RESUMEN
Reciprocal interactions between breast cancer cells and the tumor microenvironment (TME) are important for cancer progression and metastasis. We report here that the deletion or inhibition of sphingosine kinase 2 (SphK2), which produces sphingosine-1-phosphate (S1P), markedly suppresses syngeneic breast tumor growth and lung metastasis in mice by creating a hostile microenvironment for tumor growth and invasion. SphK2 deficiency decreased S1P and concomitantly increased ceramides, including C16-ceramide, in stromal fibroblasts. Ceramide accumulation suppressed activation of cancer-associated fibroblasts (CAF) by upregulating stromal p53, which restrained production of tumor-promoting factors to reprogram the TME and to restrict breast cancer establishment. Ablation of p53 in SphK2-deficient fibroblasts reversed these effects, enabled CAF activation and promoted tumor growth and invasion. These data uncovered a novel role of SphK2 in regulating non-cell-autonomous functions of p53 in stromal fibroblasts and their transition to tumor-promoting CAFs, paving the way for the development of a strategy to target the TME and to enhance therapeutic efficacy. SIGNIFICANCE: Sphingosine kinase 2 (SphK2) facilitates the activation of stromal fibroblasts to tumor-promoting cancer-associated fibroblasts by suppressing host p53 activity, revealing SphK2 as a potential target to reprogram the TME.
Asunto(s)
Fibroblastos Asociados al Cáncer , Neoplasias Mamarias Animales , Fosfotransferasas (Aceptor de Grupo Alcohol) , Microambiente Tumoral , Animales , Ratones , Fibroblastos Asociados al Cáncer/metabolismo , Fibroblastos/metabolismo , Neoplasias Pulmonares/patología , Neoplasias Pulmonares/secundario , Neoplasias Mamarias Animales/metabolismo , Neoplasias Mamarias Animales/patología , Fosfotransferasas (Aceptor de Grupo Alcohol)/genética , Fosfotransferasas (Aceptor de Grupo Alcohol)/metabolismo , Microambiente Tumoral/fisiología , Proteína p53 Supresora de Tumor/genéticaRESUMEN
The bioactive sphingolipid metabolites ceramide and sphingosine-1-phosphate (S1P) are a recent addition to the lipids accumulated in obesity and have emerged as important molecular players in metabolic diseases. Here we summarize evidence that dysregulation of sphingolipid metabolism correlates with pathogenesis of metabolic diseases in humans. This review discusses the current understanding of how ceramide regulates signaling and metabolic pathways to exacerbate metabolic diseases and the Janus faces for its further metabolite S1P, the kinases that produce it, and the multifaceted and at times opposing actions of S1P receptors in various tissues. Gaps and limitations in current knowledge are highlighted together with the need to further decipher the full array of their actions in tissue dysfunction underlying metabolic pathologies, pointing out prospects to move this young field of research toward the development of effective therapeutics.
Asunto(s)
Enfermedades Metabólicas/etiología , Esfingolípidos/fisiología , Animales , Humanos , Metabolismo de los Lípidos/fisiología , Enfermedades Metabólicas/metabolismo , Enfermedades Metabólicas/patología , Redes y Vías Metabólicas/fisiología , Obesidad/etiología , Obesidad/metabolismo , Obesidad/patología , Esfingolípidos/metabolismoRESUMEN
The serine palmitoyltransferase (SPT) complex catalyzes the rate-limiting step in the de novo biosynthesis of ceramides, the precursors of sphingolipids. The mammalian ORMDL isoforms (ORMDL1-3) are negative regulators of SPT. However, the roles of individual ORMDL isoforms are unclear. Using siRNA against individual ORMDLs, only single siORMDL3 had modest effects on dihydroceramide and ceramide levels, whereas downregulation of all three ORMDLs induced more pronounced increases. With the CRISPR/Cas9-based genome-editing strategy, we established stable single ORMDL3 KO (ORMDL3-KO) and ORMDL1/2/3 triple-KO (ORMDL-TKO) cell lines to further understand the roles of ORMDL proteins in sphingolipid biosynthesis. While ORMDL3-KO modestly increased dihydroceramide and ceramide levels, ORMDL-TKO cells had dramatic increases in the accumulation of these sphingolipid precursors. SPT activity was increased only in ORMDL-TKO cells. In addition, ORMDL-TKO but not ORMDL3-KO dramatically increased levels of galactosylceramides, glucosylceramides, and lactosylceramides, the elevated N-acyl chain distributions of which broadly correlated with the increases in ceramide species. Surprisingly, although C16:0 is the major sphingomyelin species, it was only increased in ORMDL3-KO, whereas all other N-acyl chain sphingomyelin species were significantly increased in ORMDL-TKO cells. Analysis of sphingoid bases revealed that although sphingosine was only increased 2-fold in ORMDL-TKO cells, levels of dihydrosphingosine, dihydrosphingosine-1-phosphate, and sphingosine-1-phosphate were hugely increased in ORMDL-TKO cells and not in ORMDL3-KO cells. Thus, ORMDL proteins may have a complex, multifaceted role in the biosynthesis and regulation of cellular sphingolipids.
