Brain Vitamin E Deficiency During Development Is Associated With Increased Glutamate Levels and Anxiety in Adult Mice

International audienceVitamin E, the most important lipophilic radical scavenging antioxidant in vivo, has a pivotal role in brain. In an earlier study, we observed that adult mice with a defect in the gene encoding plasma phospholipid transfer protein (PLTP) display a moderate reduction in cerebral vitamin E levels, and exacerbated anxiety despite normal locomotion and memory functions. Here we sought to determine whether dietary vitamin E supplementation can modulate neurotransmitter levels and alleviate the increased anxiety phenotype of PLTP-deficient (PLTP -/-) mice. To address this question, a vitamin E-enriched diet was used, and two complementary approches were implemented: (i) "early supplementation": neurotransmitter levels and anxiety were assessed in 6 months old PLTP -/- mice born from vitamin E-supplemented parents; and (ii) "late supplementation": neurotransmitter levels and anxiety were assessed in 6 months old PLTP -/- mice fed a vitamin E-enriched diet from weaning. Our results show for the first time that an inadequate supply of vitamin E during development, due to moderate maternal vitamin E deficiency, is associated with reduced brain vitamin E levels at birth and irreversible alterations in brain glutamate levels. They also suggest this deficiency is associated with increased anxiety at adulthood. Thus, the present study leads to conclude on the importance of the micronutrient vitamin E during pregnancy

Phospholipid transfer protein (PLTP), lipopolysaccharides detoxification and metabolic diseases: a connection ?

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Phospholipid transfer protein (PLTP) and metabolic diseases

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Deletion of plasma Phospholipid Transfer Protein (PLTP) increases microglial phagocytosis and reduces cerebral amyloid-β deposition in the J20 mouse model of Alzheimer's disease

International audiencePlasma phospholipid transfer protein (PLTP) binds and transfers a number of amphipathic compounds, including phospholipids, cholesterol, diacylglycerides, tocopherols and lipopolysaccharides. PLTP functions are relevant for many pathophysiological alterations involved in neurodegenerative disorders (especially lipid metabolism, redox status, and immune reactions), and a significant increase in brain PLTP levels was observed in patients with Alzheimer's disease (AD) compared to controls. To date, it has not been reported whether PLTP can modulate the formation of amyloid plaques, i.e. one of the major histopathological hallmarks of AD. We thus assessed the role of PLTP in the AD context by breeding PLTP-deficient mice with an established model of AD, the J20 mice. A phenotypic characterization of the amyloid pathology was conducted in J20 mice expressing or not PLTP. We showed that PLTP deletion is associated with a significant reduction of cerebral Aβ deposits and astrogliosis, which can be explained at least in part by a rise of Aβ clearance through an increase in the microglial phagocytic activity and the expression of the Aβ-degrading enzyme neprilysin. PLTP arises as a negative determinant of plaque clearance and over the lifespan, elevated PLTP activity could lead to a higher Aβ load in the brain

Influence de la protéine plasmatique de transfert des phospholipides sur la translocation intestinale des LPS, leur prise en charge par les lipoprotéines, et leurs effets métaboliques

