Recent advances in drug delivery systems based on polypeptides prepared fromN-carboxyanhydrides

Recent years have seen a dramatic increase in interest in the use of ring-opening polymerization of N-carboxyanhydride monomers as a method to prepare well-defined polypeptides and peptide hybrid materials. The resulting molecules are often capable of assembling into a variety of different structures, including micelles, vesicles, nanoparticles and hydrogels, and therefore have been explored as novel drug delivery systems. Peptides are attractive in this regard due to their rich chemical functionality and ability to assemble through the formation of secondary structures (e.g. α-helices and β-sheets). In addition, they are inherently biodegradable and biocompatible. This review describes recent advances in the field, covering aspects such as improved methods with which to prepare better-defined polypeptides, crosslinking of assemblies to enhance biostability, the preparation of materials that respond to a variety of stimuli (including light and intra- or extracellular redox conditions), functionalization with targeting ligands to enhance cellular uptake, assemblies for small interfering RNA delivery and approaches to theranostic systems. © 2014 Society of Chemical Industr

New tools for visualising nanoparticle drug delivery

Encapsulating drugs in polymeric nanoparticles (NPs) is becoming increasingly popular for targeted and sustained drug delivery. NP drug delivery systems can increase the lifetime of therapeutics in vivo, they can improve safety by allowing a lower dose to be administered, and they are able to pass the blood brain barrier. Chapter 1 will discuss the anatomy of the brain, how it can be affected by multiple sclerosis (MS) and why NP drug delivery has an important part to play in treating this disease. Due to the nanoscale size of these drug delivery systems, it is challenging to image their uptake, distribution and fate in a biological environment. Raman scattering is a vibrational technique which can probe the chemical bonds in a sample, however, it is a very weak effect. Stimulated Raman scattering (SRS) is a dual laser technique which increases the observed Raman signal and has been used to image biological samples at video-rate. Intracellular contrast can be increased further by introducing spectroscopically bioorthogonal chemical labels to the NPs, which appear in the so called cell-silent region of the Raman spectrum. Previous work to image NPs with fluorescence and Raman microscopies will be discussed in Chapter 1. In this thesis, Raman spectroscopy was used to image bioorthogonally labelled polymeric NPs in in vitro cellular and ex vivo tissue models of the brain. The polymer poly(lactic acid-co-glycolic acid) (PLGA) was chosen as a biocompatible and biodegradable polymer widely used in drug delivery. Chapter 2 describes the synthesis of PLGA with both carbon-deuterium and alkyne modifications which both produce Raman peaks in the cell-silent region. The optimisation of NP synthesis from these labelled polymers by the emulsification-evaporation method is discussed in Chapter 3. In Chapter 4, the NPs synthesised from both the deuterium and alkyne analogues of PLGA were imaged with SRS in biological models of the brain. Tuning to the bioorthogonal peaks allowed imaging of the NPs without cellular background. Both NP analogues were imaged in primary rat microglia, the macrophages of the brain, and additionally the alkyne labelled NPs were imaged in ex vivo cortical mouse brain slices. Immunohistochemical analysis of these brain slices confirmed that the NPs were selectively taken up into microglia

Block copolypeptide nanoparticles for the delivery of ocular therapeutics

Self-assembling block copolypeptides were prepared by sequential ring-opening polymerization of N-carboxyanhydride (NCA) derivatives of γ-benzyl-L-glutamic acid and ε-carbobenzyloxy-L-lysine, followed by selective deprotection of the benzyl glutamate block. The synthesized polymers had number average molecular weights close to theoretical values, and had low dispersities (ĐM = 1.15–1.28). Self-assembly of the amphiphilic block copolymers into nanoparticles was achieved using the “solvent-switch” method, whereby the polymer was dissolved in THF and water and the organic solvent removed by rotary evaporation. The type of nanostructures formed varied from spherical micelles to a mixture of spherical and worm-like micelles, depending on copolymer composition. The spherical micelles had an average diameter of 43 nm by dynamic light scattering, while the apparent diameter of the mixed phase system was around 200nm. Reproducibility of nanoparticle preparation was demonstrated to be excellent; almost identical DLS traces were obtained over three repeats. Following qualitative dye-solubilization experiments, the nanoparticles were loaded with the ocular anti-inflammatory drug dexamethasone. Loading efficiency of the nanoparticles was 90% and the cumulative drug release was 94% over 16 d, with a 20% burst release in the first 24 h.mabi201400471-gra-000

