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  • About
  • The Global ETD Search service is a free service for researchers to find electronic theses and dissertations. This service is provided by the Networked Digital Library of Theses and Dissertations.
    Our metadata is collected from universities around the world. If you manage a university/consortium/country archive and want to be added, details can be found on the NDLTD website.
1

Investigation into synthetic protease-sensitive hydrogels as siRNA nanoparticle delivery vehicles.

Doubell, Emma 18 February 2019 (has links)
RNA interference (RNAi) is receiving increasing attention as a form of gene regulation able to temporarily silence gene expression through post transcriptional mechanisms. RNAi agents have shown promise in targeting a range of ailments from cancers to myocardial infarction. However, RNAi based therapeutics have not passed the clinical trial stage of development, in part due to lack of optimal delivery mechanisms. One means towards improving delivery is the development of localised and sustained delivery systems for the RNAi molecule. Such approaches are important as it has previously been found that systemic delivery often leads to off target effects and rapid clearance. The aim of this project has been to assess the utility of an enzymatically degradable polyethylene glycol (PEG) hydrogel as a localised delivery vehicle. Enzymatically degradable PEG hydrogels that rely on cellular invasion for their degradation might enable the release of entrapped RNAi agents to surrounding cells as well as transfection of invading cells. The cationic polymer poly(ethyleneimine) (PEI), was used as a tool to investigate the PEG hydrogel as a localised delivery vehicle. Thus, an initial objective of this study was the establishment and characterisation of the PEI/siRNA nanoparticle technology in our group. siRNA was found to be fully complexed at a PEI nitrogen to siRNA phosphate ratios of 5:1 and higher. A 10:1 ratio and higher were able to protect the siRNA from RNases present in serum for 7 days with up to 65% of RNA still intact. A 40:1 ratio was found to be cytotoxic. A 20:1 ratio was found to be the most effective at gene knock down and determined to be optimal. As there is a relatively limited volume available in the PEG hydrogel system used here for loading with PEI/siRNA nanoparticles, a study was conducted to assess the maximum concentration of siRNA at which effective nanoparticles could be formed. Somewhat unexpectedly it was found that at a concentration of 7.5 µM siRNA, nanoparticles showed a significant 40% reduction in transfection efficacy of cells cultured in 2D. This finding for PEI nanoparticles indicating a limitation in the dosage attainable in the PEG hydrogel system. The influence of PEG encapsulation for PEI/siRNA nanoparticles on RNase protection was assessed by exposing hydrogels with entrapped nanoparticles to serum RNases. The PEG hydrogel was found to significantly improve PEI based protection from RNase degradation in the initial 24 hours and this protection persisted for up to 5 days. PEI/siRNA nanoparticles were retained after encapsulation within PEG hydrogels for up to 7 days whilst siRNA alone was entirely released after 24 hours. It is possible that this is a limitation of the system. A 3D invasion assay was developed to more closely mimic the in vivo scenario where cells could invade a PEG hydrogel with entrapped nanoparticles but are not initially exposed to high concentrations of nanoparticles prior to hydrogel polymerisation. The assay involved the formation of dermal equivalents (cells entrapped within contracted collagen) that were encapsulated within a PEG hydrogel. Cells were observed invading the hydrogel within 1 hour of polymerising and continued to do so for up to 7 days. Invading cells were seen to take up the fluorescent siRNA although this uptake was scant and challenging to quantify. When commercial cell death siRNA sequence nanoparticles were encapsulated, a trend towards higher death levels was observed. In conclusion, PEI/siRNA nanoparticles and related RNAi based assays have been formally established in our laboratory and can be used in the future for other applications. A 3D cell invasion assay was developed which more closely mimics the in vivo scenario where hydrogels bearing siRNA are polymerised within tissue. The increased RNase protection though relatively slight is important for the use of hydrogels as localised delivery depots in an in vivo environment. Potential dose limitations of the PEG hydrogel system when using PEI nanoparticles and their apparent very tight encapsulation in the PEG hydrogels suggests the need for modifications of both nanoparticles and hydrogels to optimise efficacy. Future investigations into scaffold based localised siRNA delivery should be facilitated by the methodologies and assays established in this study.
2

