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1.
Adv Exp Med Biol ; 1223: 129-153, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32030688

RESUMO

Sphingosine-1-phosphate (S1P), together with other phosphosphingolipids, has been found to regulate complex cellular function in the tumor microenvironment (TME) where it acts as a signaling molecule that participates in cell-cell communication. S1P, through intracellular and extracellular signaling, was found to promote tumor growth, angiogenesis, chemoresistance, and metastasis; it also regulates anticancer immune response, modulates inflammation, and promotes angiogenesis. Interestingly, cancer cells are capable of releasing S1P and thus modifying the behavior of the TME components in a way that contributes to tumor growth and progression. Therefore, S1P is considered an important therapeutic target, and several anticancer therapies targeting S1P signaling are being developed and tested in clinics.


Assuntos
Lisofosfolipídeos/metabolismo , Neoplasias/metabolismo , Transdução de Sinais , Esfingosina/análogos & derivados , Microambiente Tumoral , Humanos , Neoplasias/patologia , Esfingosina/metabolismo
2.
Cell Physiol Biochem ; 53(6): 1015-1028, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31854953

RESUMO

BACKGROUND/AIMS: Pulmonary infections with Pseudomonas aeruginosa (P. aeruginosa) or Staphylococcus aureus (S. aureus) are of utmost clinical relevance in patients with cystic fibrosis, chronic obstructive pulmonary disease, after trauma and burn, upon ventilation or in immuno-compromised patients. Many P. aeruginosa and S. aureus strains are resistant to many known antibiotics and it is very difficult or often impossible to eradicate the pathogens in patient´s lungs. We have recently shown that the sphingoid base sphingosine very efficiently kills many pathogens, including for instance P. aeruginosa, S. aureus or Acinetobacter baumannii, in vitro. In vivo experiments of our group on cystic fibrosis mice indicated that inhalation of sphingosine prevents or eliminates existing acute or chronic pneumonia with P. aeruginosa or S. aureus in these mice. We also demonstrated that sphingosine is safe to use for inhalation up to high doses, at least in mice. To facilitate development of sphingosine to an anti-bactericidal drug that can be used in humans for inhalation, safety data on non-rodents, larger animals are absolutely required. METHODS: Here, we inhaled mini pigs with increasing doses of sphingosine for 10 days and analyzed the uptake of sphingosine into epithelial cells of bronchi as well as into the trachea and lung and the systemic circulation. Moreover, we measured the generation of ceramide and sphingosine 1-phosphate that potentially mediate inflammation, the influx of leukocytes, epithelial cell death and disruption of the epithelial cell barrier. RESULTS: We demonstrate that inhalation of sphingosine results in increased levels of sphingosine in the luminal membrane of bronchi and the trachea, but not in systemic accumulation. Inhaled sphingosine had no side effects up to very high doses. CONCLUSION: In summary, we demonstrate that inhalation of sphingosine results in an increase of sphingosine concentrations in the luminal plasma membrane of tracheal and bronchial epithelial cells. The inhalation has no systemic or local side effects.


Assuntos
Antibacterianos/metabolismo , Esfingosina/metabolismo , Administração por Inalação , Animais , Antibacterianos/farmacologia , Brônquios/metabolismo , Brônquios/patologia , Ceramidas/análise , Humanos , Pulmão/patologia , Lisofosfolipídeos/análise , Espectrometria de Massas , Pseudomonas aeruginosa/efeitos dos fármacos , Esfingosina/análogos & derivados , Esfingosina/análise , Esfingosina/farmacologia , Staphylococcus aureus/efeitos dos fármacos , Suínos , Porco Miniatura , Traqueia/metabolismo , Traqueia/patologia
3.
Int J Mol Sci ; 20(14)2019 Jul 10.
Artigo em Inglês | MEDLINE | ID: mdl-31295813

RESUMO

Wound healing starts with the recruitment of inflammatory cells that secrete wound-related factors. This step is followed by fibroblast activation and tissue construction. Sphingosine-1-phosphate (S1P) is a lipid mediator that promotes angiogenesis, cell proliferation, and attracts immune cells. We investigated the roles of S1P in skin wound healing by altering the expression of its biogenic enzyme, sphingosine kinase-1 (SphK1). The murine excisional wound splinting model was used. Sphingosine kinase-1 (SphK1) was highly expressed in murine wounds and that SphK1-/- mice exhibit delayed wound closure along with less angiogenesis and inflammatory cell recruitment. Nanoparticle-mediated topical SphK1 overexpression accelerated wound closure, which associated with increased angiogenesis, inflammatory cell recruitment, and various wound-related factors. The SphK1 overexpression also led to less scarring, and the interaction between transforming growth factor (TGF)-ß1 and S1P receptor-2 (S1PR2) signaling is likely to play a key role. In summary, SphK1 play important roles to strengthen immunity, and contributes early wound healing with suppressed scarring. S1P can be a novel therapeutic molecule with anti-scarring effect in surgical, trauma, and chronic wound management.


