Host cell invasion is a major feature of Staphylococcus aureus and contributes to infection development. The intracellular metabolically active bacteria can induce host cell activation and death but they can also persist for long time periods. In this study a comparative analysis was performed of different well-characterized S. aureus strains in their interaction with a variety of host cell types. Staphylococcus aureus (strains 6850, USA300, LS1, SH1000, Cowan1) invasion was compared in different human cell types (epithelial and endothelial cells, keratinocytes, fibroblasts, osteoblasts). The number of intracellular bacteria was determined, cell inflammation was investigated, as well as cell death and phagosomal escape of bacteria. To explain strain-dependent differences in the secretome, a proteomic approach was used. Barrier cells took up high amounts of bacteria and were killed by aggressive strains. These strains expressed high levels of toxins, and possessed the ability to escape from phagolysosomes. Osteoblasts and keratinocytes ingested less bacteria, and were not killed, even though the primary osteoblasts were strongly activated by S. aureus. In all cell types S. aureus was able to persist. Strong differences in uptake, cytotoxicity, and inflammatory response were observed between primary cells and their corresponding cell lines, demonstrating that cell lines reflect only partially the functions and physiology of primary cells. This study provides a contribution for a better understanding of the pathomechanisms of S. aureus infections. The proteomic data provide important basic knowledge on strains commonly used in the analysis of S. aureus-host cell interaction.

Early metastasis leads to poor prognosis of lung cancer patients, whose 5-year survival rate is only 15%. We could recently show that the Ca2+ sensitive K+ channel KCa3.1 promotes aggressive behavior of non-small cell lung cancer (NSCLC) cells and that it can serve as a prognostic marker in NSCLC. Since NSCLC patients die of metastases, we investigated whether KCa3.1 channels contribute to poor patient prognosis by regulating distinct steps of the metastatic cascade. We investigated the extravasation of NSCLC cells and focused on their adhesion to endothelial cells and on transendothelial migration. We quantified the adhesion forces between NSCLC cells and endothelial cells by applying single cell force spectroscopy, and we monitored transendothelial migration using live-cell imaging. Inhibition of KCa3.1 channels with senicapoc or KCa3.1 silencing increases the adhesion force of A549 lung cancer cells to human microvascular endothelial cells (HMEC-1). Western blotting, immunofluorescence staining and biotinylation assays indicate that the elevated adhesion force is due to increased expression of ICAM-1 in both cell lines when KCa3.1 channels are downregulated. Consistent with this interpretation, an anti-ICAM-1 blocking antibody abolishes the KCa3.1-dependent increase in adhesion. Senicapoc inhibits transendothelial migration of A549 cells by 50%. Selectively silencing KCa3.1 channels in either NSCLC or endothelial cells reveals that transendothelial migration depends predominantly on endothelial KCa3.1 channels.In conclusion, our findings disclose a novel function of KCa3.1 channels in cancer. KCa3.1 channels regulate ICAM-1 dependent cell-cell adhesion between endothelial and cancer cells that affects the transmigration step of the metastatic cascade.

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