In vitro investigation of the intercellular cross-talk between opportunistic bacteria and eukaryotic cells
Abstract number: R2247
Bleotu C., Balotescu Chifiriuc M., Iordache C., Dracea O., Bucur M., Larion C., Matei L., Banu O., Cernat R., Lazar V.
Objective: to investigate the potential functional responses of eukariotic mammalian cells to the exposure to Pseudomonas (P.) aeruginosa and Staphylococcus (S.) aureus.
Material and Methods: HeLa cells were grown in 6 multiwell plates in DMEM supplemented with 2% foetal calf serum and 24 h later standardised microbial suspensions were added over the monolayer in an interior filter well. The logarithmic phase microbial cultures supernatants (MLCA) as such/MLCA inactivated at 100 C degrees for 20 minutes/MLCA diluted 1:2 in fresh culture medium were added directly over the HeLa cells monolayer. After 24 h of incubation, cells from the monolayer were harvested, stained with annexin V and propidium iodide and analysed by flow cytometry. The extra-cellular level of IL-1, IL-2, II-4, IL-6, IL-8, IL-10, IL-12, TNF-alpha and IFN-gamma produced by HeLa cells in the presence of bacterial cultures supernatants was assessed by ELISA.
Results: Our studies have indicated that HeLa cells respond to bacterial cultures and their respective supernatants in specific ways. The necrosis rate of HeLa cells at 24 h was significantly increased in the presence of P. aeruginosa culture supernatants, comparatively with the whole bacterial culture, demonstrating that the bacterial soluble factors could accelerate the killing of host cells, probably by inducing a strong pro-inflammatory effect demonstrated by the stimulation of IL-6 production correlated with the inhibition of IL-10. The killing activity was preserved after thermic inactivation. In exchange, in the case of S. aureus, the death rate of HeLa cells was much more increased in the presence of whole bacterial cultures, comparatively with the respective supernatants. The decreased necrosis rate is sustained also by the anti-inflammatory effect produced by the soluble staphylococcal antigens and demonstrated by the increased level of IL-10. However, in both bacterial infection models, the HeLa cells necrosis rate was significantly reduced after decreasing the soluble mediators concentrations by adding fresh medium.
Conclusion: Our results demonstrate that eukaryotic cells are able to detect bacterial chemical signals and to respond appropriately. The understanding of the mechanism of crosstalk between the bacterial products and host cells may contribute to the elaboration of an efficient strategy for controlling the severity of tissue damages and autoimmune diseases, associated or consecutive to bacterial infections.
|Session name:||18th European Congress of Clinical Microbiology and Infectious Diseases|
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