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Gentamicin is an antibiotic often essential for the treatment of severe infections due to Gram-negative bacteria, but they cause nephrotoxic reactions. This adverse effect has been attributed to their uptake by endocytosis by proximal tubular cells, their huge accumulation inside the lysosomes of these cells and the development of an array of alterations in proximal tubule epithelium leading to apoptosis. This process was observed after the treatment of rats at low, therapeutically relevant doses but has been reproduced on renal cultured cells (LLC-PK1, MDCK) (El Mouedden et al., 200. Tox. Sci. 56: 229-239; ElMouedden et al., 2000. Antimicrob. Agents Chemother. 44/665-675).
In the present work, we have investigated activation of caspases-3,8 and 9 occurring upon gentamicin-induced apoptosis in LLC-PK1 and the potential role of lysosomal proteases in this process.
We first confirmed that gentamicin induces apoptosis in a dose (1-3mM) and time (4-72h) dependent fashion by measuring the activation of executioner caspase-3. Whatever the concentration of gentamicin used, the activation of caspase-3 at 16 hours.
We then explored the implication of the extrinsic and the intrinsic pathways, by measuring the activation of caspase-8 and 9 respectively. In the presence of 2 mM gentamicin, an activation of caspase-9 only was detected after 12th of incubation, suggesting the involvement of mitochondria pathway on the apoptosis induced by gentamicin upstream to the caspase-3 activation.
Finally, based on the facts that gentamicin is accumulated in the lysosomes and is able to permeabilize the lysosomal membrane ( Servais et al, en preparation), we evaluated, using the DAPI technique, the potential role of lysosomal proteases like cathepsins D and B on the percentage of apoptotic cells in comparison with that observed on cells treated with gentamicin. Using pepstatin A as inhibitor of cathepsin D and and leupeptin as inhibitor of cathepsin B, we showed that inhibition of cathepsin D party reduced the number of apoptotic cells which, in contrast, was slightly increased in presence of an inhibitor of cathepsin B.
Together, these data suggest that gentamicin-induced apoptosis implies both the mitochondrial pathway (resulting in the activation of caspase-9) and the lysosomal pathway (through the pro-apoptotic effect of cathepsin D which is probably released from lysosomes of cells incubated with gentamicin) La gentamicine est un antibiotique appartenant à la famille des aminoglycosides qui est utilisée en thérapeutique pour faire face aux infections graves à bactéries Gram-. Après filtration glomérulaire, la gentamicine est réabsorbée, par un processus d’endocytose, au niveau des cellules du tubule proximal où elle s’accumule dans les lysosomes. A des doses équivalentes à des doses thérapeutiques, la gentamicine induit de l’apoptose in vivo dans les reins de rats traités. Ce processus a pu être reproduit sur plusieurs types cellulaires dont les cellules rénales LLC-PK1 et MDCK.
Des études antérieures ont démontré l’importance de l’accumulation cellulaire de la gentamicine pour le développement du processus apoptique ainsi qu’un effet du temps d’incubation (1 à 3 jours) et de la concentration extracellulaire en gentamicine (1 à 3 mM). Pour évaluer le processus d’apoptose induit par la gentamicine, nous avons dosé l’activité de la caspase-3 (caspase exécutrice) pour différents temps d’incubation (de 4 à 72 h) avec des concentrations en gentamicine de 1 à 3 mM et avions confirmé l’effet temps et l’effet concentration. Quelque soit la concentration en gentamicine utilisée, m’activation des caspases-3 débute à 16 heures.
Par la suite, nous avons cherché à savoir quelle(s) voie(s) signalétique(s) étai(en)t responsable(s) de l’activation de la caspase-3. Les résultats obtenus ont montré, dés 12 heures, une activation de la caspase-9, caspase’ initiatrice de la voie mitochondriale/ intrinsèque. Afin de déterminer le mécanisme par lequel cette voie était activé, nous avons étudié le rôle potentiel des cathepsines B et D dans l’apoptose induite par la gentamicine. Des études menées en parallèle ont montré la capacité de la gentamicine à perméabiliser le lysosome, suggérant que des protéases lysosomiales pourraient être relarguées dans le cytosol et activer directement ou indirectement la mitochondrie. Les résultats obtenus en utilisant la leupeptine et la pepsatine A comme inhibiteurs des cathepsines B et D semblent indiquer que la cathepsine D jouerait un rôle dans l’apoptose induite par le gentamicine alors que la cathepsine B préviendrait ce processus.
Sur base de nos travaux et d’observations réalisées en parallèle au laboratoire, nous proposons que l’apoptose induite par la gentamicine dans les cellules LLC-PK1 serait initiées par une perméabilisassions de la membrane du lysosome, entraînant un relargage de protéases lysosomiales et de gentamicine qui seraient responsables de l’activation directe ou indirecte de la mitochondrie. Cette cascade entraînerait l’activation de la caspase-3 menant aux phénomènes d’apoptose induit par la gentamicine

