Fresh peritoneal tissue was obtained directly from the operating room, with informed consent from the patients, and transferred immediately to the laboratory in E199 medium (Biochrom AG, Berlin, Germany). The tissue samples were then incubated for 15 min in phosphate-buffered saline (PBS) containing penicillin/streptomycin and amphotericin B (Sigma-Aldrich Chemie, St. Louis, MO, USA). Extraperitoneal fat was removed, and 7 × 7 mm tissue pieces were inserted between two stainless steel rings, which are available in various sizes, and thus enable large variations in the setting of these experiments. Moreover, stainless steel rings are more stable than plastic rings. In this setup, the ring system allows for tissue orientation with the mesothelial cells pointing upwards, thereby better reflecting the peritoneal cavity in the in vivo setting. The peritoneum was cultured in E199 medium containing penicillin/streptomycin, L-glutamine (Biochrom AG), FCS (Thermo Fisher Scientific), hydrocortisone (Sigma-Aldrich Chemie), fibroblast growth factor (PeproTech GmbH, Hamburg, Germany), and heparin (Biochrom AG), as described previously [12 (link)].
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