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  Glial and vascular contributions to neurodegenerative diseases
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ASSESSMENT OF HEPATIC APOE RELEASE USING HEPATOCARCINOMA CELL LINES
Veronica Cartocci1, Andrew David Lutsky1, Henrietta Nielsen1
1Department of Biochemistry and Biophysics, Stockholm University, Stockholm, Sweden, Stockholm, Sweden
 Background: Apolipoprotein E (APOE) is an important lipid transporter playing a key role in lipid metabolism [2] in which cholesterol loading onto APOE is regulated by the ATP-binding cassette transporter A1 (ABCA1) [5]. APOE and ABCA1 transcription is controlled by Liver X (LXRs) and retinoic X receptors (RXRs) [4-6]. Lack of ABCA1 is related to decreased brain APOE levels and cognitive impairment in a mouse model of Alzheimer´s disease (AD) [3]. Although high levels are synthesized in the brain, APOE is mainly synthesized in the liver. Low plasma APOE is associated with an increased risk of AD and liver-derived APOE4 levels were linked to numerous brain pathological changes [1- 7]. Using APOEε3 homozygous hepatocarcinoma cells, we studied basal and induced hepatic APOE synthesis and release.
Materials and Methods: A combination of cell pharmacology, Western Blot and ELISA was used to study APOE synthesis and release from APOEε3/ε3 HepG2 and Huh7 cells. Effects of LXR and RXR agonists, T0901917 or GW3965 and Bexarotene or 9-cis-retinoic acid on APOE and ABCA1 levels in supernatants and cell pellets were evaluated after 24 and 48hrs.
Results: LXR agonist T0901317 (1-10 μM) induced a near 80% upregulation in APOE secretion without altering intracellular APOE levels in both cell lines. ABCA1 protein levels increased by nearly 60% in both cell lines. Only in HepG2 cells, APOE release increased after GW3965 exposure (2 μM).
Discussion: Liver-derived plasma APOE levels are known to be APOE genotype-dependent. Here, we found that LXR agonism significantly upregulated APOE release and ABCA1 levels in APOEε3/ε3 hepatocarcinoma cells. Whether LXR-induced liver-secretion of APOE differs between APOE genotypes remains to be investigated. Conclusion: Our results evidence in vitro modulation of hepatic APOE and ABCA1 synthesis by LXR/RXR agonism which can be used as tools to understand the regulation of liver-secreted APOE and ABCA1.
(AD) progression, especially in patients carrying the apolipoprotein E ε4 (APOE4) isoform. The drug FTY720 mimics S1P bioactivity, but its efficacy in treating AD is unclear.
Methods: Two doses of FTY720 (0.1 mg/kg and 0.5 mg/kg daily) were given by oral gavage for 15 weeks to transgenic mouse models of familial AD carrying human apolipoprotein E (APOE) APOE3 (E3FAD) or APOE4 (E4FAD). After 12 weeks of treatment, animals were subjected to behavioral tests for memory, locomotion, and anxiety. Blood was withdrawn at different time points and brains were collected for sphingolipids analysis by mass spectrometry, gene expression by RT- PCR and Aβ quantification by ELISA.
Results: We discovered that low levels of S1P in the plasma is associated with a higher probability of failing the memory test and that FTY720 prevents memory impairments in E4FAD. The beneficial effect of FTY720 was induced by a shift of the sphingolipid metabolism in the brain towards a lower production of toxic metabolites, like ceramide d18:1/16:0 and d18:1/22:0, and reduction of amyloid-β burden and inflammation.
Conclusion: We provide further evidence of the druggability of the sphingolipid system in AD.
Keywords: 5xFAD; APOE3; APOE4; Anxiety; Ceramide; FTY720; Memory; S1P; Sphingomyelin.
