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1.
Int J Mol Sci ; 22(19)2021 Sep 30.
Artigo em Inglês | MEDLINE | ID: mdl-34638955

RESUMO

Sphingosine 1 phosphate (S1P) lyase (Sgpl1) catalyses the irreversible cleavage of S1P and thereby the last step of sphingolipid degradation. Loss of Sgpl1 in humans and mice leads to accumulation of sphingolipids and multiple organ injuries. Here, we addressed the role of hepatocyte Sgpl1 for regulation of sphingolipid homoeostasis by generating mice with hepatocyte-specific deletion of Sgpl1 (Sgpl1HepKO mice). Sgpl1HepKO mice had normal body weight, liver weight, liver structure and liver enzymes both at the age of 8 weeks and 8 months. S1P, sphingosine and ceramides, but not glucosylceramides or sphingomyelin, were elevated by ~1.5-2-fold in liver, and this phenotype did not progress with age. Several ceramides were elevated in plasma, while plasma S1P was normal. Interestingly, S1P and glucosylceramides, but not ceramides, were elevated in bile of Sgpl1HepKO mice. Furthermore, liver cholesterol was elevated, while LDL cholesterol decreased in 8-month-old mice. In agreement, the LDL receptor was upregulated, suggesting enhanced uptake of LDL cholesterol. Expression of peroxisome proliferator-activated receptor-γ, liver X receptor and fatty acid synthase was unaltered. These data show that mouse hepatocytes largely compensate the loss of Sgpl1 by secretion of accumulating sphingolipids in a specific manner into blood and bile, so that they can be excreted or degraded elsewhere.


Assuntos
Aldeído Liases/genética , Aldeído Liases/metabolismo , Bile/metabolismo , Fígado/metabolismo , Esfingolipídeos/sangue , Animais , Células Cultivadas , Ceramidas/metabolismo , LDL-Colesterol/metabolismo , Técnicas de Inativação de Genes , Hepatócitos/metabolismo , Homeostase/genética , Lisofosfolipídeos/metabolismo , Camundongos , Camundongos Knockout , Fenótipo , Receptores de LDL/metabolismo , Esfingosina/análogos & derivados , Esfingosina/metabolismo
2.
Molecules ; 26(19)2021 Sep 27.
Artigo em Inglês | MEDLINE | ID: mdl-34641404

RESUMO

Tomatoes are widely consumed, however, studies on tomato seeds are limited. In this study, we isolated 11 compounds including saponins and flavonol glycosides from tomato seeds and evaluated their effects on epidermal hydration. Among the isolated compounds, tomato seed saponins (10 µM) significantly increased the mRNA expression of proteins related to epidermal hydration, including filaggrin, involucrin, and enzymes for ceramide synthesis, by 1.32- to 1.91-fold compared with the control in HaCaT cells. Tomato seed saponins (10 µM) also decreased transepidermal water loss by 7 to 13 g/m2·h in the reconstructed human epidermal keratinization (RHEK) models. Quantitative analysis of the ceramide content in the stratum corneum (SC) revealed that lycoperoside H (1-10 µM) is a promising candidate to stimulate ceramide synthesis via the upregulation of ceramide synthase-3, glucosylceramide synthase, and ß-glucocerebrosidase, which led to an increase in the total SC ceramides (approximately 1.5-fold) in concert with ceramide (NP) (approximately 2-fold) in the RHEK models. Evaluation of the anti-inflammatory and anti-allergic effects of lycoperoside H demonstrated that lycoperoside H is suggested to act as a partial agonist of the glucocorticoid receptor and exhibits anti-inflammatory effects (10 mg/kg in animal test). These findings indicate that lycoperoside H can improve epidermal dehydration and suppress inflammation by increasing SC ceramide and steroidal anti-inflammatory activity.


Assuntos
Anti-Inflamatórios/farmacologia , Ceramidas/metabolismo , Epiderme/efeitos dos fármacos , Glicosídeos/farmacologia , Queratinócitos/efeitos dos fármacos , Lycopersicon esculentum/química , Esteroides/farmacologia , Animais , Desidratação , Epiderme/metabolismo , Glomerulonefrite/tratamento farmacológico , Glomerulonefrite/metabolismo , Glomerulonefrite/patologia , Cobaias , Humanos , Queratinócitos/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos , Saponinas/farmacologia , Sementes/química
3.
Int J Mol Sci ; 22(17)2021 Aug 27.
Artigo em Inglês | MEDLINE | ID: mdl-34502192

RESUMO

The release of exosomes can lead to cell-cell communication. Nutrients such as vitamin D3 and sphingolipids have important roles in many cellular functions, including proliferation, differentiation, senescence, and cancer. However, the specific composition of sphingolipids in exosomes and their changes induced by vitamin D3 treatment have not been elucidated. Here, we initially observed neutral sphingomyelinase and vitamin D receptors in exosomes released from HN9.10 embryonic hippocampal cells. Using ultrafast liquid chromatography tandem mass spectrometry, we showed that exosomes are rich in sphingomyelin species compared to whole cells. To interrogate the possible functions of vitamin D3, we established the optimal conditions of cell treatment and we analyzed exosome composition. Vitamin D3 was identified as responsible for the vitamin D receptor loss, for the increase in neutral sphingomyelinase content and sphingomyelin changes. As a consequence, the generation of ceramide upon vitamin D3 treatment was evident. Incubation of the cells with neutral sphingomyelinase, or the same concentration of ceramide produced in exosomes was necessary and sufficient to stimulate embryonic hippocampal cell differentiation, as vitamin D3. This is the first time that exosome ceramide is interrogated for mediate the effect of vitamin D3 in inducing cell differentiation.


