Percorrer por autor "Guerreiro, Filipa"
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- Dual antibiotherapy of tuberculosis mediated by inhalable locust bean gum microparticlesPublication . Rodrigues, Susana; Alves, Ana D.; Cavaco, Joana S.; Pontes, Jorge Filipe; Guerreiro, Filipa; Rosa Da Costa, Ana; Buttini, Francesca; Grenha, AnaDespite the existence of effective oral therapy, tuberculosis remains a deadly pathology, namely because of bacterial resistance and incompliance with treatments. Establishing alternative therapeutic approaches is urgently needed and inhalable therapy has a great potential in this regard. As pathogenic bacteria are hosted by alveolar macrophages, the co-localisation of antitubercular drugs and pathogens is thus potentiated by this strategy. This work proposes inhalable therapy of pulmonary tuberculosis mediated by a single locust bean gum (LBG) formulation of microparticles associating both isoniazid and rifabutin, complying with requisites of the World Health Organisation of combined therapy. Microparticles were produced by spray-drying, at LBG/INH/RFB mass ratio of 10/1/0.5. The aerodynamic characterisation of microparticles revealed emitted doses of more than 90% and fine particle fraction of 38%, thus indicating the adequacy of the system to reach the respiratory lung area, thus partially the alveolar region. Cytotoxicity results indicate moderate toxicity (cell viability around 60%), with a concentration-dependent effect. Additionally, rat alveolar macrophages evidenced preferential capture of LBG microparticles, possibly due to chemical composition comprising mannose and galactose units that are specifically recognised by macrophage surface receptors. (C) 2017 Elsevier B.V. All rights reserved.
- Engineering of konjac glucomannan into respirable microparticles for delivery of antitubercular drugsPublication . Guerreiro, Filipa; Swedrowska, Magda; Patel, Roshnee; Floréz- Fernández, Noelia; Torres, María Dolores; Rosa Da Costa, Ana M.; Forbes, Ben; Grenha, AnaFew medically-approved excipients are available for formulation strategies to endow microcarriers with improved performance in lung drug targeting. Konjac glucomannan (KGM) is a novel, biocompatible material, comprising mannose units potentially inducing macrophage uptake for the treatment of macrophage-mediated diseases. This work investigated spray-dried KGM microparticles as inhalable carriers of model antitubercular drugs, isoniazid (INH) and rifabutin (RFB). The polymer was characterised and different polymer/drug ratios tested in the production of microparticles for which respirability was assessed in vitro. The swelling of KGM microparticles and release of drugs in simulated lung fluid were characterised and the biodegradability in presence of beta-mannosidase, a lung hydrolase, determined. KGM microparticles were drug loaded with 66-91% association efficiency and had aerodynamic diameter around 3 mu m, which enables deep lung penetration. The microparticles swelled upon liquid contact by 40-50% but underwent size reduction (>62% in 90 min) in presence of beta-mannosidase, indicating biodegradability. Finally, drug release was tested showing slower release of RFB compared with INH but complete release of both within 24 h. This work identifies KGM as a biodegradable polymer of natural origin that can be engineered to encapsulate and release drugs in respirable microparticles with physical and chemical macrophage-targeting properties.
- Fucoidan from Fucus Vesiculosus: Evaluation of the impact of the Sulphate content on Nanoparticle production and Cell toxicityPublication . FLÓREZ-FERNÁNDEZ, NOELIA; Pontes, Jorge Filipe; Guerreiro, Filipa; Afonso, Inês T.; Lollo, Giovanna; Torres, Maria Dolores; Domínguez, Herminia; M Rosa, Ana; Grenha, AnaThe composition of seaweeds is complex, with vitamins, phenolic compounds, minerals, and polysaccharides being some of the factions comprising their structure. The main polysaccharide in brown seaweeds is fucoidan, and several biological activities have been associated with its structure. Chitosan is another marine biopolymer that is very popular in the biomedical field, owing to its suitable features for formulating drug delivery systems and, particularly, particulate systems. In this work, the ability of fucoidan to produce nanoparticles was evaluated, testing different amounts of a polymer and using chitosan as a counterion. Nanoparticles of 200–300 nm were obtained when fucoidan prevailed in the formulation, which also resulted in negatively charged nanoparticles. Adjusting the pH of the reaction media to 4 did not affect the physicochemical characteristics of the nanoparticles. The IC50 of fucoidan was determined, in both HCT−116 and A549 cells, to be around 160 µg/mL, whereas it raised to 675–100 µg/mL when nanoparticles (fucoidan/chitosan = 2/1, w/w) were tested. These marine materials (fucoidan and chitosan) provided features suitable to formulate polymeric nanoparticles to use in biomedical applications.
