats were repeatedly slower than controls, suggesting that birth hypoxia is associated to moderate but persistent impairments of functional capacities.
The development of novel therapeutic strategies for Alzheimer's disease (AD) represents one of the biggest unmet medical needs today. Application of neurotrophic factors able to modulate neuronal ...survival and synaptic connectivity is a promising therapeutic approach for AD. We aimed to determine whether the loco-regional delivery of ciliary neurotrophic factor (CNTF) could prevent amyloid-beta (Abeta) oligomer-induced synaptic damages and associated cognitive impairments that typify AD. To ensure long-term administration of CNTF in the brain, we used recombinant cells secreting CNTF encapsulated in alginate polymers. The implantation of these bioreactors in the brain of Abeta oligomer-infused mice led to a continuous secretion of recombinant CNTF and was associated with the robust improvement of cognitive performances. Most importantly, CNTF led to full recovery of cognitive functions associated with the stabilization of synaptic protein levels in the Tg2576 AD mouse model. In vitro as well as in vivo, CNTF activated a Janus kinase/signal transducer and activator of transcription-mediated survival pathway that prevented synaptic and neuronal degeneration. These preclinical studies suggest that CNTF and/or CNTF receptor-associated pathways may have AD-modifying activity through protection against progressive Abeta-related memory deficits. Our data also encourage additional exploration of ex vivo gene transfer for the prevention and/or treatment of AD.
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