Use of donor bladder tissues for in vitro research

Abstract

Objectives To evaluate deceased non‐heart beating (DNHB) donors and deceased heart beating (DHB) brain‐stem dead donors, as sources of viable urological tissue for use in biomedical research. To identify sources of viable human bladder tissue as an essential resource for cell biological research aimed at understanding human diseases of the bladder and for developing new tissue engineering and regenerative medicine strategies for bladder reconstruction. Typically, normal human urinary tract tissue is obtained from adult or paediatric surgical patients with benign urological conditions, but few surgical procedures yield useful quantities of healthy bladder tissue for research. Patients and Methods Research ethics committee approval was obtained for collection of donor bladder tissue. Consent for DHB donors was undertaken by the Donor Transplant Coordinators. Tissue Donor Coordinators were responsible for consent for DNHB donors and the retrieval of bladders was coordinated through the National Blood Service Tissue Banking Service. All retrievals were performed by practicing urologists and care was taken to maintain sterility and to minimise bacterial contamination. Two bladders were retrieved from DNHB donors and four were retrieved from DHB donors. Results By histology, DNHB donor bladder tissue exhibited marked urothelial tissue damage and necrosis, with major loss or absence of urothelium. No cell cultures could be established from these specimens, as the urothelial cells were not viable in primary culture. Bladder urothelium from DHB donors was intact, but showed some damage, including loss of superficial cells and variable separation from the basement membrane. All four DHB bladder specimens yielded viable urothelial cells that attached in primary culture, but cell growth was slow to establish and cultures showed a limited capacity to form a functional barrier epithelium and a propensity to senesce early. Conclusions We have shown that normal human bladder urothelial cell cultures can be established and serially propagated from DHB donor bladders. However, our study suggests that rapid post‐mortem changes to the bladder affect the quality and viability of the urothelium, rendering tissue from DNHB donors an inadequate source for urothelial cell culture. Our experience is that whereas patients are willing to donate surgical tissue for research, there is a barrier to obtaining consent from next of kin for retrieved tissues to be used for research purposes.

Uroplakin traffic through the Golgi apparatus induces its fragmentation: new insights from novel in vitro models

“…Together our data show that normal porcine UCs in culture reach a high differentiation stage that is comparable to the differentiation stage of superficial UCs of normal urothelium in vivo in various species 34 – 36 and therefore can be used as a suitable model for studies of UP transport.…”

Co‐culturing porcine normal urothelial cells, urinary bladder fibroblasts and smooth muscle cells for tissue engineering research

“…For culturing of urothelial cells, researchers use three types of media. The first one is KSFMc, which is KSFM (low calcium-0.09 mM keratinocyte serumfree medium containing recombinant epidermal growth factor and bovine pituitary extract) (Cilento et al, 1994) supplemented with 30 ng/mL cholera toxin and different concentrations of calcium (0.09-2.5 mM) (Southgate et al, 1994;Ludwikowski et al, 1999;Bisson et al, 2002;Cross et al, 2005;Garthwaite et al, 2014). The second one is DMEM and Ham's F12 supplemented with 5%, 10%, or 20% fetal bovine serum (FBS) (Fujiyama et al, 1995;Ehmann and Terris, 2002;Fossum et al, 2003;Larsson et al, 2014).…”

Artificial urine and FBS supplemented media in cytocompatibility assays for PLGA‐PEG‐based intravesical devices using the urothelium cell line UROtsa

“…29 However, obtaining urological tissue for primary urothelial cell cultures is not feasible, since no healthy subject would provide informed consent for the required hospitalisation and handling for the donation of bladder tissue. 30 It is possible to culture urothelial cells from bladder washing or donated urine, but such cell cultures only retain their stability for a few passages and they quickly stop proliferating. 31,32 The UROtsa cell line used in the present study were homogeneous, it required no scaffold or feeder layer and continued to grow at high passage numbers.…”