Assuntos
Cicatriz/metabolismo , Lisofosfolipídeos/metabolismo , Neovascularização Fisiológica , Pele/metabolismo , Esfingosina/análogos & derivados , Cicatrização , Animais , Biomarcadores , Proliferação de Células , Cicatriz/genética , Cicatriz/patologia , Modelos Animais de Doenças , Expressão Gênica , Granuloma/etiologia , Granuloma/metabolismo , Granuloma/patologia , Inflamação/etiologia , Inflamação/metabolismo , Inflamação/patologia , Camundongos , Camundongos Knockout , Neovascularização Fisiológica/genética , Fosfotransferases (Aceptor do Grupo Álcool)/genética , Fosfotransferases (Aceptor do Grupo Álcool)/metabolismo , Pele/lesões , Pele/patologia , Esfingosina/metabolismo , /metabolismo , Cicatrização/genética
4.
J Vet Med Sci ; 81(9): 1249-1258, 2019 Sep 03.
Artigo em Inglês | MEDLINE | ID: mdl-31341112

RESUMO

Inflammatory bowel disease (IBD) is a common gastrointestinal disease in dogs. Decreased production of intestinal immunoglobulin A (IgA) has been suggested as a possible pathogenesis in a subset of canine IBD; however, the underlying cause remains unclear. Sphingosine-1-phosphate (S1P) is a lipid mediator that regulates intestinal IgA production by controlling lymphocyte trafficking in mice. The objectives of this study were to clarify the role of S1P in IgA production in dogs and to evaluate the expression of S1P-related molecules in dogs with IBD. First, an S1P receptor antagonist was administrated to five healthy dogs. The S1P receptor antagonist significantly decreased the IgA concentration in sera and feces but did not affect the IgG concentration. Moreover, the immunoreactivity of intestinal IgA was significantly decreased by S1P signal blockade. These results indicate that S1P signaling specifically regulates the intestinal IgA production in dogs. Subsequently, the intestinal S1P concentration and the expression of S1P-related molecules were measured in dogs with IBD and healthy dogs. The intestinal concentration of S1P was significantly lower in dogs with IBD than in healthy dogs. In addition, the gene expression levels of S1P receptor (S1P1) and S1P synthase (SK1) were significantly lower in dogs with IBD than in healthy dogs. Taken together, these observations suggest that decreased S1P production, likely caused by a lower expression of S1P synthetase, leads to attenuation of S1P/S1P1 signaling pathway and the production of intestinal IgA in dogs with IBD.


Assuntos
Doenças do Cão/metabolismo , Imunoglobulina A/metabolismo , Doenças Inflamatórias Intestinais/veterinária , Lisofosfolipídeos/metabolismo , Esfingosina/análogos & derivados , Animais , Doenças do Cão/patologia , Cães , Fezes/química , Cloridrato de Fingolimode/administração & dosagem , Expressão Gênica , Imunoglobulina A/sangue , Imunoglobulina A/imunologia , Doenças Inflamatórias Intestinais/imunologia , Doenças Inflamatórias Intestinais/metabolismo , Intestinos/química , Masculino , Esfingosina/metabolismo , /genética
5.
mBio ; 10(3)2019 06 04.
Artigo em Inglês | MEDLINE | ID: mdl-31164462

