The Formyl Peptide Receptor 2 Regulates Microglial Phenotype Through Immunometabolism: Implications for Alzheimer’s Disease

Microglia are key players in the pathology of Alzheimer’s disease (AD), driving chronic inflammation, oxidative stress, and the altered metabolism seen in the brains of patients. With clinical trials continuing to fail, new approaches towards drug development are critical. Strategies to reduce microglial activation may therefore be a viable therapeutic approach to tackling AD. Formyl peptide receptor 2 (Fpr2), which drives peripheral inflammatory resolution, is expressed in microglia. However, its functional role in neuroinflammation is unclear. This thesis provides evidence to support the peripheral findings of Fpr2 stimulation, wherein it may also hold promise for exploitation as a therapeutic for neurodegenerative disorders, including AD. We also highlight novel findings surrounding the modulation of both oxidative stress and microglial metabolism associated with Fpr2 activation. Under inflammatory conditions, we report that selective agonists for Fpr2 modulate the microglial inflammatory response, actively shifting from a pro-inflammatory to a pro-resolving phenotype, emphasised by the reduction of pro-inflammatory cytokines and concomitant increases in both pro-resolving cytokines and phagocytosis. Metabolic shifting away from glycolysis was also observed for pro-resolving microglia. Moreover, we describe for the first time that Fpr2 completely reverses reactive oxygen species (ROS) production from the mitochondria and NADPH oxidase enzymes following an inflammatory stimulus. We also highlight that the toxic oligomeric amyloid (oAβ) facilitates microglial ROS production and subsequent metabolic changes without triggering an inflammatory response. oAβ facilitated NADPH oxidase activation, which in turn resulted in the activation of glucose 6-phosphate dehydrogenase (G6PD), the rate limiting step for the pentose phosphate pathway. This metabolic pathway is responsible for producing NADPH, which in turn NADPH oxidases exploit for further ROS production. These changes resulted in noticeable reductions in both microglial glycolysis and oxidative phosphorylation. We present data underlining that Fpr2/3 stimulation reverses oAβ-induced ROS production, with a resultant reduction in G6PD activity and the return of homeostatic glycolysis. These oAβ-induced microglial changes triggered the apoptosis of SH-SY5Y cells in co-culture with BV-2 microglia. However, supporting our interest in Fpr2/3 for therapeutic approaches to neurodegenerative diseases, post-treatment with a select agonist for the receptor successfully prevented apoptosis of these neuronal like SH-SY5Y cells. This original data unveils novel functions of Fpr2/3 in the central nervous system (CNS), supplementing the well-established pro-resolving functions the receptor facilitates within the periphery. The combination of pro-resolving, anti-oxidative, immunometabolic and anti-apoptotic functions of Fpr2/3 support the exploitation of this receptor for therapeutic research into multiple different CNS disorders, including AD

Banach lattices of L-weakly and M-weakly compact operators

We give conditions for the linear span of the positive L-weakly compact (resp. M-weakly compact) operators to be a Banach lattice under the regular norm, for that Banach lattice to have an order continuous norm, to be an AL-space or an AM-space.Scientific and Technological Research Council of Turkey (TUBITAK)Turkiye Bilimsel ve Teknolojik Arastirma Kurumu (TUBITAK) [2219]; Research Foundation of Namik Kemal UniversityNamik Kemal University [NKUBAP.00.10.AR.15.04]The first author was supported by The Scientific and Technological Research Council of Turkey (TUBITAK) within the context of 2219-Post Doctoral Fellowship Program and by the Research Foundation of Namik Kemal University (Project No. NKUBAP.00.10.AR.15.04.)

Activation of the pro-resolving receptor Fpr2 attenuates inflammatory microglial activation

Poster number: P-T099 Theme: Neurodegenerative disorders & ageing Activation of the pro-resolving receptor Fpr2 reverses inflammatory microglial activation Authors: Edward S Wickstead - Life Science & Technology University of Westminster/Queen Mary University of London Inflammation is a major contributor to many neurodegenerative disease (Heneka et al. 2015). Microglia, as the resident immune cells of the brain and spinal cord, provide the first line of immunological defence, but can become deleterious when chronically activated, triggering extensive neuronal damage (Cunningham, 2013). Dampening or even reversing this activation may provide neuronal protection against chronic inflammatory damage. The aim of this study was to determine whether lipopolysaccharide (LPS)-induced inflammation could be abrogated through activation of the receptor Fpr2, known to play an important role in peripheral inflammatory resolution. Immortalised murine microglia (BV2 cell line) were stimulated with LPS (50ng/ml) for 1 hour prior to the treatment with one of two Fpr2 ligands, either Cpd43 or Quin-C1 (both 100nM), and production of nitric oxide (NO), tumour necrosis factor alpha (TNFα) and interleukin-10 (IL-10) were monitored after 24h and 48h. Treatment with either Fpr2 ligand significantly suppressed LPS-induced production of NO or TNFα after both 24h and 48h exposure, moreover Fpr2 ligand treatment significantly enhanced production of IL-10 48h post-LPS treatment. As we have previously shown Fpr2 to be coupled to a number of intracellular signaling pathways (Cooray et al. 2013), we investigated potential signaling responses. Western blot analysis revealed no activation of ERK1/2, but identified a rapid and potent activation of p38 MAP kinase in BV2 microglia following stimulation with Fpr2 ligands. Together, these data indicate the possibility of exploiting immunomodulatory strategies for the treatment of neurological diseases, and highlight in particular the important potential of resolution mechanisms as novel therapeutic targets in neuroinflammation. References Cooray SN et al. (2013). Proc Natl Acad Sci U S A 110: 18232-7. Cunningham C (2013). Glia 61: 71-90. Heneka MT et al. (2015). Lancet Neurol 14: 388-40

