Paula Coutinho

Thermoresponsive chitosan-agarose hydrogel for skin regeneration.

Healing enhancement and pain control are critical issues on wound management. So far, different wound dressings have been developed. Among them, hydrogels are the most applied. Herein, a thermoresponsive hydrogel was produced using chitosan (deacetylation degree 95%) and agarose. Hydrogel bactericidal activity, biocompatibility, morphology, porosity and wettability were characterized by confocal microscopy, MTS assay and SEM. The performance of the hydrogel in the wound healing process was evaluated through in vivo assays, during 21 days. The attained results revealed that hydrogel has a pore size (90-400 μm) compatible with cellular internalization and proliferation. A bactericidal activity was observed for hydrogels containing more than 188 μg/mL of chitosan. The improved healing and the lack of a reactive or a granulomatous inflammatory reaction in skin lesions treated with hydrogel demonstrate its suitability to be used in a near future as a wound dressing.

Optimization of liquid overlay technique to formulate heterogenic 3D co-cultures models

Three‐dimensional (3D) cell culture models of solid tumors are currently having a tremendous impact in the in vitro screening of candidate anti‐tumoral therapies. These 3D models provide more reliable results than those provided by standard 2D in vitro cell cultures. However, 3D manufacturing techniques need to be further optimized in order to increase the robustness of these models and provide data that can be properly correlated with the in vivo situation. Therefore, in the present study the parameters used for producing multicellular tumor spheroids (MCTS) by liquid overlay technique (LOT) were optimized in order to produce heterogeneous cellular agglomerates comprised of cancer cells and stromal cells, during long periods. Spheroids were produced under highly controlled conditions, namely: (i) agarose coatings; (ii) horizontal stirring, and (iii) a known initial cell number. The simultaneous optimization of these parameters promoted the assembly of 3D characteristic cellular organization similar to that found in the in vivo solid tumors. Such improvements in the LOT technique promoted the assembly of highly reproducible, individual 3D spheroids, with a low cost of production and that can be used for future in vitro drug screening assays. Biotechnol. Bioeng. 2014;111: 1672–1685.

Evaluation of Nanoparticle Uptake in Co-culture Cancer Models

Co-culture models are currently bridging the gap between classical cultures and in vivo animal models. Exploring this novel approach unlocks the possibility to mimic the tumor microenvironment in vitro, through the establishment of cancer-stroma synergistic interactions. Notably, these organotypic models offer a perfect platform for the development and pre-clinical evaluation of candidate nanocarriers loaded with anti-tumoral drugs in a high throughput screening mode, with lower costs and absence of ethical issues. However, this evaluation was until now limited to co-culture systems established with precise cell ratios, not addressing the natural cell heterogeneity commonly found in different tumors. Therefore, herein the multifunctional nanocarriers efficiency was characterized in various fibroblast-MCF-7 co-culture systems containing different cell ratios, in order to unravel key design parameters that influence nanocarrier performance and the therapeutic outcome. The successful establishment of the co-culture models was confirmed by the tissue-like distribution of the different cells in culture. Nanoparticles incubation in the various co-culture systems reveals that these nanocarriers possess targeting specificity for cancer cells, indicating their suitability for being used in this illness therapy. Additionally, by using different co-culture ratios, different nanoparticle uptake profiles were obtained. These findings are of crucial importance for the future design and optimization of new drug delivery systems, since their real targeting capacity must be addressed in heterogenous cell populations, such as those found in tumors.

Recent advances on antimicrobial wound dressing: A review

Skin and soft tissue infections (SSTIs) have high rates of morbidity and mortality associated. Despite the successful treatment of some SSTIs, those affecting the subcutaneous tissue, fascia, or muscle delay the healing process and can lead to life-threatening conditions. Therefore, more effective treatments are required to deal with such pathological situations. Recently, wound dressings loaded with antimicrobial agents emerged as viable options to reduce wound bacterial colonization and infection, in order to improve the healing process. In this review, an overview of the most prominent antibacterial agents incorporated in wound dressings along with their mode of action is provided. Furthermore, the recent advances in the therapeutic approaches used in the clinic and some future perspectives regarding antibacterial wound dressings are also discussed.

