Caspase-10-Dependent Cell Death in Fas/CD95 Signalling Is Not Abrogated by Caspase Inhibitor zVAD-fmk

Upon CD95/Fas ligation, the initiator caspase-8 is known to activate effector caspases leading to apoptosis. In the presence of zVAD-fmk, a broad-spectrum caspase inhibitor, Fas engagement can also trigger an alternative, non-apoptotic caspase-independent form of cell death, which is initiated by RIP1. Controversy exists as to the ability of caspase-10 to mediate cell death in response to FasL (CD95L or CD178). Herein, the role of caspase-10 in FasL-induced cell death has been re-evaluated

Sphingomyelin Synthase 1 (SMS1) Downregulation Is Associated With Sphingolipid Reprogramming and a Worse Prognosis in Melanoma

Sphingolipid (SL) metabolism alterations have been frequently reported in cancer including in melanoma, a bad-prognosis skin cancer. In normal cells, de novo synthesized ceramide is mainly converted to sphingomyelin (SM), the most abundant SL, by sphingomyelin synthase 1 (SMS1) and, albeit to a lesser extent, SMS2, encoded by the SGMS1 and SGMS2 genes, respectively. Alternatively, ceramide can be converted to glucosylceramide (GlcCer) by the GlcCer synthase (GCS), encoded by the UGCG gene. Herein, we provide evidence for the first time that SMS1 is frequently downregulated in various solid cancers, more particularly in melanoma. Accordingly, various human melanoma cells displayed a SL metabolism signature associated with (i) a robust and a low expression of UGCG and SGMS1/2, respectively, (ii) higher in situ enzyme activity of GCS than SMS, and (iii) higher intracellular levels of GlcCer than SM. SMS1 was expressed at low levels in most of the human melanoma biopsies. In addition, several mutations and increased CpG island methylation in the SGMS1 gene were identified that likely affect SMS1 expression. Finally, low SMS1 expression was associated with a worse prognosis in metastatic melanoma patients. Collectively, our study indicates that SMS1 downregulation in melanoma enhances GlcCer synthesis, triggering an imbalance in the SM/GlcCer homeostasis, which likely contributes to melanoma progression. Evaluating SMS1 expression level in tumor samples might serve as a biomarker to predict clinical outcome in advanced melanoma patients

Guidelines for the use and interpretation of assays for monitoring autophagy (3rd edition)

In 2008 we published the first set of guidelines for standardizing research in autophagy. Since then, research on this topic has continued to accelerate, and many new scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding. Accordingly, it is important to update these guidelines for monitoring autophagy in different organisms. Various reviews have described the range of assays that have been used for this purpose. Nevertheless, there continues to be confusion regarding acceptable methods to measure autophagy, especially in multicellular eukaryotes. A key point that needs to be emphasized is that there is a difference between measurements that monitor the numbers or volume of autophagic elements (e.g., autophagosomes or autolysosomes) at any stage of the autophagic process vs. those that measure flux through the autophagy pathway (i.e., the complete process); thus, a block in macroautophagy that results in autophagosome accumulation needs to be differentiated from stimuli that result in increased autophagic activity, defined as increased autophagy induction coupled with increased delivery to, and degradation within, lysosomes (in most higher eukaryotes and some protists such as Dictyostelium) or the vacuole (in plants and fungi). In other words, it is especially important that investigators new to the field understand that the appearance of more autophagosomes does not necessarily equate with more autophagy. In fact, in many cases, autophagosomes accumulate because of a block in trafficking to lysosomes without a concomitant change in autophagosome biogenesis, whereas an increase in autolysosomes may reflect a reduction in degradative activity. Here, we present a set of guidelines for the selection and interpretation of methods for use by investigators who aim to examine macroautophagy and related processes, as well as for reviewers who need to provide realistic and reasonable critiques of papers that are focused on these processes. These guidelines are not meant to be a formulaic set of rules, because the appropriate assays depend in part on the question being asked and the system being used. In addition, we emphasize that no individual assay is guaranteed to be the most appropriate one in every situation, and we strongly recommend the use of multiple assays to monitor autophagy. In these guidelines, we consider these various methods of assessing autophagy and what information can, or cannot, be obtained from them. Finally, by discussing the merits and limits of particular autophagy assays, we hope to encourage technical innovation in the field

La cardiotoxicité des anthracyclines : mécanismes et cibles pharmacologiques de prévention

Les anthracyclines sont des agents antitumoraux très largement utilisés en cancérologie. Toutefois, un certain nombre d'effets secondaires, et notamment sur la fonction cardiaque, limitent leur utilisation. La cardiotoxicité consiste en une insuffisance cardiaque d'apparition retardée dont la fréquence est proportionnelle à la dose cumulée. Au niveau cellulaire, les mécanismes par lesquels les anthracyclines exercent leur cardiotoxicité sont encore mal compris. Dans cette revue, nous aborderons l'état des connaissances actuelles concernant la production d'espèces réactives de l'oxygène, la formation de métabolites toxiques et la mort cellulaire programmée induites par les anthracyclines au niveau cardiaque. Puis nous donnerons quelques exemples de molécules utilisées in vitro et in vivo dans le but de protéger les myocytes cardiaques des altérations induites par les anthracyclines

