Medium chain length polyhydroxyalkanoates as potential matrix materials for peripheral nerve regeneration.

Polyhydroxyalkanoates are natural, biodegradable, thermoplastic and sustainable polymers with a huge potential in fabrication of bioresorbable implantable devices for tissue engineering. We describe a comparative evaluation of three medium chain length polyhydroxyalkanoates (mcl-PHAs), namely poly(3-hydroxyoctanoate), poly(3-hydroxyoctanoate-co-3-hydoxydecanoate) and poly(3-hydroxyoctanoate-co-3-hydroxydecanoate-co-3-hydroxydodecanoate), one short chain length polyhydroxyalkanoate, poly(3-hydroxybutyrate), P(3HB) and synthetic aliphatic polyesters (polycaprolactone and polylactide) with a specific focus on nerve regeneration, due to mechanical properties of mcl-PHAs closely matching nerve tissues. In vitro biological studies with NG108-15 neuronal cell and primary Schwann cells did not show a cytotoxic effect of the materials on both cell types. All mcl-PHAs supported cell adhesion and viability. Among the three mcl-PHAs, P(3HO-co-3HD) exhibited superior properties with regards to numbers of cells adhered and viable cells for both cell types, number of neurite extensions from NG108-15 cells, average length of neurite extensions and Schwann cells. Although, similar characteristics were observed for flat P(3HB) surfaces, high rigidity of this biomaterial, and FDA-approved polymers such as PLLA, limits their applications in peripheral nerve regeneration. Therefore, we have designed, synthesized and evaluated these materials for nerve tissue engineering and regenerative medicine, the interaction of mcl-PHAs with neuronal and Schwann cells, identifying mcl-PHAs as excellent materials to enhance nerve regeneration and potentially their clinical application in peripheral nerve repair.

Aligned Polyhydroxyalkanoate Blend Electrospun Fibers as Intraluminal Guidance Scaffolds for Peripheral Nerve Repair.

The use of nerve guidance conduits (NGCs) to treat peripheral nerve injuries is a favorable approach to the current "gold standard" of autografting. However, as simple hollow tubes, they lack specific topographical and mechanical guidance cues present in nerve grafts and therefore are not suitable for treating large gap injuries (30-50 mm). The incorporation of intraluminal guidance scaffolds, such as aligned fibers, has been shown to increase neuronal cell neurite outgrowth and Schwann cell migration distances. A novel blend of PHAs, P(3HO)/P(3HB) (50:50), was investigated for its potential as an intraluminal aligned fiber guidance scaffold. Aligned fibers of 5 and 8 µm diameter were manufactured by electrospinning and characterized using SEM. Fibers were investigated for their effect on neuronal cell differentiation, Schwann cell phenotype, and cell viability in vitro. Overall, P(3HO)/P(3HB) (50:50) fibers supported higher neuronal and Schwann cell adhesion compared to PCL fibers. The 5 µm PHA blend fibers also supported significantly higher DRG neurite outgrowth and Schwann cell migration distance using a 3D ex vivo nerve injury model.

Enhanced production of biobased, biodegradable, Poly(3-hydroxybutyrate) using an unexplored marine bacterium Pseudohalocynthiibacter aestuariivivens, isolated from highly polluted coastal environment.

The production and disposal of plastics from limited fossil reserves, has prompted research for greener and sustainable alternatives. Polyhydroxyalkanoates (PHAs) are biocompatible, biodegradable, and thermoprocessable polyester produced by microbes. PHAs found several applications but their use is limited due to high production cost and low yields. Herein, for the first time, the isolation and characterization of Pseudohalocynthiibacter aestuariivivens P96, a marine bacterium able to produce surprising amount of PHAs is reported. In the best growth condition P96 was able to reach a maximum production of 4.73 g/L, corresponding to the 87 % of total cell dry-weight. Using scanning and transmission microscopy, lab-scale fermentation, spectroscopic techniques, and genome analysis, the production of thermoprocessable polymer Polyhydroxybutyrate P(3HB), a PHAs class, endowed with mechanical and thermal properties comparable to that of petroleum-based plastics was confirmed. This study represents a milestone toward the use of this unexplored marine bacterium for P(3HB) production.

