Dina M. Silva

Development of a hybrid dextrin hydrogel encapsulating dextrin nanogel as protein delivery system.

Dextrin, a glucose polymer with low molecular weight, was used to develop a fully resorbable hydrogel, without using chemical initiators. Dextrin was first oxidized (oDex) with sodium periodate and then cross-linked with adipic acid dihidrazide, a nontoxic cross-linking molecule. Furthermore, a new bidimensional composite hydrogel, made of oxidized dextrin incorporating dextrin nanogels (oDex-nanogel), was also developed. The oDex hydrogels showed good mechanical properties and biocompatibility, allowing the proliferation of mouse embryo fibroblasts 3T3 cultured on top of the gel. The gelation time may be controlled selecting the concentrations of the polymer and reticulating agent. Both the oDex and oDex-nanogel hydrogels are biodegradable and present a 3-D network with a continuous porous structure. The obtained hybrid hydrogel enables the release of the dextrin nanogel over an extended period of time, paralleling the mass loss curve due to the degradation of the material. The dextrin nanogel allowed the efficient incorporation of interleukin-10 and insulin in the oDex hydrogel, providing a sophisticated system of controlled release. The new hydrogels present promising properties as an injectable carrier of bioactive molecules. Both proteins and poorly water-soluble low-molecular-weight drugs are efficiently encapsulated in the nanogel, which performs as a controlled release system entrapped in the hydrogel matrix.

Effect of Sterilization Methods on Electrospun Poly(lactic acid) (PLA) Fiber Alignment for Biomedical Applications

Medically approved sterility methods should be a major concern when developing a polymeric scaffold, mainly when commercialization is envisaged. In the present work, poly(lactic acid) (PLA) fiber membranes were processed by electrospinning with random and aligned fiber alignment and sterilized under UV, ethylene oxide (EO), and γ-radiation, the most common ones for clinical applications. It was observed that UV light and γ-radiation do not influence fiber morphology or alignment, while electrospun samples treated with EO lead to fiber orientation loss and morphology changing from cylindrical fibers to ribbon-like structures, accompanied to an increase of polymer crystallinity up to 28%. UV light and γ-radiation sterilization methods showed to be less harmful to polymer morphology, without significant changes in polymer thermal and mechanical properties, but a slight increase of polymer wettability was detected, especially for the samples treated with UV radiation. In vitro results indicate that both UV and γ-radiation treatments of PLA membranes allow the adhesion and proliferation of MG 63 osteoblastic cells in a close interaction with the fiber meshes and with a growth pattern highly sensitive to the underlying random or aligned fiber orientation. These results are suggestive of the potential of both γ-radiation sterilized PLA membranes for clinical applications in regenerative medicine, especially those where customized membrane morphology and fiber alignment is an important issue.

Exploiting the Sequence of Naturally Occurring Elastin: Construction, Production and Characterization of a Recombinant Thermoplastic Protein-Based Polymer

Genetic engineering was used to produce an elastin-like polymer (ELP) with precise amino acid composition, sequence and length, resulting in the absolute control of MW and stereochemistry. A synthetic monomer DNA sequence encoding for (VPAVG)20, was used to build a library of concatemer genes with precise control on sequence and size. The higher molecular weight polymer with 220 repeats of VPAVG was biologically produced in Escherichia coli and purified by hot and cold centrifugation cycles, based on the reversible inverse temperature transition property of ELPs. The use of low cost carbon sources like lactose and glycerol for bacteria cells culture media was explored using Central Composite Design approach allowing optimization of fermentation conditions. Due to its self-assembling behaviour near 33 °C stable spherical microparticles with a size ~ 1µm were obtained, redissolving when a strong undercooling is achieved. The polymer produced showed hysteresis behaviour with thermal absorbing/releasing components depending on the salt concentration of the polymer solution.

Structural analysis of dextrins and characterization of dextrin-based biomedical hydrogels.

The characterization of several commercial dextrins and the analysis of the potential of dextrin derived hydrogels for biomedical applications were performed in this work. The structural characterization of dextrins allowed the determination of the polymerization and branching degrees, which ranged from 6 to 17 glucose residues and 2 to 13%, respectively. Tackidex, a medical grade dextrin was choosen for further characterization. The combination of hydrogel with a dextrin nanogel and urinary bladder matrix was achieved without compromising the mechanical properties or microstructure. The encapsulation of cells, preserving its viability, confirms the biocompatibility of the injectable hydrogels, which have therefore great potential for biomedical applications.

