RESUMEN
Since the 1980s, it has been known that the administration of ganglioside GM1 to cultured cells induced or enhanced neuronal differentiation. GM1 mechanism of action relies on its direct interaction and subsequent activation of the membrane tyrosine kinase receptor, TrkA, which naturally serves as NGF receptor. This process is mediated by the sole oligosaccharide portion of GM1, the pentasaccharide ß-Gal-(1-3)-ß-GalNAc-(1-4)-[α-Neu5Ac-(2-3)]-ß-Gal-(1-4)-ß-Glc. Here we detailed the minimum structural requirements of the oligosaccharide portion of GM1 for mediating the TrkA dependent neuritogenic processing. By in vitro and in silico biochemical approaches, we demonstrated that the minimal portion of GM1 required for the TrkA activation is the inner core of the ganglioside's oligosaccharide ß-Gal-(1-3)-ß-GalNAc-(1-4)-[α-Neu5Ac-(2-3)]-ß-Gal. The addition of a sialic acid residue at position 3 of the outer galactose of the GM1 oligosaccharide, which forms the oligosaccharide of GD1a, prevented the interaction with TrkA and the resulting neuritogenesis. On the contrary, the addition of a fucose residue at position 2 of the outer galactose, forming the Fucosyl-GM1 oligosaccharide, did not prevent the TrkA-mediated neuritogenesis.
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Gangliósido G(M1) , Galactosa , Gangliósido G(M1)/química , Ácido N-Acetilneuramínico , Oligosacáridos/químicaRESUMEN
In this work, we describe the association of a novel homozygous VPS11 variant with adult-onset generalized dystonia, providing a detailed clinical report and biological evidence of disease mechanism. Vps11 is a subunit of the homotypic fusion and protein sorting (HOPS) complex, which promotes the fusion of late endosomes and autophagosomes with the lysosome. Functional studies on mutated fibroblasts showed marked lysosomal and autophagic abnormalities, which improved after overexpression of the wild type Vps11 protein. In conclusion, a deleterious VPS11 variant, damaging the autophagic and lysosomal pathways, is the probable genetic cause of a novel form of generalized dystonia. ANN NEUROL 2021;89:834-839.
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Distonía/genética , Proteínas de Transporte Vesicular/genética , Adulto , Edad de Inicio , Secuencia de Aminoácidos , Autofagia/genética , Encéfalo/diagnóstico por imagen , ADN/genética , Distonía/diagnóstico por imagen , Distonía/etiología , Endosomas/patología , Fibroblastos/patología , Variación Genética , Homocigoto , Humanos , Lisosomas/patología , Imagen por Resonancia Magnética , Mutación , Linaje , Fagosomas/patología , Secuenciación del ExomaRESUMEN
BACKGROUND: To date, variants in the GBA gene represent the most frequent large-effect genetic factor associated with Parkinson's disease (PD). However, the reason why individuals with the same GBA variant may or may not develop neurodegeneration and PD is still unclear. OBJECTIVES: Therefore, we evaluated the contribution of rare variants in genes responsible for lysosomal storage disorders (LSDs) to GBA-PD risk, comparing the burden of deleterious variants in LSD genes in PD patients versus asymptomatic subjects, all carriers of deleterious variants in GBA. METHODS: We used a custom next-generation sequencing panel, including 50 LSD genes, to screen 305 patients and 207 controls (discovery cohort). Replication and meta-analysis were performed in two replication cohorts of GBA-variant carriers, of 250 patients and 287 controls, for whom exome or genome data were available. RESULTS: Statistical analysis in the discovery cohort revealed a significantly increased burden of deleterious variants in LSD genes in patients (P = 0.0029). Moreover, our analyses evidenced that the two strongest modifiers of GBA penetrance are a second variation in GBA (5.6% vs. 1.4%, P = 0.023) and variants in genes causing mucopolysaccharidoses (6.9% vs. 1%, P = 0.0020). These results were confirmed in the meta-analysis, where we observed pooled odds ratios of 1.42 (95% confidence interval [CI] = 1.10-1.83, P = 0.0063), 4.36 (95% CI = 2.02-9.45, P = 0.00019), and 1.83 (95% CI = 1.04-3.22, P = 0.038) for variants in LSD genes, GBA, and mucopolysaccharidosis genes, respectively. CONCLUSION: The identification of genetic lesions in lysosomal genes increasing PD risk may have important implications in terms of patient stratification for future therapeutic trials. © 2022 The Authors. Movement Disorders published by Wiley Periodicals LLC on behalf of International Parkinson Movement Disorder Society.
