Our aim was to detect the performance characteristics of multiparametric magnetic resonance imaging (mp-MRI) in patients with clinical suspicion, or previous diagnosis, of prostate cancer. Mp-MRI (index test) comprised of T2-weighted, diffusion weighted and dynamic contrast enhanced imaging. Radiologists used Likert score 1-5 based on the likelihood of the presence of prostate cancer. Concordance was made between results of mp- MRI and template prostate mapping (TPM) biopsy (reference standard). This retrospective study included patients that had both the index test and reference standard between January 2007 to January 2011 at either University College London Hospital or London Urology Associates. These were patients with; a) no prior prostate biopsy (n=129), b) prior negative prostate biopsy (n=54), c) previous positive prostate biopsy (n=194) and d) biochemical failure after radiotherapy (n=37). A set of target conditions was used and varied between the four groups of patients. These were either based on Gleason scoring, maximum cancer core length or a combination of both. In the first group, mp-MRI showed encouraging diagnostic performance results in ruling out clinically significant prostate cancer with sensitivity and negative predictive value (NPV) up to 94% and 89%, respectively. Accuracy figures were similar in the second group with sensitivity and NPV reaching up to 90% and 95%, respectively. In patients that underwent mp-MRI before reclassification TPM biopsy (third group), NPV for predicting that cancer remained low risk (as detected on previous TRUS-guided biopsy) reached up to 100%. Positive predictive value for upgrade of prostate cancer disease on subsequent TPM biopsy reached up to 75% with diagnostic odds ratio up to 2.86. In the last group, a combination of T2-weighted + high b-value showed optimum mp-MRI performance. These results suggest that mp-MRI can be used as a triage test among different patient populations, to select patients that can avoid biopsy and those that need re-biopsy before entering an active surveillance program. Time and cost can be saved by using only certain MRI sequences in patients with biochemical failure after radiotherapy.

Skin protects our bodies for a lifetime and extensive loss of this barrier renders the individual susceptible to infections and death. Clinically available treatment options, however, are limited in establishing both functional and cosmetic satisfaction. The work described in this thesis is therefore concerned with the development and characterization of a novel biodegradable nanocomposite system displaying suitable properties as skin tissue engineering scaffolds. A novel family of segmented polyurethanes (PU) with increasing hard segment content based on a poly(ε-caprolactone urea)urethane backbone incorporating POSS nanoparticles was synthesized and analysed in terms of material characteristics and biocompatibility. Incorporation of POSS nanoparticles into the PU backbone yielded mostly amorphous materials as corroborated by distinct glass transitions visible on differential scanning calorimetry spectra. With incrementally increasing hard segment content, ultimate tensile strength increased from ~10 to 21 MPa accompanied by increased values for elastic moduli from 0.03 to 0.06 MPa. Number average molecular weights (Mn) decreased with increasing hard segment content due to a corresponding decrease in the proportion of PCL. Sterilization studies raised fundamental concerns regarding the suitability of conventionally available techniques since hydrolytically and temperature labile polymers are susceptible to degradation. The results obtained suggest 70 % ethanol to be a suitable laboratory-based disinfectant which was further demonstrated to have favourable effects on skin cell compatibility. Degradation studies revealed hard segment-dependent modulation of the degradation rate and the materials’ viscoelasticity. In vivo implantation studies of porous scaffolds demonstrated firm integration with the subcutaneous tissue and extensive vascularization. The results obtained in this work highlight (i) the ability to control scaffold degradation rates and mechanical properties and (ii) cellular as well as in vivo biocompatibility, all of which fundamental in the development of a versatile skin tissue engineering scaffold.