Reversal of beta-Amyloid-Induced Microglial Toxicity In Vitro by Activation of Fpr2/3

Microglial inflammatory activity is thought to be a major contributor to the pathology of neurodegenerative conditions such as Alzheimer’s disease (AD), and strategies to restrain their behaviour are under active investigation. Classically, anti-inflammatory approaches are aimed at suppressing proinflammatory mediator production, but exploitation of inflammatory resolution, the endogenous process whereby an inflammatory reaction is terminated, has not been fully investigated as a therapeutic approach in AD. In this study, we sought to provide proof-of-principle that the major proresolving actor, formyl peptide receptor 2, Fpr2, could be targeted to reverse microglial activation induced by the AD-associated proinflammatory stimulus, oligomeric β-amyloid (oAβ). The immortalised murine microglial cell line BV2 was employed as a model system to investigate the proresolving effects of the Fpr2 ligand QC1 upon oAβ-induced inflammatory, oxidative, and metabolic behaviour. Cytotoxic behaviour of BV2 cells was assessed through the use of cocultures with retinoic acid-differentiated human SH-SY5Y cells. Stimulation of BV2 cells with oAβ at 100 nM did not induce classical inflammatory marker production but did stimulate production of reactive oxygen species (ROS), an effect that could be reversed by subsequent treatment with the Fpr2 ligand QC1. Further investigation revealed that oAβ-induced ROS production was associated with NADPH oxidase activation and a shift in BV2 cell metabolic phenotype, activating the pentose phosphate pathway and NADPH production, changes that were again reversed by QC1 treatment. Microglial oAβ-stimulated ROS production was sufficient to induce apoptosis of bystander SH-SY5Y cells, an effect that could be prevented by QC1 treatment. In this study, we provide proof-of-concept data that indicate exploitation of the proresolving receptor Fpr2 can reverse damaging oAβ-induced microglial activation. Future strategies that are aimed at restraining neuroinflammation in conditions such as AD should examine proresolving actors as a mechanism to harness the brain’s endogenous healing pathways and limit neuroinflammatory damage

Exploiting formyl peptide receptor 2 to promote microglial resolution: a new approach to Alzheimer's disease treatment.

Alzheimer’s disease and dementia are among the most significant current healthcare challenges given the rapidly growing elderly population, and the almost total lack of effective therapeutic interventions. Alzheimer’s disease pathology has long been considered in terms of accumulation of amyloid beta and hyperphosphorylated tau, but the importance of neuroinflammation in driving disease has taken greater precedence over the last 15–20 years. Inflammatory activation of the primary brain immune cells, the microglia, has been implicated in Alzheimer’s pathogenesis through genetic, preclinical, imaging and postmortem human studies, and strategies to regulate microglial activity may hold great promise for disease modification. Neuroinflammation is necessary for defence of the brain against pathogen invasion or damage but is normally self-limiting due to the engagement of endogenous pro-resolving circuitry that terminates inflammatory activity, a process that appears to fail in Alzheimer’s disease. Here, we discuss the potential for a major regulator and promoter of resolution, the receptor FPR2, to restrain pro-inflammatory microglial activity, and propose that it may serve as a valuable target for therapeutic investigation in Alzheimer’s disease

