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The phosphodiesterase 4 (PDE4) enzyme family plays a pivotal role in regulating levels of the second messenger cAMP. Consequently, PDE4 inhibitors have been investigated as a therapeutic strategy to enhance cAMP signaling in a broad range of diseases, including several types of cancers, as well as in various neurologic, dermatological, and inflammatory diseases. Despite their widespread therapeutic potential, the progression of PDE4 inhibitors into the clinic has been hampered because of their related relatively small therapeutic window, which increases the chance of producing adverse side effects. Interestingly, the PDE4 enzyme family consists of several subtypes and isoforms that can be modified post-translationally or can engage in specific protein-protein interactions to yield a variety of conformational states. Inhibition of specific PDE4 subtypes, isoforms, or conformational states may lead to more precise effects and hence improve the safety profile of PDE4 inhibition. In this review, we provide an overview of the variety of PDE4 isoforms and how their activity and inhibition is influenced by post-translational modifications and interactions with partner proteins. Furthermore, we describe the importance of screening potential PDE4 inhibitors in view of different PDE4 subtypes, isoforms, and conformational states rather than testing compounds directed toward a specific PDE4 catalytic domain. Lastly, potential mechanisms underlying PDE4-mediated adverse effects are outlined. In this review, we illustrate that PDE4 inhibitors retain their therapeutic potential in myriad diseases, but target identification should be more precise to establish selective inhibition of disease-affected PDE4 isoforms while avoiding isoforms involved in adverse effects. SIGNIFICANCE STATEMENT: Although the PDE4 enzyme family is a therapeutic target in an extensive range of disorders, clinical use of PDE4 inhibitors has been hindered because of the adverse side effects. This review elaborately shows that safer and more effective PDE4 targeting is possible by characterizing 1) which PDE4 subtypes and isoforms exist, 2) how PDE4 isoforms can adopt specific conformations upon post-translational modifications and protein-protein interactions, and 3) which PDE4 inhibitors can selectively bind specific PDE4 subtypes, isoforms, and/or conformations.

Oligodendrocyte precursor cells (OPCs) account for 5% of the resident parenchymal central nervous system glial cells. OPCs are not only a back-up for the loss of oligodendrocytes that occurs due to brain injury or inflammation-induced demyelination (remyelination) but are also pivotal in plastic processes such as learning and memory (adaptive myelination). OPC differentiation into mature myelinating oligodendrocytes is controlled by a complex transcriptional network and depends on high metabolic and mitochondrial demand. Mounting evidence shows that OPC dysfunction, culminating in the lack of OPC differentiation, mediates the progression of neurodegenerative disorders such as multiple sclerosis, Alzheimer’s disease and Parkinson’s disease. Importantly, neurodegeneration is characterised by oxidative and carbonyl stress, which may primarily affect OPC plasticity due to the high metabolic demand and a limited antioxidant capacity associated with this cell type. The underlying mechanisms of how oxidative/carbonyl stress disrupt OPC differentiation remain enigmatic and a focus of current research efforts. This review proposes a role for oxidative/carbonyl stress in interfering with the transcriptional and metabolic changes required for OPC differentiation. In particular, oligodendrocyte (epi)genetics, cellular defence and repair responses, mitochondrial signalling and respiration, and lipid metabolism represent key mechanisms how oxidative/carbonyl stress may hamper OPC differentiation in neurodegenerative disorders. Understanding how oxidative/carbonyl stress impacts OPC function may pave the way for future OPC-targeted treatment strategies in neurodegenerative disorders.

Synapses are the functional units of the brain. They form specific contact points that drive neuronal communication and are highly plastic in their strength, density, and shape. A carefully orchestrated balance between synaptogenesis and synaptic pruning, i.e., the elimination of weak or redundant synapses, ensures adequate synaptic density. An imbalance between these two processes lies at the basis of multiple neuropathologies. Recent evidence has highlighted the importance of glia-neuron interactions in the synaptic unit, emphasized by glial phagocytosis of synapses and local excretion of inflammatory mediators. These findings warrant a closer look into the molecular basis of cell-signaling pathways in the different brain cells that are related to synaptic plasticity. In neurons, intracellular second messengers, such as cyclic guanosine or adenosine monophosphate (cGMP and cAMP, respectively), are known mediators of synaptic homeostasis and plasticity. Increased levels of these second messengers in glial cells slow down inflammation and neurodegenerative processes. These multi-faceted effects provide the opportunity to counteract excessive synapse loss by targeting cGMP and cAMP pathways in multiple cell types. Phosphodiesterases (PDEs) are specialized degraders of these second messengers, rendering them attractive targets to combat the detrimental effects of neurological disorders. Cellular and subcellular compartmentalization of the specific isoforms of PDEs leads to divergent downstream effects for these enzymes in the various central nervous system resident cell types. This review provides a detailed overview on the role of PDEs and their inhibition in the context of glia-neuron interactions in different neuropathologies characterized by synapse loss. In doing so, it provides a framework to support future research towards finding combinational therapy for specific neuropathologies.

The brain's endogenous capacity to restore damaged myelin deteriorates during the course of demyelinating disorders. Currently, no treatment options are available to establish remyelination. Chronic demyelination leads to damaged axons and irreversible destruction of the central nervous system (CNS). We identified two promising therapeutic candidates which enhance remyelination: oncostatin M (OSM), a member of the interleukin-6 family, and downstream mediator tissue inhibitor of metalloproteinases-1 (TIMP-1). While remyelination was completely abrogated in OSMR beta knockout (KO) mice, OSM overexpression in the chronically demyelinated CNS established remyelination. Astrocytic TIMP-1 was demonstrated to play a pivotal role in OSM-mediated remyelination. Astrocyte-derived TIMP-1 drove differentiation of oligodendrocyte precursor cells into mature oligodendrocytes in vitro. In vivo, TIMP-1 deficiency completely abolished spontaneous remyelination, phenocopying OSMR beta KO mice. Finally, TIMP-1 was expressed by human astrocytes in demyelinated multiple sclerosis lesions, confirming the human value of our findings. Taken together, OSM and its downstream mediator TIMP-1 have the therapeutic potential to boost remyelination in demyelinating disorders. We thank Dr. Tom Struys, Katrien Wauterickx, and Joke Vanhoof for excellent technical assistance. This work was financially supported by grants from the Research Foundation of Flanders (FWO Vlaanderen, G04441N, G050617N, G0A5716N, and 1106817N), the Interuniversity Attraction Poles (IUAP-P7-39), the Belgian MS-Liga and the Charcot Foundation of Belgium, Methusalem NEURONET, the European FP7 project, HEALTH-F2-2011-278850 (INMiND), and Bijzonder Onderzoeksfonds (BOF)-UHasselt. Hellings, N (reprint author), Hasselt Univ, Biomed Res Inst, Dept Immunol, B-3590 Diepenbeek, Belgium. niels.hellings@uhasselt.be