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Enfermedad de Parkinson , Humanos , Glucosilceramidasa/genética , Heterocigoto , Lisosomas , Mutación , Enfermedad de Parkinson/complicaciones , Enfermedad de Parkinson/genéticaRESUMEN
The Cost Action "Innovation with glycans: new frontiers from synthesis to new biological targets" (INNOGLY) hosted the Workshop "Neuroglycoproteins in health and disease", in Alicante, Spain, on March 2022. This event brought together an european group of scientists that presented novel insights into changes in glycosylation in diseases of the central nervous system and cancer, as well as new techniques to study protein glycosylation. Herein we provide the abstracts of all the presentations.
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Neoplasias , Polisacáridos , Glicosilación , Humanos , Polisacáridos/metabolismoRESUMEN
Glycans have been shown to function as versatile molecular signals in cells. This prompted us to look at their roles in endocytosis, endolysosomal system and autophagy. We start by introducing the cell biological aspects of these pathways, the concept of the sugar code, and provide an overview on the role of glycans in the targeting of lysosomal proteins and in lysosomal functions. Moreover, we review evidence on the regulation of endocytosis and autophagy by glycans. Finally, we discuss the emerging concept that cytosolic exposure of luminal glycans, and their detection by endogenous lectins, provides a mechanism for the surveillance of the integrity of the endolysosomal compartments, and serves their eventual repair or disposal.
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Autofagia/fisiología , Endocitosis/fisiología , Lisosomas/fisiología , Polisacáridos/metabolismo , Regulación de la Expresión Génica , Proteínas/genética , Proteínas/metabolismoRESUMEN
It is well over a century that glycosphingolipids are matter of interest in different fields of research. The hydrophilic oligosaccharide and the lipid moiety, the ceramide, both or separately have been considered in different moments as the crucial portion of the molecule, responsible for the role played by the glycosphingolipids associated to the plasma-membranes or to any other subcellular fraction. Glycosphingolipids are a family of compounds characterized by thousands of structures differing in both the oligosaccharide and the ceramide moieties, but among them, the nervous system monosialylated glycosphingolipid GM1, belonging to the group of gangliosides, has gained particular attention by a multitude of Scientists. In recent years, a series of studies have been conducted on the functional roles played by the hydrophilic part of GM1, its oligosaccharide, that we have named "OligoGM1". These studies allowed to shed new light on the mechanisms underlying the properties of GM1 defining the role of the OligoGM1 in determining precise interactions with membrane proteins instrumental for the neuronal functions, leaving to the ceramide the role of correctly positioning the GM1 in the membrane crucial for the oligosaccharide-protein interactions. In this review we aim to report the recent studies on the cascade of events modulated by OligoGM1, as the bioactive portion of GM1, to support neuronal differentiation and trophism together with preclinical studies on its potential to modify the progression of Parkinson's disease.
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Gangliósido G(M1)/química , Gangliósido G(M1)/metabolismo , Enfermedades Neurodegenerativas/tratamiento farmacológico , Oligosacáridos/química , Animales , Diferenciación Celular , Gangliósido G(M1)/farmacología , Humanos , Mitocondrias/metabolismo , Enfermedades Neurodegenerativas/metabolismo , Neuronas/citología , Neuronas/metabolismo , Oligosacáridos/síntesis química , Oligosacáridos/metabolismo , Receptor trkA/metabolismoRESUMEN
Cardiac fibrosis is a key physiological response to cardiac tissue injury to protect the heart from wall rupture. However, its progression increases heart stiffness, eventually causing a decrease in heart contractility. Unfortunately, to date, no efficient antifibrotic therapies are available to the clinic. This is primarily due to the complexity of the process, which involves several cell types and signaling pathways. For instance, the transforming growth factor beta (TGF-ß) signaling pathway has been recognized to be vital for myofibroblasts activation and fibrosis progression. In this context, complex sphingolipids, such as ganglioside GM3, have been shown to be directly involved in TGF-ß receptor 1 (TGF-R1) activation. In this work, we report that an induced up-regulation of sialidase Neu3, a glycohydrolytic enzyme involved in ganglioside cell homeostasis, can significantly reduce cardiac fibrosis in primary cultures of human cardiac fibroblasts by inhibiting the TGF-ß signaling pathway, ultimately decreasing collagen I deposition. These results support the notion that modulating ganglioside GM3 cell content could represent a novel therapeutic approach for cardiac fibrosis, warranting for further investigations.
