In vitro chondrogenic commitment of human Wharton's jelly stem cells by co-culture with human articular chondrocytes.

Wharton's jelly stem cells (WJSCs) are a potential source of transplantable stem cells in cartilage-regenerative strategies, due to their highly proliferative and multilineage differentiation capacity. We hypothesized that a non-direct co-culture system with human articular chondrocytes (hACs) could enhance the potential chondrogenic phenotype of hWJSCs during the expansion phase compared to those expanded in monoculture conditions. Primary hWJSCs were cultured in the bottom of a multiwell plate separated by a porous transwell membrane insert seeded with hACs. No statistically significant differences in hWJSCs duplication number were observed under either of the culture conditions during the expansion phase. hWJSCs under co-culture conditions show upregulations of collagen type I and II, COMP, TGFβ1 and aggrecan, as well as of the main cartilage transcription factor, SOX9, when compared to those cultured in the absence of chondrocytes. Chondrogenic differentiation of hWJSCs, previously expanded in co-culture and monoculture conditions, was evaluated for each cellular passage using the micromass culture model. Cells expanded in co-culture showed higher accumulation of glycosaminoglycans (GAGs) compared to cells in monoculture, and immunohistochemistry for localization of collagen type I revealed a strong detection signal when hWJSCs were expanded under monoculture conditions. In contrast, type II collagen was detected when cells were expanded under co-culture conditions, where numerous round-shaped cell clusters were observed. Using a micromass differentiation model, hWJSCs, previously exposed to soluble factors secreted by hACs, were able to express higher levels of chondrogenic genes with deposition of cartilage extracellular matrix components, suggesting their use as an alternative cell source for treating degenerated cartilage.

Phenanthroimidazole derivatives as fluorimetric chemosensors in aqueous medium

In recent years the research of sensors with good sensitivity and good selectivity in aqueous medium has been of great interest. Chemosensors soluble in aqueous media are very interesting, because of the importance in revealing a number of biological processes, disease states and environmental pollutions. 2,4,5-Triaryl-imidazoles are versatile compounds with application in medicine, due to their biological activity, and materials sciences, for their interesting optical properties. These properties can be tuned by careful selection of substituents at positions 2, 4 and 5: replacement of the aryl group by a heterocyclic group results in larger π-conjugated systems with improved optical properties for application in nonlinear optics, OLEDs, DNA intercalators, and chemosensors. In this communication, we report the synthesis of new phenanthroimidazoles, substituted at position 2 with (hetero)aryl groups of different electronic character, in order to evaluate their photophysical properties and chemosensory ability. The new derivatives were characterized by the usual techniques and a detailed photophysical study was undertaken. The evaluation of the compounds as fluorimetric chemosensors was carried out by performing titrations in acetonitrile and acetonitrile/water in the presence of relevant organic and inorganic anions, and of alkaline, alkaline-earth and transition metal cations.

Skin-derived cell-sheets as powerful tools to engineer skin analogues

Cell/cell-extracellular matrix (ECM) dynamic interactions appear to have a major role in regulating communication through soluble signaling, directing cell binding and activating substrates that participate in the highly organized wound healing process. Moreover, these interactions are also crucial for in vitro mimicking cutaneous physiology. Herein we explore cell sheet (CS) engineering to create cellular constructs formed by keratinocytes (hKC), fibroblasts (hDFB) and dermal microvascular endothelial cells (hDMEC), to target skin wound healing but also the in vitro recreation of relevant models. Taking advantage of temperature-responsive culture surfaces, which allow harvesting cultured cells as intact sheets along with the deposited native ECM, varied combinations of homotypic and heterotypic three-dimensional (3-D) CS-based constructs were developed. Constructs combining one CS of keratinocytes as an epidermis-like layer plus a vascularized dermis composed by hDFB and hDMECs were assembled as skin analogues for advancing in vitro testing. Simultaneously both hKC and hDMEC were shown to significantly contribute to the re-epithelialization of full-thickness mice skin wounds by promoting an early epithelial coverage, while hDMEC significantly lead to increased vessels density, incorporating the neovasculature. Thus, although determined by the cellular nature of the constructs, these outcomes demonstrated that CS engineering appear as an unique technology that open the possibility to create numerous combinations of 3D constructs to target defective wound healing as well as the construction of in vitro models to further mimic cutaneous functions crucial for drug screening and cosmetic testing assays.

