Agnès Nadjar

Short-Term Long Chain Omega3 Diet Protects from Neuroinflammatory Processes and Memory Impairment in Aged Mice

Regular consumption of food enriched in omega3 polyunsaturated fatty acids (ω3 PUFAs) has been shown to reduce risk of cognitive decline in elderly, and possibly development of Alzheimers disease. Docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA) are the most likely active components of ω3-rich PUFAs diets in the brain. We therefore hypothesized that exposing mice to a DHA and EPA enriched diet may reduce neuroinflammation and protect against memory impairment in aged mice. For this purpose, mice were exposed to a control diet throughout life and were further submitted to a diet enriched in EPA and DHA during 2 additional months. Cytokine expression together with a thorough analysis of astrocytes morphology assessed by a 3D reconstruction was measured in the hippocampus of young (3-month-old) and aged (22-month-old) mice. In addition, the effects of EPA and DHA on spatial memory and associated Fos activation in the hippocampus were assessed. We showed that a 2-month EPA/DHA treatment increased these long-chain ω3 PUFAs in the brain, prevented cytokines expression and astrocytes morphology changes in the hippocampus and restored spatial memory deficits and Fos-associated activation in the hippocampus of aged mice. Collectively, these data indicated that diet-induced accumulation of EPA and DHA in the brain protects against neuroinflammation and cognitive impairment linked to aging, further reinforcing the idea that increased EPA and DHA intake may provide protection to the brain of aged subjects.

Inactivation of the cerebral NFκB pathway inhibits interleukin-1β-induced sickness behavior and c-Fos expression in various brain nuclei

The behavioral effects of peripherally administered interleukin-1β (IL-1β) are mediated by the production of cytokines and other proinflammatory mediators at the level of the blood–brain interface and by activation of neural pathway. To assess whether this action is mediated by NFκB activation, rats were injected into the lateral ventricle of the brain with a specific inhibitor of NFκB activation, the NEMO Binding Domain (NBD) peptide that has been shown previously to abolish completely IL-1β-induced NFκB activation and Cox-2 synthesis in the brain microvasculature. NFκB pathway inactivation significantly blocked the behavioral effects of intraperitoneally administered IL-1β in the form of social withdrawal and decreased food intake, and dramatically reduced IL-1β-induced c-Fos expression in various brain regions as paraventricular nucleus, supraoptic nucleus, and lateral part of the central amygdala. These findings strongly support the hypothesis that IL-1β-induced NFκB activation at the blood–brain interface is a crucial step in the transmission of immune signals from the periphery to the brain that underlies further events responsible of sickness behavior.

Nuclear factor κB nuclear translocation as a crucial marker of brain response to interleukin-1. A study in rat and interleukin-1 type I deficient mouse

The signalling pathways that mediate early central effects of interleukin‐1 (IL‐1) during the acute phase reaction have been poorly elucidated. Interaction of IL‐1β to its specific receptor interleukin‐1 receptor type I (IL‐1RI) leads to nuclear factor kappa B (ΝFκB) nuclear translocation and a robust transcriptional activation of inhibitor of kappa B alpha (IκBα) within the rat brain. Indeed, we demonstrated that IL‐1RI expressed in blood brain barrier (BBB) cells and in circumventricular organs (CVOs) is crucial for p65‐NFκB translocation induced by peripheral injection of IL‐1β. Moreover, it has been previously shown that monitoring IκBα mRNA synthesis is an effective tool to investigate the activity of the transcription factor NFκB into the CNS. However in the present study we observed time‐related and cell‐type differences between IκBα mRNA synthesis and p65‐NFκB translocation. This indicates that the expression of IκBα mRNA does not strictly parallel p65‐NFκB nuclear translocation, suggesting that these markers are not interchangeable to investigate NFκB activity but must be studied together. Thus, we hypothesize that IL‐1β reached the brain across the CVOs that lack a BBB and endothelial cells all over the brain and interacted with its receptors to induce NFκB translocation. The study of the consequences of the impairment of NFκB pathway activation in in vivo experimentation should bring important clues about the precise role of this transcription factor.

NFκB activates in vivo the synthesis of inducible Cox-2 in the brain

Interleukin-1β (IL-1β) induces cyclooxygenase-2 (Cox-2) expression in many of its cellular targets resulting in production and release of prostaglandins. Although IL-1β-induced Cox-2 expression most likely requires activation of nuclear transcription factor kappa B (NFκB) pathway, this has never been formally demonstrated in vivo. We tested this using a specific inhibitor of NFκB activation, the NEMO binding domain (NBD) peptide, that has been shown previously to be effective in various in vivo models of acute inflammation. Incubation of rat glioma cells with the NBD peptide blocked IL-1β-induced NFκB nuclear translocation. Furthermore, after injection of a biotinylated version of the NBD peptide into the lateral ventricle of the brain, we found that it readily diffused to its potential cellular targets in vivo. To test the effects of the peptide on NFκB activation and Cox-2 expression in the brain, we injected it intracerebroventricularly (36 μg/rat) into rats before intraperitoneal injection of IL-1β (60 μg/kg). Treatment with NBD peptide completely abolished IL-1β-induced NFκB activation and Cox-2 synthesis in microvasculature. In contrast, the peptide had no effect on constitutive neuronal Cox-2. These findings strongly support the hypothesis that IL-1β-induced NFκB activation plays a major role in transmission of immune signals from the periphery to the brain.