3D Disease Modelling of Hard and Soft Cancer Using PHA-Based Scaffolds.

Tumour cells are shown to change shape and lose polarity when they are cultured in 3D, a feature typically associated with tumour progression in vivo, thus making it significant to study cancer cells in an environment that mimics the in vivo milieu. In this study we established hard (MCF7 and MDA-MB-231, breast cancer) and soft (HCT116, colon cancer) 3D cancer tumour models utilizing a blend of P(3HO-co-3HD) and P(3HB). P(3HO-co-3HD) and P(3HB) belong to a group of natural biodegradable polyesters, PHAs, that are synthesised by microorganisms. The 3D PHA scaffolds produced, with a pore size of 30 to 300 µm, allow for nutrients to diffuse within the scaffold and provide the cells with the flexibility to distribute evenly within the scaffold and grow within the pores. Interestingly, by Day 5, MDA-MB-231 showed dispersed growth in clusters, and MCF7 cells formed an evenly dispersed dense layer, while HCT116 formed large colonies within the pockets of the 3D PHA scaffolds. Our results show Epithelial Mesenchymal Transition (EMT) marker gene expression profiles in the hard tumour cancer models. In the 3D-based PHA scaffolds, MDA-MB-231 cells expressed higher levels of Wnt-11 and mesenchymal markers, such as Snail and its downstream gene Vim mRNAs, while MCF7 cells exhibited no change in their expression. On the other hand, MCF7 cells exhibited a significantly increased E-Cadherin expression as compared to MDA-MB-231 cells. The expression levels of EMT markers were comparative to their expression reported in the tumour samples, making them good representative of cancer models. In future these models will be helpful in mimicking hypoxic tumours, in studying gene expression, cellular signalling, angiogenesis and drug response more accurately than 2D and perhaps other 3D models.

Controlled Delivery of Pan-PAD-Inhibitor Cl-Amidine Using Poly(3-Hydroxybutyrate) Microspheres.

This study deals with the process of optimization and synthesis of Poly(3-hydroxybutyrate) microspheres with encapsulated Cl-amidine. Cl-amidine is an inhibitor of peptidylarginine deiminases (PADs), a group of calcium-dependent enzymes, which play critical roles in a number of pathologies, including autoimmune and neurodegenerative diseases, as well as cancer. While Cl-amidine application has been assessed in a number of in vitro and in vivo models; methods of controlled release delivery remain to be investigated. P(3HB) microspheres have proven to be an effective delivery system for several compounds applied in antimicrobial, wound healing, cancer, and cardiovascular and regenerative disease models. In the current study, P(3HB) microspheres with encapsulated Cl-amidine were produced in a size ranging from ~4-5 µm and characterized for surface morphology, porosity, hydrophobicity and protein adsorption, in comparison with empty P(3HB) microspheres. Cl-amidine encapsulation in P(3HB) microspheres was optimized, and these were found to be less hydrophobic, compared with the empty microspheres, and subsequently adsorbed a lower amount of protein on their surface. The release kinetics of Cl-amidine from the microspheres were assessed in vitro and expressed as a function of encapsulation efficiency. There was a burst release of ~50% Cl-amidine in the first 24 h and a zero order release from that point up to 16 days, at which time point ~93% of the drug had been released. As Cl-amidine has been associated with anti-cancer effects, the Cl-amidine encapsulated microspheres were assessed for the inhibition of vascular endothelial growth factor (VEGF) expression in the mammalian breast cancer cell line SK-BR-3, including in the presence of the anti-proliferative drug rapamycin. The cytotoxicity of the combinatorial effect of rapamycin with Cl-amidine encapsulated P(3HB) microspheres was found to be 3.5% more effective within a 24 h period. The cells treated with Cl-amidine encapsulated microspheres alone, were found to have 36.5% reduction in VEGF expression when compared with untreated SK-BR-3 cells. This indicates that controlled release of Cl-amidine from P(3HB) microspheres may be effective in anti-cancer treatment, including in synergy with chemotherapeutic agents. Using controlled drug-delivery of Cl-amidine encapsulated in Poly(3-hydroxybutyrate) microspheres may be a promising novel strategy for application in PAD-associated pathologies.