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Attenuation of progressive Alzheimer’s Disease-like Pathology by Pharmacologic inhibition and Genetic deletion of Bach1 in the APP/PS1 mouse
Debashis Dutta1,2, Christopher Ye In Kwon10, Manuj Ahuja1,2,7, Navneet Ammal Kaidery1,2, Joyce Meints10,Otis C Attucks8, Carmen Valcarce8, Sudarshana Sharma5,6, Mitsuyo Matsumoto9, Kazuhiko Igarashi9, Michael K Lee10, Bobby Thomas1,2,3,4
1Darby Children’s Research Institute, Medical University of South Carolina, Charleston,
USA, 2Departments of Pediatrics, Medical University of South Carolina, Charleston, USA, 3Department of Neuroscience, Medical University of South Carolina, Charleston, USA, 4Department of Drug Discovery, Medical University of South Carolina, Charleston, USA, 5Department of Biochemistry, Medical University of South Carolina, Charleston, USA, 6Hollings Cancer Center, Medical University of South Carolina, Charleston, USA, 7Department of Pharmaceutical Sciences, The State University of New York at Buffalo, Buffalo, USA, 8vTv Therapeutics LLC, High Point, USA, 9Department of Biochemistry, Tohoku University, Graduate School of Medicine, Sendai, Japan, 10Department of Neuroscience, University of Minnesota, Minneapolis, USA
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FTY720 Decreases Ceramide Levels in the Brain and Prevents Memory Impairments in a Mouse Model of Familial Alzheimer's Disease Expressing APOE4
Daan van Kruining1, Simone Crivelli1,2, Qian Luo1, Caterina Giovagnoni1, Marina Mané-Damas1, Sandra De Hoedt3, Dusan Berkes4, Helga De Vries5, Monique Mulder3, Jochen Walter6, Etienne Waelkens7, Rita Derua7, Johannes Swinnen8, Jonas Dehairs8, Erwin Wijnands9, Erhard Bieberich2, Mario Losen1, Pilar Martinez-Martinez1 1Department of Psychiatry and Neuropsychology, Maastricht University, Maastricht ,
the Netherlands, 2Department of Physiology, University of Kentucky College of Medicine, Lexington, USA, 3Department of Internal Medicine, Erasmus MC University Medical
Center, Rotterdam, the Netherlands, 4Department of Organic Chemistry, Slovak University of Technology, Bratislava, Slovak Republic, 5Department of Molecular Cell Biology and Immunology, Amsterdam UMC, Amsterdam, the Netherlands, 6Department of Neurology, University Hospital Bonn, Bonn, Germany, 7Laboratory of Protein Phosphorylation and Proteomics, KU Leuven, Leuven, Belgium, 8Laboratory of Lipid Metabolism and Cancer,
KU Leuven, Leuven , Belgium, 9Department of Pathology, Maastricht University, Maastricht, the Netherlands
 Background: The protection mediated by the bioactive sphingolipid sphingosine-1-phosphate (S1P) declines during Alzheimer's disease
Background: A decline in the expression of the redox-dependent transcription factor nuclear-factor-erythroid 2-related factor 2 (Nrf2) is observed in humans and animal models of Alzheimer’s disease (AD) [1]. Though activation of Nrf2 is a promising therapeutic strategy [2-4], FDA- approved electrophilic Nrf2 activators cause irreversible alkylation of cysteine residues in various cellular proteins resulting in side effects [5- 6]. We observed significant upregulation of the transcriptional repressor of Nrf2, BTB and CNC homology 1 (Bach1) in AD suggesting that Bach1 inhibition might be neuroprotective. We tested therapeutic potential of a non-electrophilic Bach1 inhibitor (HPPE) and genetic deletion of Bach1 in APP/PS1 model of AD.
Materials and Methods: Cohorts of APP/PS1 transgenic mice were administered with HPPE for 45 days (20 mg/kg twice a day, 12 h apart) starting at 11 months and 15 months of age. APP/PS1 mice were crossed with Bach1-deficient mice to genetically ablate Bach1 in APP/PS1 mice. Behavioral analyses were conducted by Barnes maze and novel object recognition tests. Amyloid pathology and gliosis in the brain was monitored by immunohistochemistry of Aβ and Iba1 respectively. Integrity of cortical monoaminergic afferents was examined by tyrosine hydroxylase (TH) immunostaining coupled with unbiased stereology. Neuroprotective pathways were identified by RNA-seq analysis. Results: The HPPE treatment significantly reduced Aβ deposits, microgliosis, and attenuated the progressive loss of cortical TH afferents resulting in significant improvement in cognitive performance of the
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