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Fibroblastos/metabolismo , Gangliósido G(M3)/metabolismo , Regulación Enzimológica de la Expresión Génica , Miocardio/metabolismo , Neuraminidasa/biosíntesis , Regulación hacia Arriba , Fibroblastos/patología , Fibrosis , Humanos , Miocardio/patología , Receptor Tipo I de Factor de Crecimiento Transformador beta/metabolismoRESUMEN
Glycosphingolipids are amphiphilic plasma membrane components formed by a glycan linked to a specific lipid moiety. In this chapter we report on these compounds, on their role played in our cells to maintain the correct cell biology.In detail, we report on their structure, on their metabolic processes, on their interaction with proteins and from this, their property to modulate positively in health and negatively in disease, the cell signaling and cell biology.
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Glicoesfingolípidos , Lípidos , Membrana Celular , Transducción de SeñalRESUMEN
The favorable outcome of in vivo and ex vivo gene therapy approaches in several Lysosomal Storage Diseases suggests that these treatment strategies might equally benefit GM2 gangliosidosis. Tay-Sachs and Sandhoff disease (the main forms of GM2 gangliosidosis) result from mutations in either the HEXA or HEXB genes encoding, respectively, the α- or ß-subunits of the lysosomal ß-Hexosaminidase enzyme. In physiological conditions, α- and ß-subunits combine to generate ß-Hexosaminidase A (HexA, αß) and ß-Hexosaminidase B (HexB, ßß). A major impairment to establishing in vivo or ex vivo gene therapy for GM2 gangliosidosis is the need to synthesize the α- and ß-subunits at high levels and with the correct stoichiometric ratio, and to safely deliver the therapeutic products to all affected tissues/organs. Here, we report the generation and in vitro validation of novel bicistronic lentiviral vectors (LVs) encoding for both the murine and human codon optimized Hexa and Hexb genes. We show that these LVs drive the safe and coordinate expression of the α- and ß-subunits, leading to supranormal levels of ß-Hexosaminidase activity with prevalent formation of a functional HexA in SD murine neurons and glia, murine bone marrow-derived hematopoietic stem/progenitor cells (HSPCs), and human SD fibroblasts. The restoration/overexpression of ß-Hexosaminidase leads to the reduction of intracellular GM2 ganglioside storage in transduced and in cross-corrected SD murine neural progeny, indicating that the transgenic enzyme is secreted and functional. Importantly, bicistronic LVs safely and efficiently transduce human neurons/glia and CD34+ HSPCs, which are target and effector cells, respectively, in prospective in vivo and ex vivo GT approaches. We anticipate that these bicistronic LVs may overcome the current requirement of two vectors co-delivering the α- or ß-subunits genes. Careful assessment of the safety and therapeutic potential of these bicistronic LVs in the SD murine model will pave the way to the clinical development of LV-based gene therapy for GM2 gangliosidosis.
Asunto(s)
Gangliosidosis GM2/metabolismo , Terapia Genética/métodos , Células Madre Hematopoyéticas/metabolismo , Células-Madre Neurales/metabolismo , Cadena alfa de beta-Hexosaminidasa/metabolismo , Cadena beta de beta-Hexosaminidasa/metabolismo , Animales , Gangliosidosis GM2/genética , Vectores Genéticos , Humanos , Lentivirus , Ratones , Cadena alfa de beta-Hexosaminidasa/genética , Cadena beta de beta-Hexosaminidasa/genéticaRESUMEN
Human primary bronchial epithelial cells differentiated in vitro represent a valuable tool to study lung diseases such as cystic fibrosis (CF), an inherited disorder caused by mutations in the gene coding for the Cystic Fibrosis Transmembrane Conductance Regulator. In CF, sphingolipids, a ubiquitous class of bioactive lipids mainly associated with the outer layer of the plasma membrane, seem to play a crucial role in the establishment of the severe lung complications. Nevertheless, no information on the involvement of sphingolipids and their metabolism in the differentiation of primary bronchial epithelial cells are available so far. Here we show that ceramide and globotriaosylceramide increased during cell differentiation, whereas glucosylceramide and gangliosides content decreased. In addition, we found that apical plasma membrane of differentiated bronchial cells is characterized by a higher content of sphingolipids in comparison to the other cell membranes and that activity of sphingolipids catabolic enzymes associated with this membrane results altered with respect to the total cell activities. In particular, the apical membrane of CF cells was characterized by high levels of ceramide and glucosylceramide, known to have proinflammatory activity. On this basis, our data further support the role of sphingolipids in the onset of CF lung pathology.