Avoiding the Pitfalls of siRNA Delivery to the Retinal Pigment Epithelium with Physiologically Relevant Cell Models

Inflammation is involved in the pathogenesis of several age-related ocular diseases, such as macular degeneration (AMD), diabetic retinopathy, and glaucoma. The delivery of anti-inflammatory siRNA to the retinal pigment epithelium (RPE) may become a promising therapeutic option for the treatment of inflammation, if the efficient delivery of siRNA to target cells is accomplished. Unfortunately, so far, the siRNA delivery system selection performed in dividing RPE cells in vitro has been a poor predictor of the in vivo efficacy. Our study evaluates the silencing efficiency of polyplexes, lipoplexes, and lipidoid-siRNA complexes in dividing RPE cells as well as in physiologically relevant RPE cell models. We find that RPE cell differentiation alters their endocytic activity and causes a decrease in the uptake of siRNA complexes. In addition, we determine that melanosomal sequestration is another significant and previously unexplored barrier to gene silencing in pigmented cells. In summary, this study highlights the importance of choosing a physiologically relevant RPE cell model for the selection of siRNA delivery systems. Such cell models are expected to enable the identification of carriers with a high probability of success in vivo, and thus propel the development of siRNA therapeutics for ocular disease.Peer reviewe

Avoiding the Pitfalls of siRNA Delivery to the Retinal Pigment Epithelium with Physiologically Relevant Cell Models

Inflammation is involved in the pathogenesis of several age-related ocular diseases, such as macular degeneration (AMD), diabetic retinopathy, and glaucoma. The delivery of anti-inflammatory siRNA to the retinal pigment epithelium (RPE) may become a promising therapeutic option for the treatment of inflammation, if the efficient delivery of siRNA to target cells is accomplished. Unfortunately, so far, the siRNA delivery system selection performed in dividing RPE cells in vitro has been a poor predictor of the in vivo efficacy. Our study evaluates the silencing efficiency of polyplexes, lipoplexes, and lipidoid-siRNA complexes in dividing RPE cells as well as in physiologically relevant RPE cell models. We find that RPE cell differentiation alters their endocytic activity and causes a decrease in the uptake of siRNA complexes. In addition, we determine that melanosomal sequestration is another significant and previously unexplored barrier to gene silencing in pigmented cells. In summary, this study highlights the importance of choosing a physiologically relevant RPE cell model for the selection of siRNA delivery systems. Such cell models are expected to enable the identification of carriers with a high probability of success in vivo, and thus propel the development of siRNA therapeutics for ocular disease

Spatial scale influences how people value and perceive green open space

It is important for landscape planners and managers to understand how urban residents value and interact with green open spaces. However, the effect of spatial scale on values and perceptions of green open spaces has to date received little attention. This study explored the influence of spatial scale using Public Participation GIS (PPGIS) methods in the Lower Hunter region of Australia. By asking respondents to assign markers denoting various values and preferences to green spaces displayed on maps of their suburb and municipality, the influence of scale could be assessed experimentally. A greater abundance and diversity of value markers were consistently assigned at the suburb scale, yet this pattern was more pronounced for some values (e.g. physical activity) than others (e.g. nature, cultural significance). The strength of this relationship was related to socio-demographic variables such as education and income. These results have implications for understanding human-environment relationships and the use of PPGIS techniques to inform environmental planning

Modelling maternal obesity: the effects of a chronic high-fat, high-cholesterol diet on uterine expression of contractile-associated proteins and ex vivo contractile activity during labour in the rat

