Bladder epithelial cell studies

Human bladder epithelial cells, designated 5637 (ATCC HTB-9) cells, were obtained from the American Type Culture Collection and maintained in RPMI 1640 supplemented with heat-inactivated 10% (v/v) fetal bovine serum at 37°C in the presence of 5% CO₂. Confluent, serum-starved 5637 cells in 24-well plates were infected with UTI89 strains at a multiplicity of infection of 10. After 30 min, culture media were replaced either by fresh culture media or by media with gentamicin sulfate (120 μg/ml) (Sigma-Aldrich) to kill extracellular bacteria. Cells were further incubated for 1 hour, washed rigorously, solubilized with 1% Triton X-100, quenched with PBS, and plated for bacterial CFU quantification.