137,747 research outputs found
Overexpression and silencing of nicotinamide N-methyltransferase in human cancer cell lines
Get PDFIl carcinoma polmonare rappresenta la principale causa di morte per cancro a livello mondiale.
Il carcinoma orale rappresenta l’undicesima neoplasia più diffusa nel mondo. Nonostante i progressi effettuati in campo terapeutico, il tasso di sopravvivenza dei pazienti affetti da tali patologie rimane scarsa. Pertanto, risulta di fondamentale importanza l’identificazione di biomarcatori per una diagnosi precoce di tali neoplasie.
Oggetto del presente lavoro di ricerca è l’enzima nicotinamide N-metiltrasferasi (NNMT). Al fine di esplorare il ruolo svolto dall’enzima nel metabolismo della cellula tumorale, i livelli di espressione dell’NNMT sono stati esaminati nella linea cellulare di carcinoma polmonare A549 ed è stato valutato l’effetto del silenziamento dell’enzima sulla tumorigenicità mediante il saggio MTT ed il saggio in soft-agar. Successivamente, è stato esaminato l’effetto dell’upregolazione dell’enzima sulla proliferazione cellulare nella linea cellulare di carcinoma orale HSC-2. Inoltre, è stato esplorato l’effetto dell’overespressione dell’NNMT sui livelli di espressione di geni coinvolti nel controllo del ciclo cellulare e dell’apoptosi, quali la β-catenina, Ki-67 e la survivina.
Il silenziamento dell’NNMT nelle cellule A549 ha portato ad una significativa inibizione della proliferazione cellulare e della capacità di formare colonie in soft-agar, mentre l’overespressione dell’enzima nelle cellule HSC-2 ha determinato un significativo aumento della crescita cellulare. Inoltre, l’isoforma ΔEx3 della survivina è risultata significativamente upregolata nelle cellule HSC-2 overesprimenti l’NNMT. I risultati ottenuti hanno dimostrato il coinvolgimento dell’NNMT nella proliferazione della cellula tumorale, candidando l’enzima quale interessante bersaglio per una terapia antineoplastica. Inoltre, la correlazione osservata tra i livelli di espressione dell’NNMT e dell’isoforma ΔEx3 della survivina sembra suggerire un potenziale ruolo dell’NNMT nell’apoptosi
Increased biosynthesis of glycosphingolipids in CDG-Ia fibroblasts
No full textCongenital disorder of glycosylation Ia (CDG-Ia) is an autosomal recessive disease, characterized by the impaired biosynthesis of the N-linked oligosaccharide chains of proteins due to a deficiency of phosphomannomutase (PMM), the enzyme converting mannose-6-phosphate into mannose-1-phosphate. We investigated the consequences of the altered N-linked glycoprotein (GP) biosynthesis on the quantity and quality of glycosphingolipids (GSLs) in fibroblasts of CDG-Ia patients. First, we found that CDG-Ia fibroblasts contain an increased amount of total GSLs when compared with normal fibroblasts. Further, we assessed by metabolic labeling of CDG-Ia fibroblasts with radioactive sugar precursors, including galactose and N-acetylmannosamine, that a diminished biosynthesis of cellular GPs is antagonized by an increased biosynthesis of GSLs. An increased GSL biosynthesis was also observed by means of radio-labeled lipid precursors including sphingosine and lactosylceramide. Notably, also the degradation of GLSs is slowed down in CDG-Ia fibroblasts. Finally, when we labeled normal human fibroblasts and CHO cells with radioactive galactose in the presence and absence of deoxymannojirimycin (dMM), an inhibitor of N-glycan processing, we found that this cellular model mimics what occurs in CDG-Ia fibroblasts. Since an inverse relationship between GP expression and GSL content does exist, we assume that increased glycosphingolipid biosynthesis is secondary to protein hypoglycosylation. Altogether, our data suggest that the cell metabolic machinery may be able to partially re-equilibrate protein hypoglycosylation with increased biosynthesis of glycosphingolipids, possibly to preserve the overall physico-chemical equilibrium of the outer layer of the plasma membrane
Photoemission study of SiC films grown on Si wafers by using C-60 precursors
No full textOne possible way to grow crystalline silicon carbide (SiC) on Si (100) is to deposit fullerene molecules followed by an annealing at temperatures higher than 800 degrees C, where the C-60 molecule breaks. In this paper we present the photoemission study results on these films. X-ray photoelectron spectroscopy (XPS) and ultraviolet photoelectron spectroscopy (UPS) show the SiC formation and evolution, when the annealing temperature is varied between 800 degrees C and 850 degrees C. Crystalline and relaxed films were obtained by annealing at 850 degrees C. For these films, the valence band spectra given by UPS measurements show similar features as for crystalline SiC bulk
SiC formation on Si(100) via C60 precursors