RESUMO

Aspergillus fumigatus is a leading cause of invasive fungal infections. Resistance to first-line triazole antifungals has led to therapy with echinocandin drugs. Recently, we identified several high-minimum-effective-concentration (MEC) A. fumigatus clinical isolates from patients failing echinocandin therapy. Echinocandin resistance is known to arise from amino acid substitutions in ß-(1,3)-d-glucan synthase encoded by the fks1 gene. Yet these clinical isolates did not contain mutations in fks1, indicating an undefined resistance mechanism. To explore this new mechanism, we used a laboratory-derived strain, RG101, with a nearly identical caspofungin (CAS) susceptibility phenotype that also does not contain fks1 mutations. Glucan synthase isolated from RG101 was fully sensitive to echinocandins. Yet exposure of RG101 to CAS during growth yielded a modified enzyme that was drug insensitive (4 log orders) in kinetic inhibition assays, and this insensitivity was also observed for enzymes isolated from clinical isolates. To understand this alteration, we analyzed whole-enzyme posttranslational modifications (PTMs) but found none linked to resistance. However, analysis of the lipid microenvironment of the enzyme with resistance induced by CAS revealed a prominent increase in the abundances of dihydrosphingosine (DhSph) and phytosphingosine (PhSph). Exogenous addition of DhSph and PhSph to the sensitive enzyme recapitulated the drug insensitivity of the CAS-derived enzyme. Further analysis demonstrated that CAS induces mitochondrion-derived reactive oxygen species (ROS) and that dampening ROS formation by antimycin A or thiourea eliminated drug-induced resistance. We conclude that CAS induces cellular stress, promoting formation of ROS and triggering an alteration in the composition of plasma membrane lipids surrounding glucan synthase, rendering it insensitive to echinocandins.IMPORTANCE Resistance to first-line triazole antifungal agents among Aspergillus species has prompted the use of second-line therapy with echinocandins. As the number of Aspergillus-infected patients treated with echinocandins is rising, clinical observations of drug resistance are also increasing, indicating an emerging global health threat. Our knowledge regarding the development of clinical echinocandin resistance is largely derived from Candida spp., while little is known about resistance in Aspergillus. Therefore, it is important to understand the specific cellular responses raised by A. fumigatus against echinocandins. We discovered a new mechanism of resistance in A. fumigatus that is independent of the well-characterized FKS mutation mechanism observed in Candida This study identified an off-target effect of CAS, i.e., ROS production, and integrated oxidative stress and sphingolipid alterations into a novel mechanism of resistance. This stress-induced response has implications for drug resistance and/or tolerance mechanisms in other fungal pathogens.


Assuntos
Antifúngicos/farmacologia , Aspergillus fumigatus/efeitos dos fármacos , Farmacorresistência Fúngica/genética , Equinocandinas/farmacologia , Glucosiltransferases/genética , Estresse Fisiológico , Aspergilose/microbiologia , Aspergillus fumigatus/enzimologia , Proteínas Fúngicas/genética , Humanos , Testes de Sensibilidade Microbiana , Estresse Oxidativo , Processamento de Proteína Pós-Traducional , Espécies Reativas de Oxigênio/metabolismo , Esfingosina/análogos & derivados , Esfingosina/metabolismo
6.
Nat Chem Biol ; 15(6): 623-631, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-31036923

RESUMO

Sphingosine-1-phosphate (S1P) plays important roles as a signaling lipid in a variety of physiological and pathophysiological processes. S1P signals via a family of G-protein-coupled receptors (GPCRs) (S1P1-5) and intracellular targets. Here, we report on photoswitchable analogs of S1P and its precursor sphingosine, respectively termed PhotoS1P and PhotoSph. PhotoS1P enables optical control of S1P1-3, shown through electrophysiology and Ca2+ mobilization assays. We evaluated PhotoS1P in vivo, where it reversibly controlled S1P3-dependent pain hypersensitivity in mice. The hypersensitivity induced by PhotoS1P is comparable to that induced by S1P. PhotoS1P is uniquely suited for the study of S1P biology in cultured cells and in vivo because it exhibits prolonged metabolic stability compared to the rapidly metabolized S1P. Using lipid mass spectrometry analysis, we constructed a metabolic map of PhotoS1P and PhotoSph. The formation of these photoswitchable lipids was found to be light dependent, providing a novel approach to optically probe sphingolipid biology.


Assuntos
Lisofosfolipídeos/metabolismo , Esfingosina/análogos & derivados , Animais , Lisofosfolipídeos/química , Camundongos , Modelos Moleculares , Estrutura Molecular , Imagem Óptica , Processos Fotoquímicos , Esfingosina/química , Esfingosina/metabolismo
7.
Int J Mol Sci ; 20(7)2019 Apr 02.
Artigo em Inglês | MEDLINE | ID: mdl-30987025

RESUMO

BACKGROUND: S1P has been shown to improve the endothelialization of decellularized vascular grafts in vitro. Here, we evaluated the potential of tissue-engineered vascular grafts (TEVGs) constructed by ECs and S1P on decellularized vascular scaffolds in a rat model. METHODS: Rat aorta was decellularized mainly by 0.1% SDS and characterized by histology. Rat ECs, were seeded onto decellularized scaffolds, and the viability of the ECs was evaluated by biochemical assays. Then, we investigated the in vivo patency rate and endothelialization for five groups of decellularized vascular grafts (each n = 6) in a rat abdominal aorta model for 14 days. The five groups included (1) rat allogenic aorta (RAA); (2) decellularized RAA (DRAA); (3) DRAA with S1P (DRAA/S1P); (4) DRAA with EC recellularization (DRAA/EC); and (5) DRAA with S1P and EC recellularization (DRAA/EC/S1P). RESULTS: In vitro, ECs were identified by the uptake of Dil-Ac-LDL. S1P enhanced the expression of syndecan-1 on ECs and supported the proliferation of ECs on decellularized vascular grafts. In vivo, RAA and DRAA/EC/S1P both had 100% patency without thrombus formation within 14 days. Better endothelialization, more wall structure maintenance and less inflammation were noted in the DRAA/EC/S1P group. In contrast, there was thrombus formation in the DRAA, DRAA/S1P and DRAA/EC groups. CONCLUSION: S1P could inhibit thrombus formation to improve the patency rate of EC-covered decellularized vascular grafts in vivo and may play an important role in the construction of TEVGs.