Assuntos
Diferenciação Celular , Ceramidas/metabolismo , Colecalciferol/farmacologia , Exossomos/metabolismo , Hipocampo/metabolismo , Vitaminas/farmacologia , Células Cultivadas , Exossomos/efeitos dos fármacos , Hipocampo/efeitos dos fármacos , Hipocampo/embriologia , Humanos , Receptores de Calcitriol/metabolismo , Esfingomielina Fosfodiesterase/metabolismo
4.
Cells ; 10(9)2021 08 24.
Artigo em Inglês | MEDLINE | ID: mdl-34571822

RESUMO

As viruses are obligatory intracellular parasites, any step during their life cycle strictly depends on successful interaction with their particular host cells. In particular, their interaction with cellular membranes is of crucial importance for most steps in the viral replication cycle. Such interactions are initiated by uptake of viral particles and subsequent trafficking to intracellular compartments to access their replication compartments which provide a spatially confined environment concentrating viral and cellular components, and subsequently, employ cellular membranes for assembly and exit of viral progeny. The ability of viruses to actively modulate lipid composition such as sphingolipids (SLs) is essential for successful completion of the viral life cycle. In addition to their structural and biophysical properties of cellular membranes, some sphingolipid (SL) species are bioactive and as such, take part in cellular signaling processes involved in regulating viral replication. It is especially due to the progress made in tools to study accumulation and dynamics of SLs, which visualize their compartmentalization and identify interaction partners at a cellular level, as well as the availability of genetic knockout systems, that the role of particular SL species in the viral replication process can be analyzed and, most importantly, be explored as targets for therapeutic intervention.


Assuntos
Esfingolipídeos/metabolismo , Viroses , Transporte Biológico , Membrana Celular/química , Ceramidas/metabolismo , Sistemas de Liberação de Medicamentos , HIV/crescimento & desenvolvimento , Interações entre Hospedeiro e Microrganismos , Membranas Intracelulares/química , SARS-CoV-2/crescimento & desenvolvimento , Vírion , Replicação Viral , Vírus/crescimento & desenvolvimento
5.
J Neurochem ; 159(3): 574-589, 2021 11.
Artigo em Inglês | MEDLINE | ID: mdl-34482548

RESUMO

A contributing factor to the development of obesity is the consumption of a diet high in saturated fatty acids, such as palmitate. These fats induce hypothalamic neuroinflammation, which dysregulates neuronal function and induces orexigenic neuropeptide Y (Npy) to promote food intake. An inflammatory cytokine array identified multiple candidates that could mediate palmitate-induced up-regulation of Npy mRNA levels. Of these, visfatin or nicotinamide phosphoribosyltransferase (NAMPT), macrophage migratory inhibitory factor (MIF), and IL-17F were chosen for further study. Direct treatment of the neuropeptide Y/agouti-related peptide (NPY/AgRP)-expressing mHypoE-46 neuronal cell line with the aforementioned cytokines demonstrated that visfatin could directly induce Npy mRNA expression. Preventing the intracellular metabolism of palmitate through long-chain acyl-CoA synthetase (ACSL) inhibition was sufficient to block the palmitate-mediated increase in Npy gene expression. Furthermore, thin-layer chromatography revealed that in neurons, palmitate is readily incorporated into ceramides and defined species of phospholipids. Exogenous C16 ceramide, dipalmitoyl-phosphatidylcholine, and dipalmitoyl-phosphatidylethanolamine were sufficient to significantly induce Npy expression. This study suggests that the intracellular metabolism of palmitate and elevation of metabolites, including ceramide and phospholipids, are responsible for the palmitate-mediated induction of the potent orexigen Npy. Furthermore, this suggests that the regulation of Npy expression is less reliant on inflammatory cytokines per se than palmitate metabolites in a model of NPY/AgRP neurons. These lipid species likely induce detrimental downstream cellular signaling events ultimately causing an increase in feeding, resulting in an overweight phenotype and/or obesity.