Maternal obesity is associated with prolonged and dysfunctional labour and emergency caesarean section, but the mechanisms are unknown. The present study investigated the effects of an adiposity-inducing high fat, high-cholesterol (HFHC) diet on uterine contractile associated protein (CAP) expression and ex vivo uterine contractility in term non-labouring (TNL) and term labouring (TL) rats. Female rats were fed either control chow (CON n = 20) or HFHC (n = 20) diet 6 weeks before conception and during pregnancy. On gestational day 21(TNL) or day 22 (TL) CON and HFHC (n = 10) rats were killed to determine plasma cholesterol, triacylglycerol and progesterone concentrations and collection of myometrium for contractility studies and expression of CAPs caveolin-1 (Cav-1), connexin-43 (CX-43) and it’s phosphorylated form (pCX-43), oxytocin receptor (OXTR) and cyclooxygenase-2 (COX-2). HFHC feeding increased visceral fat (P 0.001), plasma cholesterol (P 0.001) and triacylglycerol (P = 0.039) concentrations. Stage of labour effected uterine expression of CAV-1 (P < 0.02), pCX43 and COX-2 (both P < 0.03). CAV-1 and pCX43 decreased but COX-2 increased with parturition. Significant diet- and labour-stage interactions were evident for CX-43 and pCX43 (P < 0.03 and P < 0.004 respectively). CX-43 decreased with TL in HFHC animals but was unaltered in CON. pCX-43 fell with labour in CON but remained high in HFHC. OXTR expression was significantly higher in HFHC compared with CON animals (P < 0.03). Progesterone was higher in HFHC rats at term (P < 0.014) but fell significantly with labour to similar concentrations as CON. Contractility studies identified synchronous contractions of stable amplitude in lean animals, but unstable asynchronous contractions with obesity. Uterine dose response to oxytocin was blunted during labour in HFHC rats with a log EC50 of −8.84 compared with −10.25 M in CON for integral activity (P < 0.05). In conclusion, our adiposity model exhibits adverse effects on contractile activity during labour that can be investigated further to unravel the mechanisms causing uterine dystocia in obese women

Capturing residents' values for urban green space: mapping, analysis and guidance for practice

Planning for green space is guided by standards and guidelines but there is currently little understanding of the variety of values people assign to green spaces or their determinants. Land use planners need to know what values are associated with different landscape characteristics and how value elicitation techniques can inform decisions. We designed a Public Participation GIS (PPGIS) study and surveyed residents of four urbanising suburbs in the Lower Hunter region of NSW, Australia. Participants assigned dots on maps to indicate places they associated with a typology of values (specific attributes or functions considered important) and negative qualities related to green spaces. The marker points were digitised and aggregated according to discrete park polygons for statistical analysis. People assigned a variety of values to green spaces (such as aesthetic value or social interaction value), which were related to landscape characteristics. Some variables (e.g. distance to water) were statistically associated with multiple open space values. Distance from place of residence however did not strongly influence value assignment after landscape configuration was accounted for. Value compatibility analysis revealed that some values co-occurred in park polygons more than others (e.g. nature value and health/therapeutic value). Results highlight the potential for PPGIS techniques to inform green space planning through the spatial representation of complex human-nature relationships. However, a number of potential pitfalls and challenges should be addressed. These include the non-random spatial arrangement of landscape features that can skew interpretation of results and the need to communicate clearly about theory that explains observed patterns

FU19 Nephrops Grounds 2023 UWTV Survey Report and catch scenarios for 2024

This report provides the main results of the fourteenth underwater television survey of the various Nephrops patches in Functional Unit 19. The survey was multi disciplinary in nature collecting UWTV and other ecosystem data. In 2023 a total 42 UWTV stations were successfully completed. The mean density estimates varied considerably across the different patches. The 2023 raised abundance estimate showed a 15% decrease from the 2022 estimate and at 220 million burrows is below the MSY Btrigger reference point (430 million). Using the 2023 estimate of abundance and updated stock data implies catch in 2024 that correspond to the F ranges in the EU multi annual plan for Western Waters are between 224 and 248 tonnes (assuming that discard rates and fishery selection patterns do not change from the average of 2020–2022). One species of sea pen was observed; Virgularia mirabilis which has been observed on previous surveys of FU19. Trawl marks were observed at 10% of the stations surveyed.Marine Institut

Blood protein predictors of brain amyloid for enrichment in clinical trials?

BACKGROUND: Measures of neocortical amyloid burden (NAB) identify individuals who are at substantially greater risk of developing Alzheimer's disease (AD). Blood-based biomarkers predicting NAB would have great utility for the enrichment of AD clinical trials, including large-scale prevention trials. METHODS: Nontargeted proteomic discovery was applied to 78 subjects from the Australian Imaging, Biomarkers and Lifestyle Flagship Study of Ageing with a range of NAB values. Technical and independent replications were performed by immunoassay. RESULTS: Seventeen discovery candidates were selected for technical replication. α2-Macroglobulin, fibrinogen γ-chain (FGG), and complement factor H-related protein 1 were confirmed to be associated with NAB. In an independent cohort, FGG plasma levels combined with age predicted NAB had a sensitivity of 59% and specificity of 78%. CONCLUSION: A single blood protein, FGG, combined with age, was shown to relate to NAB and therefore could have potential for enrichment of clinical trial populations