Asunto(s)
Diferenciación Celular/genética , Fibrosis Quística/genética , Hidrolasas/genética , Esfingolípidos/genética , Bronquios/enzimología , Membrana Celular/enzimología , Membrana Celular/genética , Ceramidas/genética , Fibrosis Quística/enzimología , Fibrosis Quística/metabolismo , Fibrosis Quística/patología , Glucosilceramidas/genética , Humanos , Hidrolasas/química , Cultivo Primario de Células , Esfingolípidos/metabolismoRESUMEN
Apolipoprotein L1 (APOL1) wild type (G0) plays a role in the metabolism of sphingolipids, glycosphingolipids, sphingomyelin and ceramide, which constitute bioactive components of the lipid rafts (DRM). We asked whether APOL1 variants (APOL1-Vs) G1 and G2 carry the potential to alter the metabolism of sphingolipids in human podocytes. The sphingolipid pattern in HPs overexpressing either APOL1G0 or APOL1-Vs was analysed by using a thin mono- and bi-dimensional layer chromatography, mass-spectrometry and metabolic labelling with [1-3H]sphingosine. HP G0 and G1/G2-Vs exhibit a comparable decrease in lactosylceramide and an increase in the globotriaosylceramide content. An analysis of the main glycohydrolases activity involved in glycosphingolipid catabolism showed an overall decrease in the activeness of the tested enzymes, irrespective of the type of APOL1-Vs expression. Similarly, the high throughput cell live-based assay showed a comparable increased action of the plasma membrane glycosphingolipid-glycohydrolases in living cells independent of the genetic APOL1 expression profile. Importantly, the most significative modification of the sphingolipid pattern induced by APOL1-Vs occurred in DRM resulted with a drastic reduction of radioactivity associated with sphingolipids. G1/G2-Vs present a decrease amount of globotriaosylceramide and globopentaosylceramide compared to G0. Additionally, ceramide at the DRM site and lactosylceramide in general, showed a greatest fall in G1/G2 in comparison with G0. Additionally, the levels of glucosylceramide decreased only in the DRM of human podocytes overexpressing G1/G2-Vs. These findings suggest that altered sphingolipidsprofiles may contribute to the deranged functionality of the plasma membrane in APOL1 risk milieu.
Asunto(s)
Apolipoproteína L1/genética , Glicósido Hidrolasas/genética , Podocitos/metabolismo , Esfingolípidos/genética , Membrana Celular/genética , Membrana Celular/metabolismo , Regulación de la Expresión Génica/genética , Humanos , Metabolismo , Polimorfismo Genético/genética , Esfingolípidos/metabolismoRESUMEN
Gangliosides constitute a subgroup of glycosphingolipids characterized by the presence of sialic acid residues in their structure. As constituents of cellular membranes, in particular of raft microdomains, they exert multiple functions, some of them capital in cell homeostasis. Their presence in cells is tightly regulated by a balanced expression and function of the enzymes responsible for their biosynthesis, ganglioside synthases, and their degradation, glycosidases. The dysregulation of their abundance results in rare and common diseases. In this review, we make a point on the relevance of gangliosides and some of their metabolic precursors, such as ceramides, in the function of podocytes, the main cellular component of the glomerular filtration barrier, as well as their implications in podocytopathies. The results presented in this review suggest the pertinence of clinical lipidomic studies targeting these metabolites.
