Using a mouse model of multiple sclerosis (MS), experimental autoimmune encephalitis (EAE), we evaluated the role of gut microbiota in modulating chronic-progressive (CP) versus relapse-remitting ...(RR) forms of the disease. We hypothesized that clinical courses of EAE may be shaped by differential gut microbiota. Metagenomic sequencing of prokaryotic 16S rRNA present in feces from naïve mice and those exhibiting CP-EAE or RR-EAE revealed significantly diverse microbial populations. Microbiota composition was considerably different between naïve strains of mice, suggesting microbial components present in homeostatic conditions may prime mice for divergent courses of disease. Additionally, there were differentially abundant bacteria in CP and RR forms of EAE, indicating a potential role for gut microbiota in shaping tolerant or remittance-favoring, and pathogenic or pro-inflammatory-promoting conditions. Furthermore, immunization to induce EAE led to significant alterations in gut microbiota, some were shared between disease courses and others were course-specific, supporting a role for gut microbial composition in EAE pathogenesis. Moreover, using Linear Discriminant Analysis (LDA) coupled with effect size measurement (LEfSe) to analyze microbial content, biomarkers of each naïve and disease states were identified. Our findings demonstrate for the first time that gut microbiota may determine the susceptibility to CP or RR forms of EAE.
Sphingosine 1-phosphate (S1P) is an important bioactive sphingolipid metabolite that has been implicated in numerous physiological and cellular processes. Not only does S1P play a structural role in ...cells by defining the components of the plasma membrane, but in the last 20years it has been implicated in various significant cell signaling pathways and physiological processes: for example, cell migration, survival and proliferation, cellular architecture, cell–cell contacts and adhesions, vascular development, atherosclerosis, acute pulmonary injury and respiratory distress, inflammation and immunity, and tumorogenesis and metastasis 1,2. Given the wide variety of cellular and physiological processes in which S1P is involved, it is immediately obvious why the mechanisms governing S1P synthesis and degradation, and the manner in which these processes are regulated, are necessary to understand. In gaining more knowledge about regulation of the sphingosine kinase (SK)/S1P pathway, many potential therapeutic targets may be revealed. This review explores the roles of the SK/S1P pathway in disease, summarizes available SK enzyme inhibitors and examines their potential as therapeutic agents. This article is part of a Special Issue entitled Advances in Lysophospholipid Research.
► The SK/S1P pathway is dysregulated in numerous pathologies and disease states. ► Only recently have potent and specific inhibitors of SK begun to be developed. ► Further development of SK inhibitors may provide novel disease treatments.
Resveratrol (3,5,4′-trihydroxy-
trans-
stilbene) (RES) is a naturally-derived phytoestrogen found in the skins of red grapes and berries and has potential as a novel and effective therapeutic agent. ...In the current study, we investigated the role of microRNA (miRNA) in RES-mediated attenuation of experimental autoimmune encephalomyelitis (EAE), a murine model of multiple sclerosis. Administration of RES effectively decreased disease severity, including inflammation and central nervous system immune cell infiltration. miRNA microarray analysis revealed an altered miRNA profile in encephalitogenic CD4+ T cells from EAE mice exposed to RES treatment. Additionally, bioinformatics and in silico pathway analysis suggested the involvement of RES-induced miRNA in pathways and processes that regulated cellular proliferation. Additional studies confirmed that RES affected cell cycle progression and apoptosis in activated T cells, specifically in the brain. RES treatment significantly upregulated miR-124 during EAE, while suppressing associated target gene, sphingosine kinase 1 (SK1), and this too was specific to mononuclear cells in the brains of treated mice, as peripheral immune cells remained unaltered upon RES treatment. Collectively, these studies demonstrate that RES treatment leads to amelioration of EAE development through mechanism(s) potentially involving suppression of neuroinflammation via alteration of the miR-124/SK1 axis, thereby halting cell-cycle progression and promoting apoptosis in activated encephalitogenic T cells.
Graphical Abstract
Resveratrol alters the miR-124/sphingosine kinase 1 (SK1) axis in encephalitogenic T cells, promotes cell-cycle arrest and apoptosis, and decreases neuroinflammation in experiemental autoimmune encephalomyelitis (EAE).