As the annual incidence of revision total hip replacement (THR) rises so too do the technical, biomechanical and socioeconomic challenges it presents. This thesis investigated many of the key clinical and pre-clinical aspects of this surgery. Improving implant stability was addressed through biomechanical analyses and the cost and complications of revision THR were explored. The biomechanical investigation compared two designs of tapered, fluted, modular, titanium-alloy stems. The Redapt® stem featured a novel flute configuration and chamfered tip. It was hypothesised that this would improve axial and rotational stability compared to the Modular Restoration® control stem. Each stem was implanted into one of 7 matched pairs of human cadaveric femora with simulated proximal bony defects. A photoelastic coating compared surface strains in the medial femoral cortex for the intact and operated femora. Under incremental static loads each operated bone showed marked stress-shielding with a statistically significant reduction in strain. This effect was diminished with the Redapt® stem because of reduced distal endosteal contact (‘fill’) as confirmed by radiographic analysis. Primary stability was measured using micromotion transducers and radiostereometric analysis. Under cyclical loading both stems were stable by agreed standards at x1 body weight. As load increased 85% of the Redapt stems remained stable compared to 100% of the Restoration (p=0.055). Overall transducer recorded axial subsidence was 0.1 mm for the Redapt and 0.17 mm for the Restoration. Both stems achieved results commensurate with their expected successful application in revision cases with extensive bony defects. Clinical and financial data was collected from 305 consecutive revision THRs between 1999-2008 performed at our institution. Analysis revealed a large variation in costs by indication from £10893 (SD £5476) for dislocation to £21937 (SD £10965) for septic revisions. A large shortfall in reimbursement was found questioning the ability of smaller units to continue providing this service.

Bowel symptoms (constipation and/or faecal incontinence) affect the vast majority of patients with multiple sclerosis (MS), but the pathophysiology is unclear and treatment remains empirical. The primary hypothesis of this thesis is that involvement of the spinal cord by the disease is central to the development of bowel symptoms, and this is tested in the first two studies: 1. A study of the overlap in prevalence of bladder symptoms in patients with MS and bowel symptoms. 2. A study of rectal compliance, as an important reflection of both the gut’s neural tone and its ability to hold content, in comparison to patients with supraconal spinal cord injury and normal controls. The secondary hypothesis is that residual spinal cord function can represent a potential target of treatment, and this is tested in studies 3 and 4: 3. A prospective observational study of bowel biofeedback in symptomatic MS patients. 4. A prospective observational study of transanal irrigation in symptomatic MS patients. Study 1 shows that the prevalence of bladder symptoms – determined by spinal cord disease - is higher in patients with bowel symptoms than in the general population of MS sufferers. The second study shows that rectal compliance - as an index of the spinal reflex activity regulating autonomic rectal function – is altered in patients with MS according to the clinical degree of spinal cord involvement by the disease. A similar pattern is followed for symptoms of constipation, but not faecal incontinence. The two treatment studies showed that: Biofeedback improves bowel symptoms and 5-seconds-endurance squeeze pressure. Improvement of sphincter pressure could be the result of behavioural changes, inducing physiological changes through residual efferent pathways in the spinal cord. Transanal irrigation is effective to treat bowel symptoms in patients who fail biofeedback.

The current clinical pathway for localised prostate cancer firstly involves serum PSA testing, through screening programmes or ad hoc testing in highrisk or symptomatic men. If the PSA is raised, a transrectal-ultrasound guided biopsy is performed for histological diagnosis for suspected cancer. However, serum Prostate Specific Antigen (PSA) levels are inaccurate in selecting an ‘at-risk’ group for malignancy, and transrectal-ultrasound guided biopsy is a morbid procedure in which approximately 30% of men are under-diagnosed (through missing the disease) or misdiagnosed (through underestimating the true disease burden), therefore misinforming on appropriate management. A proportion of men are ‘over-diagnosed’ with low risk cancers on biopsy that may have little to no metastatic potential. The current management options of low- to intermediate-risk disease includes active surveillance with regular repeat serum PSA and biopsy tests and their inherent limitations. The alternative, which is radical treatment, may cause significant morbidity and reduce quality of life, despite considerable technical advances. An imaging test that is able to detect, characterise and localise prostate cancer may allow better diagnosis through improved risk stratification to the current standard tests, and targeted sampling of any suspicious areas, with more effective grading of disease burden. Additionally, the morbidity associated with the current therapeutic options could be reduced through the identification and targeted treatment of cancer lesions (‘focal therapy’), rather than the whole prostate. This thesis addresses the adoption of multi-parametric (mp)MRI for these diagnostic and therapeutic purposes. Whilst mpMRI has already demonstrated high accuracy rates for the detection of clinically significant prostate cancer, the current lack of standardisation of conduct and reporting has reduced comparability between studies and resulted in poor external validity. I present the rationale for the use of a new prostate mpMRI scoring system and the consensus outputs from a European expert group on standardising conduct and reporting, and their applications to date. I also present studies that apply the information obtained from mpMRI on the location and burden of clinically significant disease, to plan, conduct and follow-up focal treatment.