Evaluation of cellular interactions with functionalized scaffolds for cardiovascular tissue engineering

Wimberley, Olivia 22 March 2022 (has links)
Introduction: Cardiovascular disease is a leading cause of death worldwide, with heart valve disease in particular becoming a rising problem in developing countries. Tissue engineering offers the next step in heart valve tissue replacement surgery. Synthetic scaffolds used in tissue engineering often do not have sufficient cell adhesion, thus the addition of biosignals is crucial. Surface modifications can be used to improve desired cell adhesion and proliferation. To covalently attach cell adhesion peptides/biosignals, synthetic hydrogel spacers are often used in an interim grafting step. Poly (acrylic acid) (PAA) is a non-toxic, FDA approved hydrogel that has been shown in preliminary studies (previous research and own experience) to improve cell adhesion even without the addition of specific cell-binding compounds. This project aims to show the effect of systematically increasing the concentration of PAA modified 2D and 3D polyurethanes (biodegradable and biostable) on cell adhesion, persistence, and proliferation. Experimental Methods: In Part 1, 2D nondegradable Pellethane® films were surface modified by varying the poly (acrylic acid-co-acrylamide) (P(AA-co-AM)) comonomer feed ratio from 0 to 100 % PAA in 20 % increments, using poly (acrylamide) (PAM) as a copolymer (unmodified and collagen coated controls). Surface properties were analysed using SEM imaging, staining, energy-dispersive X-ray spectrometry (SEM-EDS) and toluidine blue carboxyl assays (TBCA). Endothelial cells were isolated from human saphenous veins using an enzymatic digestion method, and identified by staining with DAPI and Cy3 against CD31. Isolated endothelial cells and human dermal fibroblasts (Cell bank: R039/2016) were seeded onto Pellethane® films (8 000 cells/film). Live/dead staining and XTT cell viability assays were performed over 24 and 72 hrs, respectively. Following this, XTT cell viability assays were performed at 7 and 24 hrs post-seeding on endothelial cells cultured under serum-free conditions (20 000 cells/film; unmodified and 80 % PAA). In Part 2, both Pellethane® and DegraPol® (degradable) 2D films and 3D electrospun scaffolds were used. The polymer samples were surface modified with 0, 40, and 80 % PAA. All samples were imaged using SEM prior to in vitro cell culture evaluation. Endothelial cells were seeded (8 000 cells/film) onto surface modified polymer samples, and XTT cell viability assays were performed over 72 hrs. Three-dimensional scaffolds seeded with endothelial cells (20 000 and 50 000 cells/film; unmodified and 80 % PAA) were immunocytochemically stained (Hoechst and CD31) at Day 1, 3, and 7 post-seeding. Results and discussion: In Part 1, SEM imaging and preliminary staining confirmed the addition of PAA to polymer surfaces. Systematically increasing [AA] in the P(AA-co-AM) comonomer ratio resulted in the expected increase in surface-COOH functional groups (TBCA and SEM-EDS). The number of COOH groups increased as [PAA] increased from 0-40 % (R 2=0.76; P 0.0001) before plateauing (TBCA). This was further confirmed by a decreasing N/O ratio with increasing [AA] monomer (R2=0.70; P< 0.001) (SEM-EDS). An increase in [PAA] resulted in a linear increase in endothelial cell adhesion and persistence (R 2=0.92 (live/dead staining) and 0.96 (XTT cell viability assays); P< 0.05). Endothelial cell viability on surfaces modified with 80 and 100 % PAA was comparable to that achieved on the collagen positive control. High concentrations of PAA also showed improved fibroblast adhesion (R2= 0.71 (live/dead staining) and 0.54 (XTT cell viability assays); P< 0.05) but did not display any persistence or viability close to that obtained on the collagen. Collagen coated surfaces displayed the highest cell adhesion and proliferation for both cell types (XTT cell viability assays and live/dead staining). Endothelial cell adhesion was improved by both the addition of PAA to the polymer surface, and FBS to the cell culture medium (P< 0.05) (cells cultured under serum-free conditions). In Part 2, the improvement of endothelial cell adhesion on PAA modified 2D Pellethane® films was confirmed and additionally shown on 2D DegraPol® (P ≤0.05) (XTT cell viability assays). However, the endothelial cell persistence seen in earlier assays was not observed. The positive effect of increasing [PAA] did not translate to 3D scaffolds, and cell behaviour was improved on unmodified surfaces in comparison to any of the PAA modified groups (XTT cell viability assays and immunocytochemical staining). This discrepancy is proposed to be a difference in grafting efficiency on the degradable materials and the 3D structure of the electrospun scaffolds. Conclusions: An increase in PAA surface modification on polyurethane can improve endothelial cell adhesion and persistence on nondegradable 2D polyurethane scaffolds. These results did not translate to electrospun scaffolds, probably due to the complex 3D cell environment. Further investigation is required for use in TEHV and other applications.
3