Sphingolipids and their synthetic enzymes are emerging as important mediators in inflammatory responses and as regulators of immune cell functions. In particular, sphingosine kinase (SK) and its ...product sphingosine-1-phosphate (S1P) have been extensively implicated in these processes. SK catalyzes the phosphorylation of sphingosine to S1P and exists as two isoforms, SK1 and SK2. SK1 has been shown to be activated by cytokines including tumor necrosis factor-alpha (TNF-α) and interleukin1-β (IL1-β). The activation of SK1 in this pathway has been shown to be, at least in part, required for mediating TNF-α and IL1-β inflammatory responses in cells, including induction of cyclo-oxygenase 2 (COX2). In addition to their role in inflammatory signaling, SK and S1P have also been implicated in various immune cell functions including, mast cell degranulation, migration of neutrophils, and migration and maturation of lymphocytes. The involvement of sphingolipids and sphingolipid metabolizing enzymes in inflammatory signaling and immune cell functions has implicated these mediators in numerous inflammatory disease states as well. The contribution of these mediators, specifically SK1 and S1P, to inflammation and disease are discussed in this review.
Previously we demonstrated that the sphingolipids ceramide and S1P (sphingosine 1-phosphate) regulate phosphorylation of the ERM (ezrin/radixin/moesin) family of cytoskeletal proteins Canals, ...Jenkins, Roddy, Hernande-Corbacho, Obeid and Hannun (2010) J. Biol. Chem. 285, 32476-3285. In the present article, we show that exogenously applied or endogenously generated S1P (in a sphingosine kinase-dependent manner) results in significant increases in phosphorylation of ERM proteins as well as filopodia formation. Using phosphomimetic and non-phosphorylatable ezrin mutants, we show that the S1P-induced cytoskeletal protrusions are dependent on ERM phosphorylation. Employing various pharmacological S1PR (S1P receptor) agonists and antagonists, along with siRNA (small interfering RNA) techniques and genetic knockout approaches, we identify the S1PR2 as the specific and necessary receptor to induce phosphorylation of ERM proteins and subsequent filopodia formation. Taken together, the results demonstrate a novel mechanism by which S1P regulates cellular architecture that requires S1PR2 and subsequent phosphorylation of ERM proteins.
Ezrin, radixin, and moesin (ERM) proteins link cortical actin to the plasma membrane and coordinate cellular events that require cytoskeletal rearrangement, including cell division, migration, and ...invasion. While ERM proteins are involved in many important cellular events, the mechanisms regulating their function are not completely understood. Our laboratory previously identified reciprocal roles for the sphingolipids ceramide and sphingosine‐1‐phosphate (S1P) in the regulation of ERM proteins. We recently showed that ceramide‐induced activation of PP1α leads to dephosphorylation and inactivation of ERM proteins, while S1P results in phosphorylation and activation of ERM proteins. Following these findings, we aimed to examine known inducers of the SK/S1P pathway and evaluate their ability to regulate ERM proteins. We examined EGF, a known inducer of the SK/S1P pathway, for its ability to regulate the ERM family of proteins. We found that EGF induces ERM c‐terminal threonine phosphorylation via activation of the SK/S1P pathway, as this was prevented by siRNA knockdown or pharmacological inhibition of SK. Using pharmacological, as well as genetic, knockdown approaches, we determined that EGF induces ERM phosphorylation via activation of S1PR2. In addition, EGF led to cell polarization in the form of lamellipodia, and this occurred through a mechanism involving S1PR2‐mediated phosphorylation of ezrin T567. EGF‐induced cellular invasion was also found to be dependent on S1PR2‐induced T567 ezrin phosphorylation, such that S1PR2 antagonist, JTE‐013, and expression of a dominant‐negative ezrin mutant prevented cellular invasion toward EGF. In this work, a novel mechanism of EGF‐stimulated invasion is unveiled, whereby S1P‐mediated activation of S1PR2 and phosphorylation of ezrin T567 is required.—Orr Gandy, K. A., Adada, M., Canals, D., Carroll, B., Roddy, P., Hannun, Y. A.Obeid, L. M., Epidermal growth factor‐induced cellular invasion requires sphingosine‐1‐phosphate/sphingosine‐1‐phosphate 2 receptor‐mediated ezrin activation. FASEB J. 27, 3155–3166 (2013). www.fasebj.org