International audienceIntroduction et but de l’étudeLes LPS (lipopolysaccharides, endotoxines) provenant des bactéries à Gram négatif peuvent interagir avec la barrière intestinale, altérer son intégrité et ainsi passer dans le compartiment sanguin. L’endotoxémie qui résulte de cette translocation intestinale peut provoquer une inflammation métabolique à bas bruit associée aux maladies métaboliques telles que l’obésité et le diabète de type 2. Dans le compartiment vasculaire, certaines protéines telles que la PLTP (protéine plasmatique de transfert des phospholipides) peuvent transférer les LPS aux lipoprotéines circulantes et ainsi favoriser leur détoxification. L’objectif de ce projet était d’étudier la cinétique de translocation de ces LPS ainsi que l’influence de la PLTP sur ce phénomène dans un contexte inflammatoire.Matériel et méthodesDes LPS (E. coli 055:B5) ont été marqués avec un traceur fluorescent (DOTAGA-Bodipy), avant administration par voie orale à des souris C57 BL/6J type-sauvage (WT) ou dont le gène de la PLTP a été invalidé (PLTPKO). Les différents segments intestinaux ont ensuite été prélevés afin de quantifier des LPS marqués et déterminer l’expression de gènes d’intérêt dans le métabolisme des lipoprotéines. Nous avons également mesuré la concentration plasmatique en LPS et de marqueurs de l’inflammation (cytokines et glucagon-like peptide-1 [GLP-1]). L’effet de premier passage hépatique et l’association des LPS au niveau des différentes fractions lipoprotéiques permettant leur détoxification ont été évalués après dosage des LPS dans le sang portal mais également le sang périphérique chez nos deux groupes de souris une heure après administration orale de LPS marqués. Enfin, nous avons étudié la contribution possible des facteurs impliqués dans l’assemblage (apoB, MTP…) et la clairance des lipoprotéines triches en triglycérides (lipoprotéine lipase [LPL] ainsi que ses modulateurs, les apolipoprotéines CII et CIII), sur la détoxification de ces toxines bactériennes ainsi que sur le métabolisme des lipoprotéines circulantes après gavage.RésultatsLes LPS sont captés de manière très précoce (entre 30 et 60 minutes après gavage) au niveau des segments proximaux de l’intestin grêle (duodénum et le jéjunum) tandis qu’un pic de concentration de LPS marqués dans le plasma est quant à lui visible 60 minutes post-gavage. L’absence de PLTP favorise la translocation intestinale des LPS provoquant ainsi une accumulation plus marquée de ces derniers dans le compartiment plasmatique, ceci malgré une expression significativement moins importante du gène et de la protéine apoB, nécessaire à l’assemblage des lipoprotéines riches en triglycérides (notamment les chylomicrons). Les souris PLTPKO présente un défaut de production de GLP1 mais une expression accrue de cytokines inflammatoires (interleukine 6, TNF-α) en réponse à un gavage de LPS par rapport à des souris WT. Après translocation intestinale, la majorité des LPS sont associés aux lipoprotéines circulantes, notamment les lipoprotéines riches en triglycérides et les lipoprotéines de haute densité (HDL) chez les souris WT alors que la majorité des LPS reste dans la fraction libre chez les souris déficientes en PLTP. L’effet bénéfique du premier passage hépatique est quant à lui plus marqué chez les souris ayant une PLTP active. Enfin, l’activité LPL chez les souris PLTPKO est plus faible que chez les souris WT, en parallèle avec des niveaux accrus d’ApoCIII, inhibiteur de la LPL. Ces données suggèrent ainsi la présence d’un défaut de clairance des triglycérides conduisant, in fine, à une forte accumulation des LPS plasmatiques.ConclusionLa déficience en PLTP favorise le transfert des LPS de la lumière intestinale vers la circulation, où son association aux lipoprotéines plasmatique est diminuée en parallèle avec un défaut de clairance des lipoprotéines riches en triglycérides. L’absence de PLTP induit une forte incapacité à détoxifier efficacement les LPS d’origine intestinale, conduisant à une augmentation de la réponse inflammatoire. La PLTP possède donc un rôle majeur dans la translocation intestinale des LPS et sur leur devenir métabolique

Quantitative lipopolysaccharide analysis using HPLC/MS/MS and its combination with the limulus amebocyte lysate assay

International audienceQuantitation of plasma lipopolysaccharides (LPSs) might be used to document Gram-negative bacterial infection. In the present work, LPS-derived 3-hydroxymyristate was extracted from plasma samples with an organic solvent, separated by reversed phase HPLC, and quantitated by MS/MS. This mass assay was combined with the limulus amebocyte lysate (LAL) bioassay to monitor neutralization of LPS activity in biological samples. The described HPLC/MS/MS method is a reliable, practical, accurate, and sensitive tool to quantitate LPS. The combination of the LAL and HPLC/MS/MS analyses provided new evidence for the intrinsic capacity of plasma lipoproteins and phospholipid transfer protein to neutralize the activity of LPS. In a subset of patients with systemic inflammatory response syndrome, with documented infection but with a negative plasma LAL test, significant amounts of LPS were measured by the HPLC/MS/MS method. Patients with the highest plasma LPS concentration were more severely ill. HPLC/MS/MS is a relevant method to quantitate endotoxin in a sample, to assess the efficacy of LPS neutralization, and to evaluate the proinflammatory potential of LPS in vivo

Plasma phospholipid transfer protein (PLTP) modulates adaptive immune functions through alternation of T helper cell polarization

International audienceObjective: Plasma phospholipid transfer protein (PLTP) is a key determinant of lipoprotein metabolism, and both animal and human studies converge to indicate that PLTP promotes atherogenesis and its thromboembolic complications. Moreover, it has recently been reported that PLTP modulates inflammation and immune responses. Although earlier studies from our group demonstrated that PLTP can modify macrophage activation, the implication of PLTP in the modulation of T-cell-mediated immune responses has never been investigated and was therefore addressed in the present study. Approach and results: In the present study, we demonstrated that PLTP deficiency in mice has a profound effect on CD4(+) Th0 cell polarization, with a shift towards the anti-inflammatory Th2 phenotype under both normal and pathological conditions. In a model of contact hypersensitivity, a significantly impaired response to skin sensitization with the hapten-2,4-dinitrofluorobenzene (DNFB) was observed in PLTP-deficient mice compared to wild-type (WT) mice. Interestingly, PLTP deficiency in mice exerted no effect on the counts of total white blood cells, lymphocytes, granulocytes, or monocytes in the peripheral blood. Moreover, PLTP deficiency did not modify the amounts of CD4(+) and CD8(+) T lymphocyte subsets. However, PLTP-deficiency, associated with upregulation of the Th2 phenotype, was accompanied by a significant decrease in the production of the pro-Th1 cytokine interleukin 18 by accessory cells. Conclusions: For the first time, this work reports a physiological role for PLTP in the polarization of CD4(+) T cells toward the pro-inflammatory Th1 phenotype