Stimulation of angiogenesis through growth factor delivery from synthetic heparinised hydrogels

Chokoza, Cindy January 2017 (has links)
Objectives: Vascular diseases are one of the leading causes of death. Due to minimal regenerative capability of the heart, alternative therapies have been sought after with engineered biomaterials being extensively investigated in this area. In this study, enzymatically degradable heparinised polyethylene glycol (PEG-Hep) hydrogels were synthesized and characterised for the binding and controlled release of vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF), as well as their bioactivity and angiogenic potential in vivo. Methodology: VEGF and bFGF were combined into 4% (m/v) PEG-Hep hydrogels. The binding and release rates of VEGF and BFGF were analysed via an immunosorbent assay. Released growth factor bioactivity was measured using an XTT metabolic assay on human saphenous vein endothelial cells and human dermal fibroblasts. Neovascularisation was quantified in a subcutaneous rat angiogenesis model in which hydrogel growth factor combinations were implanted within porous polyurethane discs and analysed after a 4 week period. A novel hybrid hydrogel able to degrade proteolytically and hydrolytically was further developed, characterised and analysed as above. Results: PEG-Hep hydrogels demonstrated substantial growth factor binding ability (500-600 ng) and allowed sustained release (10-20 ng/day) for up to 28 days. Bioactivity of the growth factors was retained throughout the release period. The degradation rate of the hydrogels could be controlled in vivo by varying the ratio of monomers capable of forming either hydrolytically or proteolytically degradable crosslinks. Qualitative and quantitative analysis demonstrated a pronounced and significant angiogenic response in vivo (p<0.05). Conclusion: Heparinised PEG hydrogels show significant promise as controlled release vehicles for growth factors and warrant further examination in a myocardial infarction model.
4