Asunto(s)
Sistemas CRISPR-CasRESUMEN
Severity of cardiovascular disease increases markedly in elderly patients. In addition, many therapeutic strategies that decrease cardiac injury in adult patients are invalid in elderly patients. Thus, it is a challenge to protect the aged heart in the context of underlying chronic or acute cardiac diseases including ischemia-reperfusion injury. The cause(s) of this age-related increased damage remain unknown. Aging impairs the function of the mitochondrial electron transport chain (ETC), leading to decreased energy production and increased oxidative stress due to generation of reactive oxygen species (ROS). Additionally, ROS-induced oxidative stress can increase cardiac injury during ischemia-reperfusion by potentiating mitochondrial permeability transition pore (MPTP) opening. Aging leads to increased endoplasmic reticulum (ER) stress, which contributes to mitochondrial dysfunction, including reduced function of the ETC. The activation of both cytosolic and mitochondrial calcium-activated proteases termed calpains leads to mitochondrial dysfunction and decreased ETC function. Intriguingly, mitochondrial ROS generation also induces ER stress, highlighting the dynamic interaction between mitochondria and ER. Here, we discuss the role of ER stress in sensitizing and potentiating mitochondrial dysfunction in response to ischemia-reperfusion, and the promising potential therapeutic benefit of inhibition of ER stress and / or calpains to attenuate cardiac injury in elderly patients.
Asunto(s)
Envejecimiento/metabolismo , Calpaína/metabolismo , Estrés del Retículo Endoplásmico/fisiología , Mitocondrias Cardíacas/fisiología , Daño por Reperfusión , Calpaína/antagonistas & inhibidores , Descubrimiento de Drogas , Estrés del Retículo Endoplásmico/efectos de los fármacos , Humanos , Miocardio/metabolismo , Daño por Reperfusión/metabolismo , Daño por Reperfusión/prevención & controlRESUMEN
Niemann-Pick type C (NPC) disease is a lysosomal storage disorder arising from mutations in the cholesterol-trafficking protein NPC1 (95%) or NPC2 (5%). These mutations result in accumulation of low-density lipoprotein-derived cholesterol in late endosomes/lysosomes, disruption of endocytic trafficking, and stalled autophagic flux. Additionally, NPC disease results in sphingolipid accumulation, yet it is unique among the sphingolipidoses because of the absence of mutations in the enzymes responsible for sphingolipid degradation. In this work, we examined the cause for sphingosine and sphingolipid accumulation in multiple cellular models of NPC disease and observed that the activity of sphingosine kinase 1 (SphK1), one of the two isoenzymes that phosphorylate sphingoid bases, was markedly reduced in both NPC1 mutant and NPC1 knockout cells. Conversely, SphK1 inhibition with the isotype-specific inhibitor SK1-I in WT cells induced accumulation of cholesterol and reduced cholesterol esterification. Of note, a novel SphK1 activator (SK1-A) that we have characterized decreased sphingoid base and complex sphingolipid accumulation and ameliorated autophagic defects in both NPC1 mutant and NPC1 knockout cells. Remarkably, in these cells, SK1-A also reduced cholesterol accumulation and increased cholesterol ester formation. Our results indicate that a SphK1 activator rescues aberrant cholesterol and sphingolipid storage and trafficking in NPC1 mutant cells. These observations highlight a previously unknown link between SphK1 activity, NPC1, and cholesterol trafficking and metabolism.