Nutritional n-3 PUFAs deficiency during perinatal periods alters brain innate immune system and neuronal plasticity-associated genes

Low dietary intake of the n-3 polyunsaturated fatty acids (PUFAs) is a causative factor of neurodevelopmental disorders. However the mechanisms linking n-3 PUFAs low dietary intake and neurodevelopmental disorders are poorly understood. Microglia, known mainly for their immune function in the injured or infected brain, have recently been demonstrated to play a pivotal role in regulating maturation of neuronal circuits during normal brain development. Disruption of this role during the perinatal period therefore could significantly contribute to psychopathologies with a neurodevelopmental neurodevelopmental component. N-3 PUFAs, essential lipids and key structural components of neuronal membrane phospholipids, are highly incorporated in cell membranes during the gestation and lactation phase. We previously showed that in a context of perinatal n-3 PUFAs deficiency, accretion of these latter is decreased and this is correlated to an alteration of endotoxin-induced inflammatory response. We thus postulated that dietary n-3 PUFAs imbalance alters the activity of microglia in the developing brain, leading to abnormal formation of neuronal networks. We first confirmed that mice fed with a n-3 PUFAs deficient diet displayed decreased n-3 PUFAs levels in the brain at post-natal days (PND)0 and PND21. We then demonstrated that n-3 PUFAs deficiency altered microglia phenotype and motility in the post-natal developing brain. This was paralleled by an increase in pro-inflammatory cytokines expression at PND21 and to modification of neuronal plasticity-related genes expression. Overall, our findings show for the first time that a dietary n-3 PUFAs deficiency from the first day of gestation leads to the development of a pro-inflammatory condition in the central nervous system that may contribute to neurodevelopmental alterations.

n-3 LCPUFA improves cognition: The young, the old and the sick

Due to the implication of docosahexaenoic acid (DHA) in neurogenesis, synaptogenesis, neurite outgrowth and to its high incorporation into the brain, this n-3 long chain polyunsaturated fatty acid (LCPUFA) is considered as crucial in the development and maintenance of the learning memory performance throughout life. In the present chapter we aimed at reviewing data investigating the relation between DHA and cognition during the perinatal period, young adult- and adulthood and neurodegenerative diseases such as Alzheimer disease (AD). In Humans, dietary DHA supplementation from the perinatal period to adulthood does not reveal a clear and consistent memory improvement whereas it is the case in animal studies. The positive effects observed in animal models may have been enhanced by using n-3 PUFA deficient animal models as controls. In animal models of AD, a general consensus on the beneficial effects of n-3 LCPUFA in attenuating cognitive impairment was established. These studies make DHA a potential suitable micronutrient for the maintenance of cognitive performance at all periods of life.

Transgenic Increase in n-3/n-6 Fatty Acid Ratio Protects Against Cognitive Deficits Induced by an Immune Challenge through Decrease of Neuroinflammation

Polyunsaturated fatty acids (PUFAs) display immunomodulatory properties in the brain, n-3 PUFAs being able to reduce inflammation whereas n-6 PUFAs are more pro-inflammatory. It has been extensively demonstrated that exposure to a peripheral immune challenge leads to the production and release of inflammatory mediators in the brain in association with cognitive deficits. The question arises whether n-3 PUFA supplementation could downregulate the brain inflammatory response and subsequent cognitive alterations. In this study, we used a genetically modified mouse line carrying the fat-1 gene from the roundworm Caenorhabditis elegans, encoding an n-3 PUFA desaturase that catalyzes conversion of n-6 into n-3 PUFA. Consequently, these mice display endogenously elevated n-3 PUFA tissue contents. Fat-1 mice or wild-type (WT) littermates were injected peripherally with lipopolysaccharide (LPS), a bacterial endotoxin, to induce an inflammatory episode. Our results showed that LPS altered differently the phenotype of microglia and the expression of cytokines and chemokines in Fat-1 and WT mice. In Fat-1 mice, pro-inflammatory factors synthesis was lowered compared with WT mice, whereas anti-inflammatory mechanisms were favored 24 h after LPS treatment. Moreover, LPS injection impaired spatial memory in WT mice, whereas interestingly, the Fat-1 mice showed normal cognitive performances. All together, these data suggest that the central n-3 PUFA increase observed in Fat-1 mice modulated the brain innate immune system activity, leading to the protection of animals against LPS-induced pro-inflammatory cytokine production and subsequent spatial memory alteration.

