1,721,084 research outputs found
Infrared microspectroscopy of live cells in microfluidic devices (MD-IRMS): Toward a powerful label-free cell-based assay
No full textUntil nowadays most infrared microspectroscopy (IRMS) experiments on biological specimens (i.e., tissues or cells) have been routinely carried out on fixed or dried samples in order to circumvent water absorption problems. In this paper, we demonstrate the possibility to widen the range of in-vitro IRMS experiments to vibrational analysis of live cellular samples, thanks to the development of novel biocompatible IR-visible transparent microfluidic devices (MD). In order to highlight the biological relevance of IRMS in MD (MD-IRMS), we performed a systematic exploration of the biochemical alterations induced by different fixation protocols, ethanol 70% and formaldehyde solution 4%, as well as air-drying on U937 leukemic monocytes by comparing their IR vibrational features with the live U937 counterpart. Both fixation and air-drying procedures affected lipid composition and order as well as protein structure at a different extent while they both induced structural alterations in nucleic acids. Therefore, only IRMS of live cells can provide reliable information on both DNA and RNA structure and on their cellular dynamic. In summary, we show that MD-IRMS of live cells is feasible, reliable, and biologically relevant to be recognized as a label-free cell-based assay
Effects of prolonged treatment with decarbazine on tumor metastatic potentialin mice bearing Lewis lung carcinoma.
No full textAnalysis of the cytotoxicity of synthetic antimicrobial peptides on mouse leucocytes: implications for systemic use.
No full textWe have analysed the toxicity of highly cationic, artificial alpha-helical antimicrobial peptides on blood cells to assess their suitability for systemic application. Flow cytometric methods, based on the uptake of propidium iodide, were used to obtain a rapid and quantitative estimate of membrane damage to resting and concanavalin A-activated mouse lymphocytes, which was further confirmed by morphological changes as observed by scanning electron microscopy. Membrane permeabilization appeared to correlate with structural characteristics, so that the peptide L-19(9/B), which contains helix-stabilizing aminoisobutyric acid (Aib) residues and is a potent antimicrobial, was also the most lytic towards both mouse lymphocytes and human erythrocytes. Reducing the propensity for helix formation in P19(8) resulted in a marked reduction in in vitro cytotoxicity. Changing the helical sense in D-P19(9/B) also resulted in a significant decrease in cytolytic activity towards both erythrocytes and leucocytes. A limited assessment in BALB/c mice confirmed a lower in vivo toxicity of P19(8) than L-P19(9/B). In a study of the systemic antimycotic activity of P19(8) in a mouse protection model, a modest prolongation in survival of Candida albicans-infected animals after intravenous administration was observed at 5 mg/kg peptide but not at higher doses. The implications of these observations for the systemic use of this type of peptide are discussed.
PMID:12205058[PubMed - indexed for MEDLINE] Free full tex
The marine toxin palytoxin induces necrotic death in HaCaT cells through a rapid mitochondrial damage
No full textPalytoxin (PLTX) is one of the most toxic algal biotoxin known so far. It transforms the Na+/K+-ATPase into a cationic channel inducing a massive intracellular Na+ influx. However, from a mechanistic point of view, the features and the intracellular pathways leading to PLTX-induced cell death are still not completely characterized. This study on skin HaCaT keratinocytes demonstrates that PLTX induces necrosis since propidium iodide uptake was observed already after 1 h toxin exposure, an effect that was not lowered by toxin removal. Furthermore, necrotic-like morphological alterations were evidenced by confocal microscopy. Apoptosis occurrence was excluded since no caspases 3/7, caspase 8, and caspase 9 activation as well as no apoptotic bodies formation were recorded. Necrosis was preceded by a very early mitochondrial damage as indicated by JC-1 fluorescence shift, recorded already after 5min toxin exposure. This shift was totally abolished when Na+ and Ca2+ ions were withdrawn from culture medium, whereas cyclosporine-A was ineffective, excluding the occurrence of a controlled biochemical response. These results clearly establish necrosis as the primary mechanism for PLTX-induced cell death in HaCaT cells. The rapidity of mitochondrial damage and the consequent irreversible necrosis rise serious concerns about the very fast onset of PLTX toxic effects
Rapid and reliable detection of antimicrobial peptide penetration into Gram-negative bacteria based on fluorescence quenching.
No full textIn this paper, we describe a rapid flow cytometry method to identify antimicrobial peptides that are
internalized into bacterial cells and differentiate them from those that are membrane active. The method was
applied to fluorescently labeled Bac71-35 and polymyxin B, whose mechanisms of action are, respectively, based
on cell penetration and on membrane binding and permeabilization. Identification of peptides with the former
mechanism is of considerable interest for the intracellular delivery of membrane-impermeant drugs
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