Preclinical study of peripheral nerve regeneration using nerve guidance conduits based on polyhydroxyalkanaotes.

Nerve guidance conduits (NGCs) are used as an alternative to the "gold standard" nerve autografting, preventing the need for surgical intervention required to harvest autologous nerves. However, the regeneration outcomes achieved with the current NGCs are only comparable with autografting when the gap is short (less than 10 mm). In the present study, we have developed NGCs made from a blend of polyhydroxyalkanoates, a family of natural resorbable polymers. Hollow NGCs made from a 75:25 poly(3-hydroxyoctanoate)/poly(3-hydroxybutyrate) blend (PHA-NGCs) were manufactured using dip-molding. These PHA-NGCs showed appropriate flexibility for peripheral nerve regeneration. In vitro cell studies performed using RT4-D6P2T rat Schwann cell line confirmed that the material is capable of sustaining cell proliferation and adhesion. PHA-NGCs were then implanted in vivo to repair 10 mm gaps of the median nerve of female Wistar rats for 12 weeks. Functional evaluation of the regenerated nerve using the grasping test showed that PHA-NGCs displayed similar motor recovery as the autograft, starting from week 7. Additionally, nerve cross-sectional area, density and number of myelinated cells, as well as axon diameter, fiber diameter, myelin thickness and g-ratio obtained using the PHA-NGCs were found comparable to an autograft. This preclinical data confirmed that the PHA-NGCs are indeed highly promising candidates for peripheral nerve regeneration.

Silver Nanoparticle-Coated Polyhydroxyalkanoate Based Electrospun Fibers for Wound Dressing Applications.

Wound dressings are high performance and high value products which can improve the regeneration of damaged skin. In these products, bioresorption and biocompatibility play a key role. The aim of this study is to provide progress in this area via nanofabrication and antimicrobial natural materials. Polyhydroxyalkanoates (PHAs) are a bio-based family of polymers that possess high biocompatibility and skin regenerative properties. In this study, a blend of poly(3-hydroxybutyrate) (P(3HB)) and poly(3-hydroxyoctanoate-co-3-hydroxy decanoate) (P(3HO-co-3HD)) was electrospun into P(3HB))/P(3HO-co-3HD) nanofibers to obtain materials with a high surface area and good handling performance. The nanofibers were then modified with silver nanoparticles (AgNPs) via the dip-coating method. The silver-containing nanofiber meshes showed good cytocompatibility and interesting immunomodulatory properties in vitro, together with the capability of stimulating the human beta defensin 2 and cytokeratin expression in human keratinocytes (HaCaT cells), which makes them promising materials for wound dressing applications.

Harnessing Polyhydroxyalkanoates and Pressurized Gyration for Hard and Soft Tissue Engineering.

Organ dysfunction is a major cause of morbidity and mortality. Transplantation is typically the only definitive cure, challenged by the lack of sufficient donor organs. Tissue engineering encompasses the development of biomaterial scaffolds to support cell attachment, proliferation, and differentiation, leading to tissue regeneration. For efficient clinical translation, the forming technology utilized must be suitable for mass production. Herein, uniaxial polyhydroxyalkanoate scaffolds manufactured by pressurized gyration, a hybrid scalable spinning technique, are successfully used in bone, nerve, and cardiovascular applications. Chorioallantoic membrane and in vivo studies provided evidence of vascularization, collagen deposition, and cellular invasion for bone tissue engineering. Highly efficient axonal outgrowth was observed in dorsal root ganglion-based 3D ex vivo models. Human induced pluripotent stem cell derived cardiomyocytes exhibited a mature cardiomyocyte phenotype with optimal calcium handling. This study confirms that engineered polyhydroxyalkanoate-based gyrospun fibers provide an exciting and unique toolbox for the development of scalable scaffolds for both hard and soft tissue regeneration.