Full thickness burns and trauma resulting in extensive full thickness skin loss or devitalisation gives rise to the need for skin replacement therapy. Cultured epithelial autologous keratinocytes application has been the main stay treatment over the last three decades. Different methods for cultured epithelial autologous keratinocytes delivery exist with thin epithelial sheet application and sprayed cell culture the most commonly used methods; each method has its advantages and drawbacks. Microcarrier beads for the culture of cells have been utilised in a number of different applications over the last forty years as they allow rapid cell culture and expansion in a controlled environment. More recently microcarrier commercially available gelatin microcarrier beads “Cultispher G®” have demonstrated the potential to support keratinocyte cell culture and proliferation in vitro. Wound contraction is a physiological component of wound healing and occurs within the proliferation phase of wound healing between 4 days to 3 weeks following the inflammatory phase. Wound contraction is a function of myofibroblasts leading to approximation of the wound margins and reduction in wound size. The migration of keratinocytes from the intact epithelium around the wound edges leads to epithelial closure and completes the process of wound healing. This study aimed to assess the use of microcarriers for supporting keratinocyte growth in vitro and evaluate the effect of keratinocyte delivery using microcarriers on wound contraction using a porcine wound model. In this study, in vitro assessment of keratinocyte expansion on microcarriers demonstrated sustainable expansion rates with data being comparable to traditional keratinocyte cell culture. Animal experiments utilising an in vivo porcine model have shown that keratinocytes delivered to the wound bed using gelatin microcarriers migrated off the beads and were shown to survive on the wound bed. In the same animal model, autologous keratinocytes cultured on microcarrier beads reduced wound contraction when applied to full thickness wounds in combination with widely meshed autologous split skin graft compared with split thickness skin graft (STSG) alone or control wounds. The use of allogeneic cultured keratinocytes on microcarriers in combination with dermal regeneration template “Integra®” demonstrated a reduction in wound contraction compared with Integra® and STSG or Integra® alone. The use of autologous cultured keratinocytes and fibroblasts on microcarriers in combination with Integra® reduced wound contraction compared with Integra and cultured keratinocytes or Integra and STSG. The reduction in wound contraction maintained a large area of the original wound surface area, hence reducing the possibility of contracture formation. The use of microcarrier beads for culture and delivery of keratinocytes has the potential to overcome the disadvantages of the traditional methods of keratinocyte culture and delivery. It can also play a role in the reduction of wound contraction resulting in the retention of skin mobility and providing favourable functional and aesthetic outcomes.

