Novel 3D organotypic urothelial cell culture model for identification of new therapeutic approaches in urological infections

Abstract

Purpose: 3D organotypic cell cultures offer the possibility to study cell growth in a more in vivo like situation. To our knowledge no 3D culture of primary urothelial cells has been established yet. BK Polyomavirus (BKPyV), replicating in urothelial cells, may cause haemorrhagic cystitis in immunocompromised patients.

Primary endpoints of this study: Establishment of a 3D organotypic cell culture of primary urothelial cells and fibroblasts; use of this model as infection model for archetype BKPyV; description of first parts of viral life cycle with identification of therapeutic targets.

Methods: This is an experimental study. Primary urothelial cells were purchased from CellnTec, Bern, Switzerland; fibroblasts were isolated from the ureter of patients with no urothelial malignancy in their medical history. As main methods we used quantitative real-time PCR and immunohistochemistry. Outcomes were analysed using SPSS 23.0.

Results: We were able to develop a 3D organotypic culture for primary urothelium. An infection with archetype BKPyV was established in this model with virus replication rates up to 6.41 × 108 copies/ml on day 9 following Infection. Interestingly, proliferation rate of the urothelial cells is significantly (p = 0.049 at day 6 following infection) elevated while cells are losing differentiation under infection. Phosphorylated STAT3 is also significantly elevated (p < 0.0001) during infection.

Conclusions: The established of urothelial 3D cultures is a new method to study several urothelial diseases. The archetype BKPyV infection model is novel and the first method to study archetype viral life cycle. The STAT3 pathway might be an interesting target for the development of a causal therapy.

Keywords: 3D cell culture; BK polyomavirus; STAT3 pathway; Urinary tract infection; Urothelial cell.