Gentamicins --- Caspases --- Cathepsins --- Apoptosis

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This Special Issue of Cancers focuses on new advances in the treatment of renal cell carcinoma, both surgical and pharmacological (and combinations of these), and novel approaches to tackle treatment resistance and improve our understanding of this phenomenon.

renal cell carcinoma --- autophagy --- hydroxychloroquine --- chloroquine --- ROC-325 --- cysteine cathepsins --- cysteine cathepsin inhibitors --- lysosome --- renal cancer --- metastatic renal cell carcinoma --- immune-based combination therapies --- network meta-analysis --- PD-L1 --- predictive --- biomarker --- treatment --- TKIs --- mRCC --- biomarkers --- soluble factors --- immunotherapy --- renal cell carcinoma (RCC) --- sunitib resistance --- artesunate (ART) --- Traditional Chinese Medicine (TCM) --- growth inhibition --- ferroptosis --- reactive oxygen species (ROS) --- clear cell renal cell carcinoma --- ccRCC --- RCC --- kidney cancer --- evolution --- evolutionary trajectory --- metastatic --- second line therapy --- renal cell cancer --- immune checkpoint inhibitors --- tyrosine kinase inhibitors --- individualization --- genomic signature --- transcriptomic analysis

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Once viewed solely as fat storage cells, adipocytes and their adipokines have now been proven to be central for human health. Understanding that overweight and obesity may increase the risk for various diseases requires detailed characterization of adipokine function. Weight gain, weight regain, and fasting affect adipocyte health and accordingly their secretome. Different adipose tissue deposits exist and they vary in cellular composition and function. The evidence is strong of a role of adipokines in cancer, reproductive function, neurological diseases, cardiovascular diseases ,and rheumatoid arthritis. Adipokines are considered useful biomarkers for adipose tissue and metabolic health, and may be used as diagnostic tools in rheumatoid arthritis, cancer, or sepsis. This book contains 10 original articles and 9 review articles focusing on these bioactive peptides. Several articles deal with chemerin, an adipokine discovered more than 20 years ago. Data so far have resulted in promising insights related to its biological function. We are only beginning to understand the multiple roles of chemerin, the mechanisms regulating its activity, and the signaling pathways used by this chemokine. Adipokine receptor agonists and antagonists may result in the formulation of novel drugs and ultimately may lead to new therapeutic targets to be used in clinical practice.

n/a --- lipids --- cathepsins --- neurodegeneration --- tocilizumab --- SGBS adipocytes --- chemerin receptors --- cholesterol --- metabolically healthy obese --- energy metabolism --- adipose-brain axis --- EP3 receptor --- leptin --- rheumatoid arthritis --- excessive gestational weight gain --- in vitro fat regain --- secreted frizzled-related protein 5 --- PCOS --- EP4 receptor --- leukocyte --- exchange protein directly activated by cAMP isoform 2 (EPAC2) --- extracellular remodeling --- insulin --- osteoarthritis --- lipid metabolism --- fat mass --- Tango bioassay --- fatty liver --- free fatty acids --- label-free proteomic profiling --- interleukin(IL)-33 --- sick fat --- polycystic ovary syndrome --- early-life programming --- inflammation --- gestational diabetes --- glucose restriction --- adipokines --- neonatal anthropometry --- epicardial adipose tissue (EAT) --- oestrous cycle --- Cardiovascular Diseases (CVDs) --- triglycerides --- G protein-coupled receptor 1 --- testicular pathologies --- prognosis --- ovary --- ICU --- biologic activity --- preeclempsia --- adipokine --- critical illness --- early pregnancy --- myokine --- liver steatosis --- C-C chemokine receptor-like 2 --- testis --- stimulating growth factor 2 (ST2) --- pig --- fitness --- human granulosa cells --- obesity --- proteolysis --- annexins --- adipose tissue --- biomarker --- ghrelin --- brain health --- sepsis --- adiponectin --- rheumatic diseases --- chemokine-like receptor 1 --- follicular fluid --- glucose homeostasis --- resistin --- Nonalcoholic fatty liver disease --- prostaglandin E2 (PGE2) --- visceral fat --- microglia --- weight regain --- complement factors --- alpha-fetoprotein --- polycystic ovary morphology --- chemerin --- cancer --- depression --- hypertension --- hypothalamus