Orthotopic liver transplantation has become the preferred treatment for a variety of end-stage liver disease. As competency and survival rates increase, so does increasing demand, which puts greater strain on a static donor pool. This increases pressure to accept more marginal grafts, e.g. non-heart-beating and steatotic donors. However, graft dysfunction post-transplantation contributes considerably to postoperative morbidity and mortality. Proton nuclear magnetic resonance (1HNMR) spectroscopy is a powerful technique to explore the biochemical composition of biological fluids; it is rapid, non-invasive, nondestructive and it can detect metabolites at millimolar concentrations. In this study, 1HNMR assessed the changes in bile composition during liver ischaemia/reperfusion. The primary hypothesis was that bile composition changes during liver ischaemia reperfusion injury (IRI). The aims were to document these changes, identify biliary markers for liver IRI using 1HNMR, validate these markers using known modulators, determine if the same was true for steatotic livers and attempt to understand the mechanisms by which these changes happen in relation to the redox state of the liver. Materials and methods: A rabbit model of hepatic lobar IRI was used. In most experiments 3 groups were used (n=6): Sham group (laparotomy alone), I/R group (1hr ischaemia and 6hrs reperfusion), and a modulation group similar to I/R with the addition of N-acetylcysteine (I/R+NAC: 150mg/kg of NAC) and glycine (I/R+glycine: 5mg/kg glycine). Steatosis was induced by feeding with a 2% cholesterol diet for 8 weeks. Experiments were repeated on steatotic animals with Sham, I/R, I/R+NAC and IPC+I/R (5min ischaemia followed by 10min of reperfusion before prolonged ischaemia was induced) groups. The following parameters were measured: portal blood flow, bile flow (BF) and bile 1HNMR spectroscopy, hepatic microcirculation, intracellular tissue oxygenation, serum ALT, AST and ICG clearance were measured at 1, 2, 5 and 7 hours following reperfusion. Results: Bile spectroscopy demonstrated significant changes in bile composition following I/R and alterations with NAC, glycine and IPC. These changes are evident despite a constant post-reperfusion rate of BF. They were also present in steatotic livers, and were modulated by NAC and IPC. In experiment 1: BF, COX and biliary acetate decreased following I/R while AST, ALT, biliary PC and lactate increased along with PMN accumulation in sinusoids, KC hypertrophy, necrosis and apoptosis in normal livers. In experiment 2: BF, COX and biliary acetate decreased following I/R while ALT, PC, conjugated bile acids and lactate increased in the I/R group in normal livers. NAC administration attenuated the increase in ALT and lactate following I/R in the NAC+I/R group compared to the I/R group. Changes in conjugated bile acids seem to reflect changes in BF. In experiment 3: I/R+glycine was associated with increased BF, bile acid, acetate, pyruvate, glucose, acetoacetate, and decreased bile lactate and PC levels in normal livers. In experiment 4: NAC administration in steatotic livers reduced the extent of IRI, increased portal blood flow and liver parenchymal perfusion. NAC increased BF, biliary acetate and pyruvate and reduced acute liver injury, ALT and biliary PC. In experiment 5: IPC protected the steatotic liver from IRI and maintained hepatic oxygenation, tissue perfusion and mitochondrial redox state. COX activity was decreased by IRI in the fatty liver, but can was protected by IPC. Conclusions: This thesis has demonstrated changes in bile composition during warm normal liver I/R and its modulation with NAC and glycine. It has also demonstrated changes in bile composition during warm steatotic liver I/R and its modulation with NAC and IPC. It has noted several metabolites that are consistently changed, as well as the bile redox ratio of metabolites that provides a clearer indication of liver redox state than individual metabolites.

Inefficient intracellular drug delivery is a significant limiting factor to success in cancer therapeutics. Photochemical internalisation (PCI) utilises the fundamental principles of photodynamic therapy (PDT): photosensitiser plus light and oxygen, at sub-lethal level to facilitate targeted intracellular drug delivery. This effect is mediated by reactive oxygen species (ROS). This thesis investigates the mechanisms underpinning sulfonated meso-tetraphenylporphine (TPPS2a) mediated PCI to enhance the delivery of two cytotoxins, saporin or mitoxantrone, and a novel Small Molecule Carrier (SMoC) in vitro. PCI of saporin was also assessed in a 3D-tumour model. In vitro experiments using 4T1 murine breast adenocarcinoma cells were performed to investigate which factors determined the likelihood of PDT versus PCI predominant cytotoxicity. The role of the intracellular REDOX environment in PDT/PCI was assessed using a free radical potentiator and quenchers. The results suggested that the localisation and total amount of ROS produced exerts the greatest influence in determining the likelihood of PDT versus PCI induced cell kill. In addition, PCI further enhanced SMoC-aided delivery of siRNA in MCF7 human breast cancer cells. A compressed collagen scaffold, embedded with 4T1 cells, was used to investigate TPPS2a-mediated PCI of saporin in a 3D tumour model. The results indicated that a 3D-model is potentially a useful tool for pre-clinical assessment of PCI. Bioluminescent PDT studies were also carried out on MCF7 cells transduced with luciferase and the 4T1-luc2 cell line, which is stably transfected with luciferase. These studies demonstrated that bioluminescence can be used to activate a photosensitiser for a cytotoxic effect (PDT). Overall, this thesis demonstrated that by further understanding the mechanisms that underpin PCI it is possible to further enhance its facilitative effects for drug delivery. The ongoing phase II clinical trials into PCI show its translational potential as a means to improve the therapeutic effectiveness of anti-cancer drugs.