Human cholesteryl ester transfer protein lacks lipopolysaccharide transfer activity, but worsens inflammation and sepsis outcomes in mice

International audienceBacterial lipopolysaccharides (LPSs or endotoxins) can bind most proteins of the lipid transfer/LPS-binding protein (LT/LBP) family in host organisms. The LPS-bound LT/LBP proteins then trigger either an LPS-induced proinflammatory cascade or LPS binding to lipoproteins that are involved in endotoxin inactivation and detoxification. Cholesteryl ester transfer protein (CETP) is an LT/LBP member, but its impact on LPS metabolism and sepsis outcome is unclear. Here, we performed fluorescent LPS transfer assays to assess the ability of CETP to bind and transfer LPS. The effects of intravenous (iv) infusion of purified LPS or polymicrobial infection (cecal ligation and puncture [CLP]) were compared in transgenic mice expressing human CETP and wild-type mice naturally having no CETP activity. CETP displayed no LPS transfer activity in vitro, but it tended to reduce biliary excretion of LPS in vivo. The CETP expression in mice was associated with significantly lower basal plasma lipid levels and with higher mortality rates in both models of endotoxemia and sepsis. Furthermore, CETPTg plasma modified cytokine production of macrophages in vitro. In conclusion, despite having no direct LPS binding and transfer property, human CETP worsens sepsis outcomes in mice by altering the protective effects of plasma lipoproteins against endotoxemia, inflammation, and infection

Plasma cholesterol level determines in vivo prion propagation

International audienceTransmissible spongiform encephalopathies are fatal neurodegenerative diseases with an urgent need for therapeutic and prophylactic strategies. At the time when the blood-mediated transmission of prions was demonstrated, in vitro studies indicated a high binding affinity of the scrapie prion protein (PrPSc) with apoB-containing lipoproteins, i.e., the main carriers of cholesterol in human blood. The aim of the present study was to explore the relationship between circulating cholesterol-containing lipoproteins and the pathogenicity of prions in vivo. We showed that, in mice with a genetically engineered deficiency for the plasma lipid transporter, phospholipid transfer protein (PLTP), abnormally low circulating cholesterol concentrations were associated with a significant prolongation of survival time after intraperitoneal inoculation of the 22L prion strain. Moreover, when circulating cholesterol levels rose after feeding PLTP-deficient mice a lipid-enriched diet, a significant reduction in survival time of mice together with a marked increase in the accumulation rate of PrPSc deposits in their brain were observed. Our results suggest that the circulating cholesterol level is a determinant of prion propagation in vivo and that cholesterol-lowering strategies might be a successful therapeutic approach for patients suffering from prion diseases.Copyright © 2017 by the American Society for Biochemistry and Molecular Biology, Inc

Development of Abdominal Aortic Aneurysm Is Decreased in Mice with Plasma Phospholipid Transfer Protein Deficiency

International audiencePlasma phospholipid transfer protein (PLTP) increases the circulating levels of proatherogenic lipoproteins, accelerates blood coagulation, and modulates inflammation. The role of PLTP in the development of abdominal aortic aneurysm (AAA) was investigated by using either a combination of mechanical and elastase injury at one site of mouse aorta (elastase model) or continuous infusion of angiotensin II in hyperlipidemic ApoE-knockout mice (Ang II model). With the elastase model, complete PLTP deficiency was associated with a significantly lower incidence and a lesser degree of AAA expansion. With the Ang II model, findings were consistent with those in the elastase model, with a lower severity grade in PLTP-deficient mice, an intermediate phenotype in PLTP-deficient heterozygotes, and a blunted effect of the PLTP-deficient trait when restricted to bone marrow-derived immune cells. The protective effect of whole-body PLTP deficiency in AAA was illustrated further by a lesser degree of adventitia expansion, reduced elastin degradation, fewer recruited macrophages, and less smooth muscle cell depletion in PLTP-deficient than in wild-type mice, as evident from comparative microscopic analysis of aorta sections. Finally, cumulative evidence supports the association of PLTP deficiency with reduced expression and activity levels of matrix metalloproteinases, known to degrade elastin and collagen. We conclude that PLTP can play a significant role in the pathophysiology of AAA