Asunto(s)
Enfermedad de Niemann-Pick Tipo C/metabolismo , Fosfotransferasas (Aceptor de Grupo Alcohol)/metabolismo , Esfingosina/metabolismo , Animales , Proteínas Portadoras/metabolismo , Línea Celular , Colesterol/metabolismo , Ésteres del Colesterol/metabolismo , Endosomas/metabolismo , Fibroblastos , Humanos , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Lisosomas/metabolismo , Glicoproteínas de Membrana/metabolismo , Ratones , Proteína Niemann-Pick C1/genética , Proteína Niemann-Pick C1/metabolismo , Enfermedad de Niemann-Pick Tipo C/fisiopatología , Cultivo Primario de Células , Transporte de Proteínas , Esfingolípidos/metabolismo , Esfingosina/genética , Proteínas de Transporte Vesicular/genética , Proteínas de Transporte Vesicular/metabolismoRESUMEN
Hepatocellular carcinoma (HCC) is primarily diagnosed in the latter stages of disease progression and is the third leading cause of cancer deaths worldwide. Thus, there is a need to find biomarkers of early HCC as well as the development of more effective treatments for the disease. Sphingosine-1-phosphate (S1P) is a pleiotropic lipid signaling molecule produced by two isoforms of sphingosine kinase (SphK1 and SphK2) that is involved in regulation of many aspects of mammalian physiology and pathophysiology, including inflammation, epithelial and endothelial barrier function, cancer, and metastasis, among many others. Abundant evidence indicates that SphK1 and S1P promote cancer progression and metastasis in multiple types of cancers. However, the role of SphK/S1P in HCC is less well studied. Here, we review the current state of knowledge of SphKs and S1P in HCC, including evidence for the correlation of SphK1 expression and S1P levels with progression of HCC and negative outcomes, and discuss how this information could lead to the design of more effective diagnostic and treatment modalities for HCC.
Asunto(s)
Carcinoma Hepatocelular/enzimología , Neoplasias Hepáticas/enzimología , Lisofosfolípidos/análisis , Fosfotransferasas (Aceptor de Grupo Alcohol)/metabolismo , Esfingosina/análogos & derivados , Animales , Carcinoma Hepatocelular/patología , Neoplasias Hepáticas/patología , Esfingosina/análisisRESUMEN
How plasma membrane (PM) cholesterol is controlled is poorly understood. Ablation of the gene encoding the ER stress steroidogenic acute regulatory-related lipid transfer domain (StarD)5 leads to a decrease in PM cholesterol content, a decrease in cholesterol efflux, and an increase in intracellular neutral lipid accumulation in macrophages, the major cell type that expresses StarD5. ER stress increases StarD5 expression in mouse hepatocytes, which results in an increase in accessible PM cholesterol in WT but not in StarD5-/- hepatocytes. StarD5-/- mice store higher levels of cholesterol and triglycerides, which leads to altered expression of cholesterol-regulated genes. In vitro, a recombinant GST-StarD5 protein transfers cholesterol between synthetic liposomes. StarD5 overexpression leads to a marked increase in PM cholesterol. Phasor analysis of 6-dodecanoyl-2-dimethylaminonaphthalene fluorescence lifetime imaging microscopy data revealed an increase in PM fluidity in StarD5-/- macrophages. Taken together, these studies show that StarD5 is a stress-responsive protein that regulates PM cholesterol and intracellular cholesterol homeostasis.
Asunto(s)
Proteínas Adaptadoras del Transporte Vesicular/metabolismo , Membrana Celular/metabolismo , Macrófagos Peritoneales/metabolismo , Proteínas Adaptadoras del Transporte Vesicular/genética , Animales , Células CHO , Células Cultivadas , Colesterol/metabolismo , Cricetulus , Retículo Endoplásmico/metabolismo , Femenino , Homeostasis/genética , Homeostasis/fisiología , Immunoblotting , Metabolismo de los Lípidos/genética , Metabolismo de los Lípidos/fisiología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Microscopía Fluorescente , ARN Mensajero , Triglicéridos/metabolismoRESUMEN
Transient, reversible blockade of complex I during early reperfusion after ischemia limits cardiac injury. We studied the cardioprotection of high dose of metformin in cultured cells and mouse hearts via the novel mechanism of acute downregulation of complex I. The effect of high dose of metformin on complex I activity was studied in isolated heart mitochondria and cultured H9c2 cells. Protection with metformin was evaluated in H9c2 cells at reoxygenation and at early reperfusion in isolated perfused mouse hearts and in vivo regional ischemia reperfusion. Acute, high-dose metformin treatment inhibited complex I in ischemia-damaged mitochondria and in H9c2 cells following hypoxia. Accompanying the complex I modulation, high-dose metformin at reoxygenation decreased death in H9c2 cells. Acute treatment with high-dose metformin at the end of ischemia reduced infarct size following ischemia reperfusion in vitro and in vivo, including in the AMP kinase-dead mouse. Metformin treatment during early reperfusion improved mitochondrial calcium retention capacity, indicating decreased permeability transition pore (MPTP) opening. Acute, high-dose metformin therapy decreased cardiac injury through inhibition of complex I accompanied by attenuation of MPTP opening. Moreover, in contrast to chronic metformin treatment, protection by acute, high-dose metformin is independent of AMP-activated protein kinase activation. Thus, a single, high-dose metformin treatment at reperfusion reduces cardiac injury via modulation of complex I.