Evaluation of antibacterial activity of caffeic acid encapsulated by -cyclodextrins

Context: Caffeic acid is described as antibacterial, but this bioactive molecule has some issues regarding solubility and stability to environmental stress. Thus, encapsulation devices are required. Objective: The aim of this work was to study the effect of the caffeic acid encapsulation by cyclodextrins on its antibacterial activity. Materials and methods: The interactions between the caffeic acid and three cyclodextrins (-cyclodextrin (CD), 2-hydroxypropyl--cyclodextrin (HPCD) and methyl--cyclodextrin were study. Results and discussion: The formation of an aqueous soluble inclusion complex was confirmed for CD and HPCD with a 1:1 stoichiometry. The CD/caffeic acid complex showed higher stability than HPCD/caffeic acid. Caffeic acid antibacterial activity was similar at pH 3 and pH 5 against the three bacteria (K. pneumoniae, S. epidermidis and S. aureus). Conclusions: The antibacterial activity of the inclusion complexes was described here for the first time and it was shown that the caffeic acid activity was remarkably enhanced by the cyclodextrins encapsulation.

Postharvest shelf life extension of blueberries using a chitosan-based edible coating containing aloe vera juice

Poster

Effect of surface plasmon resonance in TiO2/Au thin films on the fluorescence of self-assembled CdTe QDs structure

The exceptional properties of localised surface plasmons (LSPs), such as local field enhancement and confinement effects, resonant behavior, make them ideal candidates to control the emission of luminescent nanoparticles. In the present work, we investigated the LSP effect on the steady-state and time-resolved emission properties of quantum dots (QDs) by organizing the dots into self-assembled dendrite structures deposited on plasmonic nanostructures. Self-assembled structures consisting of water-soluble CdTe mono-size QDs, were developed on the surface of co-sputtered TiO2 thin films doped with Au nanoparticles (NPs) annealed at different temperatures. Their steady-state fluorescence properties were probed by scanning the spatially resolved emission spectra and the energy transfer processes were investigated by the fluorescence lifetime imaging (FLIM) microscopy. Our results indicate that a resonant coupling between excitons confined in QDs and LSPs in Au NPs located beneath the self-assembled structure indeed takes place and results in (i) a shift of the ground state luminescence towards higher energies and onset of emission from excited states in QDs, and (ii) a decrease of the ground state exciton lifetime (fluorescence quenching).

Expressão e caracterização de uma lectina recombinante de interesse biomédico

Dissertação de mestrado integrado em Engenharia Biomédica (área de especialização em Engenharia Clínica)

Development of elastin-like recombinamer films with antimicrobial activity

In the present work we explored the ABP-CM4 peptide properties from Bombyx mori for the creation of biopolymers with broad antimicrobial activity. An antimicrobial recombinant protein-based polymer (rPBP) was designed by cloning the DNA sequence coding for ABP-CM4 in frame with the N-terminus of the elastin-like recombinamer consisting of 200 repetitions of the pentamer VPAVG, here named A200. The new rPBP, named CM4-A200, was purified via a simplified nonchromatographic method, making use of the thermoresponsive behavior of the A200 polymer. ABP-CM4 peptide was also purified through the incorporation of a formic acid cleavage site between the peptide and the A200 sequence. In soluble state the antimicrobial activity of both CM4-A200 polymer and ABP-CM4 peptide was poorly effective. However, when the CM4-A200 polymer was processed into free-standing films high antimicrobial activity against Gram-positive and Gram-negative bacteria, yeasts and filamentous fungi was observed. The antimicrobial activity of CM4-A200 was dependent on the physical contact of cells with the film surface. Furthermore, CM4-A200 films did not reveal a cytotoxic effect against both normal human skin fibroblasts and human keratinocytes. Finally, we have developed an optimized ex vivo assay with pig skin demonstrating the antimicrobial properties of the CM4-A200 cast films for skin applications.

