Francesco Ragonese

Inhibitory effects of agmatine on monoamine oxidase (MAO) activity: Reconciling the discrepancies

Abstract Agmatine has been functionally characterized as an important hormone and co-neurotransmitter in mammals. Given its ability in binding Imidazoline sites, a regolatory site of monoaminoxydase, it has been suggested to be involved in many neurological aspects. However, its inhibitory effect on this enzyme still remains an unanswered question. This present study is aimed to asses whether different experimental conditions could affect the agmatine action on monoaminoxydase activity. We demonstrate that the monoaminoxydase inhibition by agmatine is obtained under alkaline conditions and a long time of incubation. No inhibitiory action was found for shorter times of reaction at elevated pH, or at neutral condition and long time of incubation. No inhibition was also detected by substituting the monoamineoxydase substrate tyramine with kynuramine, however, while in these conditions a remarkable inhibition was shown by two aminoxydase inhibitors tranylcypromine and idazoxan. Herein, we discuss a mechanism model and the functional consequences of agmatine action on monoaminoxydase.

Glioblastoma single-cell microRaman analysis under stress treatments

Glioblastoma multiforme (GBM) is the most frequent malignant brain tumor characterized by highly heterogeneous subpopulations. In order to reveal the heterogeneous cell response, single cell analysis is an essential requirement. In this study, optical microscopy and Raman microspectroscopy were used to follow the stress response of U251 single cells adherent on a silicon substrate. Cultured cells on silicon substrate were treated with hydrogen peroxide to promote apoptosis. Under these conditions expected changes occurred after a few hours and were revealed by the reduction of cytochrome c, lipid, nucleic acid and protein Raman signals: this ensured the possibility to analyse U251 cell line as grown on Si substrate, and to monitor the response of single cells to stress conditions. As a consequence, we used microRaman to monitor the effects induced by nutrient depletion: a fast change of Raman spectra showed two different sub-populations of sensible and resistant U251 cells. Furthermore, spectral variations after DMSO addition were associated to volume changes and confirmed by morphological analysis. Thus, our results highlight the sensitivity of Raman microspectroscopy to detect rapid variations of macromolecule concentration due to oxidative stress and/or cell volume changes at the single cell level.

Probing Internalization Effects and Biocompatibility of Ultrasmall Zirconium Metal-Organic Frameworks UiO-66 NP in U251 Glioblastoma Cancer Cells

The synthesis of ultrasmall UiO-66 nanoparticles (NPs) with an average size of 25 nm, determined by X-ray powder diffraction and electron microscopies analysis, is reported. The NPs were stabilized in water by dialyzing the NP from the DMF used for the synthesis. DLS measurements confirmed the presence of particles of 100 nm, which are spherical aggregates of smaller particles of 20–30 nm size. The NP have a BET surface area of 700 m2/g with an external surface area of 300 m2/g. UiO-66_N (UiO-66 nanoparticles) were loaded with acridine orange as fluorescent probe. UV-vis spectroscopy analysis revealed no acridine loss after 48 h of agitation in simulated body fluid. The biocompatibility of UiO-66_N was evaluated in human glioblastoma (GBM) cell line U251, the most malignant (IV grade of WHO classification) among brain tumors. In U251 cells, UiO-66_N are inert since they do not alter the cell cycle, the viability, migration properties, and the expression of kinases involved in cancer cell growth. The internalization process was evident after a few hours of incubation. After 24 h, UiO-66_N@Acr (UiO-66_N loaded with acridine orange) were detectable around the nuclei of the cells. These data suggest that small UiO-66 are biocompatible NP and could represent a potential carrier for drug delivery in glioblastoma therapies.

Anticarcinogenic activities of sulforaphane are influenced by Nerve Growth Factor in human melanoma A375 cells

Melanoma is a severe form of cancer, resistant to conventional therapies. According to in vitro studies, sulforaphane, a dietary component, has been considered a promising antineoplastic candidate. The present study analyzes the in vitro biological effects of sulforaphane in A375 melanoma cell line with or without the addition of Nerve Growth Factor. For the first time, our results show that a supplementation of Nerve Growth Factor partially reverses the sulforaphane-induced: i) inhibition of cell migration, ii) pro apoptotic changes in cell cycle and iii) modulation of active caspase-3. Furthermore, we report the sulforaphane-induced modulation in the expression of Nerve Growth Factor receptors TrKA and p75NTR, shifting their ratio from pro survival to pro apoptotic. In conclusion, the present study evidences that in vivo the antineoplastic effects of sulforaphane may be reduced by the contemporaneous presence of other biological elements such as Nerve Growth Factor and it contributes to a better definition of the real in vivo potentiality of sulforaphane as antineoplastic candidate.