Assuntos
Prótese Vascular , Células Endoteliais/metabolismo , Lisofosfolipídeos/metabolismo , Esfingosina/análogos & derivados , Grau de Desobstrução Vascular , Animais , Aorta/transplante , Implante de Prótese Vascular , Proliferação de Células , Forma Celular , Feminino , Macrófagos/metabolismo , Ratos Sprague-Dawley , Esfingosina/metabolismo , Análise de Sobrevida , Sindecana-1/metabolismo
8.
PLoS One ; 14(4): e0215770, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31002723

RESUMO

The purpose of this study was to determine the profile of bioactive sphingolipids in xenograft mouse tissues of head and neck squamous cell carcinoma. We utilized UHPLC-MS/MS to determine the profile of full set of ceramides, sphingosine, and sphingosine 1-phosphate in this xenograft mouse model. The tissues isolated and investigated were from brain, lung, heart, liver, spleen, kidney, bladder, tumors and blood. With the exception of equal volume of blood plasma (100ul), all tissues were studied with the same amount of protein (800ug). Results demonstrated that brain contained the highest level of ceramide and kidney had the highest level of sphingosine, whereas sphingosine 1-phosphate and dihydrosphingosine 1-phosphate were heavily presented in the blood. Brain also comprised the highest level of phospholipids. As for the species, several ceramides, usually present in very low amounts in cultured tumor cells, showed relatively high levels in certain tissues. This study highlights levels of bioactive sphingolipids profiles in xenograft mouse model of head and neck squamous cell carcinoma, and provides resources to investigate potential therapeutic targets and biomarkers that target bioactive sphingolipids metabolism pathways.


Assuntos
Carcinoma de Células Escamosas/metabolismo , Neoplasias de Cabeça e Pescoço/metabolismo , Esfingolipídeos/metabolismo , Animais , Carcinoma de Células Escamosas/patologia , Linhagem Celular Tumoral , Ceramidas/metabolismo , Cromatografia Líquida de Alta Pressão/métodos , Modelos Animais de Doenças , Neoplasias de Cabeça e Pescoço/patologia , Humanos , Lisofosfolipídeos/metabolismo , Camundongos Nus , Esfingosina/análogos & derivados , Esfingosina/metabolismo , Espectrometria de Massas em Tandem/métodos , Transplante Heterólogo
9.
Mol Neurobiol ; 56(10): 7188-7207, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-30997640

RESUMO

Sphingosine-1-phosphate (S1P) produced by sphingosine kinases (SPHK1 and SPHK2) is a signaling molecule involved in cell proliferation and formation of cellular junctions. In this study, we characterized the retinas of Sphk1 knockout (KO) mice by electron microscopy and immunocytochemistry. We also tested cultured Müller glia for their response to S1P. We found that S1P plays an important role in retinal and retinal pigment epithelial (RPE) structural integrity in aging mice. Ultrastructural analysis of Sphk1 KO mouse retinas aged to 15 months or raised with moderate light stress revealed a degenerated outer limiting membrane (OLM). This membrane is formed by adherens junctions between neighboring Müller glia and photoreceptor cells. We also show that Sphk1 KO mice have reduced retinal function in mice raised with moderate light stress. In vitro assays revealed that exogenous S1P modulated cytoskeletal rearrangement and increased N-cadherin production in human Müller glia cells. Aged mice also had morphological degeneration of the RPE, as well as increased lipid storage vacuoles and undigested phagosomes reminiscent of RPE in age-related macular degeneration. These findings show that SPHK1 and S1P play a vital role in the structural maintenance of the mammalian retina and retinal pigmented epithelium by supporting the formation of adherens junctions.


Assuntos
Junções Aderentes/metabolismo , Envelhecimento/metabolismo , Membrana Celular/metabolismo , Fosfotransferases (Aceptor do Grupo Álcool)/metabolismo , Retina/metabolismo , Junções Aderentes/ultraestrutura , Animais , Caderinas/metabolismo , Endotélio/metabolismo , Células Ependimogliais/metabolismo , Humanos , Lisofosfolipídeos/metabolismo , Camundongos Knockout , Fenótipo , Retina/ultraestrutura , Epitélio Pigmentado da Retina/metabolismo , Epitélio Pigmentado da Retina/ultraestrutura , Esfingolipídeos/metabolismo , Esfingosina/análogos & derivados , Esfingosina/metabolismo , Vacúolos/metabolismo , Vacúolos/ultraestrutura , Proteína cdc42 de Ligação ao GTP/metabolismo , Proteínas rac1 de Ligação ao GTP/metabolismo
10.
Immunol Rev ; 289(1): 173-185, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-30977198