- Hydrothermal processing of laminaria ochroleuca for the production of crude extracts used to formulate polymeric nanoparticlesPublication . Flórez-Fernández, Noelia; Álvarez-Viñas, Milena; Guerreiro, Filipa; Torres, María Dolores; Grenha, Ana; Domínguez, HerminiaA green extraction process using only water was proposed for the simultaneous extraction of alginate and bioactive compounds from Laminaria ochroleuca. Operation was carried out during non-isothermal heating up to maximal temperatures over the range of 70 °C to 100 °C. Once separated, the alginate and the crude extract were characterised and the biological activities and cytotoxicity of the extracts was studied, the latter in intestinal epithelial cells. Those alginates obtained at 90 °C exhibited the highest extraction yields and viscoelastic features of the corresponding hydrogels. The obtained results show that the extracts obtained by non-isothermal extraction were suitable to formulate nanoparticles, which showed the smallest size (≈ 250-350 nm) when the higher content of fucoidan extract was present. Given the evidenced properties, the extracts may find an application in the formulation of nanoparticulate carriers for drug delivery.
- In vitro behaviour of konjac glucomannan microparticles aimed at pulmonary tuberculosis therapyPublication . Guerreiro, Filipa; Swedrowska, Magda; Rosa Da Costa, Ana; Grenha, Ana; Forbes, BenTuberculosis is one of the highest causes of death worldwide. Long periods of treatment are required and result in some cases in therapeutic incompliance, potentiating the development of multidrug-resistant tuberculosis. Thus, new approaches to treat the infection are required as an alternative to conventional orally administered treatment. This work proposes the development of an inhalable system, which is specifically targeted to alveolar macrophages, where the Mycobacterium tuberculosis is located. The targeting is mainly driven by the selected matrix material, konjac glucomannan (KGM), a natural polymer comprising mannan units that are expected to potentiate phagocytosis. Microparticles were loaded with two antitubercular drugs, isoniazid (INH) and rifabutin (RFB). KGM/INH/RFB microparticles were produced by spray-drying to produce particles with suitable characteristics to deliver INH and RFB to the alveolar region. The KGM/INH/RFB microparticles possessed an aerodynamic diameter of approximately 3 µm, meeting the requirement of a therapy targeted to alveolar macrophages. Moreover, KGM microparticles exhibited suitable geometric size (2.24 µm) and shape (spherical) to be phagocytosed to deliver drugs to infected macrophages. INH and RFB were associated with KGM microparticles with efficiencies of 91% and 74%, respectively. Similar in vitro release profiles were observed for both drugs in simulated lung fluid (SLF) replicating the lining fluid composition found in human alveoli.
- Inhalable antitubercular therapy mediated by locust bean gum microparticlesPublication . Alves, Ana; Cavaco, Joana; Guerreiro, Filipa; Lourenço, J. P.; Costa, Ana M. Rosa da; Grenha, AnaTuberculosis remains a major global health problem and alternative therapeutic approaches are needed. Considering the high prevalence of lung tuberculosis (80% of cases), the pulmonary delivery of antitubercular drugs in a carrier system capable of reaching the alveoli, being recognised and phagocytosed by alveolar macrophages (mycobacterium hosts), would be a significant improvement to current oral drug regimens. Locust bean gum (LBG) is a polysaccharide composed of galactose and mannose residues, which may favour specific recognition by macrophages and potentiate phagocytosis. LBG microparticles produced by spray-drying are reported herein for the first time, incorporating either isoniazid or rifabutin, first-line antitubercular drugs (association efficiencies >82%). Microparticles have adequate theoretical properties for deep lung delivery (aerodynamic diameters between 1.15 and 1.67 μm). The cytotoxic evaluation in lung epithelial cells (A549 cells) and macrophages (THP-1 cells) revealed a toxic effect from rifabutin-loaded microparticles at the highest concentrations, but we may consider that these were very high comparing with in vivo conditions. LBG microparticles further evidenced strong ability to be captured by macrophages (percentage of phagocytosis >94%). Overall, the obtained data indicated the potential of the proposed system for tuberculosis therapy.
- Inhalable locust bean gum microparticles co-associating isoniazid and rifabutin: therapeutic assessment in a murine model of tuberculosis infectionPublication . Grenha, Ana; Alves, Ana D.; Guerreiro, Filipa; Pinho, Jacinta; Simões, Sandra; Almeida, António José; Gaspar, Maria ManuelaTuberculosis is a leading cause of death worldwide. Although the development of new antimycobacterial drugs is an obvious and necessary strategy to address the disease, improving the therapeutic performance of drugs already approved constitutes a valuable alternative approach. As the lung is the most affected organ, where M. tuberculosis is able to survive and proliferate, the direct pulmonary delivery of antitubercular drugs comprises a highly promising therapeutic strategy. In this work, spray-dried locust bean gum (LBG) microparticles were used to deliver a combination of two first line antitubercular drugs, isoniazid (INH) and rifabutin (RFB), to the alveolar zone, where macrophages hosting the bacteria reside. LBG is expected to mediate favoured macrophage uptake of microparticles, leading to enhanced therapeutic effect. The therapeutic effect of LBG/INH/RFB microparticles was evaluated in a murine model infected with M. tuberculosis, strain H37Rv and compared with oral co-therapy of INH and RFB in the free form. The pulmonary administration of LBG/INH/RFB microparticles 5 times per week was the only treatment schedule that provided negative growth index values in lung (-0.22), spleen (-0.14) and liver (-0.26) even using a lower therapeutic dose for both antibiotics. For the control group, the respective values were +1.95, +0.75 and +0.96.