Background Multiple sclerosis (MS) is a chronic autoimmune disease driven by sustained inflammation in the central nervous system. One of the pathological hallmarks of MS is extensive free radical production. However, the subsequent generation, potential pathological role, and detoxification of different lipid peroxidation-derived reactive carbonyl species during neuroinflammation are unclear, as are the therapeutic benefits of carbonyl quenchers. Here, we investigated the reactive carbonyl acrolein and (the therapeutic effect of) acrolein quenching by carnosine during neuroinflammation. Methods The abundance and localization of acrolein was investigated in inflammatory lesions of MS patients and experimental autoimmune encephalomyelitis (EAE) mice. In addition, we analysed carnosine levels and acrolein quenching by endogenous and exogenous carnosine in EAE. Finally, the therapeutic effect of exogenous carnosine was assessed in vivo (EAE) and in vitro (primary mouse microglia, macrophages, astrocytes). Results Acrolein was substantially increased in inflammatory lesions of MS patients and EAE mice. Levels of the dipeptide carnosine (β-alanyl-l-histidine), an endogenous carbonyl quencher particularly reactive towards acrolein, and the carnosine-acrolein adduct (carnosine-propanal) were ~ twofold lower within EAE spinal cord tissue. Oral carnosine treatment augmented spinal cord carnosine levels (up to > tenfold), increased carnosine-acrolein quenching, reduced acrolein-protein adduct formation, suppressed inflammatory activity, and alleviated clinical disease severity in EAE. In vivo and in vitro studies indicate that pro-inflammatory microglia/macrophages generate acrolein, which can be efficiently quenched by increasing carnosine availability, resulting in suppressed inflammatory activity. Other properties of carnosine (antioxidant, nitric oxide scavenging) may also contribute to the therapeutic effects. Conclusions Our results identify carbonyl (particularly acrolein) quenching by carnosine as a therapeutic strategy to counter inflammation and macromolecular damage in MS.

The initiation and progression of autoimmune disorders such as multiple sclerosis (MS) is linked to aberrant cholesterol metabolism and overt inflammation. Liver X receptors (LXR) are nuclear receptors that function at the crossroads of cholesterol metabolism and immunity, and their activation is considered a promising therapeutic strategy to attenuate autoimmunity. However, despite clear functional heterogeneity and cell-specific expression profiles, the impact of the individual LXR isoforms on autoimmunity remains poorly understood. Here, we show that LXR alpha and LXR beta have an opposite impact on immune cell function and disease severity in the experimental autoimmune encephalomyelitis model, an experimental MS model. While Lxr alpha deficiency aggravated disease pathology and severity, absence of Lxr beta was protective. Guided by flow cytometry and by using cell-specific knockout models, reduced disease severity in Lxr beta-deficient mice was primarily attributed to changes in peripheral T cell physiology and occurred independent from alterations in microglia function. Collectively, our findings indicate that LXR isoforms play functionally non-redundant roles in autoimmunity, potentially having broad implications for the development of LXR-based therapeutic strategies aimed at dampening autoimmunity and neuroinflammation.

Multiple health benefits have been ascribed to brown seaweeds that are used traditionally as dietary component mostly in Asia. This systematic review summarizes information on the impact of brown seaweeds or components on inflammation, and inflammation-related pathologies, such as allergies, diabetes mellitus and obesity. We focus on oral supplementation thus intending the use of brown seaweeds as food additives. Despite the great diversity of experimental systems in which distinct species and compounds were tested for their effects on inflammation and immunity, a remarkably homogeneous picture arises. The predominant effects of consumption of brown seaweeds or compounds can be classified into three categories: (1) inhibition of reactive oxygen species, known to be important drivers of inflammation; (2) regulation, i.e., in most cases inhibition of proinflammatory NF-κB signaling; (3) modulation of adaptive immune responses, in particular by interfering with T-helper cell polarization. Over the last decades, several inflammation-related diseases have increased substantially. These include allergies and autoimmune diseases as well as morbidities associated with lifestyle and aging. In this light, further development of brown seaweeds and seaweed compounds as functional foods and nutriceuticals might contribute to combat these challenges.

Phosphodiesterases (PDEs) have been an interesting drug target for many diseases. Although a vast number of mainly preclinical studies demonstrates beneficial effects of PDE inhibitors for central nervous system (CNS) diseases, no drugs are currently available for CNS indications. In this review, we discuss the rationale of PDE4 inhibitors for different CNS diseases, including memory impairments, striatal disorders, multiple sclerosis (MS), and acquired brain injury (ABU). However, clinical development has been problematic due to mechanism-based adverse effects of these drugs in humans. Our increased understanding of factors influencing the conformational state of the PDE4 enzyme and of how to influence the binding affinity of PDE4 subtype inhibitors, holds promise for the successful development of novel selective PDE4 inhibitors with higher efficacy and fewer adverse effects.

Alzheimer's disease (AD) is characterized by a sequential progression of amyloid plaques (A), neurofibrillary tangles (T) and neurodegeneration (N), constituting ATN pathology. While microglia are considered key contributors to AD pathogenesis, their contribution in the combined presence of ATN pathologies remains incompletely understood. As sensors of the brain microenvironment, microglial phenotypes and contributions are importantly defined by the pathologies in the brain, indicating the need for their analysis in preclinical models that recapitulate combined ATN pathologies, besides their role in A and T models only. Here, we report a new tau-seed model in which amyloid pathology facilitates bilateral tau propagation associated with brain atrophy, thereby recapitulating robust ATN pathology. Single-cell RNA sequencing revealed that ATN pathology exacerbated microglial activation towards disease-associated microglia states, with a significant upregulation of Apoe as compared to amyloid-only models (A). Importantly, Colony-Stimulating Factor 1 Receptor inhibition preferentially eliminated non-plaque-associated versus plaque associated microglia. The preferential depletion of non-plaque-associated microglia significantly attenuated tau pathology and neuronal atrophy, indicating their detrimental role during ATN progression. Together, our data reveal the intricacies of microglial activation and their contributions to pathology in a model that recapitulates the combined ATN pathologies of AD. Our data may provide a basis for microglia-targeting therapies selectively targeting detrimental microglial populations, while conserving protective populations. This work was supported by Innoviris (Attract grant, BB2B 2015–2) to KM; FRA-SAO (Fondation Recherche Alzheimer—Stichting Alzheimer Onderzoek, Belgium – 20180028 & 2020022) to IDW, VLAIO (Vlaams Agentschap Innoveren en Ondernemen) and FWO (Fonds Wetenschappelijk Onderzoek—Vlaanderen—G0C6819N & 1259621 N and the fellowship of I.S.). Acknowledgements We thank Yvon Elkrim and the VIB Nucleomics core for technical assistance, VIB Tech Watch for support regarding single-cell RNA sequencing technologies.