Asunto(s)
Membrana Celular/metabolismo , Gangliósidos/metabolismo , Barrera de Filtración Glomerular/metabolismo , Podocitos/patología , Animales , Humanos , Podocitos/metabolismoRESUMEN
Phagotherapy, the use of bacteriophages to fight bacterial infections as an alternative to antibiotic treatments, has become of increasing interest in the last years. This is mainly due to the diffusion of multi-drug resistant (MDR) bacterial infections that constitute a serious issue for public health. Phage therapy is gaining favor due to its success in agriculture and veterinary treatments and its extensive utilization for human therapeutic protocols in the Eastern world. In the last decades, some clinical trials and compassionate treatments have also been performed in the Western world, indicating that phage therapy is getting closer to its introduction in standard therapy protocols. However, several questions concerning the use of phages in human therapeutic treatments are still present and need to be addressed. In this review, we illustrate the state of art of phage therapy and examine the role of animal models to translate these treatments to humans.
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Terapia de Fagos/métodos , Investigación Biomédica Traslacional/métodos , Animales , Infecciones Bacterianas/terapia , Pollos , Modelos Animales de Enfermedad , Humanos , Nematodos , Terapia de Fagos/normas , Roedores , Investigación Biomédica Traslacional/normas , Pez CebraRESUMEN
Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) protein is expressed at the apical plasma membrane (PM) of different epithelial cells. The most common mutation responsible for the onset of cystic fibrosis (CF), F508del, inhibits the biosynthesis and transport of the protein at PM, and also presents gating and stability defects of the membrane anion channel upon its rescue by the use of correctors and potentiators. This prompted a multiple drug strategy for F508delCFTR aimed simultaneously at its rescue, functional potentiation and PM stabilization. Since ganglioside GM1 is involved in the functional stabilization of transmembrane proteins, we investigated its role as an adjuvant to increase the effectiveness of CFTR modulators. According to our results, we found that GM1 resides in the same PM microenvironment as CFTR. In CF cells, the expression of the mutated channel is accompanied by a decrease in the PM GM1 content. Interestingly, by the exogenous administration of GM1, it becomes a component of the PM, reducing the destabilizing effect of the potentiator VX-770 on rescued CFTR protein expression/function and improving its stabilization. This evidence could represent a starting point for developing innovative therapeutic strategies based on the co-administration of GM1, correctors and potentiators, with the aim of improving F508del CFTR function.
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Adyuvantes Inmunológicos/farmacología , Aminofenoles/farmacología , Aminopiridinas/farmacología , Benzodioxoles/farmacología , Fibrosis Quística/tratamiento farmacológico , Gangliósido G(M1)/farmacología , Quinolonas/farmacología , Adyuvantes Inmunológicos/química , Aminofenoles/química , Bronquios/efectos de los fármacos , Bronquios/metabolismo , Bronquios/patología , Agonistas de los Canales de Cloruro/química , Agonistas de los Canales de Cloruro/farmacología , Fibrosis Quística/genética , Fibrosis Quística/patología , Regulador de Conductancia de Transmembrana de Fibrosis Quística/genética , Células Epiteliales/efectos de los fármacos , Células Epiteliales/metabolismo , Células Epiteliales/patología , Gangliósido G(M1)/química , Humanos , Mutación , Quinolonas/química , Terapias en InvestigaciónRESUMEN
Lysosomal accumulation of undegraded materials is a common feature of lysosomal storage diseases, neurodegenerative disorders, and the aging process. To better understand the role of lysosomal storage in the onset of cell damage, we used human fibroblasts loaded with sucrose as a model of lysosomal accumulation. Sucrose-loaded fibroblasts displayed increased lysosomal biogenesis followed by arrested cell proliferation. Notably, we found that reduced lysosomal catabolism and autophagy impairment led to an increase in sphingolipids ( i.e., sphingomyelin, glucosylceramide, ceramide, and the gangliosides GM3 and GD3), at both intracellular and plasma membrane (PM) levels. In addition, we observed an increase in the lysosomal membrane protein Lamp-1 on the PM of sucrose-loaded fibroblasts and a greater release of the soluble lysosomal protein cathepsin D in their extracellular medium compared with controls. These results indicate increased fusion between lysosomes and the PM, as also suggested by the increased activity of lysosomal glycosphingolipid hydrolases on the PM of sucrose-loaded fibroblasts. The inhibition of ß-glucocerebrosidase and nonlysosomal glucosylceramidase, both involved in ceramide production resulting from glycosphingolipid catabolism on the PM, partially restored cell proliferation. Our findings indicate the existence of a new molecular mechanism underlying cell damage triggered by lysosomal impairment.-Samarani, M., Loberto, N., Soldà, G., Straniero, L., Asselta, R., Duga, S., Lunghi, G., Zucca, F. A., Mauri, L., Ciampa, M. G., Schiumarini, D., Bassi, R., Giussani, P., Chiricozzi, E., Prinetti, A., Aureli, M., Sonnino, S. A lysosome-plasma membrane-sphingolipid axis linking lysosomal storage to cell growth arrest.