Osteogenic differentiation of human mesenchymal stem cells in the absence of osteogenic supplements: a surface-roughness gradient study

The use of biomaterials to direct osteogenic differentiation of human mesenchymal stem cells (hMSCs) in the absence of osteogenic supplements is thought to be part of the next generation of orthopedic implants. We previously engineered surface-roughness gradients of average roughness (Ra) varying from the sub-micron to the micrometer range ( 0.5–4.7 lm), and mean distance between peaks (RSm) gradually varying from 214 lm to 33 lm. Here we have screened the ability of such surface-gradients of polycaprolactone to influence the expression of alkaline phosphatase (ALP), collagen type 1 (COL1) and mineralization by hMSCs cultured in dexamethasone (Dex)-deprived osteogenic induction medium (OIM) and in basal growth medium (BGM). Ra 1.53 lm/RSm 79 lm in Dex-deprived OI medium, and Ra 0.93 lm/RSm 135 lm in BGM consistently showed higher effectiveness at supporting the expression of the osteogenic markers ALP, COL1 and mineralization, compared to the tissue culture polystyrene (TCP) control in complete OIM. The superior effectiveness of specific surface-roughness revealed that this strategy may be used as a compelling alternative to soluble osteogenic inducers in orthopedic applications featuring the clinically relevant biodegradable polymer polycaprolactone.

Biomimetic supramolecular designs for the controlled release of growth factors in bone regeneration

The extracellular matrix (ECM) of tissues is an assembly of insoluble macromolecules that specifically interact with soluble bioactive molecules and regulate their distribution and availability to cells. Recapitulating this ability has been an important target in controlled growth factor delivery strategies for tissue regeneration and requires the design of multifunctional carriers. This review describes the integration of supramolecular interactions on the design of delivery strategies that encompass self-assembling and engineered affinity components to construct advanced biomimetic carriers for growth factor delivery. Several glycan- and peptide-based self-assemblies reported in the literature are highlighted and commented upon. These examples demonstrate how molecular design and chemistry are successfully employed to create versatile multifunctional molecules which self-assemble/disassemble in a precisely predicted manner, thus controlling compartmentalization, transport and delivery. Finally, we discuss whether recent advances in the design and preparation of supramolecular delivery systems have been sufficient to drive real translation towards a clinical impact. 

Biocatalysis in micellar systems

[Excert] Biocatalysis and biotransformations are environmentally friendly, and allow the development of sustainable production processes on a large scale. Thus, these processes are becoming important alternatives to conventional chemistry in the drug, biochemical, and emerging biorenewable energy industries. Biocatalysts are required to function under non-conventional conditions, such as in organic solvents, being competitive in terms of cost and efficiency. In fact, the technological utility of enzymes can be enhanced greatly by using them in the presence of organic solvents, rather than in their natural aqueous reaction media. Multiphase systems are more complex but offer a new field of possibilities. The presence of hydrophobic solvents in biocatalysis allows the conversion of poorly water soluble substrates more efficiently. The accessibility of hydrophobic substrates to enzymes or whole cells presents an interesting challenge for researchers and technologists. In this context, microemulsions are a promising tool in enzyme technology. This chapter presents an overview of the characterization of biphasic and microemulsion systems and their applications in biotransformation processes (...).

Production of biohydrogen from dark fermentation of organic wastes

Dissertação de mestrado em Bioengenharia