Pathways-based analysis will be done in collaboration with P15 (Cavalieri) to identify the intracellular target of Japanin and data will become available from this study.

Data was obtained on RNA transfected DC which proved superior in priming naïve T cells in vitro compared to DC loaded with native peptides.

We have been following up on the effort to generate proteomics map data of lysosomes purified from non-activated and activated mouse DCs. The decision to focus on the most important organelle for DC activity has been driven from the need to describe in greater detail the proteins involved in the DC response both qualitatively and quantitatively. The partner has been trying to quantify the relative amount of 400 proteins. They have also indexed better this list and are currently comparing it to a list of proteins present in macrophages lysosomes (in collaboration ...

Data is available from a re-analysis of existing microarray data; we started experimental work to assess the reliability and completeness of the DC_Eu.Gene pathway set using stimulation of DCs with S.cerevisiae cells, pseudohyphae, spores in comparison with the results obtained with LPS, R848, LPS and R848, Zymosan and Curdlan.

The importance of alveolar macrophages and more recently DC in the immune response to Mycobacterium tuberculosis (MTb) has been well documented, but the relative contribution of plasmacytoid DC during bacterial infections is largely unknown. Thus, data was obtained on the contribution of pDC in acute MTb infection. We examined whether plasmacytoid pDC are infected and/or activated by MTb, and whether they play a role in regulating bacterial clearance during acute MTb infection and if so how. We have compared these DC with myeloid DC and macrophages with respect t...

We have microarray data on macrophages and DC from different genetic backgrounds of mice, stimulated with CpG, in the presence or absence of a MEK inhibitor, and further data on the role of certain MAP kinases in the regulation of IL-10 and IL-12, IFN-gamma production. Using pharmacological inhibitors, or macrophages and DC from mice carrying a null mutation in MAP kinase signalling molecule(s), we have evidence for a role of certain MAP kinases in the regulation of IL-10 and IL-12, IFN-gamma production. The work on the role of the MAP kinases, TPL-2 and ERK in u...

We investigated and obtained data on the interactions between human monocyte derived DCs (MDDC), generated in the presence of GM-CSF and IL-4 (IL-4 DC), and antigen-stimulated circulating gamma delta T lymphocytes, bearing the Vgamma2 TCR. Co-cultures of IFN-DCs with zoledronate-stimulated gamma delta T lymphocytes were also performed. gamma delta T-cell activation, as demonstrated by their up-modulation of activation marker expression levels (e.g. CD25 and CD69), was observed. These studies demonstrated for the first time the existence of a bidirectional activatin...

Data on the role of STAT3 signalling in directing DC-mediated responses toward immunogenicity or tolerance will be generated. To this purpose, we will identify (i) genes and (ii) micro-RNA selectively controlled by STAT3 in human MDDCs treated with proinflammatory or tolerogenic stimuli (including TLR ligands, cytokines and vitamin D3). This will shed light on the role of STAT3 signalling.

We obtained data using specific inhibitors on the role of src-family tyrosine kinases in the maturation of human monocytes derived dendritic cells upon stimulation with several toll like receptor (TLR) agonists. The effect of these kinase inhibitors on the capacity of DC to be activated by a TLR2 (PAM3CSK4), TLR3 (Poly I:C), TLR5 (Flagellin), and TLR8 (Poly U) agonist was evaluated. Src kinases were found to be required for the initiation of some maturation characteristics but not of others, since their inhibition was able to dissociate cytokines and chemokines ...

Among the battery of cell surface receptors expressed by DCs, the receptor for the Fc portion of IgG (FcgammaRs) recognizes the antibodies coupled to their specific antigens, called immune complexes (IC). We have data available from our demonstration that the target of FcgammaRs with the antigen induces the internalization of the IC, leading to the maturation of CD11c+CD11b+ conventional DCs. These DCs become thus efficient to present antigenic peptides to helper CD4 T cells and cytotoxic CD8 T cells via cross-presentation pathway. This capability might then enhan...

Concerning the pathway based analysis and database we have started the curation of the TLR4-CD14 pathway in DCs and we have described the events occurring at the DC membrane level. We have also started global gene expression analyses to better difine the role of NFAT in DCs.

During 2008, we extended all the data regarding TLR3 signaling to TLR8 signaling. Cytokines assays and immunoblotting studies suggest that Src kinases play a crucial role in the control of both MyD88- and TRIF-dependent pathways. We also extended microarray analysis on human MoDC stimulated with R848 (TLR8 agonist), pretreated or not with PP2. In summary, the new generated data on TLR8 triggered gene expression are similar to previous data obtained with TLR3 stimulation, and confirmed that src kinases inhibition is associated to inhibition of key genes in the infl...

We obtained transcriptional and pathway analysis data of human monocyte-derived DCs stimulated with yeast, spheroplasts, pseudohyphae, spores, purified RNA and not purified RNA and LPS. We propose the role of cell wall in the mechanisms of DC-Pathogen interaction as an experiment that could serve as reference in bioinformatics approaches for the reconstruction of DC activation signalling pathways. We have stimulated human Monocyte derived Dendritic Cells (hMoDC) with yeast, spheroplasts, pseudohyphae, spores, purified RNA and not purified RNA, LPS. We have measur...