Multifunctional PLLA-ceramic fiber membranes for bone regeneration applications

A novel method to process electrospun poly(l-lactic acid) (PLLA) membranes incorporating glass reinforced hydroxyapatite granules (gHA) interspacially between the polymeric fibers is reported, thus increasing the surface area for cellular interactions. gHA granules (≤150μm) electrospun together with the polymer solution, lead to an average fiber diameter of 550±150nm for pristine PLLA and 440±170nm for the composite samples. An increase of the overall porosity was observed, from 79±3% for the PLLA up to 88±5% for the hybrid samples, keeping materials wettability and mechanical properties. Bone-bonding ability showed that both samples induced HA crystal nucleation, but with a distinct pattern of mineral deposition. gHA microcomposite allows a better F-actin cytoskeleton organization during the initial adhesion and spreading, favoring cell-fibers and cell-to-cell interactions and enhanced alkaline phosphatase activity, making them potential candidates for bone healing strategies.

Inflammatory response to dextrin-based hydrogel associated with human mesenchymal stem cells, urinary bladder matrix and Bonelike ® granules in rat subcutaneous implants

Increasing relevance has been attributed to hydrogels due to their ability to provide an extracellular matrix (ECM)-like environment for cellular adhesion and proliferation, acting as mechanical scaffolds for tissue remodeling or as delivery matrices. In vivo biocompatibility of a hybrid dextrin hydrogel produced from oxidized dextrin and adipic acid dihydrazide and its combinations with human mesenchymal stem cells (hMSCs), ECM from a porcine bladder (urinary bladder matrix) and ceramic granules (Bonelike®), was evaluated following ISO 10993 after subcutaneous implantation in a rat model. Histological analysis after 3 and 15 d showed typical acute and chronic inflammatory responses, respectively, with a more severe reaction exhibited whenever the ceramic granules were present. However, the dextrin hydrogel was able to stabilize granules in the implant site. Dextrin hydrogel was scored as slight irritant after 3 d, similar to its combination with UBM, and as non-irritant after 15 d. The presence of viable hMSCs in the subcutaneous tissue could be confirmed by the presence of anti-human nuclei antibody (HuNu+) cells. The production of growth factors and inflammatory and immunomodulatory cytokines by these cells was also quantified in peripheral blood confirming the successful encapsulation of hMSCs into the hydrogel matrix for cell survival promotion. The presence of hMSCs seemed to modulate the inflammatory response by accelerating its progression when compared to the acellular experimental groups. Dextrin hydrogel has proven to be a biocompatible multifunctional matrix for minimally invasive biomedical procedures, including orthopedic surgeries when associated with bone substitutes and also as a possible encapsulation matrix for cell-based therapies.

Antibacterial and Antifungal Activity of Poly(Lactic Acid)-Bovine Lactoferrin Nanofiber Membranes

Antimicrobial materials have become relevant for local therapies preventing microbial resistance induced by systemic antibiotic treatments. This work reports the development of electrospun poly(lactic acid) (PLLA) nanofiber membranes loaded with bovine lactoferrin (bLF) up to 20 wt%. The membranes present smooth and nondefective fibers with mean diameters between 717 ± 197 and 495 ± 127 nm, and an overall porosity of ≈80%. The hydrophobicity of the PLLA membranes is reduced by the presence of bLF. The release profile of bLF correlates with an anomalous transport model, with 17.7 ± 3.6% being released over 7 weeks. The nanofiber mats show no cytotoxicity on human skin fibroblasts and even promote cell proliferation after short exposure periods. Furthermore, the developed membranes display antifungal activity against Aspergillus nidulans by inhibiting spore germination and mycelial growth. These results evidence the strong potential of bLF-PLLA nanofiber membranes to be used as antifungal dressings.

Development of ciprofloxacin-loaded poly(vinyl alcohol) dry powder formulations for lung delivery

Graphical abstract Figure. No caption available. ABSTRACT Polymeric microparticles are micro carriers for the sustained drug delivery of drugs in the lungs, used as alternatives to the use of established excipients. This study aims to develop and characterize inhalable ciprofloxacin (CPx)‐loaded poly(vinyl alcohol) (PVA) microparticles by a single‐step spray‐drying procedure. The optimization of the processing parameters was achieved by an orthogonal design of the most relevant processing parameters (polymer concentration, feed rate and inlet temperature). The obtained spray‐dried particles showed a drug encapsulation efficiency higher than 90%. Furthermore, PVA‐CPx formulations, with drug contents up to 10 wt%, showed a morphology and size suitable for inhalation, with a sustained release profile over 24 h. Data from Fourier transformed infra‐red spectroscopy and differential scanning calorimetry indicated absence of interaction between the polymer matrix and the drug. Aerodynamic assessment of PVA‐CPx 10 wt% was determined by the next generation impactor (NGI), using spray‐dried CPx as a control. The results showed improved values of mass median aerodynamic diameter (5.06 ± 0.10&mgr;m) and a fine particle fraction (39.78 ± 0.98%) when comparing with the CPx alone (5.33 ± 0.39&mgr;m and 30.43 ± 1.38%). This study highlights the potential of spray‐dried PVA microparticles as drug carriers for lung local delivery of antibiotics.