We recently found that dietary supplementation with the seaweed Sargassum fusiforme, containing the preferential LXRβ-agonist 24(S)-saringosterol, prevented memory decline and reduced amyloid-β (Aβ) deposition in an Alzheimer’s disease (AD) mouse model without inducing hepatic steatosis. Here, we examined the effects of 24(S)-saringosterol as a food additive on cognition and neuropathology in AD mice. Six-month-old male APPswePS1ΔE9 mice and wildtype C57BL/6J littermates received 24(S)-saringosterol (0.5 mg/25 g body weight/day) (APPswePS1ΔE9 n = 20, C57BL/6J n = 19) or vehicle (APPswePS1ΔE9 n = 17, C57BL/6J n = 19) for 10 weeks. Cognition was assessed using object recognition and object location tasks. Sterols were analyzed by gas chromatography/mass spectrometry, Aβ and inflammatory markers by immunohistochemistry, and gene expression by quantitative real-time PCR. Hepatic lipids were quantified after Oil-Red-O staining. Administration of 24(S)-saringosterol prevented cognitive decline in APPswePS1ΔE9 mice without affecting the Aβ plaque load. Moreover, 24(S)-saringosterol prevented the increase in the inflammatory marker Iba1 in the cortex of APPswePS1ΔE9 mice (p &lt, 0.001). Furthermore, 24(S)-saringosterol did not affect the expression of lipid metabolism-related LXR-response genes in the hippocampus nor the hepatic neutral lipid content. Thus, administration of 24(S)-saringosterol prevented cognitive decline in APPswePS1ΔE9 mice independent of effects on Aβ load and without adverse effects on liver fat content. The anti-inflammatory effects of 24(S)-saringosterol may contribute to the prevention of cognitive decline.

Pharmacological phosphodiesterase 4D (PDE4D) inhibition shows therapeutic potential to restore memory function in Alzheimer's disease (AD), but will likely evoke adverse side effects. As PDE4D encodes multiple isoforms, targeting specific isoforms may improve treatment efficacy and safety. Here, we investigated whether PDE4D isoform expression and PDE4D DNA methylation is affected in AD and whether expression changes are associated with severity of pathology and cognitive impairment. In post-mortem temporal lobe brain material from AD patients (n = 42) and age-matched controls (n = 40), we measured PDE4D isoform expression and PDE4D DNA (hydroxy)methylation using quantitative polymerase chain reaction and Illumina 450k Beadarrays, respectively. Linear regression revealed increased PDE4D1, -D3, -D5, and -D8 expression in AD with concurrent (hydroxy)methylation changes in associated promoter regions. Moreover, increased PDE4D1 and-D3 expression was associated with higherplaque and tau pathology levels, higher Braak stages, and progressed cognitive impairment. Future studies should indicate functional roles of specific PDE4D isoforms and the efficacy and safety of their selective inhibition to restore memory function in AD. (C) 2020 The Authors. Published by Elsevier Inc. This work was financially supported by grants from ISAO/Alzheimer Nederland WE.03-2016-07, Young European Research Universities Network (YERUN), and the Baeter Laeve foundation. Additional funds have been provided by the Internationale Stichting Alzheimer Onderzoek (ISAO)/Alzheimer Netherlands (Award #11532; Funded by the Dorpmans-Wigmans Foundation) (DvdH), and by the Joint ProgrammeeNeurodegenerative Disease Research (JPND) for the EPI-AD consortium (http://www.neurodegenerationresearch.eu/wp-content/uploads/2015/10/Factsheet_EPI-AD.pdf).The project is supported through the following funding organizations under the aegis of JPND; The Netherlands, The Netherlands Organisation for Health Research and Development (ZonMw); United Kingdom, Medical Research Council; Germany, German Federal Ministry of Education and Research (BMBF); Luxembourg, National Research Fund (FNR). This project has received funding from the European Union's Horizon 2020 research and innovation program under Grant Agreement No. 643417. Prickaerts, J (corresponding author), Maastricht Univ, Dept Psychiat & Neuropsychol, Sch Mental Hlth & Neurosci, POB 616, NL-6200 MD Maastricht, Netherlands. jos.prickaerts@maastrichtuniversity.nl

Multiple sclerosis (MS) is an autoimmune inflammatory disease characterized by demyelination, axonal loss, and synaptic impairment in the central nervous system (CNS). The available therapies aim to reduce the severity of the pathology during the early inflammatory stages, but they are not effective in the chronic stage of the disease. In this phase, failure in endogenous remyelination is associated with the impairment of oligodendrocytes progenitor cells (OPCs) to migrate and differentiate into mature myelinating oligodendrocytes. Therefore, stimulating differentiation of OPCs into myelinating oligodendrocytes has become one of the main goals of new therapeutic approaches for MS. Different disease-modifying therapies targeting sphingosine-1-phosphate receptors (S1PRs) have been approved or are being developed to treat MS. Besides their immunomodulatory effects, growing evidence suggests that targeting S1PRs modulates mechanisms beyond immunomodulation, such as remyelination. In this context, this review focuses on the current understanding of S1PR modulators and their direct effect on OPCs and oligodendrocytes.

Background: Trapeziometacarpal arthroplasties are designed to restore an adequate level of mobility, stability, and grip strength. In this article, pain and functional and radiographic outcome of Ivory arthroplasty in male patients are investigated. Methods: Between 2005 and 2012, the Ivory arthroplasty was inserted in 21 male patients with degenerative trapeziometacarpal osteoarthritis, of which 14 patients were found eligible for inclusion. Mobility, grip strength, patient self-assessment (pain; Quick Disabilities of the Arm, Shoulder, and Hand [QuickDASH]), and radiographic outcome were measured. Twenty-two female patients who received an Ivory arthroplasty between 2005 and 2007 were included and underwent the same evaluation. Age at primary surgery, survival rate of the implant, and clinical outcome were compared between the 2 groups. Results: In both groups, QuickDASH score and mean pain sensation improved significantly. The improvement in mobility obtained significance in the female group. In the male group, 7 arthroplasties failed (mean follow-up of 65 months). In the female group, 3 of the 24 arthroplasties failed (mean follow-up of 123 months). Kaplan-Meier survival analysis demonstrated a significant lower implant survival in the male group. Conclusions: Decrease in muscle mass and decline in grip strength that postmenopausal women tend to experience might explain the significant difference in implant survival between sexes. In 4 of the 7 failed arthroplasties in the male group, no surgical revision was required. Trapeziometacarpal arthroplasty, even after radiographic failure, still served as a spacer, avoiding collapse of the thumb base. Nevertheless, the failure rate of the Ivory arthroplasty in male patients is high, and an alternative treatment should be considered.