RESUMO

The signaling lipid sphingosine 1-phosphate (S1P) plays key roles in many physiological processes. In the immune system, S1P's best-described function is to draw cells out of tissues into circulation. Here, we will review models of S1P distribution in the thymus, lymph nodes, spleen, and nonlymphoid tissues. These models have been challenging to construct, because of the lack of tools to map lipid gradients. Nonetheless, evidence to date suggests that S1P distribution is exquisitely tightly controlled, and that concentrations of signaling-available S1P cannot be predicted by standard rules of thumb. The fine regulation of S1P gradients may explain how S1P can simultaneously direct multiple cell movements both between tissues and circulation and within tissues. It may also make it feasible to develop drugs that enable spatially specific modulation of S1P signaling.


Assuntos
Imunidade Celular , Liases/metabolismo , Lisofosfolipídeos/metabolismo , Esfingosina/análogos & derivados , Animais , Circulação Sanguínea , Movimento Celular , Humanos , Imunomodulação , Lipídeos/imunologia , Transdução de Sinais , Esfingosina/metabolismo
11.
Prog Lipid Res ; 74: 145-159, 2019 04.
Artigo em Inglês | MEDLINE | ID: mdl-30951736

RESUMO

The increased adipose tissue mass of obese individuals enhances the risk of metabolic syndrome, type 2 diabetes and cardiovascular diseases. During pathological expansion of adipose tissue, multiple molecular controls of lipid storage, adipocyte turn-over and endocrine secretion are perturbed and abnormal lipid metabolism results in a distinct lipid profile. There is a role for ceramides and sphingosine 1-phosphate (S1P) in inducing adipose dysfunction. For instance, the alteration of ceramide biosynthesis, through the de-regulation of key enzymes, results in aberrant formation of ceramides (e.g. C16:0 and C18:0) which block insulin signaling and promote adipose inflammation. Furthermore, S1P can induce defective adipose tissue phenotypes by promoting chronic inflammation and inhibiting adipogenesis. These abnormal changes are discussed in the context of possible therapeutic approaches to re-establish normal adipose function and to, thereby, increase insulin sensitivity in type 2 diabetes. Such novel approaches include blockade of ceramide biosynthesis using inhibitors of sphingomyelinase or dihydroceramide desaturase and by antagonism of S1P receptors, such as S1P2.


Assuntos
Tecido Adiposo/metabolismo , Tecido Adiposo/fisiopatologia , Ceramidas/metabolismo , Lisofosfolipídeos/metabolismo , Esfingosina/análogos & derivados , Animais , Humanos , Esfingosina/metabolismo
12.
J Gen Virol ; 100(4): 629-641, 2019 04.
Artigo em Inglês | MEDLINE | ID: mdl-30869582

RESUMO

There is growing evidence of the influence of sphingosine kinase (SK) enzymes on viral infection. Here, the role of sphingosine kinase 2 (SK2), an isoform of SK prominent in the brain, was defined during dengue virus (DENV) infection. Chemical inhibition of SK2 activity using two different SK2 inhibitors, ABC294640 and K145, had no effect on DENV infection in human cells in vitro. In contrast, DENV infection was restricted in SK2-/- immortalized mouse embryonic fibroblasts (iMEFs) with reduced induction of IFN-ß mRNA and protein, and mRNA for the IFN-stimulated genes (ISGs) viperin, IFIT1, IRF7 and CXCL10 in DENV-infected SK2-/- compared to WT iMEFs. Intracranial (ic) DENV injection in C57BL/6 SK2-/- mice induced body weight loss earlier than in WT mice but DENV RNA levels were comparable in the brain. Neither SK1 mRNA or sphingosine-1-phosphate (S1P) levels were altered following ic DENV infection in WT or SK2-/- mice but brain S1P levels were reduced in all SK2-/- mice, independent of DENV infection. CD8 mRNA was induced in the brains of both DENV-infected WT and SK2-/- mice, suggesting normal CD8+ T-cell infiltration into the DENV-infected brain independent of SK2 or S1P. Thus, although SK2 may be important for replication of some viruses SK2 activity does not affect DENV infection in vitro and SK2 or S1P levels do not influence DENV infection or T-cell infiltration in the context of infection in the brain.