No full textThe interaction between C-60 molecules and the Si(100) surface and the preparation of silicon-carbide thin films by thermal reaction of C-60 molecules with the Si(100) surface have been investigated using X-ray photoelectron spectroscopy, ultraviolet photoelectron spectroscopy, reflection high-energy electron diffraction and atomic force microscopy measurements. The effects of annealing temperature and C-60 coverage on the SiC formation will be discussed. It is found that the C-60 molecules bond covalently with silicon, and the number of bonds increase upon increasing the annealing temperature. Annealing at T greater than or equal to 830 degrees C entails the formation of stoichiometric silicon carbide clusters that coalesce to form a continuous SiC layer when the C-60 coverage is greater than one monolayer. Deep pits acting as silicon diffusion channels are present with dimensions that increase with the amounts of C-60. The interaction of C-60 With the SiC surface was also investigated. It is found that a similar covalent interaction is present in the two systems C-60/Si and C-60/SiC. (C) 2000 Elsevier Science B.V. All rights reserved
Carnitine reduces the lipoperoxidative damage of the membrane and apoptosis after induction of cell stress in experimental glaucoma
No full textThe pathological damage caused by glaucoma is associated to a high intraocular pressure. The ocular hypertone is most likely due to a defective efflux of aqueous humor from the anterior chamber of the eye. Ocular hypertension causes apoptotic death of retinal ganglion cells and overexpression of molecular markers typical of cell stress response and apoptosis. In this work, we report on the neuroprotective, antiapoptotic and antioxidant action of a natural substance, L-carnitine. This compound is known for its ability to improve the mitochondrial performance. We analyze a number of cellular and molecular markers, typical of ocular hypertension and, in general, of the cell stress response. In particular, L-carnitine reduces the expression of glial fibrillary acidic protein, inducible nitric oxide synthase, ubiquitin and caspase 3 typical markers of cell stress. In addition, the morphological analysis of the optic nerve evidenced a reduction of the pathological excavation of the optic disk. This experimental hypertone protocol induces a severe lipoperoxidation, which is significantly reduced by L-carnitine. The overall interpretation is that mortality of the retinal cells is due to membrane damage. Cell Death and Disease (2010) 1, e62; doi: 10.1038/cddis.2010.40; published online 5 August 201
In vitro exploration of probiotic bacteria interactions with candida using culture techniques to model dysbiotic conditions in colonized tissues
Get PDFCandida albicans overgrowth at various mucosal sites is an ongoing and complex clinical concern involving interactions with indigenous microbiota and therapeutic or preventive measures superimposed on the pathogen-microbiome interaction. In this paper we describe the use of quantitative flow cytometry (specific to the cytometer’s sample introduction mechanism) to explore the in vitro interaction between Candida albicans, probiotic lactobacilli and a topical vaginal therapeutic. Our central hypothesis was cytometric measurements of co-cultures of yeast and bacteria could provide a useful method for exploring the dynamics of different microbial species in culture, with and without inhibitors. Two commercial products were used as exemplars for this research, a vaginal antimicrobial gel and two species of probiotic lactobacillus intended or oral administration with crystalline bovine lactoferrin to augment the vaginal gel. The cytometer forward channel height parameter distinguished yeast from bacteria in co-culture experiments in the presence of a vaginal therapeutic gel or components of its formulation including EDTA, glycogen, polydextrose as well as the host defense factor, lactoferrin. Flow cytometry showed lactobacilli influenced yeast counts in co-culture, with the technique lending itself to wide-ranging test conditions including organisms, media composition and screening of various antimicrobials. Key findings: The proprietary vaginal gel augmented the effect of lactobacilli, as did EDTA and lactoferrin. Prebiotic compounds also enhanced Candida inhibition by lactobacilli. Propidium iodide (Fluorescence channel 3) discriminated between necrotic and non-necrotic yeast and bacteria in co-cultures under various culture conditions. This research demonstrates the value of flow cytometry to evaluate the population dynamics of yeast and bacteria in co-culture using a proprietary product and its components. We discuss both the limitations of the current study and describe how methods employed here would be transferrable to the investigation of organisms present in defined cultures or at body sites colonized by fungal species and the effects of therapeutics or probiotics on Candida
ROLE OF THE MATRIX METALLO-PROTEINASES IN THE CELLULAR RE-MODELLING IN A GLAUCOMA MODEL SYSTEM IN RAT.
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