Alzheimer's disease (AD) is a chronic neurodegenerative disease. Aggregation of amyloid Î²1-42 (AÎ²1-42) peptide within the brain is considered the main cause of the neuropathological changes associated with AD, and development of compounds that target AÎ²1-42 peptide aggregation is a highly active research area. The blood brain barrier (BBB) can prevent efficient delivery of compounds to the brain, which limits treatment options for diseases of the central nervous system (CNS). Transferrin-functionalised liposomes are considered promising vectors for delivery to the brain since they overcome the barrier function of the BBB by binding to transferrin receptors expressed on the surface of the BBB endothelium. Resveratrol, melatonin, carnosine, tramiprosate and scyllo-inositol have previously been reported to reduce AÎ²1-42 peptide aggregation, attenuating AÎ²1-42 peptide-induced toxicity. In this thesis, the inhibitory effects of the selected compounds were reproduced using the thioflavin T (ThT) assay. In addition, more in-depth studies employing atomic force microscopy (AFM), matrix assisted laser desorption/ionization-time of flight (MALDI-TOF) and high performance liquid chromatography (HPLC) confirmed that resveratrol has antiaggregation properties. Resveratrol was also confirmed to have a significant inhibitory effect on the aggregation of AÎ²1-42 peptide under oxidative stress and fibril elongation conditions over ten days. This is the first study to demonstrate the anti-aggregation mechanism of resveratrol involves cleavage of AÎ²1-42 peptide into smaller fragments. The Plackett-Burman experimental approach was used to optimally design transferrinfunctionalised liposomes. The latter, composed of 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC) and cholesterol, were prepared by the thin lipid film rehydration method. Liposomes were characterised using dynamic light scattering (DLS), diffraction scanning calorimetry (DSC), transmission electron microscopy (TEM), Stewart assay, Bradford assay and small angle X-ray diffraction (SAXS). Liposomes were spherical and homogeneous with particle size of 80-100 nm and polydispersity index (PDI) of < 0.2, and were stable over 60 days. Liposomes were also confirmed to possess a single bilayer membrane, and the release profile of resveratrol was biphasic. Liposomes were successfully functionalised with transferrin and the entrapment efficacy percentage (EE%) of resveratrol was dramatically increased from 2.6 % (in passive loading) to 64.9 % (in active loading). In order to study delivery of resveratrol across BBB endothelial cells, an in vitro BBB model was generated on TranswellÂ® inserts. The model possessed high transendothelial electrical resistance (TEER), 400 Â± 150 Î©.cm2, and highly restrictive, with an apparent permeability of Lucifer Yellow of 10.6 Â± 7 x 10-7 cm.s-1. Significantly more resveratrol (14 - fold) was delivered across the BBB model using transferrin-functionalised liposomes than was delivered using non-functionalised liposomes (p â‰¤ 0.001) and free resveratrol (p â‰¤ 0.001). Transferrin-functionalised liposomes may therefore hold great potential for delivery of compounds, including therapeutic drugs, across the BBB.
|Date of Award||31 Dec 2017|
- The University of Manchester
|Supervisor||Harmesh Aojula (Supervisor) & Jeffrey Penny (Supervisor)|