Cytokine secretion by in vitro cultures of lung epithelial cells, differentiated macrophages and differentiated dendritic cells incubated with pneumococci and pneumococcal extracellular vesicles.

Streptococcus pneumoniae is an important human pathogen that can colonize the respiratory tract of healthy individuals. The respiratory tract mucosa is thus the first barrier for this pathogen. In this study, we have tested three models of the respiratory epithelium with immune cells: (i) monolayer of A549 human lung epithelial cells, (ii) A549 + macrophages differentiated from the human monocytic THP-1 cell line (dMφ) and (iii) A549 + dMφ + dendritic cells differentiated from THP-1 (dDC) using a two-chamber system. Pneumococcal strains Rx1 (non-encapsulated) and BHN418 (serotype 6B) were incubated with the cells and secretion of IL-6, IL-8, IL-1ß, TNF-α and IL-10 was evaluated. Overall, the models using co-cultures of A549 + dMφ and A549 + dMφ + dDC elicited higher levels of pro-inflammatory cytokines and the non-encapsulated strain elicited an earlier cytokine response. BHN418 pspA (pneumococcal surface protein A) and pspC (pneumococcal surface protein C) knockouts elicited similar cytokine secretion in the co-culture models, whereas BHN18 ply (pneumolysin) knockout induced much lower levels. The results are in accordance with the activation of the inflammasome by Ply. Finally, we evaluated pneumococcal extracellular vesicles (pEVs) in the co-culture models and observed secretion of pro-inflammatory cytokines in the absence of cytotoxicity. Since pEVs are being studied as vaccine candidate against pneumococcal infections, the co-cultures of A549 + dMφ and A549 + dMφ + dDC are simple models that could be used to evaluate pEV vaccine batches.

Suppression of AP-1 constitutive activity interferes with polyomavirus MT antigen transformation ability.

Polyomavirus (Py) encodes a potent oncogene, the middle T antigen (MT), that induces cell transformation by binding to and activating several cytoplasmic proteins which take part in transduction of growth factors-induced mitogenic signal to the nucleus. We have previously reported that the AP-1 transcriptional complex is a target for MT during cell transformation although, its activation was not sufficient for establishment of the transformed phenotype. Here we show that expression of a dominant-negative cJun mutant in MT transformed cell lines inhibits its transformation ability, indicating that constitutive AP-1 activity is necessary for cell transformation mediated by MT. Evidences also suggest that proliferation of MT transformed cells in low serum concentrations and their ability to form colonies in agarose are controlled by distinct mechanisms.