Assuntos
Vírus da Dengue/patogenicidade , Dengue/metabolismo , Fosfotransferases (Aceptor do Grupo Álcool)/metabolismo , Adamantano/análogos & derivados , Adamantano/farmacologia , Animais , Linfócitos T CD8-Positivos/efeitos dos fármacos , Linfócitos T CD8-Positivos/metabolismo , Linfócitos T CD8-Positivos/virologia , Linhagem Celular , Linhagem Celular Tumoral , Dengue/tratamento farmacológico , Vírus da Dengue/efeitos dos fármacos , Células HEK293 , Células Hep G2 , Humanos , Interferon beta/metabolismo , Lisofosfolipídeos/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Piridinas/farmacologia , RNA Mensageiro/metabolismo , Esfingosina/análogos & derivados , Esfingosina/metabolismo , Tiazolidinedionas/farmacologia
13.
High Alt Med Biol ; 20(1): 78-88, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-30892968

RESUMO

AIM: High altitude exposure alters biochemical, metabolic, and physiological features of heart and skeletal muscles, and hence has pathological consequences in these tissues. Central to these hypoxia-associated biochemical/metabolic shuffling are energy deficit accumulation of free radicals and ensuing oxidative damage in the tissue. Recent preclinical/clinical studies indicate sphingosine-1-phosphate (S1P) axis, comprising S1P G protein coupled receptors (S1PR1-5) and its synthesizing enzyme-sphingosine kinase (SphK) to have key regulatory roles in homeostatic cardiac and skeletal muscle biology. In view of this, the aim of the present study was to chart the initiation and progression of biochemical/metabolic shuffling and assess the coincident differential modulation of S1PR(1-5) expression and total SphK activity in cardiac and skeletal muscles from rats exposed to progressive hypobaric hypoxia (HH; 21,000 feet for 12, 24, and 48 hours). RESULTS: HH-associated responses were evident as raised damage markers in plasma, oxidative stress, decreased total tissue protein, imbalance of intermediate metabolites, and aerobic/anaerobic enzyme activities in cardiac and skeletal muscles (gastrocnemius and soleus) culminating as energy deficit. CONCLUSION: Cardiac and gastrocnemius muscles were more susceptible to hypoxic environment than soleus muscle. These differential responses were directly and indirectly coincident with temporal expression of S1PR(1-5) and SphK activity.


Assuntos
Hipóxia/metabolismo , Músculo Esquelético/metabolismo , Miocárdio/metabolismo , Fosfotransferases (Aceptor do Grupo Álcool)/metabolismo , Receptores de Lisoesfingolipídeo/metabolismo , Doença da Altitude/sangue , Doença da Altitude/metabolismo , Animais , Biomarcadores/sangue , Hipóxia/sangue , Lisofosfolipídeos/metabolismo , Oxirredução , Ratos , Ratos Sprague-Dawley , Esfingosina/análogos & derivados , Esfingosina/metabolismo
14.
PLoS One ; 14(3): e0213917, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30897151

RESUMO

Sphingolipids regulate several aspects of cell behavior and it has been demonstrated that cells adjust their sphingolipid metabolism in response to metabolic needs. Particularly, sphingosine-1-phosphate (S1P), a final product of sphingolipid metabolism, is a potent bioactive lipid involved in the regulation of various cellular processes, including cell proliferation, cell migration, actin cytoskeletal reorganization and cell adhesion. In previous work in rat renal papillae, we showed that sphingosine kinase (SK) expression and S1P levels are developmentally regulated and control de novo sphingolipid synthesis. The aim of the present study was to evaluate the participation of SK/S1P pathway in the triggering of cell differentiation by external hypertonicity. We found that hypertonicity evoked a sharp decrease in SK expression, thus activating the de novo sphingolipid synthesis pathway. Furthermore, the inhibition of SK activity evoked a relaxation of cell-cell adherens junction (AJ) with accumulation of the AJ complex (E-cadherin/ß-catenin/α-catenin) in the Golgi complex, preventing the acquisition of the differentiated cell phenotype. This phenotype alteration was a consequence of a sphingolipid misbalance with an increase in ceramide levels. Moreover, we found that SNAI1 and SNAI2 were located in the cell nucleus with impairment of cell differentiation induced by SK inhibition, a fact that is considered a biochemical marker of epithelial to mesenchymal transition. So, we suggest that the expression and activity of SK1, but not SK2, act as a control system, allowing epithelial cells to synchronize the various branches of sphingolipid metabolism for an adequate cell differentiation program.