Antibacterial Composite Materials Based on the Combination of Polyhydroxyalkanoates With Selenium and Strontium Co-substituted Hydroxyapatite for Bone Regeneration.

Due to the threat posed by the rapid growth in the resistance of microbial species to antibiotics, there is an urgent need to develop novel materials for biomedical applications capable of providing antibacterial properties without the use of such drugs. Bone healing represents one of the applications with the highest risk of postoperative infections, with potential serious complications in case of bacterial contaminations. Therefore, tissue engineering approaches aiming at the regeneration of bone tissue should be based on the use of materials possessing antibacterial properties alongside with biological and functional characteristics. In this study, we investigated the combination of polyhydroxyalkanoates (PHAs) with a novel antimicrobial hydroxyapatite (HA) containing selenium and strontium. Strontium was chosen for its well-known osteoinductive properties, while selenium is an emerging element investigated for its multi-functional activity as an antimicrobial and anticancer agent. Successful incorporation of such ions in the HA structure was obtained. Antibacterial activity against Staphylococcus aureus 6538P and Escherichia coli 8739 was confirmed for co-substituted HA in the powder form. Polymer-matrix composites based on two types of PHAs, P(3HB) and P(3HO-co-3HD-co-3HDD), were prepared by the incorporation of the developed antibacterial HA. An in-depth characterization of the composite materials was conducted to evaluate the effect of the filler on the physicochemical, thermal, and mechanical properties of the films. In vitro antibacterial testing showed that the composite samples induce a high reduction of the number of S. aureus 6538P and E. coli 8739 bacterial cells cultured on the surface of the materials. The films are also capable of releasing active ions which inhibited the growth of both Gram-positive and Gram-negative bacteria.

Bioresorbable and Mechanically Optimized Nerve Guidance Conduit Based on a Naturally Derived Medium Chain Length Polyhydroxyalkanoate and Poly(ε-Caprolactone) Blend.

Severe peripheral nerve injuries represent a large clinical problem with relevant challenges such as the development of successful synthetic scaffolds as substitutes to autologous nerve grafting. Numerous studies have reported the use of polyesters and type I collagen-based nerve guidance conduits (NGCs) to promote nerve regeneration through critical nerve defects while providing protection from external factors. However, none of the commercially available hollow bioresorbable NGCs have demonstrated superior clinical outcomes to an autologous nerve graft. Hence, new materials and NGC geometries have been explored in the literature to mimic the native nerve properties and architecture. Here, we report a novel blend of a natural medium chain length polyhydroxyalkanoate (MCL-PHA) with a synthetic aliphatic polyester, poly(ε-caprolactone) (PCL), suitable for extrusion-based high-throughput manufacturing. The blend was designed to combine the excellent ability of PHAs to support the growth and proliferation of mammalian cells with the good processability of PCL. The material exhibited excellent neuroregenerative properties and a good bioresorption rate, while the extruded porous tubes exhibited similar mechanical properties to the rat sciatic nerve. The NGCs were implanted to treat a 10 mm long sciatic nerve defect in rats, where significant differences were found between thin and thick wall thickness implants, and both electrophysiological and histological data, as well as the number of recovered animals, provided superior outcomes than the well-referenced synthetic Neurolac NGC.

Natural Biomaterials for Cardiac Tissue Engineering: A Highly Biocompatible Solution.