Current evidence indicates that foods with added plant sterols or stanols can lower serum levels of low-density lipoprotein cholesterol. This review summarizes the recent findings and deliberations of 31 experts in the field who participated in a scientific meeting in Winnipeg, Canada, on the health effects of plant sterols and stanols. Participants discussed issues including, but not limited to, the health benefits of plant sterols and stanols beyond cholesterol lowering, the role of plant sterols and stanols as adjuncts to diet and drugs, and the challenges involved in measuring plant sterols and stanols in biological samples. Variations in interindividual responses to plant sterols and stanols, as well as the personalization of lipid-lowering therapies, were addressed. Finally, the clinical aspects and treatment of sitosterolemia were reviewed. Although plant sterols and stanols continue to offer an efficacious and convenient dietary approach to cholesterol management, long-term clinical trials investigating the endpoints of cardiovascular disease are still lacking. International Plant Sterol and Stanol Association (IPSSA); National Center for Complementary and Integrative Health (NCCIH); National Center for Advancing Translational Sciences [IR13AT009270-01]

Objective:We sought to determine the liver X receptor (LXR) ligands present in human macrophages after myelin phagocytosis and whether LXRs are activated in multiple sclerosis (MS) lesions.Methods:We used real-time quantitative polymerase chain reaction (PCR) and immunohistochemistry to determine expression of LXRs and their response genes in human phagocytes after myelin phagocytosis and in active MS lesions. We used gas chromatographic/mass spectrometric analysis to determine LXR-activating oxysterols and cholesterol precursors present and formed in myelin and myelin-incubated cells, respectively.Results:Myelin induced LXR response genes ABCA1 and ABCG1 in human monocyte-derived macrophages. In active MS lesions, we found that both gene expression and protein levels of ABCA1 and apolipoprotein E ( APOE) are upregulated in foamy phagocytes. Moreover, we found that the LXR ligand 27-hydroxycholesterol (27OHC) is significantly increased in human monocyte-derived macrophages after myelin uptake.Conclusion:LXR response genes are upregulated in phagocytes present in active MS lesions, indicating that LXRs are activated in actively demyelinating phagocytes. In addition, we have shown that myelin contains LXR ligands and that 27OHC is generated in human monocyte-derived macrophages after myelin processing. This suggests that LXRs in phagocytes in active MS lesions are activated at least partially by (oxy)sterols present in myelin and the generation thereof during myelin processing.

Oligodendrocytes provide metabolic and functional support to neuronal cells, rendering them key players in the functioning of the central nervous system. Oligodendrocytes need to be newly formed from a pool of oligodendrocyte precursor cells (OPCs). The differentiation of OPCs into mature and myelinating cells is a multistep process, tightly controlled by spatiotemporal activation and repression of specific growth and transcription factors. While oligodendrocyte turnover is rather slow under physiological conditions, a disruption in this balanced differentiation process, for example in case of a differentiation block, could have devastating consequences during ageing and in pathological conditions, such as multiple sclerosis. Over the recent years, increasing evidence has shown that epigenetic mechanisms, such as DNA methylation, histone modifications, and microRNAs, are major contributors to OPC differentiation. In this review, we discuss how these epigenetic mechanisms orchestrate and influence oligodendrocyte maturation. These insights are a crucial starting point for studies that aim to identify the contribution of epigenetics in demyelinating diseases and may thus provide new therapeutic targets to induce myelin repair in the long run.

Dysfunction of the blood-brain barrier (BBB) contributes significantly to the pathogenesis of several neuroinflammatory diseases, including multiple sclerosis (MS). Potential players that regulate BBB function are the liver X receptors (LXRs), which are ligand activated transcription factors comprising two isoforms, LXR alpha, and LXR beta. However, the role of LXR alpha, and LXR beta in regulating BBB (dys)function during neuroinflammation remains unclear, as well as their individual involvement. Therefore, the goal of the present study is to unravel whether LXR isoforms have different roles in regulating BBB function under neuroinflammatory conditions. We demonstrate that LXR alpha, and not LXR beta, is essential to maintain barrier integrity in vitro. Specific knockout of LXR alpha in brain endothelial cells resulted in a more permeable barrier with reduced expression of tight junctions. Additionally, the observed dysfunction was accompanied by increased endothelial inflammation, as detected by enhanced expression of vascular cell adhesion molecule (VCAM-1) and increased transendothelial migration of monocytes toward inflammatory stimuli. To unravel the importance of LXR alpha in BBB function in vivo, we made use of the experimental autoimmune encephalomyelitis (EAE) MS mouse model. Induction of EAE in a constitutive LXR alpha knockout mouse and in an endothelial specific LXR alpha knockout mouse resulted in a more severe disease score in these animals. This was accompanied by higher numbers of infiltrating leukocytes, increased endothelial VCAM-1 expression, and decreased expression of the tight junction molecule claudin-5. Together, this study reveals that LXR alpha is indispensable for maintaining BBB integrity and its immune quiescence. Targeting the LXR alpha isoform may help in the development of novel therapeutic strategies to prevent BBB dysfunction, and thereby neuroinflammatory disorders. This work was supported by grants to NdW from ZonMw Memorabel program (project nr: 733050105), to JG a fellowship from Robert A. Welch (E-0004) and a grant from the Swedish Research Council, and to TV from FWO (project nr: 1506916N). The received funding had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Research at the Department of Molecular Cell Biology and immunology is part of the neuro-infection and neuroinflammation research program of Amsterdam Neuroscience.

Multiple sclerosis (MS) is a demyelinating autoimmune disease in which innate and adaptive immune cells infiltrate the central nervous system (CNS) and damage the myelin sheaths surrounding the axons. Upon activation, infiltrated macrophages, CNS-resident microglia, and astrocytes switch their metabolism toward glycolysis, resulting in the formation of alpha-dicarbonyls, such as methylglyoxal (MGO) and glyoxal (GO). These potent glycating agents lead to the formation of advanced glycation endproducts (AGEs) after reaction with amino acids. We hypothesize that AGE levels are increased in MS lesions due to the inflammatory activation of macrophages and astrocytes. First, we measured tissue levels of AGEs in brain samples of MS patients and controls. Analysis of MS patient and non-demented control (NDC) specimens showed a significant increase in protein-bound Nd-(5-hydro-5-methyl-4-imidazolon-2-yl)-ornithine (MG-H1), the major AGE, compared to white matter of NDCs (107 +/- 11 vs. 154 +/- 21, p < 0.05). In addition, immunohistochemistry revealed that MGO-derived AGEs were specifically present in astrocytes, whereas the receptor for AGEs, RAGE, was detected on microglia/macrophages. Moreover, in cerebrospinal fluid from MS patients, alpha-dicarbonyls and free AGEs correlated with their respective levels in the plasma, whereas this was not observed for protein-bound AGEs. Taken together, our data show that MG-H1 is produced by astrocytes. This suggests that AGEs secreted by astrocytes have paracrine effects on RAGE-positive macrophages/microglia and thereby contribute to the pathology of MS. Special Research Fund UHasselt [12N31BOF] We would like to thank Marleen van Greevenbroek for her advice regarding the statistical analysis and Marjo van de Waarenburg and Bieke Broux for technical assistance.

