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Engineering of decellularised porcine bladder patches

by Ashley James Ward

Institution: University of Leeds
Department:
Degree:
Year: 2017
Keywords:
Posted: 2/1/2018 12:00:00 AM
Record ID: 2162199
Full text PDF: http://etheses.whiterose.ac.uk/16431/


Abstract

For patients with end-stage bladder disease for which other treatment options have failed, the patient is treated surgically with either urinary diversion or bladder augmentation. Enterocystoplasty is the most common of these options, and it involves augmenting the bladder using a section of the patient's own intestine. However, there are several problems associated with the use of intestine for augmenting the bladder, and therefore an alternative augmentation material may be of benefit to patients. Numerous approaches have been used to develop tissue-engineered scaffolds capable of successfully augmenting bladders. Some of these approaches have involved the use of acellular tissue-derived materials, whereby the tissues are decellularised in order to remove immunogenic material and therefore prevent an immune reaction when transplanted allogeneically or xenogeneically. Decellularisation protocols typically involve a variety of chemical and physical processes which remove cells and other immunogenic material from the tissues.A protocol was previously developed to decellularise full-thickness porcine bladders. This material may have utility in bladder augmentation. The process involved distending the bladders with, and placing them in, a series of solutions. It was demonstrated that distending the whole organs was a necessary step in the decellularisation process. It was thought that this procedure applied biaxial strain to the wall of the tissue and reduced its thickness sufficiently for the solutions to penetrate the entire wall of the tissue. However, the method of distending whole bladders was not compatible with a scalable manufacturing process, and therefore the biomaterial was not able to be developed further. The overall aim of this project was to develop a novel method of manipulating bladder tissue which would enable bladder tissue to be decellularised in a way which would be compatible with a commercial manufacturing process.The original bladder decellularisation process used 500 ml of solutions to distend bladders. Preliminary experiments demonstrated that this volume was not always adequate to decellularise larger bladders. Filling experiments were performed to find relationships between bladder size and bladder capacity. A relationship between bladder capacity and bladder width*length was found to have a high correlation. Bladders were decellularised when filled to capacities calculated using the relationship. No signs of cellular material were observed in histological sections of these bladders, and DNA quantification indicated a removal of more than 99% of the DNA relative to native tissue.In order to determine the state of mechanical deformation of bladders during decellularisation, markers were placed on the surface of twelve bladders which were immersed in isotonic solution and slowly filled. Images taken of the bladders and markers during filling were used to calculate the strain of the tissues during the tests. The previously found relationships for bladder capacity were used to

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