TDK conference 2018

Lower urinary tract diseases caused by different pathological factors, such as bacterial urinary tract infections, neoplasia, urolithiasis or parasites are common in domestic cats worldwide. However, the factors causing the clinical signs may also be unknown, in which case the disease is called idiopathic bladder inflammation or feline idiopathic cystitis, abbreviated as FIC. Based on former studies, it is assumed that stress is significantly contributing to the development of the illness; therefore we aimed to investigate the role of norepinephrine as a stress factor in the pathogenesis of FIC. In order to assess the effect of stress regardless of other factors, we have established a feline primary urinary bladder cell culture that can serve as an in vitro model of FIC. The cell culture was prepared according to the modified protocol of Truschel et al. (1999). The bladders we used were obtained from a euthanized cats; after removal they were digested, and the isolated epithelial cells were seeded and cultured for 2 weeks. Confluence of cell cultures was verified by Giemsa staining. Thereafter cell cultures were characterized by immunofluorescence, the epithelial origin of the cells was investigated by anti pan-cytokeratin antibodies, and bladder epithelial cells were labeled with uroepithel-specific anti-uroplakin antibodies. Cell cultures were exposed to the chosen stress hormone, norepinephrine, at different concentrations for one hour. The metabolic activity of cultured cells was monitored by CCK-8 test, and the concentration of interleukin-6 (IL-6) as a pro-inflammatory cytokine was determined by sandwich ELISA. Based on our results, the metabolic activity of norepinephrine-treated cells was significantly increased compared to the non-treated control cells. The concentration of IL-6 in culture media was also significantly increased after the norepinephrine treatment. Our results are in line with former in vivo studies, where it was found that higher norepinephrine concentration in the blood and urine of the animals affected by the disease was associated with increased urinary IL-6 concentration. Based on this, it is highly expected that elevated production of pro-inflammatory cytokines through the action of norepinephrine plays an important role in the development of the disease. However, for a more accurate understanding of the pathomechanism of FIC, further research is needed, and this cell culture can serve as a good model for studying the molecular background of the disease.