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Puntos de Control del Ciclo Celular , Membrana Celular/metabolismo , Fibroblastos/metabolismo , Lisosomas/metabolismo , Esfingolípidos/metabolismo , Catepsina D/genética , Catepsina D/metabolismo , Línea Celular , Membrana Celular/genética , Fibroblastos/citología , Humanos , Proteínas de Membrana de los Lisosomas/genética , Proteínas de Membrana de los Lisosomas/metabolismo , Lisosomas/genética , Esfingolípidos/genéticaRESUMEN
Sphingolipid metabolism is an intricate network of several interdependent and co-regulated pathways. In addition to the mainstream biosynthetic and catabolic pathways, several processes, even if less important in contributing to the final tissue sphingolipid composition from the quantitative point of view, might become relevant when sphingolipid metabolism is for any reason dysregulated and concur to the onset of neuronal pathologies. The main subcellular sites involved in the mainstream metabolic pathway are represented by the Golgi apparatus (for the biosynthesis) and by the lysosomes (for catabolism). On the other hand, the minor collateral pathways are associated with the plasma membrane and membranes of other organelles, and likely play important roles in the local regulation of membrane dynamics and contribute to maintain a perfect membrane organization functional to the physiology of the cell. In this review, we will consider few aspects of the sphingolipid metabolic pathway depending by the dynamic of the membranes that seems to become relevant in neurodegenerative diseases.
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Membrana Celular/metabolismo , Aparato de Golgi/metabolismo , Lisosomas/metabolismo , Enfermedades del Sistema Nervioso/metabolismo , Neuronas/metabolismo , Esfingolípidos/metabolismo , Animales , Membrana Celular/genética , Membrana Celular/patología , Aparato de Golgi/genética , Aparato de Golgi/patología , Humanos , Lisosomas/genética , Lisosomas/patología , Enfermedades del Sistema Nervioso/genética , Enfermedades del Sistema Nervioso/patología , Neuronas/patología , Esfingolípidos/genéticaRESUMEN
Prostate cancer (PC) is one of the most common leading causes of cancer-related death in men. Currently, the main therapeutic approaches available for PC are based on the androgen deprivation and on radiotherapy. However, despite these treatments being initially effective in cancer remission, several patients undergo recurrence, developing a most aggressive and resistant PC.Emerging evidence showed that abiraterone acetate drug will reduce PC recurrence by a mechanism independent of the inhibition of Cytochrome P450 17α-hydroxylase/17,20-lyase. Here we describe the involvement in the abiraterone-mediated PC cell death of a particular class of bioactive lipids called sphingolipids (SL). Sphingolipids are components of plasma membrane (PM) that organize macromolecular complexes involved in the control of several signaling pathways including the tumor cell death induced by radiotherapy. Here, we show for the first time that both in androgen-sensitive and insensitive PC cells abiraterone and ionizing radiation induce a reorganization of the plasma membrane SL composition. This event is triggered by activation of the PM-associated glycohydrolases that induce the production of cytotoxic ceramide by the in situ hydrolyses of glycosphingolipids. Taken together our data open a new scenario on the SL involvement in the therapy of PC.
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Androstenos/farmacología , Neoplasias de la Próstata/patología , Radiación Ionizante , Esfingolípidos/química , Línea Celular Tumoral , Homeostasis , Humanos , MasculinoRESUMEN
The GBA2 gene encodes the non-lysosomal glucosylceramidase (NLGase), an enzyme that catalyzes the conversion of glucosylceramide (GlcCer) to ceramide and glucose. Mutations in GBA2 have been associated with the development of neurological disorders such as autosomal recessive cerebellar ataxia, hereditary spastic paraplegia, and Marinesco-Sjogren-Like Syndrome. Our group has previously identified the GBA2 c.1780G>C [p.Asp594His] missense mutation, in a Cypriot consanguineous family with spastic ataxia. In this study, we carried out a biochemical characterization of lymphoblastoid cell lines (LCLs) derived from three patients of this family. We found that the mutation strongly reduce NLGase activity both intracellularly and at the plasma membrane level. Additionally, we observed a two-fold increase of GlcCer content in LCLs derived from patients compared to controls, with the C16 lipid being the most abundant GlcCer species. Moreover, we showed that there is an apparent compensatory effect between NLGase and the lysosomal glucosylceramidase (GCase), since we found that the activity of GCase was three-fold higher in LCLs derived from patients compared to controls. We conclude that the c.1780G>C mutation results in NLGase loss of function with abolishment of the enzymatic activity and accumulation of GlcCer accompanied by a compensatory increase in GCase.