Assuntos
Lisofosfolipídeos/metabolismo , Fosfotransferases (Aceptor do Grupo Álcool)/metabolismo , Esfingolipídeos/biossíntese , Esfingosina/análogos & derivados , Junções Aderentes/metabolismo , Animais , Diferenciação Celular , Movimento Celular , Proliferação de Células , Cães , Inibidores Enzimáticos/farmacologia , Células Epiteliais/citologia , Células Epiteliais/metabolismo , Soluções Hipertônicas , Células Madin Darby de Rim Canino , Fosfotransferases (Aceptor do Grupo Álcool)/antagonistas & inibidores , Fosfotransferases (Aceptor do Grupo Álcool)/genética , RNA Interferente Pequeno/genética , Transdução de Sinais , Fatores de Transcrição da Família Snail/metabolismo , Esfingosina/metabolismo
15.
BMB Rep ; 52(3): 220-225, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-30885289

RESUMO

We have identified a mechanism to diminish the proliferative capacity of cells during cell expansion using human adiposederived stromal cells (hAD-SCs) as a model of replicative senescence. hAD-SCs of high-passage numbers exhibited a reduced proliferative capacity with accelerated cellular senescence. Levels of key bioactive sphingolipids were significantly increased in these senescent hAD-SCs. Notably, the transcription of sphingosine kinase 1 (SPHK1) was down-regulated in hAD-SCs at high-passage numbers. SPHK1 knockdown as well as inhibition of its enzymatic activity impeded the proliferation of hAD-SCs, with concomitant induction of cellular senescence and accumulation of sphingolipids, as seen in high-passage cells. SPHK1 knockdown-accelerated cellular senescence was attenuated by co-treatment with sphingosine-1-phosphate and an inhibitor of ceramide synthesis, fumonisin B1, but not by treatment with either one alone. Together, these results suggest that transcriptional down-regulation of SPHK1 is a critical inducer of altered sphingolipid profiles and enhances replicative senescence during multiple rounds of cell division. [BMB Reports 2019; 52(3): 220-225].


Assuntos
Células-Tronco Mesenquimais/fisiologia , Fosfotransferases (Aceptor do Grupo Álcool)/genética , Apoptose/fisiologia , Proliferação de Células/genética , Senescência Celular/genética , Regulação para Baixo , Humanos , Lisofosfolipídeos/metabolismo , Células-Tronco Mesenquimais/citologia , Células-Tronco Mesenquimais/metabolismo , Fosfotransferases (Aceptor do Grupo Álcool)/biossíntese , Fosfotransferases (Aceptor do Grupo Álcool)/metabolismo , Esfingolipídeos/genética , Esfingolipídeos/metabolismo , Esfingosina/análogos & derivados , Esfingosina/metabolismo
16.
Bull Exp Biol Med ; 166(5): 667-670, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-30903503

RESUMO

We examined the efficacy of embryoid bodies from 6-day induced pluripotent stem cells an in vivo sepsis model. Injection of embryoid bodies to septic mice improved the condition of their lungs and significantly increased their survival rate. Although embryoid bodies secretedsphingosine-1-phosphate in vitro, its serum levels in mouse plasma were significantly reduced compared to that in the control (untreated mice receiving PBS). Low concentrations of sphingosine-1-phosphate protected endothelial cells, while high concentrations disrupted endothelial barrier integrity. Therefore, exogenous sphingosine-1-phosphate secreted by embryoid bodies during early stage of sepsis might down regulate endogenous production of sphingosine-1-phosphate. Inhibition of excessive sphingosine-1-phosphate release protects against endothelial injury and suppresses a vicious cycle of inflammatory reactions. The obtained results open new prospects in induced pluripotent stem cells-based therapy for sepsis.


Assuntos
Corpos Embrioides/citologia , Células-Tronco Pluripotentes Induzidas/citologia , Peritonite/terapia , Animais , Células Endoteliais/citologia , Transplante de Células-Tronco Hematopoéticas , Lisofosfolipídeos/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Peritonite/metabolismo , Sepse , Esfingosina/análogos & derivados , Esfingosina/metabolismo
17.
J Med Food ; 22(4): 325-336, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-30864855

RESUMO

Nonalcoholic fatty liver disease is a progressive disease involving the accumulation of lipid droplets in the liver. In this study, we investigated the anti-hepatosteatosis effects of fermented Cordyceps militaris extract (CME) in AML-12 hepatocytes. Although the levels of adenosine and cordycepin were reduced in the extracts of CM grown on germinated soybean (GSCE) and fermented CM grown on germinated soybean (GSC) by Pediococcus pentosaceus ON188 (ON188E), the expression of fatty acid oxidation (FAO) genes were upregulated only by GSC-ON188E treatment in a dose-dependent manner. In contrast, a lipogenic gene, stearoyl Coenzyme A desaturase 1, was downregulated by ON188E. Formation of intracellular lipid droplets by the addition of oleic acid was reduced by ON188E to levels observed in WY14643-treated cells. When cells were treated with ON188E, sphingosine kinase 2 mainly responsible for hepatic sphingosine 1-phosphate (S1P) synthesis was upregulated and S1P was elevated. Collectively, the fermented GSC extract activates FAO through elevation of S1P synthesis and has potential as a therapeutic for hepatosteatosis.


