Development of a pressurised transmural decellularisation method for application in tissue engineering trachea

Tracheal abnormalities, congenital or acquired, represent a currently unmet clinical need. Tissue engineering has recently advanced and has been used to engineer hollow organs, including tracheae, for clinical use on a compassionate basis. However the current clinically used method for tracheal decellularisation has received mixed success and requires development to achieve GMP translation, quality manufacturing standards and ultimately routine clinical use. This thesis first examined the current clinically used detergent-enzymatic method (DEM) of decellularisation, a highly manual process that takes twenty-eight days to complete. Although the method achieved full decellularisation of non-cartilaginous regions of the tracheae, it failed to reduce the donor nuclear material sufficiently and resulted in the loss of key biochemical components, glycosaminoglycans (GAG) and collagen Type II, and the loss of biomechanical strength. A novel method for rapid, effective and non-detrimental tracheal decellularisation was required. Pressurised transmural flow was hypothesised to meet those requirements. A dual chamber bioreactor system was designed, fabricated and optimised to enable pressurised transmural decellularisation (PTD) to be investigated. Optimal PTD process parameters were ascertained and shown to produce tracheal scaffolds that achieved full decellularisation of the non-cartilaginous regions of the tracheae, a reduction of donor nuclear material (95%) which met the currently recommended levels of residual donor DNA for tissue engineered products, as well as maintaining GAG, collagen and biomechanical strength at comparable levels to the native tracheae. Additional to this, the new PTD process achieved this effective and non-detrimental decellularisation of tracheae in five working days with a ten-fold cost of goods reduction. With further development, the PTD method could become a fully automatable and closed process which could progress tissue engineered tracheae towards becoming a validated and regulated advanced therapy medicinal product (ATMP) and enable the advancement of tissue-engineered tracheae into regular clinical use

Robust identification from band-limited data

Consider the problem of identifying a scalar bounded-input/bounded-output stable transfer function from pointwise measurements at frequencies within a bandwidth. We propose an algorithm which consists of building a sequence of maps from data to models converging uniformly to the transfer function on the bandwidth when the number of measurements goes to infinity, the noise level to zero, and asymptotically meeting some gauge constraint outside. Error bounds are derived, and the procedure is illustrated by numerical experiment

Asymptotic estimates for interpolation and constrained approximation in H2 by diagonalization of Toeplitz operators

Sharp convergence rates are provided for interpolation and approximation schemes in the Hardy space H-2 that use band-limited data. By means of new explicit formulae for the spectral decomposition of certain Toeplitz operators, sharp estimates for Carleman and Krein-Nudel'man approximation schemes are derived. In addition, pointwise convergence results are obtained. An illustrative example based on experimental data from a hyperfrequency filter is provided

Approximate Carleman theorems and a Denjoy-Carleman maximum principle

We give an extension of the Denjoy-Carleman theorem, which leads to a generalization of Carleman's theorem on the unique determination of probability measures by their moments. We also give complex versions of Carleman's theorem extending Theorem 4.1 of [2]

Outcomes from deep brain stimulation targeting subthalamic nucleus and caudal zona incerta for Parkinson's disease.

Both subthalamic nucleus (STN) and caudal zona incerta (cZI) have been implicated as the optimal locus for deep brain stimulation (DBS) in Parkinson's disease (PD). We present a retrospective clinico-anatomical analysis of outcomes from DBS targeting both STN and cZI. Forty patients underwent bilateral DBS using an image-verified implantable guide tube/stylette technique. Contacts on the same quadripolar lead were placed in both STN and cZI. After pulse generator programming, contacts yielding the best clinical effect were selected for chronic stimulation. OFF-medication unified PD rating scale (UPDRS) part III scores pre-operatively and ON-stimulation at 1-2 year follow up were compared. Active contacts at follow-up were anatomically localised from peri-operative imaging. Overall, mean UPDRS part III score improvement was 55 ± 9% (95% confidence interval), with improvement in subscores for rigidity (59 ± 13%), bradykinesia (58 ± 13%), tremor (71 ± 24%) and axial features (36 ± 19%). Active contacts were distributed in the following locations: (1) within posterior/dorsal STN (50%); (2) dorsal to STN (24%); (3) in cZI (21%); and (4) lateral to STN (5%). When contacts were grouped by location, no significant differences between groups were seen in baseline or post-operative improvement in contralateral UPDRS part III subscores. We conclude that when both STN and cZI are targeted, active contacts are distributed most commonly within and immediately dorsal to STN. In a subgroup of cases, cZI contacts were selected for chronic stimulation in preference. Dual targeting of STN and cZI is feasible and may provide extra benefit compared with conventional STN DBS is some patients

Outcomes from deep brain stimulation targeting subthalamic nucleus and caudal zona incerta for Parkinson's disease.