Cardiovascular diseases (CVD) constitute a major fraction of the current major global diseases and lead to about 30% of the deaths, i.e., 17.9 million deaths per year. CVD include coronary artery disease (CAD), myocardial infarction (MI), arrhythmias, heart failure, heart valve diseases, congenital heart disease, and cardiomyopathy. Cardiac Tissue Engineering (CTE) aims to address these conditions, the overall goal being the efficient regeneration of diseased cardiac tissue using an ideal combination of biomaterials and cells. Various cells have thus far been utilized in pre-clinical studies for CTE. These include adult stem cell populations (mesenchymal stem cells) and pluripotent stem cells (including autologous human induced pluripotent stem cells or allogenic human embryonic stem cells) with the latter undergoing differentiation to form functional cardiac cells. The ideal biomaterial for cardiac tissue engineering needs to have suitable material properties with the ability to support efficient attachment, growth, and differentiation of the cardiac cells, leading to the formation of functional cardiac tissue. In this review, we have focused on the use of biomaterials of natural origin for CTE. Natural biomaterials are generally known to be highly biocompatible and in addition are sustainable in nature. We have focused on those that have been widely explored in CTE and describe the original work and the current state of art. These include fibrinogen (in the context of Engineered Heart Tissue, EHT), collagen, alginate, silk, and Polyhydroxyalkanoates (PHAs). Amongst these, fibrinogen, collagen, alginate, and silk are isolated from natural sources whereas PHAs are produced via bacterial fermentation. Overall, these biomaterials have proven to be highly promising, displaying robust biocompatibility and, when combined with cells, an ability to enhance post-MI cardiac function in pre-clinical models. As such, CTE has great potential for future clinical solutions and hence can lead to a considerable reduction in mortality rates due to CVD.

Chemical Modification of Bacterial Cellulose for the Development of an Antibacterial Wound Dressing.

Bacterial cellulose is a bacterially derived polymer with great potential for application in wound healing due to its innate properties such as high biocompatibility and biodegradability. In addition to this, it is naturally biosynthesized by bacteria as a hydrogel, which makes it an optimal substrate for the treatment of dry wounds, where additional moisture is required to facilitate the healing process. However, this polymer lacks antibacterial properties. As bacterial infections are becoming increasingly common and difficult to treat due to antimicrobial resistance, it is of crucial importance to develop strategies for the modification of cellulose to ensure protection against bacterial contamination. In this study, a green-chemistry approach was proposed for the functionalization of cellulose to introduce antibacterial functional groups. Two different active agents, namely glycidyl trimethylammonium chloride and glycidyl hexadecyl ether, were used for the covalent derivatization of the hydroxyl groups of glucose through a heterogeneous reaction in basic aqueous conditions. The modified material was chemically and mechanically characterized by solid-state techniques and rheological measurements. A biological assessment was then carried out both using bacterial cells and human keratinocytes. It was observed that the functionalization performed induced a reduction of approximately half of the bacterial population within 24 h of direct contact with Staphylococcus aureus subsp. aureus Rosenbach 6538PTM and Escherichia coli (Migula) Castellani and Chalmers ATCC® 8739TM (respectively, a reduction of 53% and 43% in the cell number was registered for the two strains). In parallel, cytotoxicity studies performed on keratinocytes (HaCaT cell line) showed cell viability in the range of 90 to 100% for up to 6 days of direct contact with both unmodified and modified samples. The morphology of the cells was also visually evaluated, and no significant difference was noted as compared to the control. Finally, the in vitro scratch assay evidenced good wound closure rates in the presence of the samples, with complete coverage of the scratched area after 5 days for both the modified cellulose and the positive control (i.e., keratinocytes growth medium). Overall, the modified hydrogel showed promising features, confirming its potential as an alternative substrate to develop a sustainable, antibacterial and biocompatible wound dressing.

Electrosprayed Chitin Nanofibril/Electrospun Polyhydroxyalkanoate Fiber Mesh as Functional Nonwoven for Skin Application.