Multiple sclerosis (MS) is a chronic demyelinating disease of the central nervous system (CNS) characterized by heterogeneous clinical symptoms including gradual muscle weakness, fatigue, and cognitive impairment. The disease course of MS can be classified into a relapsing-remitting (RR) phase defined by periods of neurological disabilities, and a progressive phase where neurological decline is persistent. Pathologically, MS is defined by a destructive immunological and neuro-degenerative interplay. Current treatments largely target the inflammatory processes and slow disease progression at best. Therefore, there is an urgent need to develop next-generation therapeutic strategies that target both neuroinflammatory and degenerative processes. It has been shown that elevating second messengers (cAMP and cGMP) is important for controlling inflammatory damage and inducing CNS repair. Phosphodiesterases (PDEs) have been studied extensively in a wide range of disorders as they breakdown these second messengers, rendering them crucial regulators. In this review, we provide an overview of the role of PDE inhibition in limiting pathological inflammation and stimulating regenerative processes in MS.

Activation of liver X receptors (LXRs) by synthetic agonists was found to improve cognition in Alzheimer’s disease (AD) mice. However, these LXR agonists induce hypertriglyceridemia and hepatic steatosis, hampering their use in the clinic. We hypothesized that phytosterols as LXR agonists enhance cognition in AD without affecting plasma and hepatic triglycerides. Phytosterols previously reported to activate LXRs were tested in a luciferase-based LXR reporter assay. Using this assay, we found that phytosterols commonly present in a Western type diet in physiological concentrations do not activate LXRs. However, a lipid extract of the 24(S)-Saringosterol-containing seaweed Sargassum fusiforme did potently activate LXRβ. Dietary supplementation of crude Sargassum fusiforme or a Sargassum fusiforme-derived lipid extract to AD mice significantly improved short-term memory and reduced hippocampal Aβ plaque load by 81%. Notably, none of the side effects typically induced by full synthetic LXR agonists were observed. In contrast, administration of the synthetic LXRα activator, AZ876, did not improve cognition and resulted in the accumulation of lipid droplets in the liver. Administration of Sargassum fusiforme-derived 24(S)-Saringosterol to cultured neurons reduced the secretion of Aβ42. Moreover, conditioned medium from 24(S)-Saringosterol-treated astrocytes added to microglia increased phagocytosis of Aβ. Our data show that Sargassum fusiforme improves cognition and alleviates AD pathology. This may be explained at least partly by 24(S)-Saringosterol-mediated LXRβ activation. This work was supported by the Internationale Stichting Alzheimer Onderzoek (ISAO)/Alzheimer Nederland (AN), Scientific Research-Flanders (FWO), Fondation Vaincre Alzheimer (LECMA), and Alzheimer Forschung Initiative (AFI). The authors like to thank Joke Vanhoof for excellent technical assistance and AstraZeneca for providing the AZ876 compound through the AstraZeneca Open Innovation program. We sincerely thank Patric Delhanty for editing the paper and Kenneth Vanbrabant for performing the GC-MS analysis.

Treatment of Alzheimer's disease (AD) patients with the antidepressant fluoxetine is known to improve memory and cognitive function. However, the mechanisms underlying these effects are largely unknown. To unravel these mechanisms, we aimed to treat APPswe/PS1dE9 mice with fluoxetine. Unexpectedly, with time, an increased number of animals displayed seizure behavior and died. Although spontaneous behavioral seizures have been reported previously in this mouse model, the observation of seizures and death consequential to fluoxetine treatment is new. Our results warrant further research on the underlying mechanisms as this may refine the treatment of AD patients.

Multiple sclerosis (MS) is a demyelinating autoimmune disease of the central nervous system (CNS). The immune response in MS patients leads to the infiltration of immune cells in the CNS and their subsequent activation. Immune cell activation induces a switch towards glycolysis. During glycolysis, the dicarbonyl product methylglyoxal (MGO) is produced. MGO is a glycating agent that can rapidly form advanced glycation endproducts (AGEs). In turn, AGEs are able to induce inflammatory responses. The glyoxalase system is the endogenous defense system of the body to reduce the burden of MGO thereby reducing AGE formation. This system consists of glyoxalase-1 and glyoxalase-2 which are able to detoxify MGO to D-lactate. We investigated whether AGE levels are induced in experimental autoimmune encephalitis (EAE), an inflammatory animal model of MS. Twenty seven days post EAE induction, MGO and AGE (Nε-(carboxymethyl)lysine (CML), Nε-(carboxyethyl)lysine (CEL), 5-hydro-5-methylimidazolone (MG-H1)) levels were significantly increased in the spinal cord of mice subjected to EAE. Yet, pyridoxamine treatment and glyoxalase-1 overexpression were unable to counteract AGE production during EAE and did not influence the clinical course of EAE. In conclusion, AGEs levels increase during EAE in the spinal cord, but AGE-modifying treatments do not inhibit EAE-induced AGE production and do not affect disease progression.

Myelin-containing macrophages and microglia are the most abundant immune cells in active multiple sclerosis (MS) lesions. Our recent transcriptomic analysis demonstrated that collectin placenta 1 (CL-P1) is one of the most potently induced genes in macrophages after uptake of myelin. CL-P1 is a type II transmembrane protein with both a collagen-like and carbohydrate recognition domain, which plays a key role in host defense. In this study we sought to determine the dynamics of CL-P1 expression on myelin-containing phagocytes and define the role that it plays in MS lesion development. We show that myelin uptake increases the cell surface expression of CL-P1 by mouse and human macrophages, but not by primary mouse microglia in vitro. In active demyelinating MS lesions, CL-P1 immunoreactivity was localized to perivascular and parenchymal myelin-laden phagocytes. Finally, we demonstrate that CL-P1 is involved in myelin internalization as knockdown of CL-P1 markedly reduced myelin uptake. Collectively, our data indicate that CL-P1 is a novel receptor involved in myelin uptake by phagocytes and likely plays a role in MS lesion development.

