These constructs were used to transfect immature DCs.

We have prepared several new constructs for the expression of (modified) adhesion receptors in DC.

We have prepared several new constructs for the expression of c-type lectin receptors in DC.

We found that LPS-stimulated mDCs are able to induce up-regulation of co-stimulatory molecules on co-cultured pDCs. Likewise, CpG-stimulated pDCs activate co-cultured mDCs. The cross talk between these two populations of DC is very efficient since it can be observed at mDC/pDC ratio ranging from 1/10 to 10/1.

Monocytes were cultured in RPMI 1640 medium (GIBCO BRL) supplemented with 2 mM L-glutamine (Sigma), 1% (vol/vol) non-essential amino acids, 100 mM sodium pyruvate, 50 U/ml of penicillin and 50 mg/ml of streptomycin (Gibco BRL) containing 10% (vol/vol) FCS (Hyclone).

100 micrograms/ml curdlan (Wako) was used (along with R848, LPS, yeast RNA and yeast cells in different conditions of culture) to induce DC activation.

Using curdlan as a specific agonist of dectin-1, we have shown that this C-type lectin couple to Syk kinase leads to activation of ERK, JNK and p38 MAPKs, as well as NF-kappaB.

CX3CR1 GFP mice, in which bmDC are green fluorescent (GFP), were analyzed according to their bmDC localization. It was found that the cells were organized in unique clusters, mainly located in the endosteum of the BM. These DC co-localize with B cells and these clusters are occupying architecturally definable niches and are reminiscent to the niches that were found in the cranial BM parenchyma.

These HIV strains were used to infect Human Hemato-Lymphoid System Rag2gc-/- mice to closely resemble HIV infection in humans.

DCs were exposed to cytochalasin D (10 microg/ml, TebuBio) for 30 minutes at 4°C.

This cytokine cocktail was used to mature human monocyte-derived dendritic cells, on which subsequently, transcriptional profiling was performed.

The cDNA’s encoding these early expressed HIV antigens have been human codon optimized and modified with lysosomal targeting sequences.

These animals were used to test the combined effects of these cytokines on DC development in vivo.

The DC were matured with inflammatory cytokines followed by electroporation with TAA encoding mRNA. They were used to compare different maturation methods.

Cytotoxic T cells proliferation was observed in human Hemato-Lymphoid System Rag2-/-gc-/- mice after being infected with EBV and mounting an immune response.

We decided to analyze the protein changes that occur at the cell surface of a maturing DC, induced by a prototypical maturation stimulus such as LPS. Results obtained from the LC-MS/MS analysis of the integral plasma membrane proteome of D1 cells stimulated for 24 hours with LPS have been compared to the results obtained from immature D1 cells.

D1 cells will be stimulated with LPS and harvested at different time-points for GeneChip analysis and for tandem mass spectrometry-based analysis of the integral plasma membrane proteome.

The kinetic of the transcriptional profiling of D1 cells treated with either DMSO or LPS was analysed.

The kinetic of the transcriptional profiling of D1 cells treated with either DMSO or LPS was analysed.

These cells were stained by Mini67-1KT or Mini26-1KT (PKH67/PKH26 green/red fluorescent cell linker mini kit for general cell membrane labelling, from Sigma) in order to analyse their uptake of dead cells.