Synthesis and characterization of a photocrosslinkable chitosan–gelatin hydrogel aimed for tissue regeneration

In the area of tissue engineering different approaches have been studied, so far, for promoting regeneration or replacement of damaged tissues. Among the different materials developed, hydrogels, due to their biocompatibility and similarities with the native extracellular matrix, have emerged as suitable candidates for being used for different therapeutic purposes. Herein, photocrosslinkable hydrogels, composed by chitosan methacrylamide (ChMA) and gelatin methacrylamide (GelMA) were crosslinked by ultraviolet (UV) light, using Irgacue 2959 as photoinitiator. The morphological, physicochemical and biological properties of the hydrogels were characterized by scanning electron microscopy, Fourier transform infrared spectroscopy and nuclear magnetic resonance. The obtained results demonstrated that the developed hydrogels possess suitable properties for being used as 3D constructs on several areas of tissue engineering. Furthermore, these properties may allow their future application as space filling agents or as delivery vehicles of bioactive molecules and cells.

Electrospun Polycaprolactone/Aloe Vera_Chitosan Nanofibrous Asymmetric Membranes Aimed for Wound Healing Applications

Today, none of the wound dressings available on the market is fully capable of reproducing all the features of native skin. Herein, an asymmetric electrospun membrane was produced to mimic both layers of skin. It comprises a top dense layer (manufactured with polycaprolactone) that was designed to provide mechanical support to the wound and a bottom porous layer (composed of chitosan and Aloe Vera) aimed to improve the bactericidal activity of the membrane and ultimately the healing process. The results obtained revealed that the produced asymmetric membranes displayed a porosity, wettability, as well as mechanical properties similar to those presented by the native skin. Fibroblast cells were able to adhere, spread, and proliferate on the surface of the membranes and the intrinsic structure of the two layers of the membrane is capable of avoiding the invasion of microorganisms while conferring bioactive properties. Such data reveals the potential of these asymmetric membranes, in the near future, to be applied as wound dressings.

Effect of the Nutritional Status of Semi-continuous Microalgal Cultures on the Productivity and Biochemical Composition of Brachionus plicatilis

The rotifer Brachionus plicatilis was cultured using the microalga Isochrysis aff. galbana clone T-ISO as feed. T-ISO was cultured semi-continuously with daily renewal rates of 10%, 20%, 30%, 40%, and 50% of the volume of cultures. The increase of renewal rate led to increasing nutrient and light availability in microalgal cultures, which caused differences in the biochemical composition of microalgal biomass. Growth rate, individual dry weight, organic content, and biomass productivity of rotifer cultures increased in response to higher growth rate in T-ISO cultures. Rotifer growth rate showed a strong negative correlation (R2 = 0.90) with the C/N ratio of microalgal biomass. Rotifer dry weight was also affected by nutrient availability of T-ISO cultures, increasing up to 50% from nutrient-limited to nutrient-sufficient conditions. Consequently, biomass productivity of rotifer cultures increased more than twofold with the increase of renewal rate of T-ISO cultures. Rotifer organic content underwent the same trend of total dry weight. Maximum content of polyunsaturated fatty acids was reached in rotifers fed T-ISO from the renewal rate of 40%, with percentages of docosahexaenoic acid (22:6ω-3, DHA) and eicosapentaenoic acid (20:5ω-3, EPA) of 11% and 5% of total fatty acids, respectively. Selecting the most appropriate conditions for microalgal culture can therefore enhance the nutritive quality of microalgal biomass, resulting in a better performance of filter feeders and their nutrient content, and may constitute a useful tool to improve the rearing of fish larvae and other aquaculture organisms that require live feed in some or all the stages of their life cycle.

New drug-eluting lenses to be applied as bandages after keratoprosthesis implantation.