Asunto(s)
Complejo I de Transporte de Electrón/antagonistas & inhibidores , Inhibidores Enzimáticos/farmacología , Metformina/farmacología , Daño por Reperfusión Miocárdica/tratamiento farmacológico , Proteínas Quinasas Activadas por AMP/metabolismo , Animales , Línea Celular , Citoprotección/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Activación Enzimática/efectos de los fármacos , Masculino , Metformina/uso terapéutico , Ratones , Ratones Endogámicos C57BL , Proteínas de Transporte de Membrana Mitocondrial/química , Proteínas de Transporte de Membrana Mitocondrial/metabolismo , Poro de Transición de la Permeabilidad Mitocondrial , Daño por Reperfusión Miocárdica/enzimología , Daño por Reperfusión Miocárdica/metabolismo , Daño por Reperfusión Miocárdica/patología , Conformación Proteica/efectos de los fármacosRESUMEN
Sphingosine-1-phosphate (S1P) is a potent bioactive signaling molecule that regulates many physiological processes important for development, epithelial and endothelial barrier integrity, and the immune system, as well as for pathologies, such as autoimmune diseases, cancer, and metastasis. Most of the well-known actions of S1P are mediated by five specific G protein-coupled receptors located on the plasma membrane. Because S1P is synthesized intracellularly by two sphingosine kinase isoenzymes, we have proposed the paradigm of inside-out signaling by S1P, suggesting that S1P must be exported out of cells to interact with its receptors. While several transporters of S1P have previously been identified, spinster homologue 2 (SPNS2), a member of the large family of non-ATP-dependent organic ion transporters, has recently attracted much attention as an S1P transporter. Here, we discuss recent advances in understanding the physiological actions of SPNS2 in regulating levels of S1P and the S1P gradient that exists between the high circulating concentrations of S1P and low tissue levels that control lymphocyte trafficking. Special emphasis is on the functions of SPNS2 in inflammatory and autoimmune diseases and its recently discovered unexpected importance in metastasis.
Asunto(s)
Proteínas de Transporte de Anión/metabolismo , Animales , Proteínas de Transporte de Anión/química , Homeostasis , Humanos , Inmunidad , Inflamación/metabolismo , Lisofosfolípidos/metabolismo , Transducción de Señal , Esfingosina/análogos & derivados , Esfingosina/metabolismoRESUMEN
In breast cancer, 17ß-estradiol (E2) plays critical roles mainly by binding to its canonical receptor, estrogen receptor (ER) α66, and eliciting genomic effects. E2 also triggers rapid, nongenomic responses. E2 activates sphingosine kinase 1 (SphK1), increasing sphingosine-1-phosphate (S1P) that binds to its receptors, leading to important breast cancer signaling. However, the E2 receptor responsible for SphK1 activation has not yet been identified. Here, we demonstrate in triple-negative breast cancer cells, which lack the canonical ERα66 but express the novel splice variant ERα36, that ERα36 is the receptor responsible for E2-induced activation of SphK1 and formation and secretion of S1P and dihydro-S1P, the ligands for S1PRs. Tamoxifen, the first-line endocrine therapy for breast cancer, is an antagonist of ERα66, but an agonist of ERα36, and, like E2, activates SphK1 and markedly increases secretion of S1P. A major problem with tamoxifen therapy is development of acquired resistance. We found that tamoxifen resistance correlated with increased SphK1 and ERα36 expression in tamoxifen-resistant breast cancer cells, in patient-derived xenografts, and in endocrine-resistant breast cancer patients. Our data also indicate that targeting this ERα36 and SphK1 axis may be a therapeutic option to circumvent endocrine resistance and improve patient outcome.