Pulsatile Electropolishing of Nitinol Stents

Cloete, Jeran Andre 20 July 2022 (has links) (PDF)
Alloys that oxidize easily such as those containing titanium or chromium present a challenge to electropolishing because the polarization that dissolves the metal species produces positive ions, these oxidize and form stable surface layers of metallic oxides that prevent further dissolution. This is usually overcome with the use of acid solutions that dissolve the metallic oxide. This thesis aims to shift the primary control of the electropolishing e_ect from electrolyte variables to a combination of potential variation and hydrodynamic interference. Traditionally this is achieved with one continuous mass removal process that operates after a steady state of dissolution is established, generally requiring hydro_uoric or phosphoric acid to achieve titanium dioxide breakdown. The resulting concentration gradient is heavily a_ected by electrolyte variables such as viscosity and electrical resistance, while the electrical polarization is constrained by the metallic oxide reaction rate which creates a complex net of interdependent variables that can be di_cult to tune. A rapidly changing electric _eld was applied to modulate the alloying element dissolution rates. In tandem with the electropolishing development, stages prior to the electropolishing step were selectively removed to simplify the process. Utilizing a three electrode system and an external potentiostat controller to permit greater _exibility, a variety of alternating current pulsatile waveforms were investigated and the resulting e_ect on surface topology was observed using SEM and AFM microscopes. Di_erential pulse voltammogram yielded a feedback parameter on surface composition, and various pulse parameters were adjusted to optimize for surface smoothness, and identify the primary control variable. An electropolishing method is presented which achieves a :50% reduction in the Sa surface roughness value to an area average of 45 nm on a laser cut tubular stent geometry. It is shown that this method can be adapted to eliminate the need for chemical etching or mechanical polishing prior to electropolishing. The resulting polished surface displays corrosion resistance equivalent or better than other electropolished Nitinol surfaces from literature with a breakdown potential >1V vs SCE, and a similarly high repassivation potential. Balancing the charge in the anodic and cathodic pulses was the key to minimizing the resulting surface roughness, and eliminating micropits. Nitinol is a nearly binary alloy of NiTi and a charge transfer ratio of 1 yielded the smoothest surfaces at current densities around :1 A/cm2. The initial surface condition was found to be irrelevant to electropolishing control with respect to oxide composition, provided enough mass was removed to fully dissolve the initial layers of mixed composition.
5

Polymeric Transcatheter Heart Valves

Coetzee, Johan 20 July 2022 (has links) (PDF)
Rheumatic heart disease (RHD) is one of the main causes of heart disease in the emerging world. Once the disease has become symptomatic valve repair or replacement is the only treatment currently available. Commercial bioprosthetic valves (surgical and transcatheter) suffer from calcification and decreased durability, especially when implanted into younger patients, while mechanical surgical valves require life-long anticoagulation. Polymeric transcatheter aortic valve insertion (TAVI) is proposed as a solution to provide long-term durability without the need for anticoagulation. A manufacturing method involving the spray coating of polyurethane solutions onto valve moulds, pre-coating a TAVI stent with a polyurethane of higher durometer, and subsequently spraying the combination of the stent and the mould to form integral attachments of the leaflets, is described. Valves were tested for leaflet thickness distribution, hydrodynamic function and accelerated durability, and subsequently implanted in an acute ovine TAVI model as proof of concept. Effective Orifice Area (EOA) >1.7 cm2, regurgitation < 10% and transvalvular pressure gradient < 10 mmHg were achieved. Leaflet thickness correlated indirectly with the EOA and directly with transvalvular pressure gradient, but not regurgitation. After iterative improvements in the manufacturing process, valves with average thicknesses ranging from 140 to 160 μm showed highest durability (>150 million, and up to 600 million cycles). Surface roughness was reduced by applying a final pure solvent coat. Implanted valves showed good function, with no apparent central or paravalvular regurgitation, perfusion of coronaries, and EOA greater than1.6 cm2. In conclusion, a polymeric TAVI valve made by a new manufacturing method showed durability up to 15 years equivalent in vitro, and good hydrodynamic function in vitro and in vivo. The devices hold many potential advantages in terms of automation and cost of manufacturing, as well as function and longevity.
6