- Locust bean gum (LBG) – a potential excipient for inhalation purposes: excipient characterisation and in vitro and in vivo toxicological evaluationPublication . Pontes, Jorge Filipe; Guerreiro, Filipa; da Silva, Joana Pinto; Almeida, Maria; Rosso, Annalisa; Rosa da Costa, Ana M; Agusti, Géraldine; Lollo, Giovanna; Gaspar, Maria Manuela; Grenha, AnaDry powders proposed for lung drug delivery typically involve excipients not approved for inhalation. The physicochemical characterisation of excipients is informative regarding their interaction with different body structures. Locust Bean Gum (LBG) has been proposed for diverse applications, including inhalation, benefiting from targeting ability towards phagocytic cells owing to the presence of mannose moieties. The aim of the present study was to characterise the physicochemical parameters of LBG and, for the first time, draw a toxicological profile for this excipient. LBG from three different suppliers was evaluated and eventual changes on polymer characteristics induced by purification and microparticle production were assessed, which were observed to not occur. The commercial samples showed identical FTIR spectra, as well as TGA and DSC profiles, Mw around 3.60 × 106 Da, and Tg near − 39 ◦C. All microparticles presented similar morphology and Feret diameters around 4 μm. In vitro assays performed on respiratory (A549) cells evidenced no impact of LBG microparticles on cell viability (> 80 %) when biorelevant concentrations (≈ 0.1 mg/mL) were used. Additionally, inhalation of LBG microparticles by mice provided indications of a safe profile, without induction of allergic reactions. The basis is laid for further exploration of this material in inhalation.
- Microencapsulation of selenium by spray-drying as a tool to improve bioaccessibility in food matrixPublication . Grenha, Ana; Guerreiro, Filipa; Lourenço, João P.; Lopes, João Almeida; Cámara-Martos, FernandoSe in the form of sodium selenite was microencapsulated by spray - drying and added to a food matrix (yogurt) to study the potential improvement of its bioaccessibility. Yogurt samples were also supplemented with Se in free salt form. Se-loaded microparticles were successfully prepared by spray-drying with production yields above 70%. The supplementation of yogurt with Se in the form of free sodium selenite had a low effect on improving the bioaccessibility of this micronutrient (1%). In turn, Se microencapsulation with mannitol or mannitol/gastro-resistant polymer (Eudragit (R)) had a strong impact on bioaccessibility results. After the gastric phase, Se bio-accessibility reached values of 21 and 40% for the microencapsulated formulations, respectively. This percentage rose to 55% at the end of intestinal phase, showing no differences between both formulations. Our results show the relevance of microencapsulation as an effective tool to improve the bioaccessibility of micronutrients when they are used in food supplementation.
- Nanocomposite sponges for enhancing intestinal residence time following oral administrationPublication . Rosso, Annalisa; Andretto, Valentina; Chevalier, Yves; Kryza, David; Sidi-Boumedine, Jacqueline; Grenha, Ana; Guerreiro, Filipa; Gharsallaoui, Adem; La Padula, Veronica; Montembault, Alexandra; David, Laurent; Briançon, Stéphanie; Lollo, GiovannaIn this work, nanocomposites that combine mucopenetrating and mucoadhesive properties in a single system are proposed as innovative strategy to increase drug residence time in the intestine following oral administration. To this aim, novel mucoadhesive chitosan (CH) sponges loaded with mucopenetrating nanoemulsions (NE) were developed via freeze-casting technique. The NE mucopenetration ability was determined studying the surface affinity and thermodynamic binding of the nanosystem with mucins. The ability of nanoparticles to penetrate across a preformed mucins layer was validated by 3D-time laps Confocal Laser Scanning Microscopy imaging. Microscopy observations (Scanning Electron Microscopy and Optical Microscopy) showed that NE participated in the structure of the sponge affecting its stability and in vitro release kinetics. When incubated with HCT 116 and Caco-2 cell lines, the NE proved to be cytocompatible over a wide concentration range. Finally, the in vivo biodistribution of the nanocomposite was evaluated after oral gavage in healthy mice. The intestinal retention of NE was highly enhanced when loaded in the sponge compared to the NE suspension. Overall, our results demonstrated that the developed nanocomposite sponge is a promising system for sustained drug intestinal delivery.