Assuntos
Citocinas/farmacologia , Neuropeptídeo Y/biossíntese , Palmitatos/farmacologia , Acil Coenzima A/metabolismo , Animais , Linhagem Celular , Ceramidas/metabolismo , Meios de Cultivo Condicionados , Dieta Hiperlipídica , Expressão Gênica/efeitos dos fármacos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Neurônios/metabolismo , Nicotinamida Fosforribosiltransferase/farmacologia , RNA Mensageiro/biossíntese , RNA Mensageiro/genética
6.
Int J Mol Sci ; 22(16)2021 Aug 20.
Artigo em Inglês | MEDLINE | ID: mdl-34445706

RESUMO

Here, we present the main features of human acid sphingomyelinase (ASM), its biosynthesis, processing and intracellular trafficking, its structure, its broad substrate specificity, and the proposed mode of action at the surface of the phospholipid substrate carrying intraendolysosomal luminal vesicles. In addition, we discuss the complex regulation of its phospholipid cleaving activity by membrane lipids and lipid-binding proteins. The majority of the literature implies that ASM hydrolyses solely sphingomyelin to generate ceramide and ignores its ability to degrade further substrates. Indeed, more than twenty different phospholipids are cleaved by ASM in vitro, including some minor but functionally important phospholipids such as the growth factor ceramide-1-phosphate and the unique lysosomal lysolipid bis(monoacylglycero)phosphate. The inherited ASM deficiency, Niemann-Pick disease type A and B, impairs mainly, but not only, cellular sphingomyelin catabolism, causing a progressive sphingomyelin accumulation, which furthermore triggers a secondary accumulation of lipids (cholesterol, glucosylceramide, GM2) by inhibiting their turnover in late endosomes and lysosomes. However, ASM appears to be involved in a variety of major cellular functions with a regulatory significance for an increasing number of metabolic disorders. The biochemical characteristics of ASM, their potential effect on cellular lipid turnover, as well as a potential impact on physiological processes will be discussed.


Assuntos
Fosfolipídeos/biossíntese , Esfingomielina Fosfodiesterase/biossíntese , Esfingomielina Fosfodiesterase/metabolismo , Transporte Biológico , Ceramidas/metabolismo , Colesterol/metabolismo , Endossomos/metabolismo , Humanos , Lisossomos/metabolismo , Lipídeos de Membrana/metabolismo , Doença de Niemann-Pick Tipo A/metabolismo , Fosfolipídeos/metabolismo , Esfingomielina Fosfodiesterase/fisiologia , Esfingomielinas/metabolismo , Fosfolipases Tipo C/metabolismo , Fosfolipases Tipo C/fisiologia
7.
Mol Cell Biol ; 41(10): e0035221, 2021 09 24.
Artigo em Inglês | MEDLINE | ID: mdl-34370553

RESUMO

Sjögren-Larsson syndrome (SLS) is an inherited neurocutaneous disorder whose causative gene encodes the fatty aldehyde dehydrogenase ALDH3A2. To date, the detailed molecular mechanism of the skin pathology of SLS has remained largely unclear. We generated double-knockout (DKO) mice for Aldh3a2 and its homolog Aldh3b2 (a pseudogene in humans). These mice showed hyperkeratosis and reduced fatty aldehyde dehydrogenase activity and skin barrier function. The levels of ω-O-acylceramides (acylceramides), which are specialized ceramides essential for skin barrier function, in the epidermis of DKO mice were about 60% of those in wild-type mice. In the DKO mice, levels of acylceramide precursors (ω-hydroxy ceramides and triglycerides) were increased, suggesting that the final step of acylceramide production was inhibited. A decrease in acylceramide levels was also observed in human immortalized keratinocytes lacking ALDH3A2. Differentiated keratinocytes prepared from the DKO mice exhibited impaired long-chain base metabolism. Based on these results, we propose that the long-chain-base-derived fatty aldehydes that accumulate in DKO mice and SLS patients attack and inhibit the enzyme involved in the final step of acylceramide production. Our findings provide insight into the pathogenesis of the skin symptoms of SLS, i.e., decreased acylceramide production, and its molecular mechanism.


Assuntos
Aldeído Desidrogenase/metabolismo , Síndrome de Sjogren-Larsson/metabolismo , Pele/metabolismo , Aldeído Desidrogenase/genética , Aldeído Oxirredutases/genética , Aldeído Oxirredutases/metabolismo , Aldeídos/metabolismo , Animais , Diferenciação Celular , Ceramidas/metabolismo , Ceramidas/fisiologia , Modelos Animais de Doenças , Epiderme/metabolismo , Epiderme/fisiopatologia , Ácidos Graxos/genética , Ácidos Graxos/metabolismo , Feminino , Queratinócitos/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Síndrome de Sjogren-Larsson/fisiopatologia
8.
Cancer Sci ; 112(11): 4570-4579, 2021 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-34459070

RESUMO

Although the inhibition of acid ceramidase (AC) is known to induce antitumor effects in various cancers, there are few reports in pancreatic cancer, and the underlying mechanisms remain unclear. Moreover, there is currently no safe administration method of AC inhibitor. Here the effects of gene therapy using siRNA and shRNA for AC inhibition with its mechanisms for pancreatic cancer were investigated. The inhibition of AC by siRNA and shRNA using an adeno-associated virus 8 (AAV8) vector had antiproliferative effects by inducing apoptosis in pancreatic cancer cells and xenograft mouse model. Acid ceramidase inhibition elicits mitochondrial dysfunction, reactive oxygen species accumulation, and manganese superoxide dismutase suppression, resulting in apoptosis of pancreatic cancer cells accompanied by ceramide accumulation. These results elucidated the mechanisms underlying the antitumor effect of AC inhibition in pancreatic cancer cells and suggest the potential of the AAV8 vector to inhibit AC as a therapeutic strategy.