Multiple sclerosis (MS) is an inflammatory auto-immune disease of the central nervous system (CNS). Serum glucose alterations and impaired glucose tolerance (IGT) are reported in MS patients, and are commonly associated with the development of cardio-metabolic co-morbidities. We previously found that a subgroup of MS patients shows alterations in their lipoprotein profile that are similar to a pre-cardiovascular risk profile. In addition, we showed that a high-intensity exercise training has a positive effect on IGT in MS patients. In this study, we hypothesize that exercise training positively influences the lipoprotein profile of MS patients. To this end, we performed a pilot study and determined the lipoprotein profile before (controls, n = 40; MS patients, n = 41) and after (n = 41 MS only) 12 weeks of medium-intensity continuous training (MIT, n = 21, ~60% of VO2max) or high-intensity interval training (HIT, n = 20, ~100–200% of VO2max) using nuclear magnetic resonance spectroscopy (NMR). Twelve weeks of MIT reduced intermediate-density lipoprotein particle count ((nmol/L); −43.4%; p < 0.01), low-density lipoprotein cholesterol (LDL-c (mg/dL); −7.6%; p < 0.05) and VLDL size ((nm); −6.6%; p < 0.05), whereas HIT did not influence the lipoprotein profile. These results show that MIT partially normalizes lipoprotein alterations in MS patients. Future studies including larger patient and control groups should determine whether MIT can reverse other lipoprotein levels and function and if these alterations are related to MS disease progression and the development of co-morbidities. This work was supported by grants from the European Molecular Biology Organization (EMBO) and the Agentschap Innovatie door Wetenschap en Technologie (IWT). Research by Alan T. Remaley was supported by the Division of Intramural Research (DIR) funds from NHLBI.

Multiple sclerosis (MS) is a demyelinating disease of the central nervous system (CNS). The activation of inflammatory cells is crucial for the development of MS and is shown to induce intracellular glycolytic metabolism in pro-inflammatory microglia and macrophages, as well as CNS-resident astrocytes. Advanced glycation endproducts (AGEs) are stable endproducts formed by a reaction of the dicarbonyl compounds methylglyoxal (MGO) and glyoxal (GO) with amino acids in proteins, during glycolysis. This suggests that, in MS, MGO-derived AGEs are formed in glycolysis-driven cells. MGO and MGO-derived AGEs can further activate inflammatory cells by binding to the receptor for advanced glycation endproducts (RAGE). Recent studies have revealed that AGEs are increased in the plasma and brain of MS patients. Therefore, AGEs might contribute to the inflammatory status in MS. Moreover, the main detoxification system of dicarbonyl compounds, the glyoxalase system, seems to be affected in MS patients, which may contribute to high MGO-derived AGE levels. Altogether, evidence is emerging for a contributing role of AGEs in the pathology of MS. In this review, we provide an overview of the current knowledge on the involvement of AGEs in MS.

Lipoproteins modulate innate and adaptive immune responses. In the chronic inflammatory disease multiple sclerosis (MS), reports on lipoprotein level alterations are inconsistent and it is unclear whether lipoprotein function is affected. Using nuclear magnetic resonance (NMR) spectroscopy, we analysed the lipoprotein profile of relapsing-remitting (RR) MS patients, progressive MS patients and healthy controls (HC). We observed smaller LDL in RRMS patients compared to healthy controls and to progressive MS patients. Furthermore, low-BMI (BMI

Brain-derived neurotrophic factor (BDNF) is a crucial regulator of neuronal survival and neuroplasticity in the central nervous system (CNS). As a result, there has been a growing interest in the role of BDNF in neuropsychiatric disorders associated with neurodegeneration, including depression and dementia. However, until now, BDNF-targeting therapies have yielded disappointing results. BDNF is thought to exert its beneficial effects on synaptic and neuronal plasticity mainly through binding to the tyrosine kinase B (TrkB) receptor. Recently, 7,8-dihydroxyflavone (7,8-DHF) was identified as the first selective TrkB agonist. In the present study the effect of 7,8-DHF on memory consolidation processes was evaluated. In healthy rats, 7,8-DHF improved object memory formation in the object recognition task when administered both immediately and 3 h after learning. In a transgenic mouse model of Alzheimer's disease, i.e. APPswe/PS1dE9 mice, spatial memory as measured in the object location task was improved after administration of 7,8-DHF. A similar memory improvement was found when their wild-type littermates were treated with 7,8-DHF. The acute beneficial effects in healthy mice suggest that effects might be symptomatic rather than curing. Nevertheless, this study suggests that 7,8-DHF might be a promising therapeutic target for dementia.

This double-blind, randomized, placebo-controlled, cross-over intervention-study was conducted in healthy volunteers to evaluate the effects of plant sterol ester supplemented margarine on cholesterol, non-cholesterol sterols and oxidative stress in serum and monocytes. Sixteen volunteers, average age 34 years, with no or mild hypercholesterolemia were subjected to a 4 week period of daily intake of 3g plant sterols per day supplied via a supplemented margarine on top of regular eating habits. After a wash-out period of one week, volunteers switched groups. Compared to placebo, a diet supplementation with plant sterols increased serum levels of plant sterols such as campesterol (+0.16±0.19mg/dL, p=0.005) and sitosterol (+0.27±0.18mg/dL, p

Background— In humans, the E4 allele of the apolipoprotein E gene is associated with increased coronary heart disease risk. Surprisingly, in rodents, apolipoprotein E4 only accelerates the atherosclerotic process when transgenic for the human low-density lipoprotein receptor (LDLR) protein. We therefore investigated whether the LDLR locus interacted with the apolipoprotein E gene genotype on coronary heart disease risk in patients clinically diagnosed with familial hypercholesterolemia with and without LDLR mutation. We investigated whether the presence of an LDLR mutation diminishing LDLR function was protective in E4/E4 carriers. Methods and Results— In a cohort of 2400 patients clinically diagnosed with familial hypercholesterolemia, we found an LDLR gene mutation in 1383 patients, whereas in 1013 patients, such mutation was not present. In 92 patients homozygous for the apolipoprotein E4 , the presence of an LDLR mutation conferred lower coronary heart disease risk (hazard ratio, 0.16; 95% CI, 0.05–0.58; P =0.005). Mirroring these results, the apolipoprotein E4/E4 genotype was also associated with lower coronary heart disease risk in patients with familial hypercholesterolemia with an LDLR mutation (hazard ratio, 0.26; hazard ratio, 0.08–0.80; P =0.02). Conclusions— LDLR function is key to the detrimental effects of apolipoprotein E4 in humans. Kinetic studies in humans are now required to study the consequences of our observation for prevention of both coronary heart disease and Alzheimer disease.

