Production of Polyhydroxyalkanoates and their application for coronary stent development

Development of coronary artery stents have revolutionised the treatment of coronary artery disease. However, the currently available stents have several limitations and side effects, which still need to be addressed. Within this group of biomaterials Polyhydroxyalkanoates (PHAs), which are biopolyesters produced by bacteria, have a great potential in coronary stent development. They are biodegradable, non-toxic and biocompatible. PHAs have a broad range of properties, which can be tailored based on the requirements of the application by an appropriate bacterial strain, carbon source or fermentation conditions. The main aim of this study was the production of PHAs and their application as a platform material for coronary stent. Poly(3-hydroxybutyrate), P(3HB), short chain length PHA (scl-PHA), was obtained from B.subtilis OK2 using glucose as the sole carbon source. P. mendocina was selected for the economical production of medium chain length PHA (mcl-PHAs) using waste frying oil as the carbon source. Produced mcl-PHA copolymer was hydrolysed in order to obtain lower molecular weight PHA (oligo-PHA), which was used as the plasticising agent for P(3HB) in order to alter material properties of the scl-PHA polymer. Novel P(3HB)/oligo-PHA blends, using different ratios of P(3HB) and oligo-PHA, which were 100:0 (P(3HB), 95:5 (P(3HB): oligo-PHA), 90:10 (P(3HB): oligo-PHA) and 80:20 (P(3HB): oligo-PHA) have been developed. Addition of oligo-PHA to P(3HB) resulted not only in the enhancement of mechanical properties of the material, but also improved surface topography and increased the biocompatibility of the scaffolds in comparison to neat P(3HB). The P(3HB)/oligo-PHA 90/10 blend was selected for the fabrication of radiopaque composites using addition of 1%, 3% and 5% wt barium sulphate. BaSO4 is well-known contrast agent, commonly used in biomedical applications. The composites were fully characterised with respect to material properties, surface topography and biocompatibility. Preliminary studies, performed by MicroCT confirmed the radiopacity of the composite material. The composites exhibited an increased surface roughness, lower contact angle values and higher protein adsorption in comparison to the neat blend without barium sulphate. The observed cell viability was significantly better in comparison to standard Tissue Culture Plastic (TCP). The P(3HB)/oligo-PHA 90:10 blend was selected as a base material for tube manufacturing by the dip moulding technique. Mechanical properties of the blend tubes were similar to those obtained for flat films, except for tensile strength, which was slightly higher in tubes. An incubation of tubes in DMEM media at 37°C for 7 weeks resulted in the decrease values of tensile strength, Young’s Modulus values and elongation at break in comparison to the tubes stored at room temperature without any media. Additionally, In vitro degradation study was carried out on the tubes. After 6 months of incubation in PBS,The P(3HB)/oligo-PHA 90/10 tubes underwent a 30% weight loss. pH of the media has dropped insignificantly, due to release of slightly acidic degradation products. Scanning Electron Microscope (SEM) images confirmed an enlarged porosity in the tube wall with an increased time of incubation. Furthermore, an investigation of the P(3HB)/oligo-PHA 90/10 tubes coated with PCL-PEG550 with incorporated drugs: rapamycin or tacrolimus was carried out for the development of a drug eluting biodegradable stent. The percentage viability of the HMEC-1 cells was significantly higher on the tubes with tacrolimus compared to the tubes with rapamycin. Controlled release of both drugs was observed within 90 days, which is an encouraging result for the development of biodegradable drug eluting stents.There was a clear cytotoxic effect of rapamycin on the HMEC-1 cells, whereas, presence of tacrolimus did not exhibit similar effect on cell attachment and viability. Haemocompatiblity studies were performed in order to investigate the reaction of blood cells after direct contact with PHAs polymer samples. Obtained results confirmed the non-haemolytic effect of PHAs on erythrocytes and low risk of thrombogenicity. Although all tested materials demonstrated slightly elevated levels of monocytes and neutrophil activation in comparison to fresh whole blood, obtained values were much lower in comparison to poly(L-lactic acid) (PL38), a well-established medical polymer currently used for coronary stent application. In conclusion, the results obtained in this work confirmed that PHAs have an excellent potential as suitable platform matrices for coronary stent application

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

Twenty Years of Student Scholarship: Celebrating the Dalhousie Journal of Legal Studies