Assuntos
Ceramidase Ácida/antagonistas & inibidores , Terapia Genética/métodos , Doenças Mitocondriais/etiologia , Estresse Oxidativo , Neoplasias Pancreáticas/terapia , RNA Interferente Pequeno/uso terapêutico , Ceramidase Ácida/metabolismo , Animais , Apoptose , Linhagem Celular Tumoral , Ceramidas/metabolismo , Dependovirus , Vetores Genéticos , Humanos , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Neoplasias Pancreáticas/metabolismo , Neoplasias Pancreáticas/patologia , Distribuição Aleatória , Espécies Reativas de Oxigênio/metabolismo , Superóxido Dismutase/antagonistas & inibidores , Ensaios Antitumorais Modelo de Xenoenxerto
9.
Int J Mol Sci ; 22(15)2021 Jul 22.
Artigo em Inglês | MEDLINE | ID: mdl-34360617

RESUMO

Atopic dermatitis (AD or eczema) is the most common chronic inflammatory skin disorder worldwide. Ceramides (Cer) maintain skin barrier functions, which are disrupted in lesional skin of AD patients. However, Cer status during the pre-lesional phase of AD is not well defined. Using a variation of human AD-like preclinical model consisting of a 7-day topical exposure to ovalbumin (OVA), or control, we observed elevation of Cer C16 and C24. Skin mRNA quantification of enzymes involved in Cer metabolism [Cer synthases (CerS) and ceramidases (Asah1/Asah2)], which revealed augmented CerS 4, 5 and 6 and Asah1. Given the overall pro-apoptotic nature of Cer, local apoptosis was assessed, then quantified using novel morphometric measurements of cleaved caspase (Casp)-3-restricted immunofluorescence signal in skin samples. Apoptosis was induced in response to OVA. Because apoptosis may occur downstream of endoplasmic reticulum (ER) stress, we measured markers of ER stress-induced apoptosis and found elevated skin-associated CHOP protein upon OVA treatment. We previously substantiated the importance of mast cells (MC) in initiating early skin inflammation. OVA-induced Cer increase and local apoptosis were prevented in MC-deficient mice; however, they were restored following MC reconstitution. We propose that the MC/Cer axis is an essential pathogenic feature of pre-lesional AD, whose targeting may prevent disease development.


Assuntos
Apoptose , Ceramidas/metabolismo , Dermatite Atópica/patologia , Eczema/patologia , Mastócitos/patologia , Pele/patologia , Animais , Dermatite Atópica/induzido quimicamente , Dermatite Atópica/tratamento farmacológico , Dermatite Atópica/metabolismo , Eczema/induzido quimicamente , Eczema/tratamento farmacológico , Eczema/metabolismo , Feminino , Mastócitos/efeitos dos fármacos , Mastócitos/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Ovalbumina/toxicidade , Pele/efeitos dos fármacos , Pele/metabolismo
10.
Int J Mol Sci ; 22(15)2021 Aug 02.
Artigo em Inglês | MEDLINE | ID: mdl-34361066

RESUMO

Ceramides, a class of sphingolipids containing a backbone of sphingoid base, are the most important and effective structural component for the formation of the epidermal permeability barrier. While ceramides comprise approximately 50% of the epidermal lipid content by mass, the content is substantially decreased in certain inflammatory skin diseases, such as atopic dermatitis (AD), causing improper barrier function. It is widely accepted that the endocannabinoid system (ECS) can modulate a number of biological responses in the central nerve system, prior studies revealed that activation of endocannabinoid receptor CB1, a key component of ECS, triggers the generation of ceramides that mediate neuronal cell fate. However, as the impact of ECS on the production of epidermal ceramide has not been studied, we here investigated whether the ECS stimulates the generation of epidermal ceramides in an IL-4-treated in vitro model of skin inflammation using N-palmitoyl serinol (PS), an analog of the endocannabinoid N-palmitoyl ethanolamine. Accordingly, an IL-4-mediated decrease in cellular ceramide levels was significantly stimulated in human epidermal keratinocytes (KC) following PS treatment through both de novo ceramide synthesis- and sphingomyelin hydrolysis-pathways. Importantly, PS selectively increases ceramides with long-chain fatty acids (FAs) (C22-C24), which mainly account for the formation of the epidermal barrier, through activation of ceramide synthase (CerS) 2 and Cer3 in IL-4-mediated inflamed KC. Furthermore, blockade of cannabinoid receptor CB1 activation by AM-251 failed to stimulate the production of total ceramide as well as long-chain ceramides in response to PS. These studies demonstrate that an analog of endocannabinoid, PS, stimulates the generation of specific ceramide species as well as the total amount of ceramides via the endocannabinoid receptor CB1-dependent mechanism, thereby resulting in the enhancement of epidermal permeability barrier function.