Asunto(s)
Neoplasias de la Mama/metabolismo , Fosfotransferasas (Aceptor de Grupo Alcohol)/metabolismo , Receptores de Estrógenos/metabolismo , Tamoxifeno/farmacología , Animales , Western Blotting , Neoplasias de la Mama/genética , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Resistencia a Antineoplásicos/genética , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Regulación Neoplásica de la Expresión Génica/genética , Humanos , Ratones , Microscopía Confocal , Fosfotransferasas (Aceptor de Grupo Alcohol)/genética , Receptores de Estrógenos/genética , Transducción de Señal/efectos de los fármacos , Transducción de Señal/genética , Espectrometría de Masas en TándemRESUMEN
Over 20 years ago, sphingosine-1-phosphate (S1P) was discovered to be a bioactive signaling molecule. Subsequent studies later identified two related kinases, sphingosine kinase 1 and 2, which are responsible for the phosphorylation of sphingosine to S1P. Many stimuli increase sphingosine kinase activity and S1P production and secretion. Outside the cell, S1P can bind to and activate five S1P-specific G protein-coupled receptors (S1PR1-5) to regulate many important cellular and physiological processes in an autocrine or paracrine manner. S1P is found in high concentrations in the blood where it functions to control vascular integrity and trafficking of lymphocytes. Obesity increases blood S1P levels in humans and mice. With the world wide increase in obesity linked to consumption of high-fat, high-sugar diets, S1P is emerging as an accomplice in liver pathobiology, including acute liver failure, metabolic syndrome, control of blood lipid and glucose homeostasis, nonalcoholic fatty liver disease, and liver fibrosis. Here, we review recent research on the importance of sphingosine kinases, S1P, and S1PRs in liver pathobiology, with a focus on exciting insights for new therapeutic modalities that target S1P signaling axes for a variety of liver diseases.
Asunto(s)
Hepatopatías/metabolismo , Hígado/metabolismo , Lisofosfolípidos/metabolismo , Fosfotransferasas (Aceptor de Grupo Alcohol)/metabolismo , Esfingosina/análogos & derivados , Animales , Hígado Graso/enzimología , Hígado Graso/patología , Humanos , Hígado/enzimología , Hígado/patología , Hepatopatías/enzimología , Fallo Hepático/enzimología , Fallo Hepático/patología , Síndrome Metabólico/enzimología , Síndrome Metabólico/patología , Esfingosina/metabolismoRESUMEN
Niemann-Pick type C (NPC) disease is a fatal neurodegenerative disorder caused by mutations in NPC1 or NPC2 with decreased functions leading to lysosomal accumulation of cholesterol and sphingolipids. FTY720/fingolimod, used for treatment of multiple sclerosis, is phosphorylated by nuclear sphingosine kinase 2, and its active phosphorylated form (FTY720-P) is an inhibitor of class I histone deacetylases. In this study, administration of clinically relevant doses of FTY720 to mice increased expression of NPC1 and -2 in brain and liver and decreased cholesterol in an SphK2-dependent manner. FTY720 greatly increased expression of NPC1 and -2 in human NPC1 mutant fibroblasts that correlated with formation of FTY720-P and significantly reduced the accumulation of cholesterol and glycosphingolipids. In agreement with this finding, FTY720 pretreatment of human NPC1 mutant fibroblasts restored transport of the cholera toxin B subunit, which binds ganglioside GM1, to the Golgi apparatus. Together, these findings suggest that FTY720 administration can ameliorate cholesterol and sphingolipid storage and trafficking defects in NPC1 mutant fibroblasts. Because neurodegeneration is the main clinical feature of NPC disease, and FTY720 accumulates in the CNS and has several advantages over available histone deacetylase inhibitors now in clinical trials, our work provides a potential opportunity for treatment of this incurable disease.-Newton, J., Hait, N. C., Maceyka, M., Colaco, A., Maczis, M., Wassif, C. A., Cougnoux, A., Porter, F. D., Milstien, S., Platt, N., Platt, F. M., Spiegel, S. FTY720/fingolimod increases NPC1 and NPC2 expression and reduces cholesterol and sphingolipid accumulation in Niemann-Pick type C mutant fibroblasts.
