Cell dishes

Experiments were performed with two immortalized human airway epithelial cell lines (16HBE14o^-, S9) and one alveolar cancer cell line (A549). With permission of D.C. Gruenert 16HBE14o^- cells were received from K. Kunzelmann (University of Regensburg, Germany). S9 cells were purchased from ATCC-LGC Standards (Wesel, Germany, S9). A549 cells were obtained from the cell collection of the Friedrich Loeffler-Institute (Riems, Germany).
Cells were cultured on 10 cm Cell+ dishes (3902300, Sarstedt, Numbrecht, Germany) in Eagle’s minimal essential medium (MEM) (P03-2950, PAN-Biotech, Aidenbach, Germany) containing 10% FBS superior (S0615, Merck, Darmstadt, Germany), 29.8 mmol/L NaHCO₃ and 1% (w/v) penicillin/streptomycin solution (P06-07050, PAN-Biotech, Aidenbach, Germany) at 37 °C and gassed with 5% CO₂. Cell culture medium of A549 cells additionally contained 1% L-glutamine solution (P04-80100, PAN-Biotech, Aidenbach, Germany). Cell culture medium was changed every 3 days. Just before cells formed confluent monolayers, they were passaged onto new cell culture dishes or used directly for experiments. All cell cultures were checked for Mycoplasma contaminations on a regular basis.

Experiments were performed using immortalized human airway epithelial cells (16HBE14o⁻) which had been derived from normal bronchial epithelial cells from a healthy donor [73 (link),74 (link)]. Cells were cultured on 10 cm Cell⁺ dishes (Sarstedt, Numbrecht, Germany) in Eagle’s MEM (PAN-Biotech, Aidenbach, Germany) containing 10% FBS superior (Merck, Darmstadt, Germany), 29.8 mmol/L NaHCO₃ and 1% (w/v) penicillin/streptomycin solution (PAN-Biotech, Aidenbach, Germany) at 37 °C and gassed with 5% CO₂. The cell culture medium was changed every 3 days. Cells were exposed to CYN for different periods as soon as they had reached coverage of 80% of the cell culture plate surface. Cells had formed confluent monolayers (Figure 1) by the end of the experiments. All cell cultures were checked for Mycoplasma contaminations regularly.