Assuntos
Ceramidas/metabolismo , Inflamação/metabolismo , Queratinócitos/metabolismo , Propanolaminas/farmacologia , Propilenoglicóis/farmacologia , Receptor CB1 de Canabinoide/metabolismo , Pele/metabolismo , Células Cultivadas , Humanos , Técnicas In Vitro , Inflamação/tratamento farmacológico , Inflamação/imunologia , Inflamação/patologia , Queratinócitos/citologia , Queratinócitos/efeitos dos fármacos , Propanolaminas/química , Propilenoglicóis/química , Pele/citologia , Pele/efeitos dos fármacos
11.
Nat Commun ; 12(1): 5073, 2021 08 20.
Artigo em Inglês | MEDLINE | ID: mdl-34417467

RESUMO

The contents of numerous membrane lipids change upon ageing. However, it is unknown whether and how any of these changes are causally linked to lifespan regulation. Acyl chains contribute to the functional specificity of membrane lipids. In this study, working with C. elegans, we identified an acyl chain-specific sphingolipid, C22 glucosylceramide, as a longevity metabolite. Germline deficiency, a conserved lifespan-extending paradigm, induces somatic expression of the fatty acid elongase ELO-3, and behenic acid (22:0) generated by ELO-3 is incorporated into glucosylceramide for lifespan regulation. Mechanistically, C22 glucosylceramide is required for the membrane localization of clathrin, a protein that regulates membrane budding. The reduction in C22 glucosylceramide impairs the clathrin-dependent autophagic lysosome reformation, which subsequently leads to TOR activation and longevity suppression. These findings reveal a mechanistic link between membrane lipids and ageing and suggest a model of lifespan regulation by fatty acid-mediated membrane configuration.


Assuntos
Caenorhabditis elegans/fisiologia , Ácidos Graxos não Esterificados/metabolismo , Glicoesfingolipídeos/metabolismo , Homeostase , Longevidade/fisiologia , Lisossomos/metabolismo , Animais , Proteínas de Caenorhabditis elegans/metabolismo , Ceramidas/metabolismo , Colesterol/metabolismo , Clatrina/metabolismo , Mutação em Linhagem Germinativa/genética , Proteínas de Fluorescência Verde/metabolismo , Larva/metabolismo , Modelos Biológicos , Interferência de RNA , Estresse Fisiológico
12.
Biomolecules ; 11(7)2021 06 25.
Artigo em Inglês | MEDLINE | ID: mdl-34202192

RESUMO

Ceramide is a bioactive sphingolipid involved in numerous cellular processes. In addition to being the precursor of complex sphingolipids, ceramides can act as second messengers, especially when they are generated at the plasma membrane of cells. Its metabolic dysfunction may lead to or be a consequence of an underlying disease. Recent reports on transcriptomics and electrospray ionization mass spectrometry analysis have demonstrated the variation of specific levels of sphingolipids and enzymes involved in their metabolism in different neurodegenerative diseases. In the present review, we highlight the most relevant discoveries related to ceramide and neurodegeneration, with a special focus on Parkinson's disease.


Assuntos
Antiparkinsonianos/administração & dosagem , Ceramidas/metabolismo , Sistemas de Liberação de Medicamentos/métodos , Metabolismo dos Lipídeos/fisiologia , Doença de Parkinson/tratamento farmacológico , Doença de Parkinson/metabolismo , Animais , Ceramidas/antagonistas & inibidores , Humanos , Metabolismo dos Lipídeos/efeitos dos fármacos , Oxirredutases/antagonistas & inibidores , Oxirredutases/metabolismo , Esfingolipídeos/antagonistas & inibidores , Esfingolipídeos/metabolismo
13.
J Oleo Sci ; 70(8): 1147-1156, 2021 Aug 05.
Artigo em Inglês | MEDLINE | ID: mdl-34248103

RESUMO

In plants and fungi, sphingolipids, characterized by the presence of a sphingoid base (SB), comprise neutral classes, including ceramide (Cer) and glucosylceramide (GlcCer), and acidic classes, including glycosyl inositol phosphoryl ceramide (GIPC). The major class of plant and fungal sphingolipids is GIPC; however, owing to their complicated extraction and analysis, there is still little information regarding the food characteristics of GIPC compounds. In the present study, we evaluated the content and SB composition of highly polar sphingolipids (HPS) in materials that had been obtained from our previous food processing study for GlcCer and Cer. This assessment was based on the changes that occur in HPS containing GIPC in sake rice (saka-mai) during the rice polishing and sake (rice wine) brewing process. In addition, we report a new investigation into the composition of sphingolipids in koji rice and sake yeast. HPS levels were the highest among the sphingolipid classes in brown rice cultivars and highly polished rice. Sake and sake lees (sake-kasu) were produced using three different starter cultures. In sake lees, Cer levels were the highest among the classes, while HPS was greatly reduced based on the amount of highly polished rice and koji rice, and these HPS were mainly composed of sphinganine (d18:0), which is a minor SB in highly polished rice, koji rice, and sake yeast. In addition, considerable levels of free SBs, mainly comprising d18:0, were detected in sake lees. The levels of HPS and free SBs in sake lees were dependent on the starter culture. These results suggest that HPS was hydrolyzed to Cer and that sake yeast also affected the levels of Cer and free SBs during brewing. One interesting question raised by these results is whether changes in the class and base compositions of sphingolipids during brewing contribute to taste of the final product and other food functions.


