scholarly journals Proteomics profiles of Cronobacter sakazakii and a fliF mutant: Adherence and invasion in mouse neuroblastoma cells

2020 ◽  
Vol 149 ◽  
pp. 104595 ◽  
Author(s):  
Esteban-Kenel Veronica ◽  
Ochoa Sara A ◽  
Curiel-Quesada Everardo ◽  
Quezada Héctor ◽  
Medina-Contreras Oscar ◽  
...  
Keyword(s):  
Neuroblastoma Cells ◽  
Cronobacter Sakazakii ◽  
Mouse Neuroblastoma

scholarly journals Proteomics Profile of Cronobacter sakazakii and fliF Mutant: Adherence and Invasion to Mouse Neuroblastoma Cells

2019 ◽  
Author(s):  
Veronica Esteban-‬Kenel ◽  
Everardo Curiel-Quesada ◽  
Héctor Quezada ◽  
Oscar Medina Contreras ◽  
Víctor Luna-Pineda ◽  
...  
Keyword(s):  
Intracellular Trafficking ◽  
Neuroblastoma Cells ◽  
Foodborne Pathogen ◽  
Lipid Transport ◽  
Cronobacter Sakazakii ◽  
Differential Protein Expression ◽  
Mouse Neuroblastoma ◽  
Carbohydrate Transport ◽  
Proteomic Study ◽  
Transduction Mechanisms

Abstract Background: Cronobacter sakazakii is an opportunistic foodborne pathogen associated with necrotizing enterocolitis, bacteraemia, and meningitis in infants. A comparative proteomic study of C . sakazakii ATCC BAA-894 (CS WT) and isogenic mutant of flagella were performed; including the ability of both strains to adhere and invade to N1E-115 cells. Results: To achieve this goal, a non-motile C. sakazakii ATCC BAA-894 fliF ::Tn5 (CS fliF ::Tn5) strain was generated using an EZ-Tn5Tnp Transposome kit. Analysis of differential protein expression showed that 81.49% (361/443) of the proteins were identified in both strains, 8.35% (37/443) were exclusively expressed in the CS WT strain and 10.16% (45/443) in the CS fliF ::Tn5 strain. The main exclusive proteins from the CS WT strain were classified into the following subcategories: “cell motility” and “signal transduction mechanisms”. In contrast, the exclusive proteins from the CS fliF ::Tn5 strain were classified into the following subcategories: “intracellular trafficking, secretion, and vesicular transport”, “replication, recombination, and repair”, “nucleotide transport and metabolism”, “carbohydrate transport and metabolism”, “coenzyme transport and metabolism”, and “lipid transport and metabolism”. Expression of the Cpa protein was shared by both strains but was more abundant in the CS WT strain than in the CS fliF ::Tn5 strain. Significant increases (p=0.0001) in adherence to N1E-115 cells were observed for the non-motile CS fliF ::Tn5 strain, with 31.3×10 6 CFU/mL, relative to the CS WT strain, with 14.5×10 6 CFU/mL. Additionally, for infection of N1E-115 cells, the CS WT strain showed a 0.17% invasion frequency, which was significantly increased (p=0.01) compared to that of the non-motile CS fliF ::Tn5 strain. Conclusion : The proteins involved in motility were mainly identified by proteomic analysis in CS WT strain when compare to CS fliF ::Tn5 strain. Our data showed that flagella are required to promote invasion to N1E-115 cells and absence of flagella significantly increases the adherence to N1E-115 cells when compare to CS WT strain.

On the Application of Cultured Neuronal Cell Lines in Neurotoxicological Studies: Cytotoxicity of Acrylamide

1983 ◽  
Vol 11 (3) ◽  
pp. 135-145
Author(s):  
Erik Walum
Keyword(s):  
Cell Lines ◽  
Culture Medium ◽  
Neuroblastoma Cells ◽  
Neuronal Cell ◽  
Leucine Incorporation ◽  
Biochemical Process ◽  
Neurite Retraction ◽  
Mouse Neuroblastoma ◽  
Neuronal Cell Lines