In this paper, we propose a novel controllable text-to-image generative adversarial network (ControlGAN), which can effectively synthesise high-quality images and also control parts of the image generation according to natural language descriptions. To achieve this, we introduce a word-level spatial and channel-wise attention-driven generator that can disentangle different visual attributes, and allow the model to focus on generating and manipulating subregions corresponding to the most relevant words. Also, a word-level discriminator is proposed to provide fine-grained supervisory feedback by correlating words with image regions, facilitating training an effective generator which is able to manipulate specific visual attributes without affecting the generation of other content. Furthermore, perceptual loss is adopted to reduce the randomness involved in the image generation, and to encourage the generator to manipulate specific attributes required in the modified text. Extensive experiments on benchmark datasets demonstrate that our method outperforms existing state of the art, and is able to effectively manipulate synthetic images using natural language descriptions. Code is available at https://github.com/mrlibw/ControlGAN.Comment: NeurIPS 201

Fredkin Gates for Finite-valued Reversible and Conservative Logics

The basic principles and results of Conservative Logic introduced by Fredkin and Toffoli on the basis of a seminal paper of Landauer are extended to d-valued logics, with a special attention to three-valued logics. Different approaches to d-valued logics are examined in order to determine some possible universal sets of logic primitives. In particular, we consider the typical connectives of Lukasiewicz and Godel logics, as well as Chang's MV-algebras. As a result, some possible three-valued and d-valued universal gates are described which realize a functionally complete set of fundamental connectives.Comment: 57 pages, 10 figures, 16 tables, 2 diagram

Production of a novel medium chain length Poly(3-hydroxyalkanoate) using unprocessed biodiesel waste and its evaluation as a tissue engineering scaffold

This study demonstrated the utilisation of unprocessed biodiesel waste as a carbon feedstock for Pseudomonas mendocina CH50, for the production of PHAs. A PHA yield of 39.5% CDM was obtained using 5% (v/v) biodiesel waste substrate. Chemical analysis confirmed that the polymer produced was poly(3-hydroxyhexanoate-co-3-hydroxyoctanoate-co-3- hydroxydecanoate-co-3-hydroxydodecanoate) or P(3HHx-3HO-3HD-3HDD). P(3HHx-3HO- 3HD-3HDD) was further characterised and evaluated for its use as a tissue engineering scaffold (TES). This study demonstrated that P(3HHx-3HO-3HD-3HDD) was biocompatible with the C2C12 (myoblast) cell line. In fact, the % cell proliferation of C2C12 on the P(3HHx-3HO-3HD-3HDD) scaffold was 72% higher than the standard tissue culture plastic confirming that this novel PHA was indeed a promising new material for soft tissue engineering

Binary Polyhydroxyalkanoate Systems for Soft Tissue Engineering

Progress in tissue engineering is dependent on the availability of suitable biomaterials. In an effort to overcome the brittleness of poly(3-hydroxybutyrate), P(3HB), a natural biodegradable polyester, and widen its biomedical applications, plasticising of P(3HB) with oligomeric substances of related structure has been studied. A biosynthesised medium-chain-length polyhydroxyalkanoate (mcl-PHA) copolymer, the plasticizer precursor, was obtained using vegetable waste frying oil as a sole carbon source. The mcl-PHA was transformed into an oligomeric derivative by acid hydrolysis. The plasticising effect of the oligomeric mcl-PHA on P(3HB) was studied via characterisation of thermal and mechanical properties of the blends in the course of ageing at ambient conditions. Addition of oligomeric mcl-PHA to P(3HB) resulted in softer and more flexible materials based entirely on PHAs. It was shown that the oligomeric mcl-PHA transformed highly crystalline P(3HB) into materials with a dominant amorphous phase when the content of oligomeric mcl-PHA exceeded 10wt%. In vitro biocompatibility studies of the new binary PHA materials showed high viability and proliferation of C2C12 myoblast cells. Thus, the proposed approach for P(3HB) plasticisation has the potential for the generation of more pliable biomaterials based on P(3HB) which can find application in unique soft tissue engineering applications where a balance between stiffness, tensile strength and ductility is required

Biosynthesis and characterization of a novel, biocompatible medium chain length polyhydroxyalkanoate by Pseudomonas mendocina CH50 using coconut oil as the carbon source

This study validated the utilization of triacylglycerides (TAGs) by Pseudomonas mendocina CH50, a wild type strain, resulting in the production of novel mcl-PHAs with unique physical properties. A PHA yield of 58% dcw was obtained using 20 g/L of coconut oil. Chemical and structural characterisation confirmed that the mcl-PHA produced was a terpolymer comprising of three different repeating monomer units, 3-hydroxyoctanoate, 3-hydroxydecanoate and 3-hydroxydodecanoate or P(3HO-3HD-3HDD). Bearing in mind the potential of P(3HO-3HD-3HDD) in biomedical research, especially in neural tissue engineering, in vitro biocompatibility studies were carried out using NG108-15 (neuronal) cells. Cell viability data confirmed that P(3HO-3HD-3HDD) supported the attachment and proliferation of NG108-15 and was therefore confirmed to be biocompatible in nature and suitable for neural regeneration

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

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