Sphingosine kinase 1 (SK1) produces the pro-survival sphingolipid sphingosine 1-phosphate and has been implicated in inflammation, proliferation, and angiogenesis. Recent studies identified TRAF2 as ...a sphingosine 1-phosphate target, implicating SK1 in activation of the NF-κB pathway, but the functional consequences of this connection on gene expression are unknown. Here, we find that loss of SK1 potentiates induction of the chemokine RANTES (regulated on activation, normal T cell expressed and secreted; also known as CCL5) in HeLa cells stimulated with TNF-α despite RANTES induction being highly dependent on the NF-κB pathway. Additionally, we find that SK1 is not required for TNF-induced IKK phosphorylation, IκB degradation, nuclear translocation of NF-κB subunits, and transcriptional NF-κB activity. In contrast, loss of SK1 prevented TNF-induced phosphorylation of p38 MAPK, and inhibition of p38 MAPK, like SK1 knockdown, also potentiates RANTES induction. Finally, in addition to RANTES, loss of SK1 also potentiated the induction of multiple chemokines and cytokines in the TNF response. Taken together, these data identify a potential and novel anti-inflammatory function of SK1 in which chemokine levels are suppressed through SK1-mediated activation of p38 MAPK. Furthermore, in this system, activation of NF-κB is dissociated from SK1, suggesting that the interaction between these pathways may be more complex than currently thought.
Background: SK1 is widely involved in promoting inflammatory diseases.
Results: Unexpectedly, loss of SK1 caused an increase in chemokines upon TNF stimulation. This occurred through regulation of p38 MAPK but independently of NF-κB.
Conclusion: SK1 negatively regulates RANTES expression through the p38 MAPK pathway.
Significance: Targeting SK1 in therapy may affect inflammatory conditions by up-regulating chemokines independently of NF-κB.
Sphingolipids have emerged as pleiotropic signaling molecules with roles in numerous cellular and biological functions. Defining the regulatory mechanisms governing sphingolipid metabolism is crucial ...in order to develop a complete understanding of the biological functions of sphingolipid metabolites. The sphingosine kinase/ sphingosine 1-phosphate pathway was originally thought to function in the irreversible breakdown of sphingoid bases; however, in the last few decades it has materialized as an extremely important signaling pathway involved in a plethora of cellular events contributing to both normal and pathophysiological events. Recognition of the SK/S1P pathway as a second messaging system has aided in the identification of many mechanisms of its regulation; however, a cohesive, global understanding of the regulatory mechanisms controlling the SK/S1P pathway is lacking. In this chapter, the role of the SK/S1P pathway as a second messenger is discussed, and its role in mediating TNF-α- and EGF-induced biologies is examined. This work provides a comprehensive look into the roles and regulation of the sphingosine kinase/ sphingosine 1-phosphate pathway and highlights the potential of the pathway as a therapeutic target.
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•Novel sphingosine analog, LCL351, inhibits SK1 in cells (10X more selective for SK1 over SK2 in vitro).•LCL351 treatment in vivo demonstrated protection from weight loss, blood loss, ...and splenomegaly in a mouse model of IBD.•LCL351 reduced induction of inflammatory cytokines and neutrophil infiltration in colon tissue in a mouse model of IBD.
Sphingosine-1-phosphate (S1P) is a biologically active sphingolipid metabolite which has been implicated in many diseases including cancer and inflammatory diseases. Recently, sphingosine kinase 1 (SK1), one of the isozymes which generates S1P, has been implicated in the development and progression of inflammatory bowel disease (IBD). Based on our previous work, we set out to determine the efficacy of a novel SK1 selective inhibitor, LCL351, in a murine model of IBD. LCL351 selectively inhibits SK1 both in vitro and in cells. LCL351, which accumulates in relevant tissues such as colon, did not have any adverse side effects in vivo. In mice challenged with dextran sodium sulfate (DSS), a murine model for IBD, LCL351 treatment protected from blood loss and splenomegaly. Additionally, LCL351 treatment reduced the expression of pro-inflammatory markers, and reduced neutrophil infiltration in colon tissue. Our results suggest inflammation associated with IBD can be targeted pharmacologically through the inhibition and degradation of SK1. Furthermore, our data also identifies desirable properties of SK1 inhibitors.
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