Assuntos
Cordyceps/química , Ácidos Graxos/metabolismo , Hepatopatia Gordurosa não Alcoólica/metabolismo , Extratos Vegetais/farmacologia , Animais , Linhagem Celular , Cordyceps/metabolismo , Fermentação , Hepatócitos/efeitos dos fármacos , Hepatócitos/metabolismo , Humanos , Lisofosfolipídeos/metabolismo , Camundongos , Hepatopatia Gordurosa não Alcoólica/tratamento farmacológico , Hepatopatia Gordurosa não Alcoólica/enzimologia , Hepatopatia Gordurosa não Alcoólica/genética , Oxirredução/efeitos dos fármacos , Pediococcus pentosaceus/metabolismo , Fosfotransferases (Aceptor do Grupo Álcool)/genética , Fosfotransferases (Aceptor do Grupo Álcool)/metabolismo , Extratos Vegetais/química , Extratos Vegetais/metabolismo , Esfingosina/análogos & derivados , Esfingosina/metabolismo , Estearoil-CoA Dessaturase/genética , Estearoil-CoA Dessaturase/metabolismo
18.
Plant Cell Physiol ; 60(5): 1109-1119, 2019 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-30796453

RESUMO

Long-chain bases (LCBs), also termed sphingobases, are building blocks of sphingolipids, which make up a significant proportion of the cellular membrane system. They are also bioactive molecules regulating intracellular processes. Elevated levels of LCBs like phytosphingosine and dihydrosphingosine can induce cell death in plants and correlate with programmed cell death (PCD) reactions after pathogen recognition. We investigated the previously hypothesized antagonism between phosphorylated and nonphosphorylated LCBs with respect to cell death in Arabidopsis thaliana. Using HPLC-MS/MS, we determined levels of phosphorylated and nonphosphorylated LCBs after cell death induction by LCB application or by Fumonisin B1 (FB1) treatment. We show that previously reported antagonistic effects of phosphorylated LCBs after simultaneous application with nonphosphorylated LCBs are linked to reduced uptake of nonphosphorylated LCBs into the tissue. Furthermore, phosphorylated LCBs did not antagonize PCD induced by avirulence protein recognition. In a functional approach, we used Arabidopsis lines with perturbed levels of phosphorylated LCBs. In these plants, the degree of FB1-induced cell death did not consistently correlate negatively with levels of phosphorylated LCBs, but positively with levels of major nonphosphorylated LCBs phytosphingosine and dihydrosphingosine. As treatment with phosphorylated LCBs did not antagonize cell death, and elevated in vivo levels of these LCB species did not reduce FB1-induced cell death, we conclude that the hypothesized general cell death-antagonizing effect of phosphorylated LCBs in plant cell death reactions should be rejected. Instead, our time-course analysis of LCB levels during cell death reactions showed a positive correlation between levels of nonphosphorylated LCBs and cell death.


Assuntos
Fumonisinas/farmacologia , Esfingolipídeos/metabolismo , Morte Celular/efeitos dos fármacos , Regulação da Expressão Gênica de Plantas/efeitos dos fármacos , Fosforilação/efeitos dos fármacos , Esfingosina/análogos & derivados , Esfingosina/metabolismo
19.
Methods Mol Biol ; 1949: 95-103, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30790251

RESUMO

Investigations into lipid localization and transport are often hampered by a lack of methods and tools to faithfully visualize lipids in the context of living cells, since fluorescent modifications drastically change lipid properties. Here, we describe the use of bifunctional as well as trifunctional sphingosine to reveal its subcellular localization via crosslinking, fixation, and specific staining by click reaction with a fluorophore. Additionally, these probes allow investigations into lipid metabolism as revealed by thin-layer chromatography.


Assuntos
Metabolismo dos Lipídeos , Lipídeos , Esfingolipídeos/metabolismo , Transporte Biológico , Cromatografia em Camada Delgada , Lipídeos/química , Estrutura Molecular , Esfingolipídeos/química , Esfingosina/metabolismo
20.
Methods Mol Biol ; 1949: 105-114, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30790252

RESUMO

Here we summarize how glycosphingolipid production can be followed using metabolic labeling with radiolabeled lipid precursors. No assays are available yet that directly would address the lipid transfer protein activity in vivo. Therefore, these approaches can serve as tools to indirectly study the lipid transfer protein activity in cells, by monitoring their impact on the glycosphingolipid homeostasis.


Assuntos
Proteínas de Transporte/metabolismo , Glicoesfingolipídeos/metabolismo , Ceramidas/metabolismo , Cromatografia em Camada Delgada , Glicolipídeos/metabolismo , Metabolismo dos Lipídeos , Redes e Vias Metabólicas , Esfingosina/análogos & derivados , Esfingosina/metabolismo
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