Nuno M. Oliveira

Two-Dimensional Open Microfluidic Devices by Tuning the Wettability on Patterned Superhydrophobic Polymeric Surface

We present a simple and economical method to produce a potential open microfluidic polymeric device. Biomimetic superhydrophobic surfaces were prepared on polystyrene using a phase separation methodology. Patterned two-dimensional channels were imprinted on the superhydrophobic substrates by exposing the surface to plasma or UV–ozone radiation. The wettability of the channels could be precisely controlled between the superhydrophobic and superhydrophilic states by changing the exposure time. The ability of superhydrophilic paths to drive liquid flows in a horizontal position was found to be significantly higher than for the case of hydrophilic paths patterned onto smooth surfaces.

Superhydrophobic Surfaces Engineered Using Diatomaceous Earth

We present a simple method to prepare superhydrophobic surfaces using siliceous exoskeleton of diatoms, a widespread group of algae. This makes diatomaceous earth an accessible and cheap natural material. A micro/nanoscale hierarchical topography was achieved by coating a glass surface with diatomaceous earth, giving rise to a superhydrophilic surface. Superhydrophobic surfaces were obtained by a further surface chemical modification through fluorosilanization. The wettability of the superhydrophobic surface can be modified by Argon plasma treatment in a controlled way by exposure time variation. The chemical surface modification by fluorosilanization and posterior fluorinated SH surface modification by plasma treatment was analyzed by XPS. Using appropriated hollowed masks only specific areas on the surface were exposed to plasma permitting to pattern hydrophilic features with different geometries on the superhydrophobic surface. We showed that the present strategy can be also applied in other substrates, including thermoplastics, enlarging the potential applicability of the resulting surfaces.

Open Fluidics: A Cell Culture Flow System Developed Over Wettability Contrast-Based Chips

Biological tissues are recurrently exposed to several dynamic mechanical forces that influence cell behavior. On this work, the focus is on the shear stress forces induced by fluid flow. The study of flow-induced effects on cells leads to important advances in cardiovascular, cancer, stem cell, and bone biology understanding. These studies are performed using cell culture flow (CCF) systems, mainly parallel plate flow chambers (PPFC), and microfluidic systems. Here, it is proposed an original CCF system based on the open fluidics concept. The system is developed using a planar superhydrophobic platform with hydrophilic paths. The paths work as channels to drive cell culture medium flows without using walls for liquid confinement. The liquid streams are controlled just based on the wettability contrast. To validate the concept, the effect of the shear stress stimulus in the osteogenic differentiation of C2C12 myoblast cells is studied. Combining bone morphogenic protein (specifically BMP-2) stimulation with this mechanical stimulus, a synergistic effect is found on osteoblast differentiation. This effect is confirmed by the enhancement of alkaline phosphatase activity, a well-known early marker of osteogenic differentiation. The suggested CCF system combines characteristics and advantages of both the PPFC and microfluidic systems.

Stock market sentiment lexicon acquisition using microblogging data and statistical measures

Lexicon acquisition is a key issue for sentiment analysis. This paper presents a novel and fast approach for creating stock market lexicons. The approach is based on statistical measures applied over a vast set of labeled messages from StockTwits, which is a specialized stock market microblog. We compare three adaptations of statistical measures, such as Pointwise Mutual Information (PMI), two new complementary statistics and the use of sentiment scores for affirmative and negated contexts. Using StockTwits, we show that the new lexicons are competitive for measuring investor sentiment when compared with six popular lexicons. We also applied a lexicon to easily produce Twitter investor sentiment indicators and analyzed their correlation with survey sentiment indexes. The new microblogging indicators have a moderate correlation with popular Investors Intelligence (II) and American Association of Individual Investors (AAII) indicators. Thus, the new microblogging approach can be used alternatively to traditional survey indicators with advantages (e.g., cheaper creation, higher frequencies). Proposal of an automatic procedure for the creation of stock market lexicons.The procedure uses diverse statistical measures on StockTwits labeled messages.The new lexicons obtain better investor sentiment indicators than general lexicons.The new Twitter sentiment indicators correlate with survey sentiment indicators.