Assuntos
Ceramidas/metabolismo , Esfingolipídeos/metabolismo , Esfingosina/metabolismo , Vinho/análise , Aspergillus oryzae/metabolismo , Ceramidas/análise , Fermentação , Manipulação de Alimentos , Hidrólise , Oryza/química , Saccharomyces cerevisiae/metabolismo , Esfingosina/análise
14.
Int J Mol Sci ; 22(14)2021 Jul 06.
Artigo em Inglês | MEDLINE | ID: mdl-34298889

RESUMO

The Acyl-CoA-binding domain-containing protein (ACBD3) plays multiple roles across the cell. Although generally associated with the Golgi apparatus, it operates also in mitochondria. In steroidogenic cells, ACBD3 is an important part of a multiprotein complex transporting cholesterol into mitochondria. Balance in mitochondrial cholesterol is essential for proper mitochondrial protein biosynthesis, among others. We generated ACBD3 knock-out (ACBD3-KO) HEK293 and HeLa cells and characterized the impact of protein absence on mitochondria, Golgi, and lipid profile. In ACBD3-KO cells, cholesterol level and mitochondrial structure and functions are not altered, demonstrating that an alternative pathway of cholesterol transport into mitochondria exists. However, ACBD3-KO cells exhibit enlarged Golgi area with absence of stacks and ribbon-like formation, confirming the importance of ACBD3 in Golgi stacking. The glycosylation of the LAMP2 glycoprotein was not affected by the altered Golgi structure. Moreover, decreased sphingomyelins together with normal ceramides and sphingomyelin synthase activity reveal the importance of ACBD3 in ceramide transport from ER to Golgi.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Complexo de Golgi/metabolismo , Proteínas de Membrana/metabolismo , Mitocôndrias/metabolismo , Transporte Biológico/fisiologia , Ceramidas/metabolismo , Colesterol/metabolismo , Glicosilação , Células HEK293 , Células HeLa , Humanos , Proteína 2 de Membrana Associada ao Lisossomo/metabolismo , Transdução de Sinais/fisiologia , Transferases (Outros Grupos de Fosfato Substituídos)/metabolismo
15.
Cells ; 10(5)2021 05 18.
Artigo em Inglês | MEDLINE | ID: mdl-34069977

RESUMO

Long-chain fatty acid oxidation disorders (lc-FAOD) are a group of diseases affecting the degradation of long-chain fatty acids. In order to investigate the disease specific alterations of the cellular lipidome, we performed undirected lipidomics in fibroblasts from patients with carnitine palmitoyltransferase II, very long-chain acyl-CoA dehydrogenase, and long-chain 3-hydroxyacyl-CoA dehydrogenase. We demonstrate a deep remodeling of mitochondrial cardiolipins. The aberrant phosphatidylcholine/phosphatidylethanolamine ratio and the increased content of plasmalogens and of lysophospholipids support the theory of an inflammatory phenotype in lc-FAOD. Moreover, we describe increased ratios of sphingomyelin/ceramide and sphingomyelin/hexosylceramide in LCHAD deficiency which may contribute to the neuropathic phenotype of LCHADD/mitochondrial trifunctional protein deficiency.


Assuntos
Ácidos Graxos/metabolismo , Fibroblastos/enzimologia , Erros Inatos do Metabolismo Lipídico/enzimologia , Lipidômica , Metaboloma , Pele/enzimologia , Acil-CoA Desidrogenase de Cadeia Longa/deficiência , Acil-CoA Desidrogenase de Cadeia Longa/genética , Cardiolipinas/metabolismo , Carnitina O-Palmitoiltransferase/deficiência , Carnitina O-Palmitoiltransferase/genética , Estudos de Casos e Controles , Células Cultivadas , Ceramidas/metabolismo , Feminino , Humanos , Erros Inatos do Metabolismo Lipídico/genética , 3-Hidroxiacil-CoA Desidrogenase de Cadeia Longa/deficiência , 3-Hidroxiacil-CoA Desidrogenase de Cadeia Longa/genética , Masculino , Erros Inatos do Metabolismo/enzimologia , Erros Inatos do Metabolismo/genética , Oxirredução , Esfingolipídeos/metabolismo , Espectrometria de Massas em Tandem
16.
Int J Mol Sci ; 22(10)2021 May 19.
Artigo em Inglês | MEDLINE | ID: mdl-34069652