Some studies on autologous chondrocyte implantation (ACI) have demonstrated little benefit over other techniques and few have demonstrated a lasting benefit. A number of factors can contribute to failure and a scientific approach to elucidate these variables has not been reported. This thesis reports on the use of a statistical approach - the Generalised Linear Model (GLM) to quantify the effect each factor has whilst considering the interplay of other variables. Data from a randomised controlled trial and several case-controlled studies will assess the efficacy of 2 different types of ACI, the influence of smoking, BMI, and physical activity. Non-modifiable risk factors that were assessed include the aetiology, site and size of the lesion, the duration of symptoms and number of previous operations prior to the index procedure and the presence of early osteoarthritis. Site had a significant effect on outcome but size did not. The GLM predicted a point increase in the Modified Cincinnati Score (MCS) before surgery (MCS 0) would lead to a further 0.5 point increase in MCS 2 years postoperatively (MCS 24) (p=0.001). Other significant non-modifiable risk factors include age and sex of the patient. When treating lesions in the patella, duration of symptoms was a significant factor, but age was not. The GLM predicted that smokers’ MCS 24 (the Modified Cincinnati Score 2 years after surgery) was likely to be 15 less than non-smokers (p=0.002). Patients playing no sports experienced an 11.4 point decrease. For each increase in BMI, the MCS 24 was 2.4 less (p=0.001). Factors that optimise outcome following surgery are; avoidance of numerous procedures prior to ACI and delay of more than one year before undergoing ACI. Current NICE guidelines prohibit the use of ACI as the first-line surgical procedure and prevent addressing the above 2 issues. Poorer results were observed in obese patients. Weight loss and active lifestyle are essential pre-operatively. Furthermore, we recommend that pre-operative counselling for smokers is essential and that all smokers be offered a cessation programme.

The 2 main unresolved issues inherent in coronary stents are in-stent restenosis (ISR) and late stent thrombosis (ST). ISR is largely due to vascular smooth muscle cell (VSMC) proliferation, and ST is attributed to a lack of re-endothelialization. This thesis describes the conceptualization and development of a biofunctionalized polyhedral oligomeric silsesquioxane poly(carbonate-urea) urethane (POSS-PCU) platform, for the express purpose of circumventing ISR and ST. A bare-metal stent is embedded between 3 layers of POSS-PCU facilitating endothelial progenitor cell (EPC) capture using antibodies, in tandem with sustained drug release using stealth liposomes. The luminal area is impregnated with anti-CD34 antibodies, covalently bonded to a POSS-PCU base platform, for EPC capture to enhance re-endothelialization. Results indicated successful antibody immobilization, with an increased propensity for EPC capture compared to controls. The abluminal area is integrated with paclitaxel-encapsulated stealth liposomes, with an ultrathin layer of POSS-PCU sprayed on top using an ultrasonic atomization spray system, for sustained drug release to inhibit VSMC proliferation. Results in this aspect demonstrated sustained drug release with augmented cell kill in a 28-day in vitro cell culture. Mechanical engineering tests performed on the finished product demonstrated superior mechanical functionality as a covered stent. Various sterilization techniques and a biodegradation model were also employed to robustly assess product viability, with results indicating that EPC capture potential and stealth liposomal drug elution were preserved. Taken together, this novel POSS-PCU covered stent can enhance re-endothelialization and inhibit VSMC proliferation, thereby addressing the issues of ST and ISR respectively. Furthermore, the covering membrane could also serve as a physical barrier against atherosclerotic plaque dislodgement to prevent thromboembolism. It is therefore hoped that the proof-of-principle demonstrated here in this thesis would serve as an impetus for further translational research, with the eventual goal of taking it from bench to bedside.