Paravalvular sealing pf percutaneous heart valves

Conradie, David Gideon 14 July 2022 (has links)
Paravalvular regurgitation (PVR), which frequently occurs after transcatheter aortic valve replacements (TAVR) can lead to adverse clinical consequences and has been shown to correlate to an increased late mortality and morbidity. Quantification, graduation and testing for PVR has proven challenging and a standardized method for pre-clinical testing is still sought. Commercial transcatheter heart valves (THV's) rely on sealing skirts made from treated pericardium or polyethylene terephthalate (PET) fabrics. The current study was aimed at developing novel electrospun skirts for the minimization of PVR in THV's. Thermoplastic polyurethane (TPU) was electrospun onto mandrels and several techniques (CO2 - laser, ultra-sonic, solvent and conductive heat bonding) used to attach the scaffolds to pre-coated TAVR stents. Attachment strength was modelled by finite element analysis (FEA) of stents in the crimped and expanded conditions and empirically determined by physical pull-off tests. PVR was evaluated for four different skirt designs (1× single layer “FLAT” and 3× double layers: Forward Flow Filling “FFF”, No Filling “NF” and Back Flow Filling “BFF”) using a pulse duplicator fitted with perforated mounting rings. Optimization of solution, process and environmental parameters yielded scaffolds with average fibre diameters of 3.17 ± 0.64 μm and average pore sizes 9.52 ± 6.90 μm. Tensile strength was found to be similar in the direction perpendicular to collector rotation [Abstract incomplete due to DSpace NOT being able to accommodate some formulas and equations in the abstract, BS]
7

A Poly-N-isopropylacrylamide Based Hydrogel as a Novel Material for Vitreous Replacement

Perry, Robyn 04 1900 (has links)
<p>Although improvements in vitreous replacement technologies have lead to more desirable outcomes in patients with vision problems such as retinal detachment and vitreous hemorrhage, there remain a number of issues with current approaches. Many common therapies are limited to specific cases, or require extensive post-operative positioning, follow-up, and even secondary surgery for removal.</p> <p>Thermally responsive materials composed of N-isopropylacrylamide, N-acryloxysuccinimide, and poly(ethylene glycol) were obtained through free-radical polymerization and were examined for the potential for use as artificial vitreous replacements. While in solution these materials were capable of transitioning from free-flowing liquids at room temperature to viscous solutions at physiological temperatures. Solutions of these materials were shown to have refractive indices similar to currently used vitreous replacements and gel-like mechanical properties at 37<sup>o</sup>C. Optical transparency studies were performed and most samples displayed adequate transmittance values in the visible spectrum. Co-monomer content and molecular weight were found to have an effect on the temperature of the phase transition, as well as the transparency and cellular compatibility of the solutions at 37<sup>o</sup>C.</p> <p>While the materials studied require further optimization, these results suggest that poly(N-isopropylacrylamide) based polymers may have potential for use in vitreous replacement therapies</p> / Master of Applied Science (MASc)
8

SILICA PARTICLES FOR HYDROPHOBIC DRUG DELIVERY FROM SILICONE HYDROGELS

Fernandes, Michelle D. 10 1900 (has links)
<p>Topical administration of therapeutic eye drops is currently the most employed method of drug delivery for treatment of various ocular conditions. The poor drug bioavailability with this method of delivery requires frequent administration and results in low patient compliance. Drug eluting contact lenses have been proposed as a means to overcome the challenges associated with topical drug delivery due to the ability to increase drug residence time on the ocular surface and consequently increase corneal absorption. In this study, silica particles were prepared using microemulsion polymerization, and were embedded into a model silicone hydrogel consisting hydroxyethylmethacrylate (HEMA) monomer and a hydroxylated silicone monomer. Silica particles were specifically selected as the drug-containing particle system for loading into a silicone hydrogel to observe whether better control over the rate of drug release was possible by loading the hydrophobic drug in the silicone domain via the use of silica particles. Although similar dexamethasone release profiles were observed for hydrogels incorporated with suspensions prepared with or without silica shell precursors, transmission electron micrographs of these hydrogels indicate particle localization in silicone domains for suspensions prepared with silica precursors. This result is promising, and optimization of the drug-containing silica particle synthesis formulation may potentially allow for control over the drug release profile.</p> / Master of Applied Science (MASc)
9

Denture soft linings : clinical and laboratory studies

Jepson, Nicholas John Arnold January 1991 (has links)
No description available.
10

Characterisation and evaluation of hydroxyapatite and bioglass-reinforced polyethylene composities for medical implants

Huang, Jie January 1997 (has links)
No description available.

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