Rôle de la sphingosine 1-phosphate dans les interactions mélanome-stroma

TOULOUSE3-BU Sciences (315552104) / SudocSudocFranceF

Essais de thérapie anti-tumorale application au mélanome par ciblage du métabolisme des sphingolipides

TOULOUSE3-BU Sciences (315552104) / SudocSudocFranceF

Untargeted lipidomic analysis of primary human epidermal melanocytes acutely and chronically exposed to UV radiation

Ultraviolet (UV) radiation present in sunlight has been related to harmful effects on skin such as premature aging and skin cancer. In order to study the effects of UV radiation on skin, many investigations have been carried out at transcriptomic and proteomic levels. However, studies on the effects of UV radiation on lipid composition are scarce. In this work, primary cultures of melanocytes were exposed to UV radiation in a similar UVA/UVB ratio to that found in solar light. The was carried out twice a week and different endpoints were investigated at 0.5 (acute exposure), 1.5 and 3 weeks. As a result, dendrite formation and a progressive reduction in cell viability were observed. Also, cell cycle arrest and a reduced E-cadherin content were detected at 0.5 and 1.5 weeks. In the second stage of the study, lipid extracts of melanocytes were analysed by liquid chromatography coupled to mass spectrometry (LC-MS) and subjected to an untargeted lipidomic approach using the ROIMCR chemometric method. Among the most important changes observed under UV irradiation, lipid raft components such as sphingomyelins and GM3 gangliosides as well as other signalling molecules such as phosphatidylinositols decreased progressively with time. These modifications indicated strong effects on important functions such as cell signalling and recognition. In contrast, triacylglycerol species, associated with energy storage, increased progressively, which could be interpreted as a survival mechanism under adverse conditions. Further studies are needed to better understand the functional implications of the changes observed.The research leading to these results received funding from the European Research Council under the European Union's Seventh Framework Programme (FP/2007-2013)/ERC Grant Agreement No. 320737. We thank Dr Thierry Levade (Toulouse, France) for critical reading and discussion.Peer reviewe

Phenotypic and lipidomic characterization of primary human epidermal keratinocytes exposed to simulated solar UV radiation

Background: Ultraviolet (UV) radiation is known to be one of the most important environmental hazards acting on the skin. The most part of UV radiation is absorbed in the epidermis, where keratinocytes are the most abundant and exposed cell type. Lipids have an important role in skin biology, not only for their important contribution to the maintenance of the permeability barrier but also for the production and storage of energy, membrane organization and cell signalling functions. However, the effects on the lipid composition of keratinocytes under UV radiation are little explored. Objective: The present work aims to explore the effects on the phenotype and lipid content of primary human keratinocytes exposed to simulated solar UV radiation. Methods: Keratinocytes were exposed to a single (acute exposure) and repeated simulated solar UV irradiations for 4 weeks (chronic exposure). Cell viability and morphology were explored, as well as the production of reactive oxygen species. Then, lipid extracts were analysed through liquid chromatography coupled to mass spectrometry (LC–MS) and the data generated was processed using the ROIMCR chemometric methodology together with partial least squares discriminant analysis (PLS-DA), to finally reveal the most relevant lipid changes that occurred in keratinocytes upon UV irradiation. Also, the potential induction of keratinocyte differentiation was explored by measuring the increase of involucrin. Results: Under acute irradiation, cell viability and morphology were not altered. However, a general increase of phosphatidylcholines (PC) phosphatidylethanolamines (PE) and phosphatidylglycerol (PG) together with a slight sphingomyelin (SM) decrease were found in UV irradiated cells, among other changes. In addition, keratinocyte cultures did not present any differentiation hallmark. Contrary to acute-irradiated cells, in chronic exposures, cell viability was reduced and keratinocytes presented an altered morphology. Also, hallmarks of differentiation, such as the increase of involucrin protein and the autophagy induction were detected. Among the main lipid changes that accompanied this phenotype, the increase of long-chain ceramides, lysoPC and glycerolipid species were found. Conclusion: Important lipid changes were detected under acute and chronic UV irradiation. The lipid profile under chronic exposure may represent a lipid fingerprint of the keratinocyte differentiation phenotype. © 2018The research leading to these results has received funding from the European Research Council under the European Union's Seventh Framework Programme (FP/2007-2013) / ERC Grant Agreement no. 320737. We thank Dr. Thierry Levade (Toulouse, France) for critical reading and discussion.Peer reviewe

Regulation of cell death by sphingosine 1-phosphate lyase.

International audienceBy controlling sphingosine 1-phosphate (S1P) catabolism, S1P lyase (SPL) represents an undeniable candidate as potential regulator of a cancer cell's fate in response to stress. Our recent study reveals that complete loss of SPL activity leads to upregulation of the anti-apoptotic proteins Bcl-2 and Bcl-xL and consequently protects against apoptosis induced by chemotherapy and nutrient starvation but not against autophagy. Here, we speculate on how S1P and disruption of S1P breakdown may regulate cell death and autophagy