Summary Acrylamide, a well known neurotoxic compound, was used in a first evaluation of cultured mouse neuroblastoma cells as an alternative to animal models for neurotoxicological studies. Hence, the effects of acrylamide on the growth, size, morphology and leucine incorporation of three neuroblastoma (41A3, N18 and N1E115), one neuroblastoma x glioma hybrid (NG108CC15), two glioma (138MG and C6) and two fibroblast (RLF and RMC) cell lines were studied. It was found that the concentration of acrylamide needed to inhibit the growth by 50% in 24 hr was similar in all cell lines, i.e. around 2 x 10-4g/ml culture medium. In the two cell lines, N1E115 and NG108CC15, acrylamide at this concentration caused neurite retraction and at higher concentrations (5 x 10-4g/ml) a decrease in cell viability. In a concentration range of 5 x 10-5 - 5 x 10-4g/ml acrylamide did not affect cell size, or at 2 x 10-4g/ml incorporation of leucine into trichloroacetic acid precipitable material. It is suggested that acrylamide interferes with a biochemical process common to all the tested cells, but of greater importance in differentiated nerve cells than in others. Whether this process is consistent with the in vivo target for the neurotoxic action of acrylamide remains to be unravelled.

Ultrastructure of multiple microtubule initiation sites in mouse neuroblastoma cells

1981 ◽  
Vol 47 (1) ◽  
pp. 1-24
Author(s):  
G.A. Sharp ◽  
M. Osborn ◽  
K. Weber
Keyword(s):  
Electron Microscopy ◽  
Electron Microscope ◽  
Immunofluorescence Microscopy ◽  
Neuroblastoma Cells ◽  
Biological Significance ◽  
Optimal Conditions ◽  
Serial Sectioning ◽  
Mouse Neuroblastoma ◽  
Microtubule Organizing Centres ◽  
Perinuclear Area

Morphologically undifferentiated and differentiated mouse neuroblastoma N115 and N18 cells were examined after serial sectioning by electron microscopy. A sizeable percentage of the cells revealed multiple centrioles, usually clustered together in the perinuclear area with 2 preferential locations, i.e. above and below the largest nuclear diameter. These results indicate that the multiple microtubule-organizing centres previously visualized by immunofluorescence microscopy with tubulin antibody in neuroblastoma cells recovering from Colcemid poisoning are most likely in majority related to multiple centrioles. This interpretation is further strengthened by experiments in which cells are first recorded in the fluorescence microscope and then after serial sectioning in the electron microscope. The results show that under optimal conditions immunofluorescence microscopy is able to visualize single centrioles. The possible biological significance of the combined electron and immunofluorescence microscopical results is discussed.

scholarly journals Modulation of calcium signaling depends on the oligosaccharide of GM1 in Neuro2a mouse neuroblastoma cells

2020 ◽  
Vol 37 (6) ◽  
pp. 713-727
Author(s):  
Giulia Lunghi ◽  
Maria Fazzari ◽  
Erika Di Biase ◽  
Laura Mauri ◽  
Sandro Sonnino ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Calcium Channels ◽  
Calcium Signaling ◽  
Intracellular Calcium ◽  
Calcium Imaging ◽  
Neuroblastoma Cells ◽  
Ganglioside Gm1 ◽  
Mouse Neuroblastoma ◽  
Neuro2a Cells ◽  
Neuronal Functions

AbstractRecently, we demonstrated that the oligosaccharide portion of ganglioside GM1 is responsible, via direct interaction and activation of the TrkA pathway, for the ability of GM1 to promote neuritogenesis and to confer neuroprotection in Neuro2a mouse neuroblastoma cells. Recalling the knowledge that ganglioside GM1 modulates calcium channels activity, thus regulating the cytosolic calcium concentration necessary for neuronal functions, we investigated if the GM1-oligosaccharide would be able to overlap the GM1 properties in the regulation of calcium signaling, excluding a specific role played by the ceramide moiety inserted into the external layer of plasma membrane. We observed, by calcium imaging, that GM1-oligosaccharide administration to undifferentiated Neuro2a cells resulted in an increased calcium influx, which turned out to be mediated by the activation of TrkA receptor. The biochemical analysis demonstrated that PLCγ and PKC activation follows the TrkA stimulation by GM1-oligosaccharide, leading to the opening of calcium channels both on the plasma membrane and on intracellular storages, as confirmed by calcium imaging experiments performed with IP3 receptor inhibitor. Subsequently, we found that neurite elongation in Neuro2a cells was blocked by subtoxic administration of extracellular and intracellular calcium chelators, suggesting that the increase of intracellular calcium is responsible of GM1-oligosaccharide mediated differentiation. These results suggest that GM1-oligosaccharide is responsible for the regulation of calcium signaling and homeostasis at the base of the neuronal functions mediated by plasma membrane GM1.

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