Polyhydroxyalkanoates (PHAs) are a family of bio-based polyesters that have found different biomedical applications. Chitin and lignin, byproducts of fishery and plant biomass, show antimicrobial and anti-inflammatory activity on the nanoscale. Due to their polarities, chitin nanofibril (CN) and nanolignin (NL) can be assembled into micro-complexes, which can be loaded with bioactive factors, such as the glycyrrhetinic acid (GA) and CN-NL/GA (CLA) complexes, and can be used to decorate polymer surfaces. This study aims to develop completely bio-based and bioactive meshes intended for wound healing. Poly(3-hydroxybutyrate)/Poly(3-hydroxyoctanoate-co-3-hydroxydecanoate), P(3HB)/P(3HO-co-3HD) was used to produce films and fiber meshes, to be surface-modified via electrospraying of CN or CLA to reach a uniform distribution. P(3HB)/P(3HO-co-3HD) fibers with desirable size and morphology were successfully prepared and functionalized with CN and CLA using electrospinning and tested in vitro with human keratinocytes. The presence of CN and CLA improved the indirect antimicrobial and anti-inflammatory activity of the electrospun fiber meshes by downregulating the expression of the most important pro-inflammatory cytokines and upregulating human defensin 2 expression. This natural and eco-sustainable mesh is promising in wound healing applications.

Toward a Closed Loop, Integrated Biocompatible Biopolymer Wound Dressing Patch for Detection and Prevention of Chronic Wound Infections.

Chronic wound infections represent a significant burden to healthcare providers globally. Often, chronic wound healing is impeded by the presence of infection within the wound or wound bed. This can result in an increased healing time, healthcare cost and poor patient outcomes. Thus, there is a need for dressings that help the wound heal, in combination with early detection of wound infections to support prompt treatment. In this study, we demonstrate a novel, biocompatible wound dressing material, based on Polyhydroxyalkanoates, doped with graphene platelets, which can be used as an electrochemical sensing substrate for the detection of a common wound pathogen, Pseudomonas aeruginosa. Through the detection of the redox active secondary metabolite, pyocyanin, we demonstrate that a dressing can be produced that will detect the presence of pyocyanin across clinically relevant concentrations. Furthermore, we show that this sensor can be used to identify the presence of pyocyanin in a culture of P. aeruginosa. Overall, the sensor substrate presented in this paper represents the first step toward a new dressing with the capacity to promote wound healing, detect the presence of infection and release antimicrobial drugs, on demand, to optimized healing.

Cytocompatibility Evaluation of a Novel Series of PEG-Functionalized Lactide-Caprolactone Copolymer Biomaterials for Cardiovascular Applications.

Although the use of bioresorbable materials in stent production is thought to improve long-term safety compared to their durable counterparts, a recent FDA report on the 2-year follow-up of the first FDA-approved bioresorbable vascular stent showed an increased occurrence of major adverse cardiac events and thrombosis in comparison to the metallic control. In order to overcome the issues of first generation bioresorbable polymers, a series of polyethylene glycol-functionalized poly-L-lactide-co-ε-caprolactone copolymers with varying lactide-to-caprolactone content is developed using a novel one-step PEG-functionalization and copolymerization strategy. This approach represents a new facile way toward surface enhancement for cellular interaction, which is shown by screening these materials regarding their cyto- and hemocompatibility in terms of cytotoxicity, hemolysis, platelet adhesion, leucocyte activation and endothelial cell adhesion. By varying the lactide-to-caprolactone polymer composition, it is possible to gradually affect endothelial and platelet adhesion which allows fine-tuning of the biological response based on polymer chemistry. All polymers developed were non-cytotoxic, had acceptable leucocyte activation levels and presented non-hemolytic (<2% hemolysis rate) behavior except for PLCL-PEG 55:45 which presented hemolysis rate of 2.5% ± 0.5. Water contact angles were reduced in the polymers containing PEG functionalization (PLLA-PEG: 69.8° ± 2.3, PCL-PEG: 61.2° ± 7.5) versus those without (PLLA: 79.5° ± 3.2, PCL: 76.4° ± 10.2) while the materials PCL-PEG550, PLCL-PEG550 90:10 and PLCL-PEG550 70:30 demonstrated best endothelial cell adhesion. PLLA-PEG550 and PLCL-PEG550 70:30 presented as best candidates for cardiovascular implant use from a cytocompatibility perspective across the spectrum of testing completed. Altogether, these polymers are excellent innovative materials suited for an application in stent manufacture due to the ease in translation of this one-step synthesis strategy to device production and their excellent in vitro cyto- and hemocompatibility.