Asunto(s)
Colesterol/metabolismo , Clorhidrato de Fingolimod/farmacología , Inhibidores de Histona Desacetilasas/farmacología , Enfermedad de Niemann-Pick Tipo C/metabolismo , Proteínas/metabolismo , Esfingolípidos/metabolismo , Proteínas de Transporte Vesicular/metabolismo , Células 3T3 , Animales , Células Cultivadas , Fibroblastos/efectos de los fármacos , Fibroblastos/metabolismo , Humanos , Péptidos y Proteínas de Señalización Intracelular , Masculino , Ratones , Ratones Endogámicos C57BL , Proteína Niemann-Pick C1 , Enfermedad de Niemann-Pick Tipo C/genética , Transporte de Proteínas , Proteínas/genética , Proteínas de Transporte Vesicular/genéticaRESUMEN
The bioactive sphingolipid metabolite, ceramide, regulates physiological processes important for inflammation and elevated levels of ceramide have been implicated in IL-1-mediated events. Although much has been learned about ceramide generation by activation of sphingomyelinases in response to IL-1, the contribution of the de novo pathway is not completely understood. Because yeast ORM1 and ORM2 proteins negatively regulate ceramide levels through inhibition of serine palmitoyltransferase, the first committed step in ceramide biosynthesis, we examined the functions of individual mammalian ORM orthologs, ORM (yeast)-like (ORMDL)1-3, in regulation of ceramide levels. In HepG2 liver cells, downregulation of ORMDL3 markedly increased the ceramide precursors, dihydrosphingosine and dihydroceramide, primarily from de novo biosynthesis based on [U-(13)C]palmitate incorporation into base-labeled and dual-labeled dihydroceramides, whereas downregulation of each isoform increased dihydroceramides [(13)C]labeled in only the amide-linked fatty acid. IL-1 and the IL-6 family cytokine, oncostatin M, increased dihydroceramide and ceramide levels in HepG2 cells and concomitantly decreased ORMDL proteins. Moreover, during irritant-induced sterile inflammation in mice leading to induction of the acute-phase response, which is dependent on IL-1, expression of ORMDL proteins in the liver was strongly downregulated and accompanied by increased ceramide levels in the liver and accumulation in the blood. Together, our results suggest that ORMDLs may be involved in regulation of ceramides during IL-1-mediated sterile inflammation.
Asunto(s)
Ceramidas/metabolismo , Inflamación/metabolismo , Proteínas de la Membrana/fisiología , Animales , Citocinas/metabolismo , Células Hep G2 , Humanos , Hígado/metabolismo , Ratones Endogámicos C57BLRESUMEN
The bioactive sphingolipid sphingosine-1-phosphate (S1P) and the kinase that produces it have been implicated in inflammatory bowel diseases in mice and humans; however, little is known about the role of the 2 S1P-specific phosphohydrolase isoforms, SGPP1 and SGPP2, which catalyze dephosphorylation of S1P to sphingosine. To elucidate their functions, we generated specific knockout mice. Deletion of Sgpp2, which is mainly expressed in the gastrointestinal tract, significantly reduced dextran sodium sulfate (DSS)-induced colitis severity, whereas deletion of ubiquitously expressed Sgpp1 slightly worsened colitis. Moreover, Sgpp1 deletion enhanced expression of multifunctional proinflammatory cytokines, IL-6, TNF-α, and IL-1ß, activation of the transcription factor signal transducer and activator of transcription 3, and immune cell infiltration into the colon. Conversely, Sgpp2-null mice failed to mount a DSS-induced systemic inflammatory response. Of interest, Sgpp2 deficiency suppressed DSS-induced intestinal epithelial cell apoptosis and improved mucosal barrier integrity. Furthermore, down-regulation of Sgpp2 attenuated LPS-induced paracellular permeability in cultured cells and enhanced expression of the adherens junction protein E-cadherin. Finally, in patients with ulcerative colitis, SGPP2 expression was elevated in colitis tissues relative to that in uninvolved tissues. These results indicate that induction of SGPP2 expression contributes to the pathogenesis of colitis by promoting disruption of the mucosal barrier function. SGPP2 may represent a novel therapeutic target in inflammatory bowel disease.-Huang, W.-C., Liang, J., Nagahashi, M., Avni, D., Yamada, A., Maceyka, M., Wolen, A. R., Kordula, T., Milstien, S., Takabe, K., Oravecz, T., Spiegel, S. Sphingosine-1-phosphate phosphatase 2 promotes disruption of mucosal integrity, and contributes to ulcerative colitis in mice and humans.