Astrocyte-derived adenosine modulates increased sleep pressure during inflammatory response.

Activation of the immune system elicits several behavioral changes collectively called sickness. Among the behavioral changes, systemic infections induce an increase in time spent in nonrapid‐eye‐movement (NREM) sleep and an increase of slow wave activity (or “sleep pressure”). Using an inducible, astrocyte‐specific transgenic dominant negative SNARE (dnSNARE) mouse line we recently demonstrated that gliotransmission plays an important role in sleep homeostasis through an adenosine receptor 1 (A1R)‐sensitive pathway. It has been hypothesized that systemic infection, mimicked by peripheral administration of lipopolysaccharide (LPS), increases sleeping behavior in part through upregulation of central adenosine levels. Moreover, as a source of immunologically relevant factors, astrocytes play a pivotal role in the central nervous system immune and inflammatory responses. However, little is known about the role of astrocytes in the CNS response to a peripheral immune challenge. We hypothesize that LPS impacts sleep homeostasis through the modulation of astrocyte‐derived adenosine accumulation. We therefore used dnSNARE mice to determine whether astrocytes contribute to the increased sleep pressure under immune challenge and whether this is a result of changes in adenosine signaling. We demonstrate that dnSNARE‐mediated gliotransmission is required for the ability of LPS to elevate sleep pressure as measured by the power of slow wave activity during NREM sleep. Moreover, in agreement with a role of astrocyte‐derived adenosine in modulating sleep homeostasis, we find that intracerebroventricular infusion of the A1R antagonist 8‐cyclopentyl‐1,3‐dimethylxanthine (CPT) mimics this dnSNARE phenotype. Taken together, our data demonstrate that astrocytic adenosine acting through A1 receptors contributes to the modulation of sleep pressure by LPS. GLIA 2013

Resolvin D1 and E1 promote resolution of inflammation in microglial cells in vitro.

Sustained inflammation in the brain together with microglia activation can lead to neuronal damage. Hence limiting brain inflammation and activation of microglia is a real therapeutic strategy for inflammatory disease. Resolvin D1 (RvD1) and resolvin E1 (RvE1) derived from n-3 long chain polyunsaturated fatty acids are promising therapeutic compounds since they actively turn off the systemic inflammatory response. We thus evaluated the anti-inflammatory activities of RvD1 and RvE1 in microglia cells in vitro. BV2 cells were pre-incubated with RvD1 or RvE1 before lipopolysaccharide (LPS) treatment. RvD1 and RvE1 both decreased LPS-induced proinflammatory cytokines (TNF-α, IL-6 and IL-1β) gene expression, suggesting their proresolutive activity in microglia. However, the mechanisms involved are distinct as RvE1 regulates NFκB signaling pathway and RvD1 regulates miRNAs expression. Overall, our findings support that pro-resolving lipids are involved in the resolution of brain inflammation and can be considered as promising therapeutic agents for brain inflammation.

Signaling pathways of interleukin-1 actions in the brain: Anatomical distribution of phospho-ERK1/2 in the brain of rat treated systemically with interleukin-1β

Interleukin-1beta is released at the periphery during infection and acts on the nervous system to induce fever, neuroendocrine activation, and behavioral changes. These effects are mediated by brain type I IL-1 receptors. In vitro studies have shown the ability of interleukin-1beta to activate mitogen-activated protein kinase signaling pathways including p38, c-Jun N-terminal kinase and extracellular signal-regulated protein kinase 1 and 2 (ERK1/2). In contrast to other mitogen-activated protein kinases, little is known about ERK1/2 activation in the rat brain in response to interleukin-1beta. The aim of the present study was therefore to investigate spatial and temporal activation of ERK1/2 in the rat brain after peripheral administration of interleukin-1beta using immunohistochemistry to detect the phosphorylated form of the kinase. In non-stimulated conditions, phosphorylated ERK1/2 immunoreactivity was observed in neurons throughout the brain. Administration of interleukin-1beta (60 microg/kg, i.p.) induced the phosphorylation of ERK1/2 in areas at the interface between brain and blood or cerebrospinal fluid: meninges, circumventricular organs, endothelial like cells of the blood vessels, and in brain nuclei involved in behavioral depression, fever and neuroendocrine activation: paraventricular nucleus of the hypothalamus, supraoptic nucleus, central amygdala and arcuate nucleus. Double labeling of phosphorylated ERK1/2 and cell markers revealed the expression of phosphorylated ERK1/2 in neurons, astrocytes and microglia. Since phosphorylated ERK1/2 was found in structures in which type I IL-1 receptor has already been identified as well as in structures lacking this receptor, activation of ERK1/2 is likely to occur in response to both direct and indirect action of interleukin-1beta on its target cells.