RESUMO

Obesity is a global health issue for which no major effective treatments have been well established. High-fat diet consumption is closely related to the development of obesity because it negatively modulates the hypothalamic control of food intake due to metaflammation and lipotoxicity. The use of animal models, such as rodents, in conjunction with in vitro models of hypothalamic cells, can enhance the understanding of hypothalamic functions related to the control of energy balance, thereby providing knowledge about the impact of diet on the hypothalamus, in addition to targets for the development of new drugs that can be used in humans to decrease body weight. Recently, sphingolipids were described as having a lipotoxic effect in peripheral tissues and the central nervous system. Specifically, lipid overload, mainly from long-chain saturated fatty acids, such as palmitate, leads to excessive ceramide levels that can be sensed by the hypothalamus, triggering the dysregulation of energy balance control. However, no systematic review has been undertaken regarding studies of sphingolipids, particularly ceramide and sphingosine-1-phosphate (S1P), the hypothalamus, and obesity. This review confirms that ceramides are associated with hypothalamic dysfunction in response to metaflammation, endoplasmic reticulum (ER) stress, and lipotoxicity, leading to insulin/leptin resistance. However, in contrast to ceramide, S1P appears to be a central satiety factor in the hypothalamus. Thus, our work describes current evidence related to sphingolipids and their role in hypothalamic energy balance control. Hypothetically, the manipulation of sphingolipid levels could be useful in enabling clinicians to treat obesity, particularly by decreasing ceramide levels and the inflammation/endoplasmic reticulum stress induced in response to overfeeding with saturated fatty acids.


Assuntos
Ceramidas/metabolismo , Metabolismo Energético/fisiologia , Ácidos Graxos/fisiologia , Animais , Ceramidas/fisiologia , Dieta Hiperlipídica/efeitos adversos , Estresse do Retículo Endoplasmático/efeitos dos fármacos , Ácidos Graxos/metabolismo , Humanos , Hipotálamo/metabolismo , Hipotálamo/fisiologia , Resistência à Insulina/fisiologia , Leptina/metabolismo , Lisofosfolipídeos/metabolismo , Obesidade/metabolismo , Transdução de Sinais/fisiologia , Esfingolipídeos/metabolismo , Esfingosina/análogos & derivados , Esfingosina/metabolismo
17.
FASEB J ; 35(7): e21732, 2021 07.
Artigo em Inglês | MEDLINE | ID: mdl-34143450

RESUMO

Acid ceramidase (murine gene code: Asah1) (50 kDa) belongs to N-terminal nucleophile hydrolase family. This enzyme is located in the lysosome, which mediates conversion of ceramide (CER) into sphingosine and free fatty acids at acidic pH. CER plays an important role in intracellular sphingolipid metabolism and its increase causes inflammation. The mammalian target of rapamycin complex 1 (mTORC1) signaling on late endosomes (LEs)/lysosomes may control cargo selection, membrane biogenesis, and exosome secretion, which may be fine controlled by lysosomal sphingolipids such as CER. This lysosomal-CER-mTOR signaling may be a crucial molecular mechanism responsible for development of arterial medial calcification (AMC). Torin-1 (5 mg/kg/day), an mTOR inhibitor, significantly decreased aortic medial calcification accompanied with decreased expression of osteogenic markers like osteopontin (OSP) and runt-related transcription factor 2 (RUNX2) and upregulation of smooth muscle 22α (SM22-α) in mice receiving high dose of Vitamin D (500 000 IU/kg/day). Asah1fl/fl /SMCre mice had markedly increased co-localization of mTORC1 with lysosome-associated membrane protein-1 (Lamp-1) (lysosome marker) and decreased co-localization of vacuolar protein sorting-associated protein 16 (VPS16) (a multivesicular bodies [MVBs] marker) with Lamp-1, suggesting mTOR activation caused reduced MVBs interaction with lysosomes. Torin-1 significantly reduced the co-localization of mTOR vs Lamp-1, increased lysosome-MVB interaction which was associated with reduced accumulation of CD63 and annexin 2 (exosome markers) in the coronary arterial wall of mice. Using coronary artery smooth muscle cells (CASMCs), Pi -stimulation significantly increased p-mTOR expression in Asah1fl/fl /SMCre CASMCs as compared to WT/WT cells associated with increased calcium deposition and mineralization. Torin-1 blocked Pi -induced calcium deposition and mineralization. siRNA mTOR and Torin-1 significantly reduce co-localization of mTORC1 with Lamp-1, increased VPS16 vs Lamp-1 co-localization in Pi -stimulated CASMCs, associated with decreased exosome release. Functionally, Torin-1 significantly reduces arterial stiffening as shown by restoration from increased pulse wave velocity and decreased elastin breaks. These results suggest that lysosomal CER-mTOR signaling may play a critical role for the control of lysosome-MVB interaction, exosome secretion and arterial stiffening during AMC.


Assuntos
Ceramidase Ácida/metabolismo , Exossomos/metabolismo , Mamíferos/metabolismo , Miócitos de Músculo Liso/metabolismo , Osteogênese/fisiologia , Sirolimo/metabolismo , Animais , Aorta/metabolismo , Cálcio/metabolismo , Ceramidas/metabolismo , Subunidade alfa 1 de Fator de Ligação ao Core/metabolismo , Vasos Coronários/metabolismo , Lisossomos/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Corpos Multivesiculares/metabolismo , Análise de Onda de Pulso/métodos , Transdução de Sinais/fisiologia , Esfingolipídeos/metabolismo , Serina-Treonina Quinases TOR/metabolismo , Calcificação Vascular/metabolismo
18.
Anticancer Res ; 41(6): 2875-2883, 2021 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-34083278