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Colitis Ulcerosa/metabolismo , Mucosa Intestinal/patología , Proteínas de la Membrana/metabolismo , Monoéster Fosfórico Hidrolasas/metabolismo , Animales , Cadherinas , Colitis Ulcerosa/inducido químicamente , Colitis Ulcerosa/genética , Sulfato de Dextran/toxicidad , Regulación hacia Abajo , Humanos , Inflamación/metabolismo , Mucosa Intestinal/enzimología , Lipopolisacáridos/toxicidad , Proteínas de la Membrana/genética , Ratones , Ratones Noqueados , Permeabilidad , Monoéster Fosfórico Hidrolasas/genéticaRESUMEN
Sphingosine-1-phosphate (S1P) is a bioactive lipid mediator that regulates many processes in inflammation and cancer. S1P is a ligand for five G-protein-coupled receptors, S1PR1 to -5, and also has important intracellular actions. Previously, we showed that intracellular S1P is involved in tumor necrosis factor alpha (TNF)-induced NF-κB activation in melanoma cell lines that express filamin A (FLNA). Here, we show that extracellular S1P activates NF-κB only in melanoma cells that lack FLNA. In these cells, S1P, but not TNF, promotes IκB kinase (IKK) and p65 phosphorylation, IκBα degradation, p65 nuclear translocation, and NF-κB reporter activity. NF-κB activation induced by S1P was mediated via S1PR1 and S1PR2. Exogenous S1P enhanced the phosphorylation of protein kinase Cδ (PKCδ), and its downregulation reduced S1P-induced the phosphorylation of IKK and p65. In addition, silencing of Bcl10 also inhibited S1P-induced IKK phosphorylation. Surprisingly, S1P reduced Akt activation in melanoma cells that express FLNA, whereas in the absence of FLNA, high phosphorylation levels of Akt were maintained, enabling S1P-mediated NF-κB signaling. In accord, inhibition of Akt suppressed S1P-mediated IKK and p65 phosphorylation and degradation of IκBα. Hence, these results support a negative role of FLNA in S1P-mediated NF-κB activation in melanoma cells through modulation of Akt.
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Filaminas/inmunología , Lisofosfolípidos/inmunología , Melanoma/inmunología , FN-kappa B/inmunología , Proteínas Proto-Oncogénicas c-akt/inmunología , Transducción de Señal , Esfingosina/análogos & derivados , Línea Celular Tumoral , Humanos , Proteína Quinasa C-delta/inmunología , Receptores de Lisoesfingolípidos/inmunología , Esfingosina/inmunología , Receptores de Esfingosina-1-FosfatoRESUMEN
Sphingosine 1-phosphate (S1P) is a pleiotropic bioactive sphingolipid metabolite that regulates numerous processes important for immune responses. S1P is made within cells and must be transported out of cells to exert its effects through activation of 5 specific cell surface GPCRs in an autocrine or paracrine fashion. Spinster 2 (Spns2) transports S1P out of cells, and its deletion in mice reduces circulating levels of S1P, alters immune cell trafficking, and induces lymphopenia. Here we examined the effects of Spns2 deletion on adaptive immune responses and in autoimmune disease models. Airway inflammation and hypersensitivity as well as delayed-type contact hypersensitivity were attenuated in Spns2(-/-) mice. Similarly, Spns2 deletion reduced dextran sodium sulfate- and oxazolone-induced colitis. Intriguingly, Spns2(-/-) mice were protected from the development of experimental autoimmune encephalopathy, a model of the autoimmune disease multiple sclerosis. Deletion of Spns2 also strongly alleviated disease development in collagen-induced arthritis. These results point to a broad role for Spns2-mediated S1P transport in the initiation and development of adaptive immune related disorders.
Asunto(s)
Proteínas de Transporte de Anión/fisiología , Enfermedades Autoinmunes/fisiopatología , Inflamación/fisiopatología , Animales , Proteínas de Transporte de Anión/genética , Modelos Animales de Enfermedad , Ratones , Ratones NoqueadosRESUMEN
BACKGROUND: Asthma, a chronic inflammatory condition defined by episodic shortness of breath with expiratory wheezing and cough, is a serious health concern affecting more than 250 million persons. Genome-wide association studies have identified ORM (yeast)-like protein isoform 3 (ORMDL3) as a gene associated with susceptibility to asthma. Although its yeast ortholog is a negative regulator of de novo ceramide biosynthesis, how ORMDL3 contributes to asthma pathogenesis is not known. OBJECTIVES: We sought to decipher the molecular mechanism for the pathologic functions of ORMDL3 in asthma and the relationship to its evolutionarily conserved role in regulation of ceramide homeostasis. METHODS: We determined the relationship between expression of ORMDL3 and ceramide in epithelial and inflammatory cells and in asthma pathogenesis in mice. RESULTS: Although small increases in ORMDL3 expression decrease ceramide levels, remarkably, higher expression in lung epithelial cells and macrophages in vitro and in vivo increased ceramide production, which promoted chronic inflammation, airway hyperresponsiveness, and mucus production during house dust mite-induced allergic asthma. Moreover, nasal administration of the immunosuppressant drug FTY720/fingolimod reduced ORMDL3 expression and ceramide levels and mitigated airway inflammation and hyperreactivity and mucus hypersecretion in house dust mite-challenged mice. CONCLUSIONS: Our findings demonstrate that overexpression of ORMDL3 regulates ceramide homeostasis in cells in a complex manner and suggest that local FTY720 administration might be a useful therapeutic intervention for the control of allergic asthma.