Both subthalamic nucleus (STN) and caudal zona incerta (cZI) have been implicated as the optimal locus for deep brain stimulation (DBS) in Parkinson's disease (PD). We present a retrospective clinico-anatomical analysis of outcomes from DBS targeting both STN and cZI. Forty patients underwent bilateral DBS using an image-verified implantable guide tube/stylette technique. Contacts on the same quadripolar lead were placed in both STN and cZI. After pulse generator programming, contacts yielding the best clinical effect were selected for chronic stimulation. OFF-medication unified PD rating scale (UPDRS) part III scores pre-operatively and ON-stimulation at 1-2 year follow up were compared. Active contacts at follow-up were anatomically localised from peri-operative imaging. Overall, mean UPDRS part III score improvement was 55 ± 9% (95% confidence interval), with improvement in subscores for rigidity (59 ± 13%), bradykinesia (58 ± 13%), tremor (71 ± 24%) and axial features (36 ± 19%). Active contacts were distributed in the following locations: (1) within posterior/dorsal STN (50%); (2) dorsal to STN (24%); (3) in cZI (21%); and (4) lateral to STN (5%). When contacts were grouped by location, no significant differences between groups were seen in baseline or post-operative improvement in contralateral UPDRS part III subscores. We conclude that when both STN and cZI are targeted, active contacts are distributed most commonly within and immediately dorsal to STN. In a subgroup of cases, cZI contacts were selected for chronic stimulation in preference. Dual targeting of STN and cZI is feasible and may provide extra benefit compared with conventional STN DBS is some patients

Nitrogen use efficiency of ¹⁵N urea applied to wheat based on fertiliser timing and use of inhibitors

Improving fertiliser nitrogen (N) use efficiency is essential to increase productivity and avoid environmental damage. Using a 15N mass balance approach, we investigated the effects of five N fertiliser management strategies to test the hypothesis that increasing uptake of applied N by wheat improves productivity and reduces loss of N in a semi-arid environment. Three experiments were conducted between 2012 and 2014. Treatments included urea application (50 kg N/ha) at sowing with and without nitrification inhibitor (3,4-dimethylpyrazole phosphate, DMPP) and surface broadcast with and without urease inhibitor (n-butyl thiophosphoric triamide, NBPT) at the end of tillering plus an unfertilised control. It was found that deferring fertiliser application until the end of tillering decreased losses of fertiliser N (35–52%) through increasing uptake by the crop and or recovery in the soil at harvest, while maintaining yield except when rainfall following application was low. In this case, deferring application reduced fertiliser uptake (− 71%) and grain yield (− 18%) and increased recovery of N in the soil (+ 121%). Use of DMPP or NBPT reduced N loss where seasonal conditions were conducive to denitrification during winter (DMPP) and volatilisation or denitrification later in the season (NBPT). Their effect on grain yield was less significant; DMPP increased yield (+ 3–31%) in all years and NBPT increased yield (+ 7–11%) in 2 of 3 years compared to unamended urea. The majority of crop N uptake was supplied from soil reserves and as a result, crop recovery of applied N was not strongly related to grain yield response

Substrate utilization during exercise performed with and without glucose ingestion in female and male endurance trained athletes

Compared to males, females oxidize proportionately more fat and less carbohydrate during endurance exercise performed in the fasted state. This study was designed to test the hypothesis that there may also be gender differences in exogenous carbohydrate (CHOexo) oxidation during exercise. Healthy, young males (n = 7) and females (n = 7) each completed 2 exercise trials (90 min cycle ergometry at 60% VO[sub2peak]), 1 week apart. Females were eumenorrheic and were tested in the midfollicular phase of their menstrual cycle. Subjects drank intermittently either 8% CHOexo (1 g glucose &sdot; kg &sdot; h[sup-1]) enriched with U-13C glucose or an artificially sweetened placebo during the trial. Whole-body substrate oxidation was determined from PER, urinary urea excretion, and the ratio of 13C:12C in expired gas during the final 60 min of exercise. During the placebo trial, fat oxidation was higher in females than in males (0.42 &plusmn; 0.07 vs. 0.32 &plusmn; 0.09 g &sdot; min[sup-1] . kg LBM[sup-1] x 10[sup-2]) at 30 min of exercise (p &lt; .05). When averaged over the final 60 min of exercise, the relative proportions of fat, total carbohydrate, and protein were similar between groups. During CHOexo ingestion, both the ratio of 13C: 12C in expired gas (p &lt; .05) and the proportion of energy derived from CHOexo relative to LBM (p &lt; .05) were higher in females compared to males at 75- and 90-min exercise. When averaged over the final 60 min of exercise, the percentage of CHOexo to the total energy contribution tended to be higher in females (14.3 + 1.2%) than in males (11.2 &plusmn; 1.2%; p = .09). The reduction in endogenous CHO oxidation with CHOexo intake was also greater in females (12.9 &plusmn; 3.1%) than in males (5.1 &plusmn; 2.0%; p = .05). Compared to males, females may oxidize a greater relative proportion of CHOexo during endurance exercise which, in turn, may spare more endogenous fuel. Based on these observations, ingested carbohydrate may be a particularly beneficial source of fuel during endurance exercise for females.<br /

Phragmén–Lindelöf Principles for Generalized Analytic Functions on Unbounded Domains

We prove versions of the Phragmén–Lindelöf strong maximum principle for generalized analytic functions defined on unbounded domains. A version of Hadamard’s three-lines theorem is also derived