Biocompatible Polymeric Microparticles Produced by a Simple Biomimetic Approach

The use of superhydrophobic surfaces to produce polymeric particles proves to be biologically friendly since it entails the pipetting and subsequent cross-linking of polymeric solutions under mild experimental conditions. Moreover, it renders encapsulation efficiencies of ∼100%. However, the obtained particles are 1 to 2 mm in size, hindering to a large extent their application in clinical trials. Improving on this technique, we propose the fabrication of polymeric microparticles by spraying a hydrogel precursor over superhydrophobic surfaces followed by photo-cross-linking. The particles were produced from methacrylamide chitosan (MA-CH) and characterized in terms of their size and morphology. As demonstrated by optical and fluorescence microscopy, spraying followed by photo-cross-linking led, for the first time, to the production of spherical particles with diameters on the order of micrometers, nominal sizes not attainable by pipetting. Particles such as these are suitable for medical applications such as drug delivery and tissue engineering.

Chitosan–alginate multilayered films with gradients of physicochemical cues

Tissues presenting continuous variations of properties in one direction have inspired the development of functional graded materials. In this work, we developed a new facile method for the development of continuous gradients in chitosan (CHIT) and alginate (ALG) polyelectrolyte multilayers (PEMs) obtained layer-by-layer based on the gradual dipping of CHIT/ALG coated glass slides in genipin solution. Stiffness gradients were produced in the cm scale by varying the reaction time with genipin. Quartz crystal microbalance, colorimetric measurements, trypan blue assay, attenuated total reflection-Fourier transform infrared spectroscopy, swelling ability, water contact angle and dynamic mechanical analysis (DMA) were used to find suitable conditions for the stiffness gradient. The PEMs can be successfully built up and cross-linked with genipin to yield surfaces with uniform physicochemical properties or with gradients of different physicochemical properties. It was found that a large reduction in the hydrophobic nature of the CHIT/ALG PEMs could be produced with higher cross-linking reaction times, regardless of the decrease in their swelling ability. Moreover, the mechanical properties were evaluated using an innovative and non-conventional DMA to monitor the cross-linking reaction in situ. The results confirm an enhancement on the tensile storage modulus with increasing reaction times from 60 to 140 MPa. In another original DMA testing protocol the local compression storage modulus was also measured directly on the films along the stiffness gradient, with results consistent with the tensile tests obtained on the freestanding membranes with different cross-linking degrees. The in vitro biological performance demonstrates that L929 adhered and spread more in the stiffer regions. This work demonstrates the versatility and feasibility of the LbL methodology to generate functional biomimetic surfaces with tuned mechanical and physicochemical properties, which hold great promise for the study of cell-substrate interactions.

Fabrication and characterization of Eri silk fibers-based sponges for biomedical application.

UNLABELLEDnCocoon-derived semi-domesticated Eri silk fibers still lack exploitation for tissue engineering applications due to their poor solubility using conventional methods. The present work explores the ability to process cocoon fibers of non-mulberry Eri silk (Samia/Philosamia ricini) into sponges through a green approach using ionic liquid (IL)--1-buthyl-imidazolium acetate as a solvent. The formation of β-sheet structures during Eri silk/IL gelation was acquired by exposing the Eri silk/IL gels to a saturated atmosphere composed of two different solvents: (i) isopropanol/ethanol (physical stabilization) and (ii) genipin, a natural crosslinker, dissolved in ethanol (chemical crosslinking). The sponges were then obtained by freeze-drying. This approach promotes the formation of both stable and ordered non-crosslinked Eri silk fibroin matrices. Moreover, genipin-crosslinked silk fibroin sponges presenting high height recovery capacity after compression, high swelling degree and suitable mechanical properties for tissue engineering applications were produced. The incorporation of a model drug--ibuprofen--and the corresponding release study from the loaded sponges demonstrated the potential of using these matrices as effective drug delivery systems. The assessment of the biological performance of ATDC5 chondrocyte-like cells in contact with the developed sponges showed the promotion of cell adhesion and proliferation, as well as extracellular matrix production within 2 weeks of culture. Sponges intrinsic properties and biological findings open up their potential use for biomedical applications.nnnSTATEMENT OF SIGNIFICANCEnThis work addresses the preparation and characterization of non-mulberry cocoon-derived Eri silk sponges. The insolubility of cocoons-derived non-mulberry silkworms impairs their processability and applications in the healthcare field. We used a green approach with ionic liquids to overcome the lack solubility of such silk fibers. The formation of beta-sheet structures into Eri-based sponges was physically and chemically induced. The sponges were obtained by freeze-drying. The developed structures exhibited flexibility to adapt and recover their shapes upon application and subsequent removal of load, high swelling degree, ability to load an anti-inflammatory drug and to promote its sustained release. They promoted in vitro cellular adhesion, proliferation and extracellular matrix production of a chondrocyte-like cell line, opening up their potential application for biomedical applications.

