Navegando por Autor "Brambilla, Isabelle de Paula Ribeiro"
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Item Assessment of the Effect of Antimicrobial Photodynamic Therapy on Multidrug- Resistant Acinetobacter baumannii Strain Biofilm - an in Vitro StudyBrambilla, Isabelle de Paula Ribeiro; Marcolino, Luciana Maria Cortez; Pinto, Juliana Guerra; Ferreira-Strixino, Juliana; Ribeirão PretoAcinetobacter baumannii is an opportunistic bacterium associated with numerous cases of hospital-acquired infections and possesses, among other characteristics that make it virulent, resistance to various antibiotics. The increasing resistance of microorganisms has led to the search for alternative methods, such as Antimicrobial Photodynamic Therapy (aPDT), inducing bacterial cell death. This study aimed to evaluate the antimicrobial action of PDT with curcumin on A. baumannii biofilm in vitro. A. baumannii biofilm was treated with curcumin for 20 min and then irradiated with a 450 nm LED (Light Emitting Diode) at 93.9 mW/cm2, with a fluence of 42.72 J/cm2. Subsequently, tests were conducted using Confocal Microscopy with the Resazurin dye, Colony-Forming Units (CFU) counting, and Scanning Electron Microscopy (SEM) imaging. Cell viability tests were also conducted using the L929 fibroblast cell line. The results show that although PDT with curcumin did not reduce bacterial growth, it caused alterations in cell metabolism. Additionally, confocal microscopy showed that curcumin was internalized by A. baumannii at the concentrations used. SEM demonstrated structural changes, although no bacterial reduction in the biofilm was observed. The parameters used in biofilm treatment were cytotoxic to L929 cells. This study showed that PDT with curcumin induces fragility in A. baumannii cells, suggesting that a possible combination with antibiotics may lead to the elimination of bacterial infection.Item Curcumin-loaded bovine serum albumin (BSA) nanoparticles: photoreactivity and photodynamic action against Acinetobacter baumannii and Staphylococcus aureus(Royal Society of Chemistry) Pinto, Juliana Guerra; Laneri, Francesca; Brambilla, Isabelle de Paula Ribeiro; Azevedo, Maria Clara Barbosa Silva de; Ambrósio, Jéssica Aparecida Ribeiro; Simioni, Andreza Ribeiro; Fraix, Aurore; Sortino, Salvatore; Ferreira-Strixino, JulianaCurcumin (CUR) is a naturally occurring pigment, poorly soluble in water and an object of intense interest due to its multifaceted therapeutic and phototherapeutic activity. In this contribution, bovine serum albumin nanoparticles (BSA-NPs) with oppositely charged surfaces have been used as suitable nanocarriers both to overcome the poor water solubility of CUR and to encourage its interaction with Gram-positive and Gram-negative bacteria. The different surface charges of the BSA-NPs do not affect either the spectroscopic or the photochemical behaviour of the encapsulated CUR which in all cases is almost exclusively present in a diketo form rather than the enolic one. This strictly dictates the response of CUR to blue light excitation which, under these conditions, exhibits the usual reactivity of carbonyl compounds toward molecules that behave as hydrogen donors such as the BSA-NP components. Steady-state and time-resolved photochemical experiments show that the encapsulated CUR under- goes photodecomposition with rate basically independent of the presence of oxygen. The photo- decomposition seems to be mediated mainly by an intermolecular H-abstraction from the components of the nanocarrier by the lowest excited triplet state of CUR with the formation of the corresponding ketyl radical occurring in less than 0.5 ms. This radical is oxidized by molecular oxygen, likely leading to peroxyl and hydroperoxyl radical species, probably responsible for the photodynamic action. Accordingly, with the short triplet lifetime, no singlet oxygen photogeneration is observed. Concentrations of 0.2, 0.3 and 0.4 mM were selected for in vitro tests. Internalization of both formulations was observed in strains of A. baumannii and S. aureus, after 15 minutes of interaction. After irradiation, a reduction in the viability of bacteria was observed, at the different concentrations tested, with concentrations of 0.4 and 0.3 mM being the most efficient.Item Simultaneous photoactivation of a fluoroquinolone antibiotic and nitric oxide with fluorescence reporting(Royal Society of Chemistry) Martins, Tassia Joi; Parisi, Cristina; Pinto, Juliana Guerra; Brambilla, Isabelle de Paula Ribeiro; Melilli, Barbara; Aleo, Danilo; Ferreira-Strixino, Juliana; Sortino, SalvatoreThe achievement of smart pharmaceuticals whose bioactivity can be spatiotemporally controlled by light stimuli is known as photopharmacology, an emerging area aimed at improving the therapeutic outcome and minimizing side effects. This is especially attractive for antibiotics, for which the inevitable development of multidrug resistance and the dwindling of new clinically approved drugs represent the main drawbacks. Here, we show that nitrosation of the fluoroquinolone norfloxacin (NF), a broad- spectrum antibiotic, leads to the nitrosated bioconjugate NF–NO, which is inactive at the typical minimum inhibitory concentration of NF. Irradiation of NF–NO with visible blue light triggers the simul- taneous release of NF and nitric oxide (NO). The photouncaging process is accompanied by the revival of the typical fluorescence emission of NF, quenched in NF–NO, which acts as an optical reporter. This permits the real-time monitoring of the photouncaging process, even within bacteria cells where antibacterial activity is switched on exclusively upon light irradiation. The mechanism of photorelease seems to occur through a two-step hopping electron transfer mediated by the lowest triplet state of NF–NO and the phosphate buffer ions or aminoacids such as tyrosine. Considering the well-known role of NO as an ‘‘unconventional’’ antibacterial, the NF–NO conjugate may represent a potential bimodal antibacterial weapon activatable on demand with high spatio-temporal control.Item Stepwise Nitric Oxide Release and Antibacterial Activity of a Nitric Oxide Photodonor Hosted within Cyclodextrin Branched Polymer Nanocarriers(ACS Publications) Martins, Tassia Joi; Parisi, Cristina; Pinto, Juliana Guerra; Brambilla, Isabelle de Paula Ribeiro; Malanga, Milo; Ferreira-Strixino, Juliana; Sortino, SalvatoreA hydrophobic nitric oxide (NO) photodonor integrating both nitroso and nitro functionalities within its chromophoric skeleton has been synthesized. Excitation of this compound with blue light triggers the release of two NO molecules from the nitroso and the nitro functionalities via a stepwise mechanism. Encapsulation of the NO photodonor within biocompatible neutral, cationic, and anionic β-cyclodextrin branched polymers as suitable carriers leads to supramolecular nanoassemblies, which exhibit the same nature of the photochemical processes but NO photorelease performances enhanced by about 1 order of magnitude when compared with the free guest. Antibacterial tests carried out with methicillin-resistant Staphylococcus aureus and Acinetobacter baumannii demonstrate an effective antibacterial activity exclusively under light activation and point out a differentiated role of the polymeric nanocarriers in determining the outcome of the antibacterial photodynamic action.