High dose daily intake of plant sterols decreases the uptake of cholesterol in the intestine by competitive mechanisms and thus leads to reduced serum levels of total and LDL-cholesterol. By this, the commercialization of plant sterol enriched 'functional food' products is rapidly increasing. Subjects using these kinds of diet present a duplication of their serum plant sterol levels after long-term intake. In analogy to cholesterol, plant sterols such as campesterol and sitosterol can be oxidized to oxyphytosterols and these may counteract the primary anti-atherosclerotic action of cholesterol lowering. In order to investigate the whole spectrum of the consequences following high plant sterol intake a highly sensitive and specific isotope dilution gas chromatography-mass spectrometry method for the analysis of 7-oxygenated campesterol/sitosterol in trace amounts in human serum is presented in this paper. The validation was based on limits for detection and quantification, recovery, precision and minimization of autoxidation during work-up. Our results show an overall coefficient of variation ≤10% for the precision. The lowest limits for detection and quantification for 7α-hydroxy-campesterol were 7 pg/mL and 23 pg/mL, respectively. Data for overall sum recovery ranged from 92% to 115%. We practically used this method for analysis of oxyphytosterols simultaneously with plant sterol concentrations in serum from healthy volunteers. Sixteen subjects were treated with plant sterol enriched margarine (3 g/day) for 28 days. The results showed a significant increase of the oxyphytosterol 7β-hydroxy-sitosterol from 1.19±0.54 (before intake) to 2.24±1.24 ng/mL (mean±SD; +86.7%; P=0.007) after intake of the margarine. There was a highly significant correlation between the serum levels of campesterol and the sum of 7-oxygenated campesterol (R(2)=0.915; P

Contains fulltext : 98057.pdf (Publisher’s version ) (Closed access) Alterations in cerebral cholesterol metabolism are thought to play a role in the progression of Alzheimer's disease (AD). Liver X receptors (LXRs) are key regulators of cholesterol metabolism. The synthetic LXR activator, T0901317 has been reported to improve memory functions in animal models for AD and to reduce amyloid-beta (Abeta) deposition in the brain. Here we provide evidence that long-term administration of T0901317 to aged, 21-month-old APPSLxPS1mut mice restores impaired memory. Cerebral cholesterol turnover was enhanced as indicated by the increased levels of brain cholesterol precursors and the upregulation of LXR-target genes Abca1, Abcg1, and Apoe. Unexpectedly, the improved memory functions in the APPSLxPS1mut mice after T0901317 treatment were not accompanied by a decrease in Abeta plaque load in the cortex or hippocampus DG, CA1 or CA3. T0901317 administration also enhanced cerebral cholesterol turnover in aged C57BL/6NCrl mice, but did not further improve their memory functions. In conclusion, long-term activation of the LXR-pathway restored memory functions in aged APPSLxPS1mut mice with advanced Abeta deposition. However the beneficial effects of T0901317 on memory in the APPSLxPS1mut mice were independent of the Abeta plaque load in the hippocampus, but were associated with enhanced brain cholesterol turnover.

In mammals, the central nervous system (CNS) is the most cholesterol rich organ by weight. Cholesterol metabolism is tightly regulated in the CNS and all cholesterol available is synthesized in situ. Deficits in cholesterol homeostasis at the level of synthesis, transport, or catabolism result in severe disorders featured by neurological disability. Recent studies indicate that a disturbed cholesterol metabolism is involved in CNS disorders, such as Alzheimer's disease (AD), multiple sclerosis (MS), and amyotrophic lateral sclerosis (ALS). In contrast to circulating cholesterol, dietary plant sterols, can cross the blood brain barrier and accumulate in the membranes of CNS cells. Plant sterols are well-known for their ability to lower circulating cholesterol levels. The finding that they gain access to the CNS has fueled research focusing on the physiological roles of plant sterols in the healthy and diseased CNS. To date, both beneficial and detrimental effects of plant sterols on CNS disorders are defined. In this review, we discuss recent findings regarding the impact of plant sterols on homeostatic and pathogenic processes in the CNS, and elaborate on the therapeutic potential of plant sterols in CNS disorders. (C) 2015 Elsevier Ltd. All rights reserved.

Amyloid-β (Aβ), major constituent of senile plaques in Alzheimer's disease (AD), is generated by proteolytic processing of the amyloid precursor protein (APP) by β- and γ-secretase. Several lipids, especially cholesterol, are associated with AD. Phytosterols are naturally occurring cholesterol plant equivalents, recently been shown to cross the blood–brain-barrier accumulating in brain. Here, we investigated the effect of the most nutritional prevalent phytosterols and cholesterol on APP processing. In general, phytosterols are less amyloidogenic than cholesterol. However, only one phytosterol, stigmasterol, reduced Aβ generation by (1) directly decreasing β-secretase activity, (2) reducing expression of all γ-secretase components, (3) reducing cholesterol and presenilin distribution in lipid rafts implicated in amyloidogenic APP cleavage, and by (4) decreasing BACE1 internalization to endosomal compartments, involved in APP β-secretase cleavage. Mice fed with stigmasterol-enriched diets confirmed protective effectsin vivo, suggesting that dietary intake of phytosterol blends mainly containing stigmasterol might be beneficial in preventing AD.

Background Foamy macrophages, containing myelin degradation products, are abundantly found in active multiple sclerosis (MS) lesions. Recent studies have described an altered phenotype of macrophages after myelin internalization. However, mechanisms by which myelin affects the phenotype of macrophages and how this phenotype influences lesion progression remain unclear. Results We demonstrate that myelin as well as phosphatidylserine (PS), a phospholipid found in myelin, reduce nitric oxide production by macrophages through activation of peroxisome proliferator-activated receptor β/δ (PPARβ/δ). Furthermore, uptake of PS by macrophages, after intravenous injection of PS-containing liposomes (PSLs), suppresses the production of inflammatory mediators and ameliorates experimental autoimmune encephalomyelitis (EAE), an animal model of MS. The protective effect of PSLs in EAE animals is associated with a reduced immune cell infiltration into the central nervous system and decreased splenic cognate antigen specific proliferation. Interestingly, PPARβ/δ is activated in foamy macrophages in active MS lesions, indicating that myelin also activates PPARβ/δ in macrophages in the human brain. Conclusion Our data show that myelin modulates the phenotype of macrophages by PPAR activation, which may subsequently dampen MS lesion progression. Moreover, our results suggest that myelin-derived PS mediates PPARβ/δ activation in macrophages after myelin uptake. The immunoregulatory impact of naturally-occurring myelin lipids may hold promise for future MS therapeutics.