Comparison of the Influence of 45S5 and Cu-Containing 45S5 Bioactive Glass (BG) on the Biological Properties of Novel Polyhydroxyalkanoate (PHA)/BG Composites.

Polyhydroxyalkanoates (PHAs), due to their biodegradable and biocompatible nature and their ability to be formed in complex structures, are excellent candidates for fabricating scaffolds used in tissue engineering. By introducing inorganic compounds, such as bioactive glasses (BGs), the bioactive properties of PHAs can be further improved. In addition to their outstanding bioactivity, BGs can be additionally doped with biological ions, which in turn extend the functionality of the BG-PHA composite. Here, different PHAs were combined with 45S5 BG, which was additionally doped with copper in order to introduce antibacterial and angiogenic properties. The resulting composite was used to produce scaffolds by the salt leaching technique. By performing indirect cell biology tests using stromal cells, a dose-depending effect of the dissolution products released from the BG-PHA scaffolds could be found. In low concentrations, no toxic effect was found. Moreover, in higher concentrations, a minor reduction of cell viability combined with a major increase in VEGF release was measured. This result indicates that the fabricated composite scaffolds are suitable candidates for applications in soft and hard tissue engineering. However, more in-depth studies are necessary to fully understand the release kinetics and the resulting long-term effects of the BG-PHA composites.

Antimicrobial Materials with Lime Oil and a Poly(3-hydroxyalkanoate) Produced via Valorisation of Sugar Cane Molasses.

A medium chain-length polyhydroxyalkanoate (PHA) was produced by Pseudomonas mendocina CH50 using a cheap carbon substrate, sugarcane molasses. A PHA yield of 14.2% dry cell weight was achieved. Chemical analysis confirmed that the polymer produced was a medium chain-length PHA, a copolymer of 3-hydroxyoctanoate and 3-hydroxydecanoate, P(3HO-co-3HD). Lime oil, an essential oil with known antimicrobial activity, was used as an additive to P(3HO-co-3HD) to confer antibacterial properties to this biodegradable polymer. The incorporation of lime oil induced a slight decrease in crystallinity of P(3HO-co-3HD) films. The antibacterial properties of lime oil were investigated using ISO 20776 against Staphylococcus aureus 6538P and Escherichia coli 8739, showing a higher activity against the Gram-positive bacteria. The higher activity of the oil against S. aureus 6538P defined the higher efficiency of loaded polymer films against this strain. The effect of storage on the antimicrobial properties of the loaded films was investigated. After one-year storage, the content of lime oil in the films decreased, causing a reduction of the antimicrobial activity of the materials produced. However, the films still possessed antibacterial activity against S. aureus 6538P.

Picosecond Laser Ablation of Polyhydroxyalkanoates (PHAs): Comparative Study of Neat and Blended Material Response.

Polyhydroxyalkanoates (PHAs) have emerged as a promising biodegradable and biocompatible material for scaffold manufacturing in the tissue engineering field and food packaging. Surface modification is usually required to improve cell biocompatibility and/or reduce bacteria proliferation. Picosecond laser ablation was applied for surface micro structuring of short- and medium-chain length-PHAs and its blend. The response of each material as a function of laser energy and wavelength was analyzed. Picosecond pulsed laser modified the surface topography without affecting the material properties. UV wavelength irradiation showed halved ablation thresholds compared to VIS wavelength, revealing a greater photochemical nature of the ablation process at UV wavelength. Nevertheless, the ablation rate and, therefore, ablation efficiency did not show a clear dependence on beam wavelength. The different mechanical behavior of the considered PHAs did not lead to different ablation thresholds on each polymer at a constant wavelength, suggesting the interplay of the material mechanical parameters to equalize ablation thresholds. Blended-PHA showed a significant reduction in the ablation threshold under VIS irradiation respect to the neat PHAs. Picosecond ablation was proved to be a convenient technique for micro structuring of PHAs to generate surface microfeatures appropriate to influence cell behavior and improve the biocompatibility of scaffolds in tissue engineering.