Corneal tissue is the most commonly transplanted tissue worldwide. This work aimed to develop a new drug-eluting contact lens that may be used as a bandage after keratoprosthesis. During this work, films were produced using poly(vinyl alcohol) (PVA) and chitosan (CS) crosslinked with glyoxal (GL). Vancomycin chlorhydrate (VA) was impregnated in these systems by soaking. Attenuated total reflectance - Fourier transform infrared spectroscopy was used to confirm crosslinking. The cytotoxic and drug release profile, hydrophilicity, thermal and biodegradation as well as swelling capacity of the samples were assessed through in vitro studies. PVA and PVA/CS films were obtained by crosslinking with GL. The films were transparent, flexible with smooth surfaces, hydrophilic and able to load and release vancomycin for more than 8h. Biodegradation in artificial lachrymal fluid (ALF) with lysozyme at 37°C showed that mass loss was higher for the samples containing CS. Also, the samples prepared with CS showed the formation of pores which were visualized by SEM. All samples revealed a biocompatible character after 24h in contact with cornea endothelial cells. As a general conclusion it was possible to determine that the 70PVA/30CS film showed to combine the necessary features to prepare vancomycin-eluting contact lenses to prevent inflammation after corneal substitution.

Innovation in Thermalism: An Example in Beira Interior Region of Portugal

Portugal is one of the richest European countries in thermal waters, and the majority of the spas are located inland in the northern and central regions. Thermal spa treatment is a touristic product that is highly recognized for its potential in the exploration of endogenous regional resources. Consequently, the development of new and competitive thermal tourism products can play an important role in the development of the inland regions, contributing effectively to addressing the economic asymmetries of the region. Based on their physico-chemical properties, thermal waters can have different therapeutic indications, including in dermatology. In the region of the Beira Interior there are two spas with thermal water presenting therapeutic dermatological applications, and the use of these waters in dermocosmetic formulations is proposed as one of the most relevant applications of innovation tools in the Thermal Spa sector. A number of cosmetic companies have encapsulated active molecules in nanoscale systems to enhance product performance. The properties of the nanocarriers systems, namely liposomes, polymeric and solid lipid nanoparticles as well nanoemulsions, are fundamental to increase the skin permeation/penetration. Furthermore, these systems have the capacity to preserve the original and unique properties of the thermal water and ensure the stability of the other active components during more time. The development of dermocosmetics based on thermal water carried in nanobiotechnological systems is proposed for its contribution to the differentiation amongst local, unique and genuine products, especially relevant in the case of products with high economic impact in tourism markets.

Lyophilized tablets for focal delivery of fluconazole and itraconazole through vaginal mucosa, rational design and in vitro evaluation

&NA; The present work deals with the rational design and in vitro evaluation of vaginal tablets for focal delivery of fluconazole (FLZ) and itraconazol (ITZ). Drug loaded liposomes with and without d‐alpha‐tocopheryl polyethylene glycol 1000 succinate (vit E TPGS) were prepared by direct sonication of the components and mixed with albumin to obtain albusomes. Tablets were obtained by direct compression of the lyophilized cake. The influence of vit E TPGS on size, zeta potential and entrapment efficiency (EE%) of liposomes and albusomes was evaluated. Tablet swelling and drug release were studied by in vitro assays. Vit E TPGS neither affected the zeta potential nor the EE% of liposomes and albusomes, but affected the liposomes size and the tablet disintegration time. A rapid erosion was observed for the tablets with the highest content of vitamin, while a slow swelling for those lacking the vitamin (swelling index = 57.76 ± 13.51%). A faster drug release profile was obtained for the former compared to the latter. The in vitro assay showed that FLZ diffused and solved in the vaginal fluid simulant while ITZ remained into the albusomes, which slowly released ITZ‐albumin complex and ITZ‐loaded liposomes, both suitable carriers for drug transport to deeper vaginal endothelium. Graphical abstract Figure. No caption available.