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Discapacidad Intelectual/genética , Discapacidad Intelectual/metabolismo , Linfocitos/metabolismo , Espasticidad Muscular/genética , Espasticidad Muscular/metabolismo , Mutación Missense , Atrofia Óptica/genética , Atrofia Óptica/metabolismo , Ataxias Espinocerebelosas/genética , Ataxias Espinocerebelosas/metabolismo , beta-Glucosidasa/genética , Alelos , Biomarcadores , Línea Celular , Activación Enzimática , Glucosilceramidasa/metabolismo , Glucosilceramidas/metabolismo , Humanos , beta-Glucosidasa/metabolismoRESUMEN
Cystic fibrosis (CF) is the most common autosomal genetic recessive disease caused by mutations of gene encoding for the cystic fibrosis transmembrane conductance regulator. Patients with CF display a wide spectrum of symptoms, the most severe being chronic lung infection and inflammation, which lead to onset of cystic fibrosis lung disease. Several studies indicate that sphingolipids play a regulatory role in airway inflammation. The inhibition and downregulation of GBA2, the enzyme catabolizing glucosylceramide to ceramide, are associated with a significant reduction of IL-8 production in CF bronchial epithelial cells. Herein, we demonstrate that GBA2 plays a role in the proinflammatory state characterizing CF cells. We also report for the first time that Pseudomonas aeruginosa infection causes a recruitment of plasma membrane-associated glycosphingolipid hydrolases into lipid rafts of CuFi-1-infected cells. This reorganization of cell membrane may be responsible for activation of a signaling cascade, culminating in aberrant inflammatory response in CF bronchial epithelial cells upon bacterial infection. Taken together, the presented data further support the role of sphingolipids and their metabolic enzymes in controlling the inflammatory response in CF.
Asunto(s)
Fibrosis Quística/metabolismo , Fibrosis Quística/microbiología , Glicósido Hidrolasas/metabolismo , Infecciones por Pseudomonas/metabolismo , Esfingolípidos/metabolismo , beta-Glucosidasa/metabolismo , Bronquios/metabolismo , Bronquios/microbiología , Línea Celular , Membrana Celular/metabolismo , Membrana Celular/microbiología , Fibrosis Quística/complicaciones , Glucosilceramidasa , Humanos , Mediadores de Inflamación/metabolismo , Microdominios de Membrana/metabolismo , Modelos Biológicos , Infecciones por Pseudomonas/complicaciones , Mucosa Respiratoria/metabolismo , Mucosa Respiratoria/microbiología , Transducción de SeñalRESUMEN
The brain is characterized by the presence of cell types with very different functional specialization, but with the common trait of a very high complexity of structures originated by their plasma membranes. Brain cells bear evident membrane polarization with the creation of different morphological and functional subcompartments, whose formation, stabilization and function require a very high level of lateral order within the membrane. In other words, the membrane specialization of brain cells implies the presence of distinct membrane domains. The brain is the organ with the highest enrichment in lipids like cholesterol, glycosphingolipids, and the most recently discovered brain membrane lipid, phosphatidylglucoside, whose collective behavior strongly favors segregation within the membrane leading to the formation of lipid-driven membrane domains. Lipid-driven membrane domains function as dynamic platforms for signal transduction, protein processing, and membrane turnover. Essential events involved in the development and in the maintenance of the functional integrity of the brain depend on the organization of lipid-driven membrane domains, and alterations in lipid homeostasis, leading to deranged lipid-driven membrane organization, are common in several major brain diseases. In this review, we summarize the forces behind the formation of lipid membrane domains and their biological roles in different brain cells. This article is part of a Special Issue entitled Brain Lipids.