Green Composites of Poly(3-hydroxybutyrate) Containing Graphene Nanoplatelets with Desirable Electrical Conductivity and Oxygen Barrier Properties.

Poly(3-hydroxybutyrate), a green polymer originating from prokaryotic microbes, has been used to prepare composites with graphene nanoplatelets (GnP) at different concentrations. The films were fabricated by drop-casting and were hot-pressed at a temperature lower than their melting point to provide the molecular chains enough energy to reorientate while avoiding melting and degradation. It was found that hot-pressing increases crystallinity and improves mechanical properties. The Young's modulus increased from 1.2 to 1.6 GPa for the poly(3-hydroxybutyrate) (P(3HB)) films and from 0.5 to 2.2 GPa for the 15 wt % P(3HB)/GnP composites. Electrical resistivity decreases enormously with GnP concentration and hot-pressing, reaching 6 Ω sq-1 for the hot-pressed 30 wt % P(3HB)/GnP composite. Finally, the hot-pressed P(3HB) samples exhibit remarkable oxygen barrier properties, with oxygen permeability reaching 2800 mL µm m-2 day-1, which becomes 895 mL µm m-2 day-1 when 15% GnP is added to the biopolymer matrix, one of the lowest values known for biopolymers and biocomposites. We propose that these biocomposites are used for elastic packaging and electronics.

Esterase-Cleavable 2D Assemblies of Magnetic Iron Oxide Nanocubes: Exploiting Enzymatic Polymer Disassembling To Improve Magnetic Hyperthermia Heat Losses.

Here, we report a nanoplatform based on iron oxide nanocubes (IONCs) coated with a bioresorbable polymer that, upon exposure to lytic enzymes, can be disassembled increasing the heat performances in comparison with the initial clusters. We have developed two-dimensional (2D) clusters by exploiting benchmark IONCs as heat mediators for magnetic hyperthermia and a polyhydroxyalkanoate (PHA) copolymer, a biodegradable polymer produced by bacteria that can be digested by intracellular esterase enzymes. The comparison of magnetic heat performance of the 2D assemblies with 3D centrosymmetrical assemblies or single IONCs emphasizes the benefit of the 2D assembly. Moreover, the heat losses of 2D assemblies dispersed in water are better than the 3D assemblies but worse than for single nanocubes. On the other hand, when the 2D magnetic beads (2D-MNBs) are incubated with the esterase enzyme at a physiological temperature, their magnetic heat performances began to progressively increase. After 2 h of incubation, specific absorption rate values of the 2D assembly double the ones of individually coated nanocubes. Such an increase can be mainly correlated to the splitting of the 2D-MNBs into smaller size clusters with a chain-like configuration containing few nanocubes. Moreover, 2D-MNBs exhibited nonvariable heat performances even after intentionally inducing their aggregation. Magnetophoresis measurements indicate a comparable response of 3D and 2D clusters to external magnets (0.3 T) that is by far faster than that of single nanocubes. This feature is crucial for a physical accumulation of magnetic materials in the presence of magnetic field gradients. This system is the first example of a nanoplatform that, upon exposure to lytic enzymes, such as those present in a tumor environment, can be disassembled from the initial 2D-MNB organization to chain-like assemblies with clear improvement of the heat magnetic losses resulting in better heat dissipation performances. The potential application of 2D nanoassemblies based on the cleavable PHAs for preserving their magnetic losses inside cells will benefit hyperthermia therapies mediated by magnetic nanoparticles under alternating magnetic fields.