Conservation of ciliary proteins in plants with no cilia

<p>Abstract</p> <p>Background</p> <p>Eukaryotic cilia are complex, highly conserved microtubule-based organelles with a broad phylogenetic distribution. Cilia were present in the last eukaryotic common ancestor and many proteins involved in cilia function have been conserved through eukaryotic diversification. However, cilia have also been lost multiple times in different lineages, with at least two losses occurring within the land plants. Whereas all non-seed plants produce cilia for motility of male gametes, some gymnosperms and all angiosperms lack cilia. During these evolutionary losses, proteins with ancestral ciliary functions may be lost or co-opted into different functions.</p> <p>Results</p> <p>Here we identify a core set of proteins with an inferred ciliary function that are conserved in ciliated eukaryotic species. We interrogate this genomic dataset to identify proteins with a predicted ancestral ciliary role that have been maintained in non-ciliated land plants. In support of our prediction, we demonstrate that several of these proteins have a flagellar localisation in protozoan trypanosomes. The phylogenetic distribution of these genes within the land plants indicates evolutionary scenarios of either sub- or neo-functionalisation and expression data analysis shows that these genes are highly expressed in <it>Arabidopsis thaliana </it>pollen cells.</p> <p>Conclusions</p> <p>A large number of proteins possess a phylogenetic ciliary profile indicative of ciliary function. Remarkably, many genes with an ancestral ciliary role are maintained in non-ciliated land plants. These proteins have been co-opted to perform novel functions, most likely before the loss of cilia, some of which appear related to the formation of the male gametes.</p

Promiscuous Receptors and Neuroinflammation: The Formyl Peptide Class

Formyl peptide receptors, abbreviated as FPRs in humans, are G-protein coupled receptors (GPCRs) mainly found in mammalian leukocytes. However, they are also expressed in cell types crucial for homeostatic brain regulation, including microglia and blood–brain barrier endothelial cells. Thus, the roles of these immune-associated receptors are extensive, from governing cellular adhesion and directed migration through chemotaxis, to granule release and superoxide formation, to phagocytosis and efferocytosis. In this review, we will describe the similarities and differences between the two principal pro-inflammatory and anti-inflammatory FPRs, FPR1 and FPR2, and the evidence for their importance in the development of neuroinflammatory disease, alongside their potential as therapeutic targets

Estrogen Promotes Pro-resolving Microglial Behavior and Phagocytic Cell Clearance Through the Actions of Annexin A1

Local production of estrogen rapidly follows brain tissue injury, but the role this hormone plays in regulating the response to neural damage or in the modulation of mediators regulating inflammation is in many ways unclear. Using the murine BV2 microglia model as well as primary microglia from wild-type and annexin A1 (AnxA1) null mice, we have identified two related mechanisms whereby estradiol can modulate microglial behavior in a receptor specific fashion. Firstly, estradiol, via estrogen receptor β (ERβ), enhanced the phagocytic clearance of apoptotic cells, acting through increased production and release of the protein AnxA1. Secondly, stimulation of either ERβ or the G protein coupled estrogen receptor GPER promoted the adoption of an anti-inflammatory/pro-resolving phenotype, an action similarly mediated through AnxA1. Together, these data suggest the hypothesis that locally produced estrogen acts through AnxA1 to exert powerful pro-resolving actions, controlling and limiting brain inflammation and ultimately protecting this highly vulnerable organ. Given the high degree of receptor selectivity in evoking these responses, we suggest that the use of selective estrogen receptor ligands may hold therapeutic promise in the treatment of neuroinflammation, avoiding unwanted generalized effects

The Expanded Kinesin-13 Repertoire of Trypanosomes Contains Only One Mitotic Kinesin Indicating Multiple Extra-Nuclear Roles

BACKGROUND: Kinesin-13 proteins have a critical role in animal cell mitosis, during which they regulate spindle microtubule dynamics through their depolymerisation activity. Much of what is known about Kinesin-13 function emanates from a relatively small sub-family of proteins containing MCAK and Kif2A/B. However, recent work on kinesins from the much more widely distributed, ancestral Kinesin-13 family, which includes human Kif24, have identified a second function in flagellum length regulation that may exist either alongside or instead of the mitotic role. METHODOLOGY/PRINCIPAL FINDINGS: The African trypanosome Trypanosoma brucei encodes 7 distinct Kinesin-13 proteins, allowing scope for extensive specialisation of roles. Here, we show that of all the trypanosomal Kinesin-13 proteins, only one is nuclear. This protein, TbKIN13-1, is present in the nucleoplasm throughout the cell cycle, but associates with the spindle during mitosis, which in trypanosomes is closed. TbKIN13-1 is necessary for the segregation of both large and mini-chromosomes in this organism and reduction in TbKIN13-1 levels mediated by RNA interference causes deflects in spindle disassembly with spindle-like structures persisting in non-mitotic cells. A second Kinesin-13 is localised to the flagellum tip, but the majority of the Kinesin-13 family members are in neither of these cellular locations. CONCLUSIONS/SIGNIFICANCE: These data show that the expanded Kinesin-13 repertoire of trypanosomes is not associated with diversification of spindle-associated roles. TbKIN13-1 is required for correct spindle function, but the extra-nuclear localisation of the remaining paralogues suggests that the biological roles of the Kinesin-13 family is wider than previously thought