RESUMO

BACKGROUND/AIM: A series of experiments on HeLa cells were conducted to provide new information concerning the anti-cancer properties of jaspine B hydrochloride (JBH). MATERIALS AND METHODS: HeLa cells treated with 0.5 µmol/l JBH for 24, 48, and 72 h underwent flow cytometric analysis of the cell cycle, and measurement of phosphatidylserine externalization, mitochondrial membrane potential (MMP), casp-3 activation, cleavage of PARP, ceramide levels, aSMase activity, and Bcl-2 release. nSMase activity was measured by a colorimetric assay. Gene expression was determined by qRT-PCR. Immunocytochemistry was performed to detect p21 and p27 expression. RESULTS: JBH-induced apoptosis in HeLa cells associated with externalization of phosphatidylserine, reduced MMP, activation of casp-3, and cleavage of PARP as well as up-regulation of TNF-α, FasL, and casp-8. Significant increase in nSMase activity, ceramide levels, Bcl-2 release (predominantly in the inactive form), and pro-apoptotic nuclear localization of p21 and p27 were also detected. CONCLUSION: JBH-induced apoptosis in HeLa cells is associated with disrupted sphingolipid homeostasis resulting in increased ceramide levels.


Assuntos
Apoptose/efeitos dos fármacos , Ceramidas/metabolismo , Esfingolipídeos/metabolismo , Esfingosina/análogos & derivados , Proliferação de Células/efeitos dos fármacos , Células HeLa , Humanos , Transdução de Sinais/efeitos dos fármacos , Esfingosina/farmacologia
19.
Aging (Albany NY) ; 13(12): 15750-15769, 2021 06 08.
Artigo em Inglês | MEDLINE | ID: mdl-34102611

RESUMO

Cellular senescence is linked to chronic age-related diseases including atherosclerosis, diabetes, and neurodegeneration. Compared to proliferating cells, senescent cells express distinct subsets of proteins. In this study, we used cultured human diploid fibroblasts rendered senescent through replicative exhaustion or ionizing radiation to identify proteins differentially expressed during senescence. We identified acid ceramidase (ASAH1), a lysosomal enzyme that cleaves ceramide into sphingosine and fatty acid, as being highly elevated in senescent cells. This increase in ASAH1 levels in senescent cells was associated with a rise in the levels of ASAH1 mRNA and a robust increase in ASAH1 protein stability. Furthermore, silencing ASAH1 in pre-senescent fibroblasts decreased the levels of senescence proteins p16, p21, and p53, and reduced the activity of the senescence-associated ß-galactosidase. Interestingly, depletion of ASAH1 in pre-senescent cells sensitized these cells to the senolytics Dasatinib and Quercetin (D+Q). Together, our study indicates that ASAH1 promotes senescence, protects senescent cells, and confers resistance against senolytic drugs. Given that inhibiting ASAH1 sensitizes cells towards senolysis, this enzyme represents an attractive therapeutic target in interventions aimed at eliminating senescent cells.


Assuntos
Ceramidase Ácida/metabolismo , Senescência Celular , Fibroblastos/citologia , Fibroblastos/enzimologia , Ceramidase Ácida/genética , Linhagem Celular , Proliferação de Células/genética , Sobrevivência Celular , Ceramidas/metabolismo , Inativação Gênica , Humanos , Metaboloma , Biossíntese de Proteínas/genética , Estabilidade de RNA/genética
20.
Cells ; 10(6)2021 05 21.
Artigo em Inglês | MEDLINE | ID: mdl-34064003

RESUMO

Ceramide and diacylglycerol (DAG) are bioactive lipids and mediate many cellular signaling pathways. Sphingomyelin synthase (SMS) is the single metabolic link between the two, while SMS2 is the only SMS form located at the plasma membrane. SMS2 functions were investigated in HepG2 cell lines stably expressing SMS2. SMS2 overexpression did not alter sphingomyelin (SM), phosphatidylcholine (PC), or ceramide levels. DAG content increased by approx. 40% and led to downregulation of DAG-dependent protein kinase C (PKC). SMS2 overexpression also induced senescence, characterized by positivity for ß-galactosidase activity and heterochromatin foci. HepG2-SMS2 cells exhibited protruded mitochondria and suppressed mitochondrial respiration rates. ATP production and the abundance of Complex V were substantially lower in HepG2-SMS2 cells as compared to controls. SMS2 overexpression was associated with inflammasome activation based on increases in IL-1ß and nlpr3 mRNA levels. HepG2-SMS2 cells exhibited lipid droplet accumulation, constitutive activation of AMPK based on elevated 172Thr phosphorylation, increased AMPK abundance, and insensitivity to insulin suppression of AMPK. Thus, our results show that SMS2 regulates DAG homeostasis and signaling in hepatocytes and also provide proof of principle for the concept that offset in bioactive lipids' production at the plasma membrane can drive the senescence program in association with steatosis and, seemingly, by cell-autonomous mechanisms.


Assuntos
Membrana Celular/metabolismo , Ceramidas/metabolismo , Diglicerídeos/metabolismo , Transferases (Outros Grupos de Fosfato Substituídos)/metabolismo , Senescência Celular , Fígado Gorduroso/metabolismo , Células Hep G2 , Humanos
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