Asunto(s)
Asma/inmunología , Ceramidas/inmunología , Regulación de la Expresión Génica/inmunología , Homeostasis/inmunología , Proteínas de la Membrana/inmunología , Animales , Asma/tratamiento farmacológico , Asma/genética , Asma/patología , Línea Celular Tumoral , Ceramidas/genética , Células Epiteliales/inmunología , Células Epiteliales/patología , Femenino , Clorhidrato de Fingolimod/farmacología , Regulación de la Expresión Génica/efectos de los fármacos , Homeostasis/efectos de los fármacos , Homeostasis/genética , Humanos , Inmunosupresores/farmacología , Macrófagos/inmunología , Macrófagos/patología , Proteínas de la Membrana/genética , Ratones , Mucosa Respiratoria/inmunología , Mucosa Respiratoria/patologíaAsunto(s)
Lisofosfolípidos/fisiología , Neoplasias/metabolismo , Esfingosina/análogos & derivados , Animales , Antineoplásicos/farmacología , Antineoplásicos/uso terapéutico , Transformación Celular Neoplásica/metabolismo , Ceramidas/fisiología , Humanos , Terapia Molecular Dirigida , Neoplasias/tratamiento farmacológico , Fosfotransferasas (Aceptor de Grupo Alcohol)/antagonistas & inhibidores , Fosfotransferasas (Aceptor de Grupo Alcohol)/metabolismo , Receptores Lisofosfolípidos/metabolismo , Transducción de Señal , Esfingosina/fisiologíaRESUMEN
The tumor necrosis factor (TNF) receptor family member CD40 plays an essential role in the activation of antigen-presenting cells, B cell maturation, and immunoglobulin (Ig) class switching critical for adaptive immunity. Although the bioactive sphingolipid metabolite sphingosine-1-phosphate (S1P) and the kinase that produces it, sphingosine kinase 1 (SphK1), have long been implicated in the actions of TNF mediated by engagement of TNFR1, nothing is yet known of their role in CD40-mediated events. We have now found that ligation of CD40 activates and translocates SphK1 to the plasma membrane, leading to generation of S1P. SphK1 inhibition in human tonsil B cells, as well as inhibition or deletion of SphK1 in mouse splenic B cells, significantly reduced CD40-mediated Ig class switching and plasma cell differentiation ex vivo. Optimal activation of downstream CD40 signaling pathways, including NF-κB, p38, and JNK, also required SphK1. In mice treated with a SphK1 inhibitor or in SphK1(-/-) mice, isotype switching to antigen-specific IgE was decreased in vivo by 70 and 55%, respectively. Our results indicate that SphK1 is important for CD40-mediated B cell activation and regulation of humoral responses and suggest that targeting SphK1 might be a useful therapeutic approach to control antigen-specific IgE production.
Asunto(s)
Antígenos CD40/metabolismo , Cambio de Clase de Inmunoglobulina , Inmunoglobulina E/genética , Lisofosfolípidos/metabolismo , Fosfotransferasas (Aceptor de Grupo Alcohol)/metabolismo , Esfingosina/análogos & derivados , Animales , Linfocitos B/citología , Linfocitos B/metabolismo , Antígenos CD40/genética , Diferenciación Celular , Membrana Celular/metabolismo , Células HEK293 , Humanos , Inmunoglobulina E/metabolismo , MAP Quinasa Quinasa 4/genética , MAP Quinasa Quinasa 4/metabolismo , Ratones , Ratones Endogámicos C57BL , FN-kappa B/genética , FN-kappa B/metabolismo , Fosfotransferasas (Aceptor de Grupo Alcohol)/genética , Transporte de Proteínas , Esfingosina/metabolismo , Proteínas Quinasas p38 Activadas por Mitógenos/genética , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismoRESUMEN
Sphingolipids are ubiquitous building blocks of eukaryotic cell membranes. Progress in our understanding of sphingolipid metabolism, state-of-the-art sphingolipidomic approaches and animal models have generated a large body of evidence demonstrating that sphingolipid metabolites, particularly ceramide and sphingosine-1-phosphate, are signalling molecules that regulate a diverse range of cellular processes that are important in immunity, inflammation and inflammatory disorders. Recent insights into the molecular mechanisms of action of sphingolipid metabolites and new perspectives on their roles in regulating chronic inflammation have been reported. The knowledge gained in this emerging field will aid in the development of new therapeutic options for inflammatory disorders.