Liquid Marbles for High‐Throughput Biological Screening of Anchorage‐Dependent Cells

Stable liquid marbles (LM) are produced by coating liquid droplets with a hydrophobic powder. The used hydrophobic powder is produced by fluorosi-lanization of diatomaceous earth, used before to produce superhydrophobic structures. Here, the use of LM is proposed for high-throughput drug screening on anchorage-dependent cells. To provide the required cell adhesion sites inside the liquid environment of LM, surface-modified poly(l-lactic acid) microparticles are used. A simple method that takes advantage from LM appealing features is presented, such as the ability to inject liquid on LM without disrupting (self-healing ability), and to monitor color changes inside of LM. After promoting cell adhesion, a cytotoxic screening test is performed as a proof of concept. Fe(3+) is used as a model cytotoxic agent and is injected on LM. After incubation, AlamarBlue reagent is injected and used to assess the presence of viable cells, by monitoring color change from blue to red. Color intensity is measured by image processing and the analysis of pictures takes using an ordinary digital camera. The proposed method is fully validated in counterpoint to an MTS (3-(4,5-dimethylthiazol-2-yl)-5-(3-carbou200bxymethoxyphenyl)-2-(4-sulfophenyl)-2H-teu200btrazolium) colorimetric assay, a well-known method used for the cytotoxicity assessment.

The potential of liquid marbles for biomedical applications: a critical review

Liquid marbles (LM) are freestanding droplets covered by micro/nanoparticles with hydrophobic/hydrophilic properties, which can be manipulated as a soft solid. The phenomenon that generates these soft structures is regarded as a different method to generate a superhydrophobic behavior in the liquid/solid interface without modifying the surface. Several applications for the LM have been reported in very different fields, however the developments for biomedical applications are very recent. At first, the LM properties are reviewed, namely shell structure, LM shape, evaporation, floatability and robustness. The different strategies for LM manipulation are also described, which make use of magnetic, electrostatic and gravitational forces, ultraviolet and infrared radiation, and approaches that induce LM self-propulsion. Then, very distinctive applications for LM in the biomedical field are presented, namely for diagnostic assays, cell culture, drug screening and cryopreservation of mammalian cells. Finally, a critical outlook about the unexplored potential of LM for biomedical applications is presented, suggesting possible advances on this emergent scientific area.

Bioinspired superamphiphobic surfaces as a tool for polymer-and solvent-independent preparation of drug-loaded spherical particles

Superamphiphobic surfaces were evaluated as a tool to prepare spherical particles from polymers and solvents of very diverse nature, under mild conditions and with 100% drug encapsulation yield. Different from bioinspired superhydrophobic surfaces suitable only for aqueous dispersions, the superamphiphobic platforms allowed the formation of spherical droplets when solvents of any polarity were deposited onto them. Spherical poly(d,l-lactide-co-glycolide) (PLGA) particles were synthesized by placing drops of PLGA/ciprofloxacin suspensions in dioxane on a superamphiphobic surface followed by solvent evaporation. The particles prepared covering a wide range of PLGA/ciprofloxacin weight ratios delivered a 20% dose in the first 24h and then sustained the release of the remaining drug for more than 1month. The particles, both freshly prepared and after being 26days in the release medium, showed efficiency against different types of microorganisms. The developed polymer- and solvent-independent approach could be useful for microencapsulation with very high efficiency of active substances of varied nature into size-tunable particles for a wide range of applications in an affordable and cost-effective manner.