Cholesterol and plant sterols are lipids which are abundantly present in a western type diet of animal and plant origin, respectively. The daily intake averages 300 mg/day each. Over the past decades, a steadily increasing consumption of plant sterol enriched dairy products (2-3 g/day) took place to lower circulating LDL cholesterol concentrations. Like all unsaturated components, plant sterols can be attacked by reactive oxygen species resulting in plant sterol oxidation products (POPs). The most widespread methods for POP determination are high-performance liquid chromatography and gas-liquid chromatography. Yet, based on the low plasma POP concentrations in normophytosterolemic subjects (POPs: ∼0.3-4.5 ng/mL), a reliable quantification yielding an appropriate limit of detection remains a challenge. While the more abundantly present cholesterol oxidation products (COPs) have elaborately been studied, research on the metabolism and biological effects of POPs is only emerging. In relation to atherogenity, biological effects including modulation of cholesterol homeostasis, membrane functioning, and inflammation are attributed to POPs. Although mostly supra-physiological concentrations are applied in in vitro assays, anti-tumor activity, cytotoxicity and estrogen-competition have been attributed to specific POPs. However, it is not obvious, if and how POPs may exert in vivo adverse or beneficial health effects similar to those attributed to COPs. In the field of nutritional science, standardized methods for the determination of POPs are required to perform relevant biological studies and to assess their presence in complex foods or biological tissues and fluids. The aim of this review is to provide an overview and evaluation of the published methods and an update on the biological effects attributed to POPs.

Plant sterols such as sitosterol and campesterol are frequently administered as cholesterol-lowering supplements in food. Recently, it has been shown in mice that, in contrast to the structurally related cholesterol, circulating plant sterols can enter the brain. We questioned whether the accumulation of plant sterols in murine brain is reversible. After being fed a plant sterol ester-enriched diet for 6 weeks, C57BL/6NCrl mice displayed significantly increased concentrations of plant sterols in serum, liver, and brain by 2- to 3-fold. Blocking intestinal sterol uptake for the next 6 months while feeding the mice with a plant stanol ester-enriched diet resulted in strongly decreased plant sterol levels in serum and liver, without affecting brain plant sterol levels. Relative to plasma concentrations, brain levels of campesterol were higher than sitosterol, suggesting that campesterol traverses the blood-brain barrier more efficiently.(jlr) In vitro experiments with brain endothelial cell cultures showed that campesterol crossed the blood-brain barrier more efficiently than sitosterol. We conclude that, over a 6-month period, plant sterol accumulation in murine brain is virtually irreversible.-Vanmierlo, T., O. Weingartner, S. van der Pold, C. Huschea, A. Kerksiek, S. Friedrichs, E. Sijbrandse, H. Steinbusch, M. Grimm, T. Hartmann, U. Laufs, M. Bohm, H. E. de Vries, M. Mulder, and D. Lutjohann. Dietary intake of plant sterols stably increases plant sterol levels in the murine brain. J. Lipid Res. 2012. 53: 726-735.

Objective Hypercholesterolemia is a major risk factor for cardiovascular disease (CVD), and diabetes mellitus and statin treatment affect cholesterol metabolism. The aim of the present study was to evaluate markers of cholesterol metabolism and determine their relationship with CVD in patients without diabetes mellitus who were not receiving statin treatment. Methods In addition to conventional CVD risk factors, plasma levels of campesterol and sitosterol (indicators of cholesterol absorption) and lathosterol (an indicator of cholesterol synthesis) were determined in 835 consecutive patients referred for coronary angiography. Coronary artery disease was evaluated by coronary angiograms, carotid atherosclerosis and peripheral vascular disease were assessed by Doppler ultrasound, and cerebrovascular accidents and transient ischemic attacks were identified by medical history. Results After excluding patients with known diabetes mellitus and those receiving statin treatment, 177 patients were included in the analysis. Compared to patients without CVDs (n = 111), patients with concomitant CVDs (n = 66) had a reduced lathosterol-to-cholesterol ratio (1.25 ± 0.61 vs. 1.38 ± 0.63, P

Transporters in the ABCG family appear to be involved in the cellular excretion of cholesterol and other sterols in a cell- and tissue-specific fashion. Overexpression of ATP-binding cassette transporters G1 (Abcg1) and G4 (Abcg4) can promote cellular cholesterol efflux to high-density lipoprotein (HDL), but the in vivo functions of Abcg4 are poorly understood. We used mice with knockouts of Abcg1 or Abcg4 singly or together to further elucidate the function of these transporters. Abcg1 and Abcg4 are highly expressed in the brain and are found in both astrocytes and neurons. Whereas Abcg1(-/-) or Abcg4(-/-) mice showed essentially normal levels of brain sterols, in Abcg1(-/-)/Abcg4(-/-) mice, levels of several sterol intermediates in the cholesterol biosynthetic pathway, namely desmosterol, lathosterol, and lanosterol, as well as 27-OH cholesterol, were increased 2- to 3-fold. Overexpression of Abcg1 or Abcg4 promoted efflux of desmosterol and cholesterol from cells to HDL, and combined deficiency of these transporters led to defective efflux and accumulation of these sterols in primary astrocytes. Consistent with defective efflux and sterol accumulation, cholesterol biosynthesis was reduced in Abcg1(-/-)/Abcg4(-/-) astrocytes. The accumulation of desmosterol, a known liver-X receptor (LXR) activator, was associated with increased expression of LXR target genes, including ATP-binding cassette transporter A1, and increased apolipoprotein E secretion in Abcg1(-/-)/Abcg4(-/-) astrocytes. Our findings provide the first in vivo demonstration of a role for Abcg4 in sterol efflux in the brain and show that Abcg1 and Abcg4 have overlapping functions in astrocytes, promoting efflux of cholesterol, desmosterol, and possibly other sterol biosynthetic intermediates to HDL.

Fibroblasts isolated from a skin biopsy of a healthy individual were infected with Sendai virus containing the Yamanaka factors to produce transgene-free human induced pluripotent stem cells (iPSCs). CRISPR/Cas9 was used to generate an isogenic cell line carrying an inactivation of ST3GAL3, a risk gene associated with neurodevelopmental and psychiatric disorders. This ST3GAL3 null mutant (ST3GAL3-/-) iPSC line, which displays the expression of pluripotency-associated markers, the ability to differentiate into cells of the three germ layers in vitro, and a normal karyotype, is a powerful tool to investigate the impact of deficient sialylation of glycoproteins in neural development and plasticity. This work was supported by ERA-NET NEURON under Grant No. 01EW1902 (DECODE!), the Horizon 2020 Research and Innovation Programme under Grant No. 728018 (Eat2beNICE), Grant No. 953327 (Serotonin and Beyond), the Horizon Europe Research and Innovation Programme under Grant No. EU 101086453 (Aqua-Synapse), the DFG (Grant No. 413657723, Clinician Scientist-Program UNION CVD), and the University of Würzburg in the funding program Open Access Publishing. We thank Julia Merk, Gabriela Ortega, and Nicole Schraud for excellent technical assistance.